Aging, Immunity, and Infection (Infectious Disease)

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Aging, Immunity, and Infection JOSEPH F. A LBRIGHT JULIA W. A LBRIGHT

Aging, Immunity, and Infection

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n f e c t i o u s . D i s e a s e SERIES EDITOR: Vassil St. Georgiev National Institute of Allergy and Infectious Diseases National Institutes of Health

Aging, Immunity, and Infection, by Joseph F. Albright and Julia W. Albright, 2003 Handbook of Cytokines and Chemokines in Infectious Diseases, edited by Malak Kotb, PhD and Thierry Calandra, MD, PhD, 2003 Opportunistic Infections: Treatment and Prophylaxis, Vassil St. Georgiev, PhD, 2003 Innate Immunity, edited by R. Alan B. Ezekowitz, MBChB, DPhil, FAAP and Jules A. Hoffmann, PhD, 2003 Pathogen Genomics: Impact on Human Health, edited by Karen Joy Shaw, PhD, 2002 Immunotherapy for Infectious Diseases, edited by Jeffrey M. Jacobson, MD, 2002 Retroviral Immunology: Immune Response and Restoration, edited by Giuseppe Pantaleo, MD and Bruce D. Walker, MD, 2001 Antimalarial Chemotherapy: Mechanisms of Action, Resistance, and New Directions in Drug Discovery, edited by Philip J. Rosenthal, MD, 2001 Drug Interactions in Infectious Diseases, edited by Stephen C. Piscitelli, PharmD and Keith A. Rodvold, PharmD, 2001 Management of Antimicrobials in Infectious Diseases: Impact of Antibiotic Resistance, edited by Arch G. Mainous III, PhD and Claire Pomeroy, MD, 2001 Infectious Disease in the Aging: A Clinical Handbook, edited by Thomas T. Yoshikawa, MD and Dean C. Norman, MD, 2001 Infectious Causes of Cancer: Targets for Intervention, edited by James J. Goedert, MD, 2000

Infectious.Disease

Aging, Immunity, and Infection By

Joseph F. Albright, PhD and

Julia W. Albright, PhD George Washington University School of Medicine, Washington, DC

Humana Press

Totowa, New Jersey

© 2003 Humana Press Inc. 999 Riverview Drive, Suite 208 Totowa, New Jersey 07512 humanapress.com All rights reserved. No part of this book may be reproduced, stored in a retrieval system, or transmitted in any form or by any means, electronic, mechanical, photocopying, microfilming, recording, or otherwise without written permission from the Publisher. Due diligence has been taken by the publishers, editors, and authors of this book to assure the accuracy of the information published and to describe generally accepted practices. The contributors herein have carefully checked to ensure that the drug selections and dosages set forth in this text are accurate and in accord with the standards accepted at the time of publication. Notwithstanding, as new research, changes in government regulations, and knowledge from clinical experience relating to drug therapy and drug reactions constantly occurs, the reader is advised to check the product information provided by the manufacturer of each drug for any change in dosages or for additional warnings and contraindications. This is of utmost importance when the recommended drug herein is a new or infrequently used drug. It is the responsibility of the treating physician to determine dosages and treatment strategies for individual patients. Further it is the responsibility of the health care provider to ascertain the Food and Drug Administration status of each drug or device used in their clinical practice. The publisher, editors, and authors are not responsible for errors or omissions or for any consequences from the application of the information presented in this book and make no warranty, express or implied, with respect to the contents in this publication. Production Editor: Mark J. Breaugh. Cover design by Patricia F. Cleary.

This publication is printed on acid-free paper. ∞ ANSI Z39.48-1984 (American National Standards Institute) Permanence of Paper for Printed Library Materials. For additional copies, pricing for bulk purchases and/or information about other Humana titles, contact Humana at the above address or any of the following numbers: Tel: 973-256-1699; Fax: 973-256-8341; E-mail: [email protected], or visit our website at www.humanapress.com Photocopy Authorization Policy: Authorization to photocopy items for internal or personal use, or the internal or personal use of specific clients, is granted by Humana Press Inc., provided that the base fee of US $20.00 per copy is paid directly to the Copyright Clearance Center at 222 Rosewood Drive, Danvers, MA 01923. For those organizations that have been granted a photocopy license from the CCC, a separate system of payment has been arranged and is acceptable to Humana Press Inc. The fee code for users of the Transactional Reporting Service is: [0-89603-644-8/03 $20.00]. Printed in the United States of America. 10 9 8 7 6 5 4 3 2 1 Library of Congress Cataloging-in-Publication Data Albright, Joseph F. Aging, immunity, and infection / authored by Joseph F. Albright and Julia W. Albright. p. ; cm. -- (Infectious disease) Includes bibliographical references and index. ISBN 0-89603-644-8 (alk. paper) e-ISBN 1-59259-402-6 1. Developmental immunology. 2. Aged. 3. Immunosuppression--Age factors. 4. Natural immunity. 5. Infection--Age factors. I. Albright, Julia W. II. Title. III. Infectious disease (Totowa, N.J.) [DNLM: 1. Immunity--Aged. 2. Aging--physiology. 3. Infection--physiopathology--Aged. QW 540 A342a 2003] QR 184.5.A43 2003 616.07'9--dc21 2002191941

Dedication We are deeply grateful to our mentors, Takashi (Mak) Makinodan and the late James D. (Jim) Ebert who introduced us to the satisfactions and occasional frustrations of biological research. "The only way to cross this Malebolge—and without Vergil as your guide—is to tell yourself that what was is; that once young, always young, once beautiful, always beautiful; once bright, always bright; that what lived cannot die." Erwin Chargaff Heraclitean Fire

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Preface The preparation of Aging, Immunity, and Infection has been a "labor of labor." When we began, there existed a huge literature—but manageable, we thought, given our years of experience in the area often referred to as immunogerontology. However, in the time that we have been at work, the new relevant literature has increased at a prodigious rate. The more we read and tried to assimilate, the farther we fell behind. In order to have any hope of completing a book on this rapidly evolving topic, we have been forced to become increasingly selective in covering new and recent publications. We dare to hope that many readers will find the book useful and only a few will dwell on the inevitable inadequacies. We consider the book a work in progress, and welcome suggestions for future editions. Five chapters cover several aspects of infection and the decline of immunity with age. The first chapter "Human Aging: Present and Future," is devoted to demographics and theories of senescence. Chapter 2 outlines the gradual breakdown of resistance to infection in the aged individual. Chapters 3 and 4 cover changes in innate and acquired immunity. The final chapter, "Nutrition, Longevity, and Integrity of the Immune System," discusses such provocative ideas as life-span extension and nutritional intervention for the delay of immunosenescence. We acknowledge with gratitude the outstanding staff of the National Cancer Institute Scientific Library at Frederick, Maryland for maintaining a first-rate library where nearly everything is available and easy to locate.

Joseph F. Albright, PhD Julia W. Albright, PhD

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Contents Dedication ............................................................................................... v Preface .................................................................................................. vii 1 Human Aging: Present and Future .....................................................1 Demographics ...................................................................................... 1 Infectious Diseases of the Aging ......................................................... 5 Limits on Life Expectancy and Future Prospects .............................. 7 Theories of Senescence ....................................................................... 11 Chapter Summary .............................................................................. 14 References ........................................................................................... 15 2 Aging and Altered Resistance to Infection ..................................... 19 Relatively Common Bacterial Infections of Aging Humans ........... 20 Selected Examples of Age-Associated Susceptibility to Bacterial Infections .................................................................... 24 Bacterial Interactions with Mucosal Surfaces.................................. 28 Antibiotic Resistance and Bacterial Variation ................................. 39 Viral Infections in Aging Humans ................................................... 42 Protozoan Parasites in Aging Subjects ............................................ 47 Fungal Infections in Aging Subjects ................................................ 50 Chapter Summary .............................................................................. 51 References ........................................................................................... 53 3 Senescence of Natural/Innate Resistance to Infection ................. 61 Pattern Recognizing Receptors of Innate Immunity ....................... 62 Phagocytic Cells: Monocytes/Macrophages ..................................... 72 Microbial Evasion of Phagocytic Destruction ................................. 80 Age-Related Changes in Macrophages ............................................. 81 Phagocytic Cells: Neutrophils ........................................................... 96 Natural Killer/Lymphokine-Activated Killer Cells ........................ 105 Chapter Summary ............................................................................ 115 References ......................................................................................... 117 4 Aging of Adaptive/Acquired Immunity .......................................135 Aging of the Thymus and Thymus-Derived (T) Cells ................... 136 The Functions and Diversity of Peripheral T Cells ....................... 145 Summary: Known and Cognizable Effects of Aging T Cells ......... 172 Differentiation, Functions, and Aging of B Cells .......................... 183 ix

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Contents Chapter Summary ............................................................................ 195 References ......................................................................................... 197 5 Nutrition, Longevity, and Integrity of the Immune System .................................................................213 RCI-Mediated Delay of Immunosenescence ................................... 214 How Does RCI Promote Life-Span Extension? ............................. 217 Dietary Restriction vs Malnutrition .............................................. 218 References ......................................................................................... 221 Epilogue ...............................................................................................225 Index .................................................................................................... 233

Human Aging: Present and Future

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1 Human Aging Present and Future Then the Lord said, “My spirit will not contend with man forever, for he is mortal; his days will be a hundred and twenty years.” —Genesis 6:3

DEMOGRAPHICS Of all the potential disasters and scourges that threaten mankind—famine, nuclear war, collision of Earth with meteoroids, and many others—none seems as impending as the aging human population. According to data and projections released recently by the Population Division of the United Nations’ Department of Economic and Social Affairs (1), the world population in 1998 was 5.9 billion. By the time this book is published, it will be well over 6 billion. The projected, “most likely” estimate for the year 2050 is 8.9 billion. It took 12 years, from 1987 to 1999, to add 1 billion people to the world’s population. In another 50 years, 3 billion more persons will be alive. What should be considered startling, indeed alarming, are the expected increases in the aging and aged segments of the population. In 1998, 66 million persons in the world were over 80 years of age (Table 1-1). That number is projected to rise almost sixfold, to 370 million persons by the year 2050. The number of centenarians will reach 2.2 million by 2050, a 16-fold increase over the number in 1998. The life expectancy of human beings has been increasing dramatically all around the world and will continue to increase in the years ahead. Table 1-2 provides data for a few “developed” countries that show the increase in life expectancy of men and women over the 25-year period from 1965 to 1990. For reasons that remain obscure, females have lived longer than males in every era From: Aging, Immunity, and Infection By J. F. Albright and J. W. Albright © Humana Press Inc., Totowa, NJ

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Aging, Immunity, and Infection Table 1-1 Present and Projected Oldest Humans Worldwide Population in millions Group

Ages

1998

2050

Octogenarian Nonagenarian Centenarian

80–89 90–99 100+

59 7 0.1

311 57 2

Data from ref. 1.

Table 1-2 Increase in Life Expectancy over the Period 1965 to 1990 Life expectancy (years) Men Country England/Wales France Japan Sweden United States

Women

1965

1990

1965

1990

68 67 67 72 67

73 72 76 75 72

74 75 73 76 73

78 80 82 81 78

Data from ref. 1.

of history. The increase that occurred in Japan during that 25 years was profound and Japan now leads the world in life expectancy at birth (2). A similar demographic shift has been occurring in the less-developed regions around the world. In those regions, the life expectancy rose from around age 40 in the early 1950s to an average of about 62 in 1990 (3). The worldwide aging of the population is the consequence of two contributing phenomena: a) the increase in life expectancy at birth, and b) the declining rate of new births. That is illustrated by the data presented in Figure 1-1. It is projected, as the figure shows, that 50 years hence the population in regions now classed as “less-developed” will comprise equal numbers of persons aged 60 and over, and aged 14 or younger. In the regions now considered developed, more than 30% of the population will be age 60 and over and approx 15% will be 14 and under. The relatively rapid shift in the demographics of the world’s population that will occur in the next half century presents formidable challenges in many respects. One major challenge, of course, is to provide the aged and aging with


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Fig. 1-1. Projected percentage of world’s population, aged 14 and younger or aged 60 and over, who will reside in regions considered “developed” or “underdeveloped.” Developed: ≥60 (䊉), ≤14 (䊊). Underdeveloped: ≥60 (䉱), ≤14 (䉭). Redrawn from data in ref. (1).

proper health care; and the attendant challenge, which is outside the scope of this book (fortunately, for us!), of how to pay for that health care. To more firmly grasp the magnitude of the challenges, consider the statement, “each month, the world sees a net gain of 800,000 people over 65, 70% of whom are in the developing world” (3). From the perspective of health care, it is the population aged 65 and over who have, far and away, the greatest needs. The greater the age, the greater the needs. To further emphasize the challenge, look again at Table 1-1, which provides a summary of the anticipated growth in the “oldestold” segment of the world’s population in the next 50 years. Future Health and Research The social and economic problems associated with the aging population in the developed nations are clearly illustrated by the current situation in Japan (reviewed by Oshima, ref. 2). The problems facing the, as yet, underdeveloped regions of the world are clearly and succinctly presented by Holden (3). In reviewing the situation in the United States, Schneider (4) has presented a compelling case for vigorous programs in aging research, leading to significant advances in preventing and treating the diseases of the elderly, as the

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Aging, Immunity, and Infection Table 1-3 Projections of the Older Population of the United States: Years 2000–2050 Number of persons (in thousands) aged Year

60–64

65–74

75–84

85–100

>100

2000 2050

45,363 99,459

34,709 78,859

16,574 44,127

4,259 18,223

72 834

Data from refs. 5 and 6.

key to coping with the impending crisis in health care for the older population. The dimensions of the problem for the United States are summarized in Table 1-3 using projections by the US Bureau of the Census in 1996 (5,6). The data presented in Table 1-3 leave little doubt that the entitlement program, Medicare, “will be stressed by the large numbers of eligible older Americans” (4). Schneider argues that through strong, adequately supported programs of research aimed at better understanding aging and the prevention and treatment of diseases of the elderly, the likely result would be that “the average health of a future 85-year-old in the year 2040 resembles that of a current 70-year-old with relatively modest needs for acute and long-term care.” Indeed, research leading to effective means to retard and prevent the debilitating effects of aging, which are neither too complex nor costly, may be the only hope for enabling much of the world’s population to age with dignity and relative independence. Research leading to good sanitation, good nutrition, and the control of communicable diseases has led to the phenomenal increase in life expectancy and is largely responsible for getting us into the present dilemna. It seems, therefore, paradoxical to assert that more research represents the best hope for the way out. The fact is, however, that the research that led to extension of life expectancy was not concerned with understanding aging. Research to elucidate the causes and possible moderation of aging is relatively new on the biomedical scene but already has made considerable progress. In this book, we consider an important outcome of aging, viz., the heightened susceptibility to infections, and explore the underlying causes, the consequences and the prevention of infectious diseases in the elderly. Infectious diseases remain an important cause of the morbidity and mortality of aging humans especially in the developing nations. Advances in the ability to cure and prevent those diseases will greatly improve the health and independence of the aging population and decrease the expenditures for health care.


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Table 1-4 Rates of Death From the 10 Leading Causes, United States, 1996 Rank

Cause of death

Deaths/100,000 population

Ages 45–64, both sexes, all races

1 2 3 4 5 6 7 8 9 10

All causes

703.6

Malignant neoplasms Heart diseases Accidents (motor vehicle & other) Cerebrovascular diseases Chronic obstructive pulmonary disease Diabetes mellitus Chronic liver disease & cirrhosis HIV infection Suicide Pneumonia and influenza

244.7 190.5 31.1 28.8 23.9 23.6 20.0 15.0 14.4 10.6

Ages >65 years, both sexes, all races 1 2 3 4 5 6 7 8 9 10

All causes Heart diseases Malignant neoplasms Cerebrovascular diseases Chronic obstructive pulmonary disease Pneumonia and influenza Diabetes mellitus Accidents (motor vehicle & other) Alzheimer’s disease Kidney diseases Septicemia

5061.1 1808.0 1131.1 414.9 270.1 221.4 137.0 91.0 62.2 61.6 51.2

From Communicable Diseases Center. Vital Statistics Report 1998;47(9):26–36.

