Discussion
From Hayflick to Walford: the role of T cell replicative senescence in human aging

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Abstract

The immunologic theory of aging, proposed more than 40 years ago by Roy Walford, suggests that the normal process of aging in man and in animals is pathogenetically related to faulty immunological processes. Since that time, research on immunological aging has undergone extraordinary expansion, leading to new information in areas spanning from molecular biology and cell signaling to large-scale clinical studies. Investigation in this area has also provided unexpected insights into HIV disease, many aspects of which represent accelerated immunological aging. This article describes the initial insights and vision of Roy Walford into one particular facet of human immunological aging, namely, the potential relevance of the well-studied human fibroblast replicative senescence model, initially developed by Leonard Hayflick, to cells of the immune system. Extensive research on T cell senescence in cell culture has now documented changes in vitro that closely mirror alterations occurring during in vivo aging in humans, underscoring the biological significance of T cell replicative senescence. Moreover, the inclusion of high proportions of putatively senescent T cells in the ‘immune risk phenotype’ that is associated with early mortality in octogenarians provides initial clinical confirmation of both the immunologic theory of aging and the role of the T cell Hayflick Limit in human aging, two areas of gerontological research pioneered by Roy Walford.

Introduction

Whereas Roy Walford is best known to the gerontology community for his work on caloric restriction, his scientific contribution is arguably equally significant in the area of immunological aging. Indeed, his 1961 NIH grant, entitled, ‘The role of immune phenomena in the aging process’, continuously funded and renewed through 1997, formed the foundation for the highly productive field of immunogerontology, a discipline that has produced an impressive array of new and unexpected insights into immunological aging. Some of the basic tenets that formed the basis for Roy's thinking in this area were first described in his 1969 book, now a classic, entitled, ‘The Immunologic Theory of Aging’ (Walford, 1969), which posits that the normal process of aging in man and all animals is pathogenetically related to faulty immunological processes. Research over nearly 50 years since Roy submitted his 1961 NIH proposal has confirmed the basic framework of his prophetic vision regarding the central role of the immune system in aging. Indeed, in humans, numerous clinical studies show significant correlations between specific immune functional traits and early mortality, irrespective of the cause of death (Wikby et al., 1998, Wayne et al., 1990). Such correlations, while not proving causality, are consistent with the potential influence of the aging immune system on basic physiological processes that affect lifespan. Interestingly, even in C. elegans, certain mutants that confer increased longevity show