INFECTIOUS DISEASES OF THE AGING Table 1-4 presents the 10 leading causes of death in the United States among humans aged 45–64 and over 65 years of age. Pneumonia and influenza are listed as 10th in the 45–64 age group and septicemia does not appear on the list. On the list of the over-65 age group, pneumonia and influenza appear as the 5th leading cause and septicemia is 10th. Among persons over 65, pneumonia and influenza and septicemia are responsible for the deaths of approximately three persons per thousand in the United States.

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Fig. 1-2. Increase in numbers of patients over age 65 diagnosed as having an infectious disorder; based on US hospital discharges over the decade 1980 (䊐) to 1990 (䊐).

Morbidity resulting from infections among US residents over age 65 is illustrated in Fig. 1-2 where the index is the rate of discharge from hospitals of patients who had been treated for one of the three infections shown. The rates are shown for the years 1980 and 1990. Notice that the rate was almost 4 times greater in 1990 than 1980 for septicemia, 2 times greater for urinary tract infection, and approx 1.5 times greater in the case of pneumonia. Those increases reflected more frequent contacts between elderly patients and physicians, more aggressive treatment regimens, and an increasing proportion of the older-old among the over-65 population. As Fig. 1-2 shows, in 1980 the rate of hospitalization of persons over 65 for three leading infections was about 745 per 10,000 elderly persons or 7.5/100. In 1990, the rate was about 1365 per 10,000 elderly persons or about 13.7/100. Thus, it is abundantly clear that if there are no new or improved programs for preventing and managing those infections among the US population over 65 years of age, the toll of suffering and the drain on medical resources will be enormous in the years ahead. To illustrate: if nothing changes in the next 50


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years except the increase in number of person over 65, in the year 2050 there will be 20 million elderly persons hospitalized for pneumonia, septicemia, and/ or urinary tract infections. That, of course, is a serious underestimate because as Table 1-4 shows, the greatest increase will occur in the oldest-old age groups in which the rate of hospitalization increases dramatically. The preceeding discussion of infections among the elderly in the United States is approximately true of other developed regions of the world. It is far from representative of the less-well-developed regions. In many of the underdeveloped regions, infections such as influenza and pneumonia are secondary to much more formidable scourges. The leading killers in most of the world are malaria, tuberculosis, leishmaniasis, and a host of diseases caused by enteric pathogens. The impact of those pathogens, which are so prevalent and to which the elderly are inordinately susceptible (discussed more fully later), on the health of emerging elderly populations is impossible to foresee. This monograph is intended to be a review of current knowledge about the susceptibility of the elderly to infections in relation to the immune and allied systems that decline in competence associated with aging. In order to deal effectively with that broad subject, it is necessary to include information drawn from the fields of nutrition, biochemistry and molecular biology, cellular and systems physiology, and others. LIMITS ON LIFE EXPECTANCY AND FUTURE PROSPECTS Because we do not understand the mechanisms of biological aging or the reasons for aging, estimating the limits of human life expectancy is highly empirical. There is a strong evolutionary and genetic influence on life expectancy (7–9). The forces of natural selection decrease with advancing age because, in natural populations, few individuals survive past the reproductive ages. Therefore, among the survivors, random mutations (alleles) will accumulate and their detrimental effects will be expressed after reproductive activity has ceased. In recognition of those ideas, Williams (10) proposed an “antagonistic pleiotropy” hypothesis, which suggests that disadvantageous genes in a population will not be selected against if they arise after the reproductive phase that is required to maintain the population. A related theory is that of the “disposable soma” theory (11), the concepts of which were summarized by Holliday (12) as follows: The environment is hostile, and individuals are competing for natural resources. This competition results in the natural selection of the fittest. In these circumstances the probability of an organism surviving and reproducing for a long period becomes very small, so potential immortality confers very little, if any, adaptive advantage. In other words, such organisms are not necessarily the fittest because resources are used to maintain the soma for a long period of time. It

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Aging, Immunity, and Infection is a better strategy for the survival of an organism’s lineage to invest resources into growth to adulthood and reproduction, rather than in long-term maintenance of the soma. Thus, the organism that evolves a soma with a limited survival time is at an advantage over one that attempts to maintain the soma indefinitely. This disposable soma theory neatly explains the early origins of aging in animals.

Finch (9) has reviewed the relationships between life-span and reproductive ability, and between life-span and hostility of the environment for a number and animal and plant species. He cites examples of organisms such as benthic fishes and bristlecone pine trees that live for hundreds or even several thousand years. In such cases, it appears that the environments to which such organisms have adapted are not threatening and that reproductive activity of those organisms is quite prolonged. In recognition of this important relationship between environment and life-span, Finch has written (9): “one may consider that the recent expansion of human life-spans parallels that of bristlecones at high altitude, and may be due, in our case, to improvements of hygiene and nutrition that adventitiously favored greater life spans.” That brings us to the questions: What is the current practical limit of human longevity? Can it be extended? For many years, the Gompertz function, or “Gompertz hazard function” (14), first formulated by Benjamin Gompertz in 1825, has seemed to describe best the relationship between human age and mortality. This relationship shows the increasing probability of death with the increasing age of a population (e.g., ref. 15). Over the range of age from approx 45 to 85, the rate of death increases steadily and can be represented graphically by a straight line. As is discussed later, data gathered in the last three decades indicate that the death rate in the oldest segment of the population (beyond age 80, approximately) has slowed considerably. As a consequence the Gompertz relationship needs to be revised and new models developed; this has become apparent as a result of the increasing human survivorship beyond age 85 (14). The development of new models is a complex undertaking owing in part to the dearth of data extending over a protracted time from which to extract factors for relative risk of mortality at given ages and how those risks may vary among different elements (cohorts) of the population. The complexity that a satisfactory model must assume is illustrated by the following (14): The model must describe the effects on mortality of internal mechanisms of physiological change with age operating under genetic constraints. It must also show how genetically constrained processes evolve with age as a result of the stochastic impact of environmental shocks, and how the operation of physiological mechanisms evolve to respond to and modify the organism’s internal environment because of those shocks.

The Gompertz formulation cannot accommodate the leveling out of the relationship between mortality and aging that has become evident in the human


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population in recent years. Data obtained from the US population over the last 25 years have shown that the major change contributing to extended life expectancy has been in the segment of the population over 50 years of age. It has been stated (16) that: “Most of the declines in mortality and gains in life expectancy during this recent mortality transition were achieved in the elderly population—a phenomenon so unexpected and unexplained that it has been referred to as a new stage in the epidemiologic history of developed nations.” Is there, at present, a realistic estimate of the mean human life expectancy? If so, how is it obtained? Several methods (reviewed in ref. 16) have provided estimates in the range of 85 to 99 years. The US Social Security Administration has forecast life expectancy in the year 2050: for women, the figure is 82.9 years, for men, 77.5 years. These figures are based on extrapolation of the data accumulated over the last quarter century (approximately). The advantages and pitfalls of projections based on extrapolation have been reviewed (17). The most reliable estimates place life expectancy at birth at around age 85. Only recently has the steady trend toward increasing life-expectancy beyond age 50 shown any indication that life-span is approaching an upper limit (16,17). As noted in ref. 16 very recent data suggest that “a biological limit to life is operating.” That suggestion comes from evidence of “mortality compression.” The latter refers to a change in shape of survival curves toward rectangularization. Figure 1-3 is a hypothetical example of a survival curve beyond age 50 as it might look today for a population with a life expectancy at birth of 80 years compared with a curve for a population in the year 2050 having a life expectancy at birth of 100 years. The second curve shows evidence of mortality compression. The rectangularization results from prolonging the phase of the curve in which few deaths occur followed by a relatively short phase in which the rate of mortality is high and the number of survivors declines precipitately. The rectangularization of the survival curve suggests that the life expectancy of the population may be approaching an upper limit. Retangularization also means that a proportion of the population will live somewhat longer than is the case when it does not occur. The combined trends of longer life expectancy at birth and longer life expectancy beyond age 50 argue strongly that the risk of death in later years has been declining over the last 20–30 years. This might be explained by a version of the antagonistic pleiotropy theorem based on the supportable conclusion that the random accumulation of mutations is not necessarily deleterious but relatively neutral. Mutations that are potentially deleterious may remain latent until some extrinsic (environmental) or intrinsic (pathogenic) event triggers their expression. Changes in any number of factors including better nutrition, reduced incidence and severity of infections, reduced exposure to certain xenobiotics (e.g., carcinogens) and others, singly or in combination, could result in improved environments for the aging population.

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Fig. 1-3. Rectangularization of a survival curve and illustration of mortality compression. Curve 1 reflects mortality from various causes such as infectious diseases, cardiovascular diseases, cancers, etc. Curve 2 indicates that aging has become a leading cause of mortality because deaths from other major causes have been reduced or eliminated.

What are the prospects for further prolongation of life expectancy? If there exists an upper limit, is it possible to extend that limit? If it could be extended, should it be? Consider, first, the consequences on life expectancy of eliminating some of the major diseases of the aged. Working with data gathered from the US population and made available by government agencies in 1985, Olshansky and associates (16) estimated the impact on life expectancy were it possible to eradicate only cancer, only ischemic heart disease, both cancer and ischemic heart disease, or all cancer, cardiovascular disease, and diabetes. The results were quite informative. The data provided in 1985 indicated life expectancy at birth for males of 71.2 years, and for females of 78.3 years. Elimination of all forms of cancer would have increased life expectancy by approx 3.2 years for both males and females. Elimination of both cancer and ischemic heart disease would have raised life expectancy by 8.1 years for males and 7.0 years for females. Elimination of all cardiovascular diseases, cancer, and diabetes would have increased life expectancy at birth by about 15.3 years for males and 15.8 years for females. Thus, elimination of those


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major diseases (responsible for 71.3% of all deaths in 1985) would have raised the average life expectancy to 86.5 years for males and 94.1 years for females. The cohort of the population aged 50 in 1985 was estimated to survive on average for another 25.5 years (males) or 30.9 years (females). Elimination of those major diseases would have raised the average remaining life expectancy by 15.1 years for males and 15.3 years for females. Thus, the best available analyses of the impact of eliminating the major diseases from the human population (16) provide little hope for extending life expectancies beyond 90–95 years of age. Major modification in the genetic and/or physiological regulation of longevity of the human probably will be required to extend life expectancy beyond age 100. There is now ample evidence that less complex forms of life—nematodes, fruit flies, medflies, yeast, and even rodents—are subject to considerable extension of longevity by genetic and physiological modifications. Those are discussed in later sections. Although the subject is outside the scope of this book, it is nevertheless appropriate to wonder whether or not it would be wise to prolong the human life-span beyond that which could be achieved by eliminating all major human diseases. Even that endeavor may not be so wise. As pointed out by Olshansky and associates (16): If improvements in risk factors for fatal degenerative diseases are responsible for the observed declines in old age mortality, then mortality and disability may exhibit commensurate declines. These declines would occur only if improvements in risk factors have the same effect on postponing the onset of morbidity and disability as they have on postponing mortality. However, advances in medical treatment, more than improvements in risk factors, may be allowing elderly persons who are frail and who suffer from fatal degenerative diseases to survive longer after the onset of the disease than was the case in the past. In this case, age-specific morbidity and disability rates and their duration would increase substantially.

Discussions such as the preceding are highly relevant to the formulation of medical, social, and fiscal plans in the developed regions of the world. They must, however, seem abstract and virtually irrelevant to the underdeveloped regions where the immediate problem is how to provide health and medical care to the increasing elderly population when such care is limited or not available. For much of the world, providing therapy and care to an elderly population suffering from infectious diseases looms as a formidable problem. THEORIES OF SENESCENCE Numerous theories to explain senescence have been promulgated. Each of them offers some promise, at least, for understanding the heightened susceptibility of the older population to infections. Here, we briefly review the more

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substantive theories and later, consider more fully the theories most relevant to infections in the elderly. The theories in their various forms are presented below under four major subheadings. Nutrition and Body Composition To date, the only known practical method of extending longevity of mammals is dietary restriction (DR; ref. 18). This was first recognized in 1935 by the nutritionist, McCay (19). Actually, it is the restriction of caloric intake that results in prolongation of life in model studies with rodents (18,20). Within reasonable limits, the greater the restriction of calories the longer life expectancy is extended (20). Although in most studies DR has been initiated at the time of weaning of juvenile rodents, DR commenced in young adult or midlife ages also results in life extension (20,21). DR results in a significant retardation of the age-associated decline in immunological competence and a significant lowering of the incidence of tumors in rodents, both of which are well correlated with increased longevity (20,22). The reasons offered to explain this apparently beneficial effect of DR are discussed in Chapter 5. The application of DR to extending longevity in humans at present is out of the question. It is far from certain that it would succeed and, if it did, that exaggerated morbidity would not occur. Furthermore, there is considerable uncertainty about the optimum body composition associated with health and longevity. For example, there are unanswered questions about the optimum proportions of lean muscle and fat masses and the rates of change of those components at different physiological and chronological ages (23–25). Free Radical and Oxidative Damage Theory of Aging Reactive radicals of nitrogen (nitric oxide and derivatives such as peroxynitrite) and of oxygen (superoxide anion, hydrogen peroxide, hydroxyl radical) can inflict considerable damage on macromolecules (proteins, nucleic acids, complex lipids), give rise to carcinogens (e.g., nitrosamines), and trigger (or sometimes prevent) apoptotic death of cells such as macrophages and vascular epithelial cells. There are mechanisms for scavenging and antagonizing those highly reactive species of molecules and for repairing damage caused by them. However, unless such mechanisms are absolutely effective, damage inflicted by free radicals may accumulate, even in a self-potentiating or exponential manner. There is evidence that the efficiency of mitochondrial electron transport and energy-generating processes deteriorate with age, resulting in increased appearance of oxidizing free radicals (26,27). Moreover, antioxidant resistance declines with age (28,29). Thus, the free radical and nitric oxide theories of aging are topics of considerable significance and research (30–32).


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Postsynthetic Modifications and Molecular Crosslinking of Proteins Contribute to Aging Following translation, proteins are susceptible to several chemical modifications including oxidation (33), prenylation (34,35), homocysteinylation (36), glycation, and the formation of crosslinks or advanced glycosylated end products (37). It is presumed that the gradual accumulation of altered proteins such as crosslinked collagens, elastins, and other structural proteins will lead to morphological and functional alterations of cells and tissues and the manifestations of senescence. There is compelling evidence of an increasing pool of oxidized, defective enzymes during aging (33) that probably parallels the increase in oxygen free radicals. As Stadtman (33) expressed it: “substantial decreases in the amounts of important enzymes and the accumulation of massive amounts of damaged protein as occurs during aging seriously compromise cellular integrity.” The gradual enlargement of intracellular pools of defective proteins, especially enzymes, could partially explain the well-known senescent decline in reserve functional potential that is characteristics of major organ systems such as the immune system, the kidneys, and the liver. Genes Influence Aging: “Gerontogenes” and “Virtual Gerontogenes” Is there a small number of dedicated genes that control senescence and impose themselves on all other genes at some predetermined rate or express themselves at some predetermined time in the lives of individuals of a species? Is there a single gene, or perhaps a few, that control the average longevity of a species? These are difficult questions to answer at the present time. There are data, however, and a plethora of opinions and interpretations. There is reasonable agreement with the conclusion that senescence and life expectancy are not controlled in a simple manner by a few genes (38). Even within inbred lines of a given species such as mice or fruit flies there is considerable variation in the apparent rate of aging and life expectancy. Several lines of evidence lead to the conclusion that aging and longevity are controlled in complex fashion by both genes and environmental influences. There are several recent demonstrations of genes that influence longevity. These include genes in Drosophila (38,39), yeast (40), the nematode Caenorhabditis elegans (41–43), and the Mediterranean fruit fly, Ceratitis capitata (44). In related work, investigators have identified genes in human cell lines that are responsible for converting cells considered to have an immortal phenotype into a senescent phenotype (45). One such gene, termed MORF4, appears to encode a transcription factor that regulates expression of several other genes that are involved in the senescent phenotype (45). The term gerontogenes has been applied to genetic factors that regulate aging (46).

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Although it is highly unlikely that a single gene, or even a very few genes, play a determining role in the rate of aging or the duration of life-span, it is possible for a single gene that affects the expression of a panel of other genes to play a considerable role. A particularly informative example is the age-1 gene in C. elegans that is involved in the resistance to various types of stress that affect life-span of the organism (47). Such genes, acting together in closely coordinated fashion, may resemble the action of a single gene and initially appear to be an “aging gene.” The several existing examples of such genes have led to the notion of “virtual gerontogenes” defined as “several genes whose functions are tightly coupled and whose combined action and interaction resemble the effect of one gene” (48). Referring to such genes as “virtual” implies that upon dissection and sequencing the individual component genes will be found to control some precise function concerned most likely with normal maintenance and repair. An immunological theory of aging was proposed (49) at a time when the only system that could be demonstrated to age in quantitative terms was the immune system. Because it was evident that the immune system plays such a central role in protection against infectious and neoplastic diseases, and because it appeared that diseases of the immune system, especially autoimmune, were associated with advanced age, the theory had merit at the time. In precise terms, the immune system plays no causal role in aging. However, as a factor in infuencing length of life in relation to disease it assumes major importance as is shown later. CHAPTER SUMMARY Aging may be viewed as a process that arose early in phylogeny in order to eliminate the competition of postreproductive individuals for limited resources. Because natural selection cannot operate on populations that have passed reproductive activity, those individuals who survived beyond reproductive competence were likely to accumulate random mutations. Although mutations frequently are deleterious, the expression of their adverse effects may require the influence of a hostile environment. Thus, aging may result from a combination of genetic and environmental influences. The remarkable extension of life expectancy in the human population over the last quarter century may reflect, especially, environmental changes coupled with advances in medicine. Recent trends in longevity may suggest that a limit on human life-span is about to be reached. During the next 50 years, there will be a major shift in demographics such that at least a third of the world’s population will require access to medical care. Such care and facilities may be available in the developed areas of the world but the sheer numbers of people who need them will create a heavy


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economic load. In the still-underdeveloped areas of the world, the medical care and facilities are likely to be inadequate and possibly prohibitively expensive. For those reasons, vigorous, adequately supported research focused on understanding the causes of senescence and the pathogenesis of diseases that afflict the elderly is essential. The diseases that are particularly debilitating in the elderly include cancer, cardiovascular and other degenerative disorders, diabetes, and infections. In many areas of the world that are experiencing a rapid increase in an aging population, infectious diseases are the foremost health problem. Those infections include tuberculosis, a variety of bacterial and viral diseases, and numerous parasitic diseases such as malaria and leishmaniasis. At present, methods for treatment of those infections are relatively unsatisfactory and expensive, and approaches to prevention are still under development. It is impossible to estimate how devastating those infections might be on populations of aging humans who lack vigorous immune systems, are difficult to immunize, and may already suffer from some other disorder. REFERENCES 1. United Nations Population Division. World Population Prospects: The 1998 Revision. New York: United Nations, 1998. 2. Oshima S. Japan: Feeling the strains of an aging population. Science 1996;273: 44–45. 3. Holden C. New populations of old add to poor nations’ burdens. Science 1996; 273:46–48. 4. Schneider EL. Aging in the third millenium. Science 1999;283:796–797. 5. US Census Bureau. US Population Estimates by Age, Sex, Race and Hispanic Origin: 1990 to 1994. Report no. PPL-21. Washington, DC: US Census Bureau, 1995. 6. US Census Bureau. Population Projections of the United States by Age, Sex, Race and Hispanic Origin: 1995 to 2050. Report no. P25-1130. Washington, DC: US Census Bureau, 1996. 7. Holliday R. Understanding Ageing. Cambridge: Cambridge University Press, 1995. 8. Finch CE, Tanzi RE. Genetics of aging. Science 1997;278:407–411. 9. Finch CE. Variations in senescence and longevity include the possibility of negligible senescence. J Gerontol Biol Sci 1998;53A:B235–B239. 10. Williams GC. Pleiotropy, natural selection, and the evolution of senescence. Evolution 1957;11:398–411. 11. Kirkwood TBL, Holliday R. The evolution of ageing and longevity. Proc Roy Soc London [Biol] 1979;205:531–546. 12. Holliday R. Causes of aging. Ann NY Acad Sci 1998;854:61–71. 13. Vaupel J. Trajectories of mortality at advanced ages. In: Wachter K, Finch CE, (eds.) Biodemography of Aging. Washington, DC: National Academy, 1977: 17–34. 14. Manton KG. Dynamic paradigms for human mortality and aging. J Gerontol Biol Sci 1999;54A:B247–B254.

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15. Harman D. Ageing: phenomena and theories. Ann NY Acad Sci 1998;854:1–7. 16. Olshansky SJ, Carnes BA, Cassel C. In search of Methuselah: Estimating the upper limits to human longevity. Science 1990;250:634–640. 17. Wilmoth JR. The future of human longevity: A demographer’s perspective. Science 1998;280:395–397. 18. Masoro ET Possible mechanisms underlying the antiaging actions of caloric restriction. Toxicol Pathol 1996;24:738–741. 19. McCay CM, Crowell MF, Maynard LA. The effect of retarded growth upon the length of the lifespan and upon the ultimate body size. J Nutr 1935;10:63–79. 20. Weindruch R. Immunogerontologic outcomes of dietary restriction started in adulthood. Nutr Rev 1995;53:S66–S71. 21. Yu BP, Maeda H, Murata I, Masoro EJ. Nutritional modulation of longevity and age-related disease. Fed Proc 1994;43:858 (abs. 3349). 22. Ross MH, Bras G. Tumor incidence patterns and nutrition in the rat. J Nutr 1965;87:245–260. 23. Losconczy KG, Harris TB, Cornoni-Huntley J, et al. Does weight loss from middle age to old age explain the inverse weight-mortality relation in old age? Am J Epidemiol 1995;141:213–221. 24. Allison DB, Gallagher D, Heo M, et al. Body mass index and all-cause mortality among people age 70 and over: The longitudinal study of aging. Internat J Obesity 1997;21:424–431. 25. Roubenoff R, Harris TB. Failure to thrive, sarcopenia, and functional decline in the elderly. Clin Geriatr Med 1997;13:613–621. 26. Sohal RS, Weindruch R. Oxidative stress, caloric restriction and aging. Science 1996;273:59–63. 27. Hagen TM, Yowe DL, Bartholomew JC, et al. Mitochondrial decay in hepatocytes from old rats: Membrane potential declines, heterogeneity and oxidants increase. Proc Natl Acad Sci USA 1997:94:3064–3069. 28. Erdineler DS, Seven A, Inci F, et al. Lipid peroxidation and antioxidant status in experimental animals: Effects of aging and hypercholesterolemic diet. Clin Chem Acta 1997;265:77–84. 29. Sanz N, Diez-Fernandez C, Alvarez A, Cascales M. Age-dependent modifications in rat hepatocyte antioxidant defense systems. J Hepatol 1997;27:524–534. 30. Harman D. Free-radical theory of aging: Increasing the functional life span. Ann NY Acad Sci 1994;717:1–15. 31. Beckman KB, Ames BN. The free radical theory of aging matures. Physiol Rev 1998;78:547–581. 32. McCann SM, Licinio J, Wang ML, et al. The nitric oxide hypothesis of ageing. Exp Gerontol 1998;33:813–826. 33. Stadtman ER. Protein oxidation and aging. Science 1992;257:1220–1224. 34. Zhang FL, Casey PJ. Protein prenylation: Molecular mechanisms and functional consequences. Ann Rev Biochem 1996;65:241–269. 35. Gelb MH. Protein prenylation, et cetera: Signal transduction in two dimensions. Science 1995;275:1750–1751. 36. Jakubowski H. Protein homocysteinylation: Possible mechanism underlying pathological consequences of elevated homocysteine levels. FASEB J 1999;13:2277–2283.


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37. Bucala R, Cerami A. Advanced glycosylation: Chemistry, biology and implications for diabetes and aging. Adv Pharmacol 1992;23:1–34. 38. Curtsinger JW, Fukui HH, Khazaeli AA, et al. Genetic variation and aging. Ann Rev Genet 1995;29:553–575. 39. Arking R, Force AG, Dudas SP, et al. Factors contributing to the plasticity of the extended longevity phenotypes of Drosophila. Exp Gerontol 1996;31:623–643. 40. Shama S, Lai CY, Antoniazzi JM, et al. Heat stress-induced life span extension in yeast. Exp Cell Res 1998;245:379–388. 41. Duhon SA, Murakami S, Johnson TE. Direct isolation of longevity mutants in the nematode, Caenorhabditis elegans. Development Genetics 1996;18:144–153. 42. Kenyon C. 1996 Ponce d’elegans: Genetic quest for the fountain of youth. Cell 1996;84:501–504. 43. Lakowski B, Hekimi S. Determination of life-span in Caenorhabditis elegans by four clock genes. Science 1996;272:1010–1013. 44. Carey JR, Liedo P, Muller H-G, et al. Relationship of age patterns of fecundity to mortality, longevity, and lifetime reproduction in a large cohort of Mediterranean fruit fly females. J Gerontol Biol Sci 1998;53A:B245–B251. 45. Bertram MT, Berube NG, Hang-Swanson X, et al. Identification of a gene that reverses the immortal phenotype of a subset of cells and is a member of a novel family of transcription factor-lilke genes. Molec Cell Biol 1999;19:1479–1485. 46. Rattan SIS. Beyond the present crisis in gerontology. Bio Essays 1985;2:226–228. 47. Lithgow GJ, Kirkwood TBL. Mechanisms and evolution of aging. Science 1996; 273:80. 48. Rattan SIS. Gerontogenes: Real or virtual? FASEB J 1995;9:284–286. 49. Walford RL. The Immunologic Theory of Ageing. Copenhagen, Munksgaard,1969.

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Aging and Altered Resistance to Infection

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2 Aging and Altered Resistance to Infection All in all, my fellow pathogens, Homo is the opportunity that ultimately can benefit us all. Aside from their prevalence in numbers, they show all the weaknesses that maximize our effective potential. Although they themselves deny that there is such a thing as a free lunch, we know better. There is a free lunch, and it is them. —Thomas Eisner and Paul R. Ehrlich, New world pathogen strategy disclosed, Science 2000; 292, Editorial.

Altogether, microbial and parasitic diseases constitute the leading cause of morbidity and mortality worldwide. They affect preferentially the very young and the elderly, the two age groups that are deficient in immunological competence. This chapter is a review of some of the organisms that are particularly devastating to the elderly. A portion of the chapter deals with the remarkable variability that those microorganisms are capable of manifesting in order to ensure their adequacy to reproduce in their hosts. Optimally, a pathogen should be sufficiently virulent to thwart the defenses of its host without overwhelming it. A host that is quickly ravaged is unsuitable for the pathogen, which has the single objective of perpetuating itself. Upon infection, a struggle develops between host and pathogen with the advantage going first to one adversary and then to the other. Microbial pathogens are, of course, capable of much more rapid variation than are their hosts. Therefore, it is in the pathogen’s self-interest to utilize sparingly the weapons of virulence in its arsenal so that there is opportunity to reproduce and allow progeny to move on to new hosts. When a microbial pathogen (or any parasite) quickly overwhelms its host, it probably indicates that an adaptive equilibrium has not been achieved. That is sure to be the case when hosts that are immunodeficient are involved, i.e., hosts that are very young, those suffering from immunodeficiency diseases or being treated with immunosuppressive agents, and the elderly. From: Aging, Immunity, and Infection By J. F. Albright and J. W. Albright © Humana Press Inc., Totowa, NJ

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Aging, Immunity, and Infection Table 2-1 Some Important Geriatric Infectious Diseases and Their Relative Mortality Rates Infection

Relative mortality rate (compared with young adults)

Pneumonia Urinary tract infection Bacterial meningitis Tuberculosis Sepsis

3 5–10 3 10a 3

aExcluding HIV-infected young adults. Adapted from ref. 1.

RELATIVELY COMMON BACTERIAL INFECTIONS OF AGING HUMANS Some important infectious diseases and their relative mortality in elderly subjects are shown in Table 2-1 (1). As expected, that list reflects the fact that there are three principal routes of infection: respiratory, urinary, and gastrointestinal (GI). The most compelling explanations of the prevalence of those diseases in the elderly are: 1) age-associated changes in the structure and function of the respiratory, urinary, and gastrointestinal organs; 2) underlying pathological changes resulting from existing disorders (comorbidity); and 3) age-associated decline (dysregulation) in innate (natural) and acquired (adaptive) imunological competence. Respiratory and Urinary Tract Infection Table 2-2 provides a list of organisms found most often in respiratory and urinary tract infections of the elderly. The most common respiratory infection is bacterial pneumonia. In about half of the community-acquired pneumonia (CAP) cases, the etiologic agent remains unidentified (2). It is estimated that 20%–30% of all CAP infections are caused by Streptococcus pneumoniae and most of the remaining cases by the other bacteria listed in Table 2-2. Upper respiratory viral infections were studied in a group of 533 subjects ages 60 to 90 years living in England (3). In that group of patients, 52% of the infections were associated with rhinoviruses, 26% with coronaviruses, 9.5% with influenza A or B, and 7% with respiratory syncytial virus, and the remainder with other agents. In the case of urinary tract infections (UTIs) in the elderly, two independent studies, separated by an interval of 12 yr, gave very similar results. One study was performed in Sweden in 1986 on a group of 1966 subjects having a mean


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Table 2-2 Pathogens Found Frequently in Elderly Subjects with Respiratory or Urinary Tract Infections Organ system Respiratory tract (upper and lower)

Pathogen found frequently Bacteria Streptococcus pneumoniae Hemophilus influenza Legionella pneumophila Chlamidia pneumoniae Viruses Rhinoviruses Coronaviruses Influenza Respiratory syncytial

Urinary tract

Bacteria Escherichia coli Proteus Klebsiella Pseudomonas aeruginosa Enterococci

age of 70 years (4). The majority of those subjects were not in hospitals or institutions. The other study occurred in England in 1998 on a group of 3119 subjects all of age greater than 65 years (5). The results of the two studies agreed that Escherichia coli was the most common organism in UTIs. Klebsiella, Proteus, Pseudomonas aeruginosa, and enterococci were found less frequently but in significant numbers of subjects. The study performed in 1998 (5) included comparisons of the organisms found in bacteremic patients with respect to: 1) where the infection was acquired, i.e., in the community or in the hospital; and 2) the patients’ ages, over 65 years or in the range 18–64 years. In both age groups, E. coli was the dominant organism in more than 70% of the community-acquired infections. In the case of hospital-acquired infections, E. coli was the principal organism in approx 40% of the patients, regardless of age. Various other organisms (Klebsiella, Proteus, P. aeruginosa) were dominant in about 60% of the hospital-acquired infections. The list of organisms associated with UTIs reflects the fact that a major portion of UTIs is caused by pathogens derived from the patients’ colonic flora that enter the bladder by the “ascending route,” i.e., via the perineum, urethra, vagina, or prostate. Viral infections of the bladder are rare.

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The purpose of presenting the lists in Tables 2-1 and 2-2 is to provide a general indication of the types of microorganisms with which the elderly must contend. Geriatric infectious diseases per se are not discussed here; they are the topic of a major, recent publication edited by Yoshikawa and Norman (6). Gastrointestinal Infections Many factors can influence the GI flora; e.g., diet, medications, malabsorption, deficient intestinal motility, lumenal pH. Several of these may be altered with age as is discussed below. There are important reasons for giving special attention to intestinal microbial flora in the elderly. First, the gut is a likely source of pathogens that cause illnesses of high mortality in older patients (e.g., infective endocarditis, cholecystitis, sepsis); second, the importance of diet (caloric restriction) on longevity (discussed in a later chapter); third, the rather common problems of malnourishment, malnutrition, and dietary deficiencies (e.g., vitamins) in the elderly; and fourth, the translocation of microbial components and products from the gut to the circulation and the adverse effects on the health of the elderly. There is no known intestinal microbial pattern that distinguishes young adult from elderly. Given that there are more than 400 bacterial species in the colonic flora of a single individual (7), it is unlikely that a catalog of intestinal flora would be a useful biomarker of senescence. However, it is possible that one or a few species might be characteristically different in the young adult and the elderly. Apparently, this possibility has not been explored. The number of bacteria and the spectrum of species in normal adults varies with the segment of the intestine, as displayed in Table 2-3 (8). There are relatively few bacteria in the stomach and jejunum; those that are present are predominantly aerobes or facultative aerobes. In contrast, the colon is lushly endowed with bacteria, as revealed by fecal examination, the majority of which are anaerobes. Only small numbers of fungi, or protozoa, are present, even in the colon. The number and variety of bacteria in the gut remain rather constant in the healthy individual and are controlled primarily by gastric acid secretion and normal intestinal motility. In the healthy, well-nourished, elderly subject, the intestinal flora appears to be similar to that of the young adult. However, there is much more variation among the elderly for reasons that are considered below. As far as is known, there are no microbial pathogens that uniquely infect the elderly. Rather, the heightened susceptibility to infections associated with aging may be viewed in the following way (9): “Diminished physiologic reserve secondary to both biologic changes of aging and coexisting chronic diseases contributes to the higher mortality and morbidity rates observed for serious infection in older compared with younger persons.”


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Table 2-3 The Normal Gastrointestinal Flora of Humans Stomach 0–103

Jejunum 0–105

Ileum 103–107

Feces 1010–1012

Aerobic or facultative anaerobic bacteria Enterobacteria Streptococci Staphylococci Lactobacilli Fungi

0–102 0–103 0–102 0–103 0–102

0–103 0–104 0–103 0–104 0–102

102–106 102–106 102–105 102–105 102–103

104–1010 105–1010 104–107 106–1010 102–106

Anaerobic bacteria Bacteroides Bifidobacteria Gram-positive coccia Clostridia Eubacteria

Rare Rare Rare Rare Rare

0–102 0–103 0–103 Rare Rare

103–107 103–105 102–105 102–104 Rare

1010–1012 108–1012 108–1011 106–1011 109–1012

Total bacterial count

aIncludes Peptostreptococcus and Peptococcus. From ref. 8.

Age-Associated Changes in Anatomical–Functional Relationships The “diminished physiologic reserve” referred to in the preceding quotation includes anatomical and functional changes associated with aging of the respiratory, urinary, and gastrointestinal systems. In the case of the respiratory system, it is well established that pulmonary function deteriorates with age (2). Some of the anatomical changes that contribute to loss of function include: (a) decreased mean broncheolar diameter; (b) increased diameter of the alveolar sacs, which become shallower; (c) decrease in elastic fibers and increase in type III collagen. Those anatomical changes contribute to the following functional changes: (a) decrease in elastic recoil; (b) decrease in oxygen diffusion capacity; (c) small airway closure resulting in air trapping; (d) decreased expiratory flow rates. Spirometric changes include decreased inspiratory reserve volume, decreased expiratory reserve volume, and decreased vital capacity. In addition, the mucociliary clearance is substantially decreased in older subjects. The net effect of these changes is an increased probability of being unable to expire or clear infectious organisms that enter the lungs. The normal oropharygeal flora is a mixture of aerobic and anaerobic bacteria and may account for a significant number of cases of CAP. In fact, it has been estimated that aspiration of oral flora is second only to S. pneumoniae in causing CAP (10).

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The healthy bladder is quite resistant to colonization by bacteria. Emptying of the bladder is the most effective way of preventing bacteria from colonizing. The elasticity of the bladder diminishes with age, which makes effective emptying more difficult. Whereas among mature adults the incidence of bacteria is much greater in females, in males and females over age 65 the incidence of bacteria is almost equal. The principal contributing factors are 1) obstructive uropathy from prostatic disease in males, 2) impaired emptying of the bladder with residual urine in females, and 3) urethral catheters and associated paraphernalia in both (5). As long as the physiological condition of the individual remains good, there are no changes in the GI system that become threatening. That is not to say that there are no changes in the GI system, rather that what changes may occur are of no serious consequence. This point was made by Saltzman and Russell (11), who wrote: “The multiorgan system that composes the gastrointestinal tract has a large reserve capacity, and thus there is little change in gastrointestinal function because of aging in the absence of disease.” That can accurately be said about many organ systems with respect to aging. Consider the large functional reserve of the liver, the necessity for only one kidney, the reserve capacity of the lungs, or the large excess capacity of the bone marrow for hematopoiesis. Certainly, there is a large excess of immunological potential in the young adult that gradually diminishes with advancing age as is discussed in Chapters 3 and 4. Indeed, it can be argued that the gradual diminution in potential declines to a point approaching the level that must be expressed to deal with an acute need or an emergency; beyond that point the effects of aging are manifested. There are functional changes that occur in the GI system with age, beginning with the fact that gastric acid secretion diminishes resulting in an increase in pH in the proximal small intestine and the potential for bacterial overgrowth. In addition, normal intestinal motility may not be maintained, a factor that also disposes to bacterial overgrowth. The latter condition can cause histological changes in the mucosa of the small bowel such as hypertrophy of villi and crypts, vesiculation of the cytoplasm of mucosal cells, swollen mitochondria, and dilated cisternae of the endoplasmic reticulum (12–14). SELECTED EXAMPLES OF AGE-ASSOCIATED SUSCEPTIBILITY TO BACTERIAL INFECTIONS Mycobacterium tuberculosis Worldwide, tuberculosis (TB) is a major cause of morbidity and mortality. In the more-developed countries, during the early 20th century, TB gradually declined and by midcentury was not considered a significant public health problem. That changed in the 1970s with the onslaught of HIV-1 infections and the associated immunodeficiency. The incidence of TB rose significantly over the next 20 years or more. Prior to 1970, it was already recognized that there was a


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clear association between advancing age and the susceptibility to TB. For example, in 1970 the incidence in the United States among persons 65 and under was approx 7 per 100,000 population and among persons over 65 about 35 per 100,000. In 1992, it was reported (15) that slightly over half of all TB cases in the United States were found in people over 65 who, at that time, constituted about 14% of the population. Research concerned with TB was at a low ebb during much of the 20th century. In the 1980s there was a resurgence of research prompted by the recognition that (a) TB was a prominent opportunistic infection among AIDS victims and (b) many cases of TB were caused by antibiotic-resistant organisms. Much has been learned in the last decade. There has been some debate concerning which experimental animal serves as a suitable model of human TB; and, further, as to whether or not aging experimental animals are more susceptible to Mycobacterium infections than young adults. It was reported that old mice were no more susceptible than young adults to M. tuberculosis (16). The levels of bacteria in target organs and the frequency of death from infection were reported to be essentially the same in young and aged mice. Systematic studies by Orme and associates have shown that there is a difference in the way mice (young and old) cope with M. tuberculosis infection depending on the route of infection and the dose (number) of bacteria provided to the animals (17–19). Aged mice were definitely more susceptible than young when a relatively high number of bacteria was given intravenously. However, when a much smaller number of bacteria was provided aerogenically (modeling a realistic human exposure) the course of infection in the lungs of young and aged mice was similar. Nevertheless, there remained important differences between young and aged mice with respect to elements of the immune system involvement in the infection. For example, T cells collected from infected aged mice failed to confer adoptive immunity on recipient mice whereas T cells from infected young mice did. In the lungs, the levels of mRNA specific for several cytokines, especially IL-12 and IFN-γ, were severalfold lower in aged than in young adult mice. In this regard, it was found that M. tuberculosis infections progressed unabated in interferon (IFN)-γ knockout mice (18). Recent work has shown that components of M. tuberculosis can block IFN-γ-induced, STAT-1 mediated gene transcription in macrophages (20). (STAT is the acronym for “signal transducer and activator of transcription.”) The dissemination of live bacteria from the lungs to form granulomas in livers of aged mice was much greater than in young mice. Orme and associates concluded that there exist (unidentified) mechanisms in the aged animals that can compensate for the impaired immune control of M. tuberculosis infection. Their finding suggested that CD4+ T cells, which play a pivotal role in the control of infection, are affected by aging.

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Recent work has revealed that T cells other than the CD4+ subset can afford protection against M. tuberculosis and probably other intracellular infections (21). T cells were isolated and cell lines generated that were CD4–CD8– (double negative) or CD4–CD8+ and were CD1-restricted. Those T-cell lines possessed αβ T-cell receptors and responded to M. tuberculosis lipid and lipoglycan antigens when the latter were presented by CD1+ macrophages. Both the double negative and the CD8+ lines could affect lysis of M. tuberculosis-infected macrophages. However, the mechanisms of lysis by the two types of T cells were entirely different. Lysis achieved by the double-negative cells was mediated by way of interaction of Fas on the infected target cells and Fas ligand on the T cells and, therefore, was an apoptotic event. Lysis by the CD8+ cells involved exocytosis of granules containing the lytic factors, perforin and granzymes, in typical cytotoxic T lymphocyte (CTL) fashion. Only the CD8+ cells were able to destroy the intracellular M. tuberculosis organisms. Thus, CD1-restricted, CD8+ T cells are candidates for the mechanism postulated by Orme and associates that compensates in the old mice for senescent CD4+ T cells. Of course, there are other candidates. It should be informative to analyze the effects, if any, of senescence on CD1-restricted T cells. Listeria monocytogenes This bacterium is a Gram-positive, human pathogen. The natural portal of entry is oral, leading to invasion of mucosal surfaces of the small intestine. However, L. monocytogenes, which is a facultative intracellular organism, can invade and replicate inside a variety of mammalian cells including those that are, and are not, typical phagocytes. Once ingested, the bacteria are incorporated into phagosomes from which they escape by lysing the phagosomal membrane. The bacteria replicate in the cytoplasm and spread from cell to cell often without becoming extracellular. Thus, they become sheltered from the humoral immune response of the host. Immune defense against L. monocytogenes is cell mediated and involves both activated phagocytic cells, especially IFN-γactivated macrophages, and cytotoxic T cells (see Chapters 3 and 4). Foci of infection may be seen in various organs, such as the liver and spleen where they appear as granulomas. One of the early reports that aged animals are more susceptible to infections than young adults, was a study of L. monocytogenes in mice (22). When mice were inoculated intravenously with a moderate number of L. monocytogenes, the course of infection was similar in young and old animals as judged by the numbers of bacteria in livers and spleens. However, when a larger inoculum was used, the numbers and persistence of bacteria in livers and spleens were substantially greater in the case of the aged mice. Moreover, adoptive immunity conferred on recipients by transfer of either spleen cells or enriched T cells


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from immunized donors was much more effective with cells from young compared to old donors. The results of this very interesting study were challenged by a report of a very similar investigation performed in the same location utilizing the same strain and ages of mice and the same strain of Listeria (23). The conclusion was drawn that aging was without detriment on the ability of mice to generate T-cell immunity to L. monocytogenes. It was found that the numbers of bacteria surviving in the livers and spleens of aged mice were considerably lower than in young mice over the first day following intravenous inoculation of the same number of bacteria. Therefore, some nonimmunological mechanism that destroyed the bacteria in aged mice prevented an optimum dose of antigen from reaching immunological tissues. When a significantly larger number of bacteria was provided to the old than to the young mice (to compensate for those destroyed), it was now found that the T-cell responses in aged mice were equivalent to those in the young. Thus, the apparent defect in T-cell responses in aged mice was in reality a matter of inadequate antigen reaching sites of immune response. It was argued that destruction/sequestration of bacteria by the more-active monocytes/macrophages of aged mice prevented antigens from stimulating the immune response. The discrepancies between the findings in the two reports (see refs. 22 and 23) remain unexplained. Whatever the explanation may be, it is clear that aged mice in the experiments of Lovik and North (23) required a larger inoculum of L. monocytogenes to generate a T-cell response equal that in the young mice. Considerably more bacteria were retained in the livers of old than of young mice. The condition of the bacteria in the livers of aged mice was not determined. It is now well-known that macrophages vary in the way ingested L. monocytogenes are handled; they may be killed or they may be retained in viable condition (24). They may not have been killed but, rather, retained alive in the Kupffer cells as occurs, for example, for 24–48 hours after intravenous inoculation of the parasite, Leishmania donovani (25). If those entrapped bacteria were subsequently released by the Kupffer cells, a large bolus of antigen might arrive at sites of immune response just in time to drive an anamnestic response. Thus, the response reported in ref. 23 might not have been an assessment of the competence of aged mice for a true primary immune response. The question of why the livers of aged mice retained bacteria more effectively than livers of young mice is a separate matter. The effects of senescence on macrophages and their ability to deal with microorganisms are discussed in Chapter 3. The need to provide aged mice with 10- to 50-fold more L. monocytogenes to obtain a T-cell response equal to that of the young, as found by Lovik and North, could be a reflection of inefficient antigen processing by dendritic cells of old mice, or a reflection of a requirement for more intense processed-anti-

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gen stimulation of senescence-altered T cells. The effects of aging on dendritic cells (DCs) and T cells are discussed in Chapter 4. At this point, it need only be mentioned that dendritic cells are the critical antigen-presenting cells that prepare microbial antigens for triggering immune responses. It appears that more attention to the effects of aging on immune responses to L. monocytogenes could be rewarding. Much is now known about the mechanisms of natural and acquired immunological resistance to this organism (26) but that knowledge has not been applied to understanding the possible effects of senescence. Salmonella typhimurium In the preceding paragraphs, the finding (23) that intravenous L. monocytogenes are trapped effectively by livers (and spleens) of aged compared to young mice was discussed. That is also true of liver (and spleen) of aged rats inoculated with S. typhimurium (27). Perhaps that is the case generally for intracellular microorganisms. If so, it is important to determine why this is so and investigate the influence of macrophage entrapment of the microbes on the immune response to their antigens. Macrophages themselves are not efficient microbial antigen-presenting cells. However, after ingestion of bacteria, macrophages may undergo apoptosis, and components of bacteria picked up by immature dendritic cells. The latter may thus acquire the bacterial antigens, which they then present to T cells (28). Uptake of apoptotic material can induce maturation of the dendritic cells and expression of new surface molecules that allow the cells to migrate to lymphoid sites where they interact with T cells (29). Studies of these events in aged mice and other animals is likely to provide much new insight into the effects of senescence on immune responses. Before leaving this discussion of S. typhimurium infections, it should be mentioned that this pathogen typically enters the body by the oropharyngeal route. It traverses the intestinal barrier by invading epithelial cells and membranous epithelial (M) cells, which overlie the lymphoid follicles (see Chapter 4). After passing through the M cells, the bacteria encounter a network of macrophages and dendritic cells where the events described in the preceding paragraph can occur. However, there is an alternative process, which involves transmigration of the macrophages bearing live S. typhimurium from the intestine into the circulation and subsequent dissemination to sites where humoral antibodies can be generated (30). This is discussed in more detail in Chapters 3 and 4. BACTERIAL INTERACTIONS WITH MUCOSAL SURFACES Whether it be in the lungs, the urinary bladder, or the intestine, the flourishing of bacteria depends upon their attachment to, and successful interac-


29

tions with, mucosal surfaces. The interactive processes in which various types of bacteria engage include: attachment and effacement, translocation across epithelial or endothelial barriers either between cells (pericellular route) or through cells (transcellular route), and invasion of host cells. Only in the last decade have these various events been elucidated. Most of the studies have been done in model, in vitro systems or in young, experimental animals. At present, little is known about how the various interactive events might differ in the case of mucosal surfaces of aged hosts. There follows a series of brief descriptions of the interactive events as currently understood. Bacterial Attachment The attachment to host cells is required for bacterial proliferation, colony formation, invasion of host cells, or translocation across endothelial or epithelial host cell layers. Both the bacteria and the host cells may be altered as a consequence of activation of genes in both. Adherence allows the bacteria to resist host defensive processes such as mucociliary sweeping. There is a clear correlation between the ability of a pathogen to adhere to host cells and the susceptibility of the host to that pathogen. For example, among individuals who experience recurring UTIs, adherence of E. coli to epithelial cells of the subjects may be as much as five times greater than in the case of subjects who remain free of infections (31). Pathogens, including bacteria, employ a variety of mechanisms for adhering to host cells. In several, well-studied cases, known adhesion molecules are involved (32). For example, outer membrane molecules of several bacteria (Yersinia spp., Bordetella pertussis), protozoa (Leishmania mexicana), and even viruses (echovirus 1, adenovirus) have been found to bind directly to integrins present on model host cells in vitro. Either β1or β2 integrins may be utilized. Several studies have revealed that in some cases bacteria such as Streptococcus spp., P. aeruginosa, and Staphylococcus aureus bind first to host cell molecules such as laminin, collagen, and fibronectin, which then associate with integrin receptors. Other pathogens such as Legionella pneumophila may bind selectively to the complement component, C3bi, which is a ligand for αmac β2 integrin. B. pertussis display several adhesive molecules of which two have been fairly well studied, viz., the filamentous hemagglutinin (FHA) and the pertussis toxin (PT) (reviewed in ref. 33). Although not a pathogen of elderly humans, what has been learned about adherence of B. pertussis to human cells is broadly instructive. Ciliated cells and macrophages are the host cells to which B. pertussis binds. Mutant strains of B. pertussis have been prepared that lack either FHA or PT or both (34). When tested in normal rabbits, wildtype strains localized to cilia in the respiratory tract and produced lesions.

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Mutants lacking both FHA and PT were cleared without inducing pathology. Mutants lacking either adhesin failed to attach to cilia but managed to pass into the alveoli where they caused pathological changes (33). FHA is a functionally complex molecule that displays several domains capable of interacting with complementary sites on host cells. These domains include: an N-terminal lectin domain that binds sialic acid and is required for hemagglutination; a lectin domain for ciliated host cells; a domain containing an arginine-glycine-asparagine (RGD) sequence that binds to the leukocyte integrin CR3 (CD11b/CD18) and two regions that resemble sites on factor X of the coagulation mechanism that also bind to leukocyte CR3 (33). This complexity allows FHA to interact with a variety of receptors on host ciliated cells, erythrocytes, and leukocytes. The PT protein is a hexamer. One monomeric subunit bears the catalytic site that affects adenosine 5'-diphosphate (ADP) ribosylation of guanine nucleotide proteins involved in host cell signal transduction and thus exerts the toxic effect of PT. The pentameric region of PT displays the binding sites that promote binding to host cells and intracellular delivery of the toxin. Those binding sites include lectin subunits (S2 and S3) of the pentamer, which are responsible for PT binding to glycoconjugates on cilia (S2) and the association of B. pertussis with macrophages (S3). Studies on the binding specificities of the S2 and S3 subunits, especially their recognition of the Lewis “a” and “x” blood group determinants, suggested that they are selectins. They possess significant sequence homology to stretches of known selectins (35) and structural studies have revealed that those stretches of homology are superimposable in the crystal structures of S3 and E selectin (36). It has been demonstrated that FHA and PT resemble natural ligands to the extent that they can act as competitive inhibitors of integrins and selectins, respectively. As a consequence these components of B. pertussis can interfere with neutrophil adherence and endothelial transmigration. This is an example of how bacterial components might exacerbate infection by interfering with a host defense mechanism. Although whooping cough is not a problem in the elderly, the preceding discussion of B. pertussis adherence to host cells highlights an important point; namely, that very little work has been done to understand whether or not adhesion receptors and ligands change with age and, if certain of them do, the consequences of such changes. That is an important consideration in the following discussion of infections with S. pneumoniae. S. pneumoniae is responsible for several localized and systemic infections such as otitis media, meningitis, sepsis, and pneumonia. There is a wellestablished relationship between pneumococcal bacteremia associated with pneumonia and mortality of aging subjects (38). Among patients with bacteremia, the fatality rate has been related to age as follows: 17–18% among


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young adults, 43% among those of age 60–69, 48% of those aged 70–79, and 60% among patients 80 years or older. S. pneumoniae enters the body by the nasopharyngeal route and pneumonia results from inspiration of the bacteria into the lower respiratory tract. Bacteria may be found in the alveoli from which they gain access to the circulation by crossing the endothelium of the alveolar capillaries. S. pneumoniae attaches primarily to cells of the nasopharynx, vascular endothelium, and other cells of the lung. The local inflammation that they induce is triggered by components of the cell wall. In fact, the pathogenesis of inflammation can be induced experimentally by mixtures of cell wall components (39,40). Those components are able to activate the complement system via the alternative pathway giving rise to leukocytosis, vascular permeability, secretion of interleukin-1 (IL-1) by macrophages, and other effects (33). The pneumococci bind to glycoconjugate moieties on host cells. Cells of the nasopharynx display glycoconjugate receptors of the neolactose type containing GlcNAc β1-3 Gal. The latter is also a component of the ABH, Lewis, and Ii blood group antigens and is present in human colostrum. In fact, colostrum can inhibit pneumococcal adherence to nasopharyngeal cells (41). The receptors on type II pneumocytes and vascular endothelial cells responsible for attachment of pneumococci are of two types; both of them differ from the receptor on nasopharyngeal cells. Saccharides that can competitively inhibit the adherence of S. pneumoniae to pneumocytes and vascular endothelial cells help to define those receptors. They include mannose, GalNAc, Gal, the glycoconjugates asialo-GM1 and GM2, and the Gal NAcβ1-3 Gal-containing Forssman glycolipid (33). It should be mentioned here that the exposure of type II pneumocytes and vascular endothelial cells to the inflammatory cytokines TNFα and IL-1 significantly elevate the glycoconjugate receptors for pneumococci (33,42). As a consequence, adherence of S. pneumoniae is markedly enhanced. Enhanced adherence entails a new receptor, viz., that for the platelet-activating factor (PAF). As pointed out (33), this is an example of an important principle of bacterial adherence; viz., the initial attachment of bacteria to resting host cells may involve one set of sugar specificities leading to activation of the cells and the expression of a new or altered receptor specificity. A virulent organism must parallel this change in the host cell by expressing a new cognate adhesion molecule. It has been emphasized that: “This action-and-reaction scenario underlies the success of virulent pathogens and illustrates the dynamic nature of the response of both partners in an adherence interaction” (33). It appears that little, if anything, is known about the possible changes that might occur in, for example, pneumocytes or endothelial cells with age that would affect attachment and transmigration of pneumococci or other pneumonia-causing pathogens.

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The ability to adhere to host enterocytes is a major determinant of virulence in the case of enteric pathogens. Most of those pathogens express adhesins that function as lectins (43). The adhesins may or may not be present on bacterial fimbriae. Adhesins present on rigid fimbriae are maintained at a distance from bacterial surface components that might interfere with adherence to host cells. Adhesins present on flexible fimbriae are allowed spatial freedom in binding to cognate receptors. Many enteric bacterial adhesins interact with and agglutinate erythrocytes from various species. They can be characterized by the spectrum of species of erythrocytes that they agglutinate. By this test, families of lectins have been identified (43); these include galactoside-specific, sialic acid-specific, and N-acetylglucosamine-specific lectins. Some adhesins fail, however, as hemagglutinins (perhaps because the saccharide moieties that they recognize on erythrocytes are inaccessible) but do interact with cells of the intestinal tract. Intestinal mucous contains numerous, potentially inhibitory saccharide moieties that may block bacterial adherence. Whether or not this occurs depends on a number of factors including the quantity and rate of formation of mucous. The wide variety of glycoconjugates with which bacterial lectins interact is shown in Table 2-4. That variety ensures the ability of many enteric bacteria to adhere at optimum sites and niches within the intestine. Intestinal E. coli strains can be categorized into several types depending on their attachment and invasive properties. Those categorized as enteropathogenic E. coli (EPEC) adhere to gut epithelial cells through intimin molecules. The latter are ligands for the bacterial complementary receptor known as Tir (translocated intimin receptor) (44,45). Following the initial adherence of EPEC to host cells, the bacteria introduce Tir, along with several other proteins, into the host cells via a type III secretion mechanism. The expression of Tir significantly enhances the adherence and, thus, promotes the secretion of effacing factors into the host cells. This is presumably a key event in the initiation of diarrhea. What is particularly interesting about this example of adherence is the fact that the bacterium provides to the host cell the receptor (Tir) to which it strongly binds. Additional interest in this interaction arises from the recent finding (46) that the ensuing inflammatory reaction that leads to pathology of the colon is dependent on the involvement of type 1 T helper (Th1) cells (CD4+) and the IFN-γ that they secrete. It appears that intimin not only interacts with Tir but also with β1 integrin molecules on T cells. The resulting hyperplastic changes in the gut provide further opportunities for EPEC colonization. This is an excellent example of the diversion of host immune defenses (T cells and their cytokines) into paths that promote the welfare of the bacterial pathogen. There are other outstanding cases such as that of S. typhimurium (47). In this case, the bacterium secretes proteins via a type III mechanism that


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Table 2-4 Some Interactions of Bacterial Lectins-glycoconjugates Bacteria Klebsiella pneumoniae Mycobacteria Salmonella Serratia marcescens Shigella flexneri Vibrio cholerae Streptococcus pyogenes Escherichia coli (urinary pathogens)

Lectin type Type 1 & 3 Mycotin

P-related (F7–F16) G-I G-II G-III

Clostridium difficile Escherichia coli Streptococcus pneumoniae Escherichia coli (sepsis pathogen)

Escherichia coli (urinary pathogen) Vibrio cholerae Helicobacter pylori

Mycoplasma galliseptum; M. pneumoniae Candida albicans Haemophilus influenzae Staphylococcus aureus Bordetella pertusis Borrelia burgdorferi (Lyme disease)

AFA (enterotoxin) G S

(enterotoxin)

PHA

Carbohydrate specificity mannose mannose, mannan mannose mannose α-mannoside fucoside galactose Galα1-4Gal in globotriaosylceramide globotetraosylceramide globopentaosylceramide (Forssman antigen) Dr blood group antigen Galα1-3Galβ1-4GlcNAc N-acetylglucosamine GlcNAcβ1-3Gal NeuAc α2-3Gal β1-3GalNAc & NeuAcα2-3Galβ1-3(NeuAcα2-6) GalNAc NeuAcα2-3Galβ1 GM 1 GM3 (NeuAcα2-3 Gal β1– 4Glc-cer)So3-GM3; Leb-group blood antigen NeuAc α2– Lewis (a) antigen Lewis (a) antigen Lewis (a) antigen Sulfated glycolipids, heparin Gal-Cer, Lac-cer; GD1a, GD1b, GM2, GM3

Modified from ref. 43.

activates signaling pathways in the host epithelial cells leading to production of IL-8 and other proinflammatory cytokines, e.g., TNF (tumor necrosis factor) α, and GM-CSF (48–50). Bacterial Type III Secretion Mechanism There are four different mechanisms utilized by bacteria to export synthesized products, especially virulence factors (51). Types I and II lead to secre-

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tion of materials directly into the surrounding milieu. In type I secretion, the mechanism employs three proteins that form a channel through the inner and outer bacterial membranes. The type II mechanism has been studied extensively in Vibrio cholerae (52). There are at least 12 proteins that appear to create a pore that bridges the inner and outer membranes and through which bacterial products are secreted. The type IV system was discovered fairly recently and the mechanism is under scrutiny (53). Among other products, the secretion of immunoglobulin A (IgA) proteases occurs via the type IV system. The type III secretion system is of particular interest owing to its complexity and the fact that it is employed by a number of Gram-negative pathogens to introduce virulence- related substances directly into the cytosol of host cells (54). Some substances are secreted into the extracellular environment rather than translocated into a host cell. The type III system is employed by both animal and plant pathogens. Among the animal pathogens are Shigella spp., Yersinia spp., Chlamydia spp., P. aeruginosa, EPEC, enterohemorrhagic E. coli, and S. typhimurium. The type III secretion mechanism involves a structure termed the needle complex, which has been isolated from interacting S. typhimurium and host cells (55). Electron microscopy showed that the structure resembles a long hollow tube attached to a cylindrical base that anchors the structure to the bacterial inner and outer membranes. Other bacteria may induce the formation of pedestals or cuplike structures between themselves and host cells. The formation of such structures is initiated upon contact and association of bacterium and host cells facilitated by adhesion molecules. Some bacteria, e.g., EPEC, induce extensive cytoskeletal reorganization in host cells leading to pedestals that contain cytoskeletal proteins (e.g., actin, α-actinin, talin). Those cytoskeletal rearrangements result in effacement, which includes loss of microvilli and the resulting diarrhea. Bacterial Invasion of Host Cells Some bacteria are obligate intracellular pathogens (e.g., Chlamydia spp.), many important pathogens are facultative intracellular organisms (e.g., Salmonella spp., L. pneumophila, Mycobacterium spp., L. monocytogenes), and others are predominantly extracellular (e.g., enterotoxigenic E. coli, Haemophilus influenzae, V. cholerae). Pathogens that penetrate epithelial barriers survive by invading and replicating in host cells. Tight junctions (zona occludens) that normally prevent penetration of epithelial cell layers also divide the epithelial cells into apical (lumenal) and basolateral surfaces. Some pathogenic bacteria such as Salmonella invade host cells from the apical surface whereas others (Yersinia, Shigella) interact with and invade through the basolateral surface. The invasion of host cells by S. typhimurium has attracted the attention of several groups of researchers (55–57), whose findings are quite significant.


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The adherence of the bacteria triggers some pronounced cytoskeletal and membrane modifications of the host cells. Those modifications are triggered by the introduction (via a type III secretion mechanism) of bacterial proteins including Sop E, which activates GTPases of the Rho subfamily, and Sip A, which binds to actin and prevents depolymerization of actin filaments. The formation of actin filaments is required for bacterial internalization. Other proteins involved in cytoskeletal changes include α-actinin, talin, and ezrin. Associated with those changes is an intracellular Ca2+ shift and activation of “mitogenactivated protein” (MAP) kinase. These and other events, still to be elucidated, in a signal transduction pathway lead to membrane ruffling, macropinocytosis, and thus to internalization of the bacteria by a host cell. The preceding descriptions of attachment, type III secretion, and invasion of bacteria ,vis-à-vis, epithelial cells are fundamental to future studies of epithelial cells that have been altered by senescence. To underscore the importance of such studies, it is worth stressing the growing evidence (discussed later) that aging results in some considerable changes in the composition and structure of membranes. As noted above, pathogens such as Yersinia and Shigella invade epithelial cells via the basolateral surface. Salmonella can also take this as well as the apical approach. How do bacteria that can only penetrate the basolateral surface gain access to that surface? This question is particularly relevant to enteric pathogens. The answer appears to be that they are transported indiscriminately through specialized M cells overlying Peyer’s patches that are scattered throughout the small intestine (56). M cells lack all but a thin layer of mucous and are nearly devoid of villi. It is agreed that the transport of macromolecules and particulate matter from the intestinal lumen through the M cell brings the transported material into proximity with macrophages and lymphocytes located in “pockets” on the antilumenal side of M cells. In this manner bacteria can reach the basolateral surfaces of epithelial cells. Shigella employ an alternate method of crossing the barrier. They can induce chemotaxis of phagocytic cells, especially polymorphonuclear leukocytes. As the latter migrate toward the bacteria they open spaces in the tight junctions through which the bacteria pass across the epithelial surface (58). It is interesting to note that S. typhimurium is representative of several bacteria that can cross the intestinal barrier either via M cells or by traversing enterocytes (56). The entrance and fate of bacteria that enter into host macrophages and dendritic cells are discussed in some detail in Chapter 3. As mentioned previously, the case of L. monocytogenes may be considered prototypic. Some macrophages readily destroy ingested L. monocytogenes whereas others lack that ability (24). Bacteria replicate in the latter and may be passed from cell to cell without becoming accessible to the host’s immune system.

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Biofilms and Quorum Sensing Individual bacteria that lodge in alveoli or attach to the epithelium of the intestine or urinary bladder are in a precarious situation. The hostility of the environment makes it unlikely they will survive for long. Therefore, in order to outmaneuver host defenses, the bacteria replicate rapidly and form microcolonies. Many species of bacteria, including a number of pathogens, form organized communities termed “biofilms.” Only in the last decade has a clear understanding of the mechanisms and significance of biofilm formation begun to emerge (59). For example, it was realized that particles of biofilm formed by L. pneumoniae and circulating in building air ducts were responsible for the notorious outbreak of Legionnaires disease in Philadelphia in 1976. In 1993–1994, hundreds of asthmatic individuals received albuterol that, although drawn from a disinfected processing tank, was contaminated with particles of P. aeruginosa biofilm (60). It is now well-known that the lungs of cystic fibrosis patients are sites where biofilms of P. aeruginosa are formed. The importance of biofilms in infectious diseases associated with aging is suggested by the contents of Table 2-5. In addition to the infections shown in that table, there is growing evidence that biofilms may develop and complicate bacterial pneumonia and intestinal infections (bacterial overgrowth) and may perpetuate the durable infections of M. tuberculosis often associated with the recurrence of tuberculosis in the elderly. The recent realization that bacteria present in biofilms are notoriously resistant to antibiotics and are protected from both humoral and cell-mediated immunity of the host are of major concern in treating infections of the elderly. The community of bacteria in biofilms is protected by an extracellular matrix of polysaccharide (termed “glycocalyx”). The chemical nature of the latter varies with the species of bacteria and whether the community is mixed or of a single species. Moreover, the community organization allows functional heterogeneity and regional specialization much like an organized tissue. Individual bacteria (called “planktonic”) may leave the biofilm and disperse to other sites somewhat resembling metastasis of tumor cells. Planktonic bacteria are susceptible to antibiotics and host immune response. The formation of biofilms by P. aeruginosa has received considerable attention because it is a critical event in the devastating disease, cystic fibrosis (59,61). This same bacterium is found in most healthy individuals in whom it causes no disease. It is an opportunistic organism that only becomes pathogenic in compromised individuals. A brief account of biofilm formation by P. aeruginosa on the epithelial linings of the lungs of cystic fibrosis patients provides a concept of the process. Attachment factors are present on hairlike appendages of the bacteria called type IV pili (62). Those pili allow a twitching


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Table 2-5 Some Human Infections Involving Biofilm Formation Infection or disease

Common biofilm bacterial species

Dental caries Periodontitis Biliary tract infection Osteomyelitis Bacterial prostatitis Native valve endocarditis

Gram-positive cocci (e.g., Streptococcus) Gram-negative anaerobic oral bacteria Enteric bacteria (e.g., E. coli) Several bacterial and fungal species E. coli and other Gram-negative bacteria Viridans group Streptococci

Nosocomial infections ICU Pneumonia Sutures Contact lens Urinary catheter cystitis Hickman catheters Vascular grafts Biliary stent blockage Orthopedic devices

Gram-negative rods Staphylococcus epidermidis and S. aureus P. aeruginosa and Gram-positive cocci E. coli and other Gram-negative rods S. epidermidis and C. albicans Gram-positive cocci Various enteric bacteria and fungi S. aureus and S. epidermidis


motion that aids the bacteria in assembling into colonies. P. aeruginosa utilizes a type III secretion system to secrete toxin into epithelial cells, which among other effects interferes with ciliary sweeping thus further aiding microcolony formation. The attached bacteria proliferate, and as their number reaches a certain minimum density, sets of genes become activated the products of which include both virulence factors (e.g., toxin A, exoenzyme S) and other substances that promote bacterial cell wall remodeling and glycocalyx formation. In the case of P. aeruginosa, the principal glycocalyx material is the polysaccharide, alginate. At this point, the bacteria embedded in the glycocalyx are protected from antibodies, cell-mediated immunity, and antibiotics. Antigens are released by bacteria in the biofilm as well as planktonic forms and high levels of immunity may prevail in the host, but to no avail. As noted in the preceding paragraph, when bacteria in the developing colony reach a certain number (density) activation of a new set of genes occurs. This event reflects the recognition of a threshold concentration of organisms termed “quorum sensing.” It was first noted in bioluminescent marine bacteria in which the intensity of luminescence increased dramatically at a certain bacterial density. What is the mechanism of quorum sensing? There appear to be several mechanisms employed by different species of bacteria as well as certain fungi

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and possibly protozoa. The best understood mechanism, at present, operates in several Gram-negative human pathogens and has been analyzed extensively, again utilizing P. aeruginosa (61,63). As the density of the proliferating bacteria increases, so does the local concentration of small molecules that the bacteria synthesize known as acylhomoserine lactones (acyl-HSL) [e.g., N-(3oxododecanoyl)-L-homoserine lactone (3OC12-HSL) and N-butyrylL-homoserine lactone (but-HSL)]. The acyl-HSL molecules are the quorumsensing signals. Two quorum-sensing systems have been identified in P. aeruginosa: one is the lasR-lasI system and the other the rhlI-rhlR system. The product of the lasI gene directs the synthesis of 3OC12-HSL and the product of lasR, in the presence of a sufficient concentration of 3OC12-HSL, activates a set of virulence genes that includes rhlI and rhlR. The gene, rhlI, is responsible for a product that directs the synthesis of but-HSL, which is involved in the activation of virulence genes. Progress in understanding quorum sensing and the variety of systems that participate in the phenomenon has been rapid (see, e.g., ref. 64). Some biofilms comprise a single species of bacteria in which case the signals utilized by that species must be distinguished from those of other species. Other biofilms are composed of mixed species and in this case organisms may need to recognize several signals but obviously not all. Signal sensing among various species may help a given species, or group of related species, to avoid competition, ascertain an appropriate niche, or perhaps produce an antimicrobial substance to minimize or eliminate local competitors. From the perspective of bacterial infections in aging subjects, biofilms would appear to be of great importance. First and perhaps foremost, biofilms render many pathogens safe from antibiotics and immune attack. Second, it is likely that biofilm formation by various bacteria that are nonpathogenic in healthy, young adults may lead to serious infections in immunocompromised elderly or those already afflicted with some disorder. Third, the widespread use of urinary catheters, the high prevalence of prostatic disease among elderly males, and the frequency of bone and joint repair and replacement in the elderly offer to microbial pathogens a range of opportunities for clinical biofilm formation. Finally, it seems important to stress that biofilms is a subject that has received very little attention in relation to the susceptibility of the elderly to infections. Are conditions in aging tissues more or less favorable for the formation of biofilms? Or unchanged? Consider the mounting evidence that changes in the cytoplasmic membranes accompany senescence of various cells; does such a change occur in epithelial cells and might that influence bacterial adherence and biofilm formation? Do senescing tissues offer more and better-sheltered niches for bacterial colonization? An opportunity seems to exist for research that could have quite significant consequences.


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ANTIBIOTIC RESISTANCE AND BACTERIAL VARIATION Resistance to antibiotics is increasing rapidly among human pathogens as pointed out by many authors (e.g., refs. 65 and 66). The reasons center around one problem: the failure to control the human use of antibiotics. Numerous studies have shown that in medicine antibiotics are frequently prescribed unnecessarily or inappropriately. For example, it was estimated that in 1992, 12 million adults who presented bronchitis or upper respiratory infections received prescriptions for antibiotics that offered little or no benefit (67). Similar studies of inappropriate antibiotics usage have focused on Canada, Europe, and Japan. In developing countries, antibiotics usage has been poorly regulated, patient compliance has been poorly monitored, and much of the supply of antibiotics is of low quality. The common use of antibiotics in veterinary medicine and in agriculture has contributed to the problem to an extent that is difficult to determine but likely to be considerable. Geriatric medicine has certainly contributed to the growing problem of microbial antibiotic resistance. As a group, elderly patients in hospitals and long-term care facilities (LTCFs) are the major recipients of antibiotics. It has been estimated that among residents of LTCFs approx 40% of prescribed drugs for systemic use are antibiotics (68). A distressing result is the promotion and spread of antibiotic-resistant microorganisms (69). The types and origins of antibiotic-resistant pathogens in LTCFs have been carefully reviewed in a recent publication (70). Information about antibiotic resistance in the population of elderly who reside in LTCFs is probably the most reliable available. As stated in the review article (70), the antibiotic resistant bacteria of greatest concern to geriatricians are: 1) β-lactam resistant organisms, especially penicillin-resistant pneumococci and aerobic Gram-negative bacilli resistant to third-generation cephalosporins; 2) vancomycin-resistant enterococci; and 3) quinolone-resistant Gram-negative and Gram-positive bacteria. As recently as 1997, it was reported that more than one-third of S. pneumoniae isolates analyzed in broad survey were resistant to penicillin and more than 13% of them were highly resistant (71). Relatively resistant isolates have been found in many regions of the world. Altered penicillin-binding proteins (enzymes involved in the final stage of bacterial cell wall formation), which have low affinity for penicillin, are responsible for resistance to that antibiotic. Other organisms noted for β-lactam antibiotic resistance are certain species of Staphylococcus (some resistant to all penicillins, cephalosporins, and carbapenems) and Enterococci, which are resistant to all cephalosporins because they lack significant penicillin-binding proteins. Enzymes known as β-lactamases are largely responsible for the ability of bacteria to resist the cephalosporins. Considerable effort to produce enzyme-

40


resistant derivatives of those antibiotics has been expended leading to so-called third-generation cephalosporins that showed considerable promise. However, a number of enzymes have now been reported (called “extended spectrum βlactamases”) that can cleave advanced-design cephalosporins and penicillins. Many of the extended spectrum β-lactamases (ESBLs) are encoded by genes borne on plasmids such as those encoding TEM-1 associated with enteric bacteria. TEM-1 is responsible for close to three-fourths of plasmid-borne β-lactamase resistance world wide (72). The TEM group of enzymes is encoded by the TnA transposon, which probably accounts for the β-lactamases present in more than one-third of H. influenzae isolates in the United States. Several pathogenic Gram-negative bacilli (Enterobacter, Citrobacter, Pseudomonas) produce βlactamases that are encoded by a chromosomal gene. Those AmpC enzymes are able to inactivate all cephalosporins. Several AmpC β-lactamases are now known to be conveyed by mobile, conjugative plasmids in E. coli and in Klebsiella species. The presence of those enzymes can result in resistance to penicillins, cephalosporins, cephamycins, and β-lactamase inhibitors (73). It should be noted that resistance to β-lactam antibiotics can occur as a result of restricted entry of the antibiotics as well as low binding affinity to the penicillin-binding proteins and destruction by β-lactamases. Drug efflux pumps, which restrict antibiotic entrance into bacteria, have become a major problem in antibiotic therapy. They result in inadequate accumulation of antibiotics inside bacterial cells to be effective. The formation of transport proteins, which bind and inactivate antibiotics or escort them out of the bacterial cell, prevents the antibiotics from reaching critical targets. Some of these efflux pumps are drug-specific such as Tet B in enteric bacteria and H. influenzae; others act in a broad, “multidrug” pattern. Tet B is plasmid encoded, although chromosomally mediated tetracycline resistance occurs in some bacteria such as Proteus. Tetracycline resistance conveyed by plasmids is situated near insertion sites and as a consequence those plasmids rather readily acquire other genetic information, which results in broadening the specificity of the resistance. It is likely that the widespread use of tetracyclines in animal feed is responsible, in part, for the existence in many regions of the world of resistant Enterobacteriaceae. The quinolone antibiotics such as nalidixic acid and ciprofloxacin bind to DNA gyrase (type II topoisomerase) in Gram-negative bacteria, and to topoisomerase IV in Gram-positive bacteria thus interfering with DNA replication. Some of the later quinolones have other actions in addition to their effects on topoisomerases and display a broad spectrum of antimicrobial activity (74). Resistance to these antibiotics may emerge as a result of mutations in bacterial topoisomerases, diminished membrane penetrability in Gram-negative bacteria, or active efflux transporter proteins. Such changes are generally


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caused by alterations in the chromosomal DNA such as point mutations in the A subunit of DNA gyrase (74). Several species of Staphylococci and Enterococci are notorious for being nosocomial infections. The antibiotic, vancomycin, has been widely used to combat these infections because for many years no resistance to this substance was reported. Resistance of entercocci was first reported in 1986, which resulted in considerable effort to elucidate the causes (75,76). Vancomycin is a glycopeptide that interferes with cell wall formation of Gram-negative bacteria. It interacts with D-alanine at the C-terminus of precursors of peptidoglycans. This creates a complex from which the precursor substances cannot be transferred by transglycosidases to the growing peptidoglycan cell wall. Resistance to vancomycin appears as a consequence of expression in bacteria of transposable genes, which encode cell-wall-synthesizing enzymes that alter the C-terminus of the peptidoglycan precursors from D-alanine to D-lactate. This change allows cell wall construction to continue even in the presence of vancomycin. There are four known phenotypes of vancomycin-resistant enterococci of which two (Van A and Van B) are associated with moderate to high resistance to the antibiotic (70). Both of those phenotypes are readily transferred on plasmid and transposon elements. The Van B phenotype is also associated with a chromosomal complex that closely resembles the organization of the Van A transposable element. The importance of vancomycin resistance to the geriatrician is the common usage of that antibiotic to treat UTIs and infected pressure ulcers, which are relatively common in LTCFs (70). Vancomycin-resistant enterocci are introduced most often into LTCFs by accepting patients who have acquired resistant organisms in hospitals. To complete this brief discussion of antibiotic resistance, a word about transposons is in order. Those genetic elements, conveyed by resistance plasmids (R-plasmids), are responsible for much of the current microbial resistance to antibiotics. There are collections of bacteria, assembled in the preantibiotic era, that display recognizable plasmids; but most of those plasmids lack antibiotic-resistance elements. This must mean that current bacterial pathogens displaying antibiotic resistance harbor familiar plasmids that have become R-plasmids by acquiring resistance transposons. That is a consequence of the excessive use of antibiotics, which has given selective advantage to bacteria possessing R-plasmids. Among the latter, those that convey multiple antibiotic resistance vary with respect to the transposons they contain. Some have a single transposon composed of multiple resistance determinants. Some have several transposons located in different sites. In some cases, there is present a complex element in which one transposon has become integrated into another. Apparently, there has been no effort to evaluate the R-plasmids present in bacterial

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isolates from older persons, especially those residing in LTCFs, to determine whether or not the plasmids and the resistance transposons differ from isolates obtained from young adults. If a significant difference exists, such information should be useful in planning judicious antibiotic therapy. The extreme variability, versatility, and adaptability of bacteria arise from two processes that are not found in eukaryotic organisms: (a) horizontal (lateral) transfer of genetic material (77); and (b) hypermutability associated with “mutator” strains (78,79). There are three mechanisms for delivering exogenous DNA into recipient bacteria: (a) transformation, which involves bacterial uptake of naked, ambient DNA; (b) transduction, in which new DNA is delivered by bacteriophages; and (c) conjugation, which requires physical contact between donor and recipient cells and, most frequently, transfer of a plasmid. Once inside the recipient cell, the DNA must become assimilated either as a stable episome or by integration into the recipient’s genome if it is to be expressed. Hypermutation, exemplified by mutator strains, appears to arise from mutations in genes that affect the synthesis, modification, or repair of DNA. A recent study (78) of the presence of mutator strains of P. aeruginosa isolates from cystic fibrosis patients has provided strong evidence for the concept that the emergence of mutator strains is a mechanism employed by bacteria for rapid adaptation to nonoptimal, even hostile conditions in their hosts. This concept (see comments in ref. 79) could be of major significance with regard to infections in the elderly in whom microenvironmental conditions in organs such as the lung, urinary bladder, and GI system may differ considerably from those in younger subjects. VIRAL INFECTIONS IN AGING HUMANS Viruses are small, composite particles of nucleic acid and protein, and are obligate, intracellular parasites; i.e., they cannot replicate outside a host cell. An individual virus contains only one type of nucleic acid, either RNA or DNA, which is protected by the associated protein from destruction by hostile substances, such as nucleases, present in its environment. Viral proteins serve two other crucial functions; first, they are responsible for attachment of virions to host cells and, second, they include a minimal array of enzymes that are necessary to cajole the host cell machinery into synthesizing new virions. Together, the nucleic acid and associated protein form the nucleocapsid. Some viruses are encased in a lipid bilayer, derived from host cell membrane, termed the “envelope.” It is often studded with outward-protruding, complex molecules of glycoprotein. Viral nucleic acid (genome) may be either RNA or DNA arranged in linear or circular fashion. The nucleic acid may occur in singlestranded or double-stranded form. If the genome is single-stranded RNA, it is considered to be in the positive (plus) sense orientation if it can serve as its


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own mRNA and in negative (minus) sense orientation if a copy must first be made by a viral RNA transcriptase, which then serves as mRNA. Most DNA viruses display a linear, double-stranded genome although a few families are characterized by linear single-stranded (parvoviruses), circular double-stranded (papovaviruses), or even circular single-stranded (circoviruses) DNA. Comprehensive introductions to viral genomes and virus replication are provided in many excellent texts (e.g., ref. 80 and 81). Viral infection begins with attachment of virions to host cells. Specific attachment that leads to virion penetration generally depends upon complementary interaction between viral protein (counter-receptor or anti-receptor) and specific receptors on host cells. Viruses may display a single species of protein counter-receptor or multiple species of counter-receptors in the case of some complex viruses such as herpes simplex. Whether or not a host cell is susceptible to a given virus depends upon the cell having receptors. Cells lacking receptors are not susceptible. If a host cell supports the complete reproduction of a given virus, it is termed “permissive.” Some host cells can be shown to be permissive but not susceptible because they lack the appropriate receptors. The cellular receptors for viruses are generally glycoproteins. Some of the receptors are familiar molecules known to be involved in other functions. Table 2-6 is a list of a few of those receptors. The ability of viruses to usurp surface molecules designed for some other purpose as receptors for themselves is well illustrated by human and simian immunodeficiency viruses. Those viruses utilize members of the chemokine superfamily (CXCR4, CCR5) along with CD4 as coreceptors for entrance into T cells and monocytes (reviewed in ref. 82). Chemokine receptors have been appropriated also by other viruses; e.g., myxoma virus can utilize CCR1, CCR5, CXCR4 for entrance into host cells (83). Myxoma virus is a poxvirus the receptors for which have been difficult to identify. The epidermal growth factor receptor is utilized by vaccinia virus another poxvirus. A poxvirus that has the human as primary host is the molluscum contagiosum virus (MCV), which causes persistent, benign, skin neoplasms in children and severe opportunistic infections in AIDS victims. Both children and AIDS victims are immunodeficient (to much different degrees, of course). Elderly individuals are immunodeficient. It seems natural, therefore, to wonder whether or not the elderly are also susceptible to MCV. If so, it has not been reported (to our knowledge). Perhaps factors other than immunodeficiency are involved in rendering subjects susceptible to MCV. It would be useful to know. The complexity of viruses is exemplified by their nomenclature, which comprises some 8 major families of DNA viruses and 14 families of RNA viruses (with more to come, no doubt). Of those 22 families, 20 include members that have medical importance (see ref. 84 for a concise overview). The first five of

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Table 2-6 Some Familiar Cell Membrane Receptors for Viruses Virus Adenovirus Epstein-Barr Herpes simplex Influenza A, B Respiratory syncytial Rhinoviruses

HIV-1, -2

Receptor Integrin (α5β3) Complement type 2 receptor (CD 21) Proteoglycans (heparin sulfate moieties) Glycoproteins of 5Ac Neu Hemagglutinin glycoprotein Intercellular adhesion molecule (ICAM) CD4, galactosyl ceramide, chemokine receptors

Representative important cell infected Respiratory epithelium B lymphocytes Oral and genital epithelium Oropharyngeal cells Respiratory epithelium Nasal epithelium

T lymphocytes

six families listed in Table 2-7 contains members that cause respiratory disorders any of which can progress to pneumonia in the elderly. The influenza viruses of family Orthomyxaviridae are of the greatest concern because the elderly are so susceptible and because each new flu season may bring an antigenic variant arising from the “drift” and “shift” in antigenic types that are so typical of influenza viruses. The far-right column of Table 2-7 is headed “Persistence.” Two families (four types) of viruses are listed to illustrate persistence. There are three types of persistence, termed “chronic” (diffuse or focal), “latent,” and “slow.” Here, we are interested in latent persistence. Latency refers to the fact that some viruses may integrate into the genomes of host cells where they persist for extended periods without replicating or killing host cells and without causing disease. An outstanding example of latency in a bacterial infection is that of the Mycobacteria, which cause tuberculosis. The four viruses listed in the table are representative of persistence. This phenomenon may be of unrecognized importance in aging humans. It should be stressed that (to our knowledge) there is no irrefutable evidence that persistent viruses afflict the elderly inordinately. However, it should be stressed equally that (to our knowledge) there have been no systematic studies of that possibility. In the following brief paragraph, evidence is marshalled to support the idea that more attention should be focused on assessing persistent viral infections in aging humans.


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Table 2-7 Significant Viruses of Aging Humans Family

Virus

Coronaviridae

Coronaviruses (two major types) Orthomyxoviridae Influenza (A,B,C) Paramyxoviridae Respiratory syncytial virus Picornoviridae Rhinovirus (>100 serotypes) Adenoviridae Adenovirus (numerous serotypes) gastroenteritis Herpesviridae Herpes simplex

Cytomegalovirus

Varicella-zoster Virus

Disorder

Persistence

Common cold (sinusitis) Influenza (pneumonia) Respiratory infections (upper and lower) Respiratory infections (common cold) Respiratory infections (colds, pneumonia) Gingivostomatitis, genital herpes, herpetic keratitis, encephalitis Mononucleosis (multiple organ infection in immunocompromised) Herpes zoster (shingles)

No No No No Yes

Yes

Yes

Yes

Consider, first, the adenoviruses. There are at least six subgenera and numerous serotypes of human adenoviruses. The principal targets of the viruses are the respiratory tract, ocular tissues and, less frequently, the GI system. The ability of a few types of human adenoviruses to induce tumors in hamsters and transform human and animal cell lines has attracted attention for many years although there is little evidence that they are oncogenic in humans. The adenoviruses present a classical example of latency. The viruses or their genomes are found in tonsils. Cells of the tonsils of individuals who have experienced infections but have been symptom-free for extended times may have whole or partial virus genomes integrated in their own genomes. It is uncertain how long the virus genomes may continue to replicate in individuals who remain symptom-free. Whether or not latent adenoviruses may be reactivated under certain conditions in aging subjects is a question that seems not to have been addressed. The establishment of latency generally involves integration of viral genomes into the host cell genome or occasionally an episome. Integration of adenovirus DNA has been demonstrated in transformed human cells and in virusinduced tumors in hamsters; and integrated viral DNA may persist for long

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periods in human tonsil cells. What restricts the viral replication in those cells, and the events or factors that trigger reactivation, are unknown. Clearly, this emergence from latency deserves careful study on the hypothesis that adenovirus and other viruses may be reactivated in the immunodeficient elderly and precipitate disease. Three of the viruses in the family of Herpesviridae that are well known for their latency are listed in Table 2-7. First, there is herpes simplex, which exists as two closely related types (HSV-1 and -2). The former is primarily responsible for gingivostomatitis in young children, the latter for genital herpes in adults. HSV-1 is the principal cause of focal, sporadic encephalitis, which in the United States occurs in approximately 1 in 150,000 population. Second is the cytomegalovirus (CMV), which when acquired congenitally (approx 1% of live births in the United States) causes severe disease in infants and young children. In adults and older children, CMV may cause a mononucleosis which resembles that caused by Epstein-Barr virus. The third herpes virus known for latency is varicella-zoster virus (VZV), the causative agent of chickenpox, which may occur in children or adults. Reactivation of VZV may produce herpes zoster (“shingles”), which appears in about 1% of individuals over age 50. HSV-1 and -2 infections occur preferentially at mucocutaneous sites. As the infection and accompanying inflammation progress, the viruses ascend peripheral sensory nerves to reach dorsal root ganglia. The viruses replicate in nervous tissue and then migrate in retrograde fashion along axons to reach other mucosal and epithelial surfaces thus spreading the infection. Latency is established in cells of the dorsal root ganglia. Herpes simplex encephalitis affects preferentially the temporal lobe of the brain and can be initiated by reactivated viruses in addition to viruses of the primary infection. Primary CMV infections occur most efficiently in salivary glands and kidneys. Persistent infections are found in those tissues and in breast, endocervix, seminal vesicle tissues, and peripheral blood leukocytes. Patients with deficient immune systems, such as bone marrow transplant recipients and those with immunodeficiency diseases, are at risk of primary or reactivated CMV infections. In those patients, infection may involve the lungs, GI system, liver, and other organs/tissues, and often becomes life-threatening. It would be interesting, and probably quite significant, to determine whether, and how frequently, CMV-induced respiratory and GI disorders occur in the aging population as a consequence of reactivation of host cell-integrated viral genomes. Similar to HSV-1 and -2, VZV assumes latency in the dorsal root ganglion. Herpes zoster appears as a result of reactivation of latent virus. An important fact about herpes zoster stands out; viz., acute neuritis is characteristic in most patients whereas the frequency of postherpetic neuralgia occurs in about half of the adults, but not in juveniles, and the frequency seems to increase in older


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patients. As noted above, herpes zoster occurs with a frequency of about 1% in adults over 50. All four of the described latent viral infections are serious problems in immunocompromised individuals in whom multiple organ sites are involved. It is well established that aging humans (and laboratory animals) are deficient in one or more aspects of immunity and that T-lymphocyte-dependent antiviral immunity is one such aspect (Chapters 3 and 4). We are not so foolish as to suggest that an elderly subject is similar to an AIDS victim or an individual under treatment with an immunosuppressive drug; however, we do suggest that lessons learned from those patients may be applicable to the elderly. Under conditions of good health and environmental circumstances, most elderly persons retain sufficient immunological potential to cope effectively with acute infections. However, when the immune potential is further reduced by illness, injury, stress, or severe xenobiotic (pollutant) exposure, many elderly subjects may become vulnerable to microbial pathogens. Those are precisely the insults and injuries that are known to activate latent viruses. The need for effective, safe antiviral drugs will continue with increasing urgency in the years ahead. One reason it has been difficult to find or develop antivirals is because viruses utilize so much of the host cell machinery for their own fabrication. Another reason is the extreme ingenuity displayed by viruses to defend and protect themselves as shown in Table 2-8 (85). A new, promising direction toward antivirals is that of interfering with, or redirecting, viral association with cellular receptors and is based on detailed structural knowledge. For example, the counter-receptor site (“knob domain”) in association with the binding domain of the cognate receptor (the Coxsackie and adenovirus receptor, or CAR) has been crystallized and analyzed to the 2.6 A resolution level (86). Whether or not the extensive viral use of receptors involved in key host cell functions will allow the development of discriminating antivirals remains to be discovered. PROTOZOAN PARASITES IN AGING SUBJECTS There are no reliable data on the relative susceptibility of aging humans to animal parasites (i.e., parasites other than microbial) or on the relative severity of parasitic infections in aged compared to young or middle-aged subjects. The principal reasons for this dearth of information are (a) parasitic infections are largely restricted to tropical climates in underdeveloped regions of the world where public health records are limited, and (b) where parasites abound, the majority of the population carries chronic infections acquired in childhood or young adulthood. Similarly, there are few data concerned with parasitic infections relative to age in natural animal populations. An example of the few published studies in animals reported an analysis of cattle infected with the parasite,

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Table 2-8 Intracellular Defense Strategies Used by Viruses Type of Virus Epstein-Barr

Host cell antivirus mechanism Apoptosis (cell death)

Virus counter strategies Homologs of bcl-2

Rabbit pox

Serpins

Simian virus 40

p 53 binding protein

Herpes virus

Intracellular signaling

Myxoma virus

Tyrosine kinase modulation Receptor mimicry

Adenovirus Cytomegalovirus

Viral antigen presentation

MHC Class I suppression

Molluscum contagiosum

Oxidative stress response

Antioxidant selenoprotein


Onchocerca ochengi (87). Those cattle lived in an area of high endemicity in the Cameroon and 71% of those studied were infected. Although there was no difference in the prevalence of infection among the three age groups studied (1.5–2.5 years, 3–5 years, >8 years of age), the parasite burden (“worm load”) was significantly greater in the group >8 years of age. In contrast, there was a significantly lower number of the immature forms (microfilariae) in the older compared to the younger cattle. Whether or not this latter finding reflected more effective immunity or some other, inimical physiological change with age could not be determined. It is necessary, therefore, to extrapolate from experimental studies in laboratory animals to gain insight concerning the abilities of elderly humans to cope with parasites. The earliest study (of which we are aware) was of infections of rats of different ages with the nematode, Trichinella spiralis (88). The data suggested that the severity of infection (parasite burden) was significantly greater in the oldest animals. Apparently, there have been no other studies with parasites other than protozoa. The work of Gardner and Remington has shown clearly that aged mice develop significantly worse infections with Toxoplasma gondii than do younger mice (89,90). T. gondii is not a natural human parasite but can infect normal infants and young children in whom it may cause serious central nervous system disorders. T. gondii is one of the major opportunistic, protozoan infections in AIDS victims. The susceptibility of aged mice was shown to be, in part, the


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result of decreased antibody production against the parasite in both the acute and chronic phases of the infection. However, of greater importance was the finding of a pronounced difference in the activation of macrophages of young and aged mice during the acute phase of infection. T. gondii is an intracellular parasite and, therefore, immunological resistance is primarily a cell-mediated process. The depressed activity of macrophages, which play key roles in natural/innate resistance and immunity to intracellular parasites, was considered responsible for the heightened infections found in aged mice. The effects of senescence on macrophages are discussed in Chapters 3 and 4. However, T cells (both CD4+ and CD8+) play roles in immunity to T. gondii (91) and those cells are significantly altered by senescence, as is discussed later. The protozoan, Trypanosoma musculi, is a natural parasite of mice. It infects all of a number of inbred strains of mice; however, the severity of infection, judged by the parasite burden, varies over a 20-fold range (approximately) among different strains (92). Regardless of the strain, however, aged mice develop significantly worse infections. This is illustrated in Figure 2-1, where the course of infection in young and aged mice of strain A is depicted. T. musculi organisms live extracellularly in the bloodstream of mice and the parasite burden can be assessed by determining the level of parasitemia, i.e., counting the numbers of parasites in blood samples. T. musculi infections are self-limiting; i.e., after a prolonged period of about 3 weeks in young adult mice the infections terminate. Thereafter the cured mice are permanently immune to reinfection. As Figure 2-1 shows, both the parasite burden (parasitemia) and the duration of infection (time before the cure) are markedly extended in aged compared to young adult mice. To demonstrate that the elevated parasitemia in aged mice was a reflection of a deficient immune response, the technique of adoptive conferral (“transfer”) of immunity was employed. The conferral of immunity to T. musculi on irradiated, immunologically incompetent mice by the transfer of a predetermined, optimum number of spleen cells from normal, infected, or cured donor mice was evaluated. After receiving the donor spleen cells, the irradiated recipient mice were inoculated with viable T. musculi and the course of infection monitored. The results of such a study, in which equivalent numbers of spleen cells were transferred from young or aged infected donor mice into irradiated young-adult recipients, are depicted in Figure 2-2. Irradiated mice, lacking a competent immune system, that were inoculated with T. musculi but given no donor spleen cells died from overwhelming T. musculi infection (Fig. 22A).The transfer of spleen cells from young donor mice on day 7 of their infection was able to protect irradiated recipients and cure their infection in about three weeks (Fig. 2-2B). In contrast, the same number of spleen cells from aged donors on day 7 of infection conferred no protection on the irradiated

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recipients (Fig. 2-2B). On day 14 of infection, spleen cells from both young and aged donor mice were able to protect irradiated, young recipients from lethal T. musculi infection. However, the cells from young donors were much more efficient than those from aged donors as shown by the marked differences in levels of parasitemia and duration of infection in the recipient mice (Fig. 2-2C). Finally 21 days after initial infection, cells from aged donors were able to protect aged recipients but only after a prolonged infection (Fig. 2-2D). The two preceding examples of the relative inability of aged mice to cope with protozoan infections provide compelling evidence that senescence cripples the immune system. In both cases, there is considerable understanding of the nature of the immune response against the parasites in young adults as is discussed in Chapters 3 and 4. It should be stressed here that the two parasites, T. gondii and T. musculi, are quite different in their life cycles and in other aspects. T. gondii are intracellular parasites whereas T. musculi are extracellular. Immunity to T. gondii is a cell-mediated process whereas immunity to T. musculi is dependent on specific antibodies, probably of IgG2a isotype (mouse) (93). T. gondii will establish infection in several hosts (cats, mice, humans) whereas T. musculi is strictly a mouse-specific parasite. Considered together, studies of these two protozoa suggest that the ability of aged mice to generate both humoral and cell-mediated immunity to pathogens is impaired. FUNGAL INFECTIONS IN AGING SUBJECTS There have been few attempts to evaluate the frequency or severity of fungal infections in aging subjects. On the other hand, it is well-established that fungal infections are rather common in other immunodeficient individuals such as AIDS victims, persons being treated with immunosuppressive drugs, patients on antibiotic therapy or suffering from burns, diabetes, or malnutrition (94). One study in particular, strongly indicates that more attention should be given to fungal infections in the aging (95). In that study, the frequency of mortality as a consequence of systemic infections with bacteria (bacteremia) or fungi (fungemia) was assessed from the medical records of 500 patients identified as having true bacteremia or fungemia. The parameters relevant to the present discussion that were considered included: 1) mortality associated with both bacteremia and fungemia; 2) the primary site of the infection; 3) body temperature; and 4) the degree of leukopenia. There was a substantial increase in the risk of death of subjects over age 50 and deaths were more frequent in males than females. The risk of death was significantly greater when the primary site of infection was a surgical wound, a burn or even untraumatized skin, an abscess, or the respiratory tract compared to other sites. There was a markedly greater (about fivefold) frequency of mortality of patients whose body temperature was less than 36°C compared to those whose temperature was over 40°C. A peripheral leukocyte count


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Fig. 2-1. Course of parasitemia in young (䊉) and aged (䊊) A/He mice following inoculation with T. musculi. Four or five samples per point. Bars represent 1 S.E.M. (From Albright JW, Albright JF. Mech Ageing Dev 1982;20:315–330.)

of less than 4000/µL, or a granulocyte count of less than 1000/µL, both correlated with substantially higher mortality. Given that (a) injury and trauma (i.e., stress) significantly alter immune responses in the elderly (see later), (b) the skin and respiratory systems of the elderly are sites of common fungi that are benign in younger individuals (e.g., ref. 96), and (c) that elderly humans are less disposed to run fevers (97), it is difficult to avoid the conclusion that fungal infections are significant problems in the elderly. Emerging, opportunistic fungi (98) and drug-resistant fungi (99) will begin to compound the problems for the elderly in the near future. CHAPTER SUMMARY There are no known microorganisms that uniquely infect elderly humans. Overt diseases caused by some pathogens (e.g., tuberculosis, pneumonia, influenza, UTIs, sepsis) are clearly more common in the elderly. The reasons

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Fig. 2-2. Course of parasitemia in irradiated A/He recipients of 3 × 107 spleen cells from : (A) uninfected donors; (B) donors on day 7 after trypanosome inoculation (*indicates parasites at this time largely transferred as contaminants of donor spleen cell preparations); (C) donors on day 14 after trypanosome inoculation; (D) donors on day 21 after trypanosome inoculation. Donor-recipient combinations: (䊊) young donor-young recipients; (䊉) young donors-aged recipients; (䊐) aged donors-young recipients; (䊏) aged donors-aged recipients. Data from one of two replicated experiments, 5–6 samples per point. Bars represent 1 SEM. (D means all were dead). (From Albright JW, Albright JF. Mech Ageing Dev 1982;20:315–330.)

for this include (a) functional and anatomical changes in organs (e.g., respiratory, urinary), (b) comorbidity, and (c) decline in immunological competence. With regard to the latter, research with experimental animals has provided compelling evidence that immunological resistance to infection declines with age. Such is the case of infections with M. tuberculosis, S. typhimurium, T. gondii, T. musculi, and probably L. monocytogenes. Bacterial infections begin with attachment of the microorganisms to host cells (epithelial, endothelial) followed by secretion of toxins and other virulence factors both outside and inside (type III secretion) host cells. Attachment involves a number of well-known cell adhesion receptors and counter-receptors as well as some that are restricted to microorganisms. In some cases, bacteria may secrete their own receptors into host cells. In order to survive in the hostile environment of the host, microorganisms often engage in colonial orga-


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nization. To form organized colonies, called biofilms, bacteria reproduce rapidly and upon reaching a certain density, lay down a matrix that protects them from host defenses, including both humoral and cell-mediated immunity. Those bacteria that can form biofilms (many, perhaps most, species can) are able to sense population density and to distinguish between self and other species, an ability termed quorum sensing. The formation of biofilms also renders colonized bacteria impervious to antibiotics. Is it possible that biofilm formation by bacteria (or fungi that also appear capable) that have been selected for resistance in the elderly is a major factor in the elevated morbidity and mortality that characterize the aging? At present, very little is known about bacterial adherence and the efficiency of secretion of virulence factors into host cells of aging subjects. There are limited data that suggest that intercellular adhesion is altered in aging humans, perhaps as a consequence of the effect of senescence on cytoplasmic membranes. If so, it is likely that changes occur in bacterial adherence to host cells. Similarly, very little attention has been given to date to evalulating the relationships between susceptibility of aging humans to infection and the formation of biofilms in aging subjects. Both persistent (latent) bacterial and persistent viral infections in the elderly require attention. Tuberculosis appearing as a result of reactivated Mycobacteria is a classical example of bacterial latency. It is probable that bacterial latency is much more important in the pathogenesis of infectious diseases than has been realized (see ref. 100). Herpes zoster (shingles) is one of several well-known examples of viral latency that result in diseases in the elderly. It is likely that the decline in immunological potential contributes in a major way to the reactivation of latent infections in aging subjects. However, this has not been afforded rigorous proof. If it is true, it becomes important to ask which of numerous other latent infections reappear in the elderly. Has sufficient attention been given to this question? To what extent are the well-known opportunistic infections in immunocompromised individuals paralleled in less severe degree in the elderly? Has this question received adequate attention? Finally, it should be pointed out that both parasitic (especially protozoan) and fungal infections in the elderly deserve much more concern than they have received to date. Both environmental pollution and global warming are likely to precipitate a significant increase in the prevalence of infections by those organisms in the years ahead. And in the case of both protozoa (e.g., malaria) and fungi (e.g., Candida spp.) drug-resistant forms are already with us. REFERENCES 1. Yoshikawa TT. “Perspective”: Aging and infectious diseases; past, present and future. J Infect Dis 1997;176:1053–1057.

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3 Senescence of Natural/Innate Resistance to Infection If an experiment does not hold out the possibility of causing one to revise one’s views, it is hard to see why it should be done at all. —Peter Medawar, Advice to a Young Scientist

It has been realized for decades that in humans and higher vertebrates there are, in addition to the adaptive immune system, accessory mechanisms and systems that contribute to overall immune defense against infectious organisms. They have been grouped under the heading “natural” or “innate” immunity. They include the reticuloendothelial system comprising “fixed” and “mobile” (circulating) monocytes (Mo’s) and macrophages (MPs), polymorphonuclear (PMN) cells (especially neutrophils and eosinophils that can discharge antimicrobial peptides such as defensins), natural killer (NK) and other “naturally cytotoxic” (NC) cells, and the complement (C) system; all of which have been preserved and handed down during the evolution of higher vertebrates from primitive vertebrates and invertebrates. The significance and importance of the innate system (sometimes called the “constitutive system”) and its complementary relationship to the adaptive (acquired) immune system have been clearly recognized only in the last decade (1,2). In the vertebrates, it is the job of the rapid-response, innate system to prevent infections from overwhelming the host before the more powerful, but more slowly developing, adaptive response is manifested. In addition, the innate response facilitates the development of the adaptive response by way of C components, cytokines and chemokines, and overlapping receptors and signaling pathways.

From: Aging, Immunity, and Infection By J. F. Albright and J. W. Albright © Humana Press Inc., Totowa, NJ

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PATTERN RECOGNIZING RECEPTORS OF INNATE IMMUNITY Certain conceptual difficulties that arose with the recognition of the roles played by the innate system in vertebrates are framed by the following questions: (a) how does the innate system, which possesses limited recognitive diversity, detect microbial substances and distinguish them from self? and (b) how can a sufficient number of reactive host cells be mobilized in a very short time (roughly, 12 h or less)? The answer to the first question lies in the use of pattern-recognizing receptors (PRRs) by the innate system to recognize microbial constituents. Those constituents comprise conserved structural cores that cannot be significantly altered by mutations without destroying the pathogenicity of the microorganism. Those constituents that form common patterns on microbes include lipopolysaccharides of Gram-negative bacteria, lipoteichoic acids of Gram-positive bacteria, lipoproteins of bacteria and parasites, glycolipids of mycobacteria, mannans of yeast, and double-stranded RNAs of viruses. It is probable that “receptors for these structures have been selected over evolutionary time to provide broad-spectrum recognition of harmful foreign materials” (1). The answer to the second question also invokes the use of PRRs. The latter are not clonally restricted; that would limit their distribution to relatively few cells. Rather they are likely to be present on the majority of certain types of cells. If all the cells that possess a given, broad-spectrum receptor can be activated into effectors quickly, rapid control of a pathogenic infection can occur. That response to infection will be quite different from the adaptive response, which begins with the activation of a small number of lymphocytes of a clone displaying cognate receptors for a precise epitope of a pathogen. The ensuing development of a controlling response will be delayed by the time required for proliferative expansion and maturation of the effector cells. There are several receptors of broad specificity for common microbial components. Four that have been most thoroughly studied are: (a) mannose-binding protein (MBP), (b) the mannose receptor (MR), (c) a group of homologous proteins termed Toll-like receptors (TLRs), and (d) a family of related proteins known as scavenger receptors (SRs). Mannose-Binding Protein (3,4) MBP is produced by the liver and released as an acute phase reactant. It belongs to a family of related proteins termed “collectins”; other members include lung surfactant proteins, SP-A and SP-D, bovine conglutinin, and bovine collectin-43. The collectins exhibit a broad range of target binding. For example, they associate with Gram-negative bacteria, yeasts, Pneumocystis carinii, influenza, and HIV viruses. SP-A and SP-D interact with airborne particulate material including pollen grains. The common feature of the target substances is their accessible carbohydrates (oligosaccharides).

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Structural information about MBP is informative with regard to pattern recognition. The seemingly broad range of target specificity does not include the carbohydrates associated with self-glycoproteins. The explanation for this important restriction lies in the particular spatial orientation of the hydroxyl groups situated at positions 3 and 4 of the hexose moiety (e.g., N-acetylglucosamine, glucose, fucose, mannose) common to many microorganisms (5,6). In comparison, the hydroxyl groups of those sugars commonly found at the termini of mammalian glycoproteins (viz., sialic acid and galactose ) are not recognized by MBP. Additional information about the structure of MBP aids in understanding its function of eliminating pathogenic organisms (7). It is particularly important that the assembled polymers of MBP molecules can activate the C system thus promoting the destruction of pathogens. This is a consequence of the structural similarity of MBP to the first component, C1q, of the C system. The individual molecules of MBP are trimers composed of a collagen domain, a neck region, and a globular carboxy-terminal, C-type lectin-binding domain. The three polypeptide chains in the neck region are properly aligned and trimerized as a result of interactions between three parallel α-helical coiled coils (8,9). Molecules of MBP, like those of SP-A, assemble as hexamers of trimers and resemble a “bunch of tulips” (10,11) (Fig. 3-1). Until recently, it was believed that MBP is able to interact with the two serine proteases, C1r and C1s, and thus to activate the classical complement pathway through C4 (12). It is now clear that two novel serine proteases, MASP-1 and MASP-2 (i.e., “mannan-binding-lectin-associated serine protease”), act similarly to C1r and C1s to cleave molecules of C4 and C2 and to effect cleavage of the pivotal component C3; or a complex of MBL-MASP-1 and another factor, Map 19, may act directly to activate C3 (7). In addition to its ability to effect complement activation, MBP can directly mediate phagocytosis of microorganisms (13), i.e., serve as an opsonin not involving complement components. Presumably, this is achieved by way of an MBP receptor that has not been clearly identified but may be complement receptor 1 (CR 1). Both SP-A and SP-D are surfactants and are particularly important in defense against infections in the lungs. Both interact with Gram-negative bacteria. SP-A has been shown to promote phagocytosis in the aveoli whereas SPD probably does not (14). Mannose Receptor The MR facilitates the phagocytic and endocytic ingestion of both particulate and soluble glycoconjugates. It comprises eight C-type, CRDs through which glylcoconjugates are bound providing they display accessible mannose, fucose, or N-acetylglucosamine residues (15) (see Table 3-1). Only one of the

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Fig. 3-1. Superimposed molecules of human (black) and rat (white) trimeric mannose binding proteins (MBPs). (A) View down the threefold axis of superimposed α-helical coiled coils of human and rat trimers. (B) Sideview of (a) rotated 90° showing resemblance to a “bunch of tulips.” (From Epstein J et al. Curr Opin Immunol 1996;8:29–35.) Table 3-1 Binding of Mannose Receptor Fragments Expressed In Vitro to Immobilized Monosaccharides Relative bindingb

Immobilized Monosaccharidea Man Fuc Glc Nac Glc Gal

CRDs 1–3

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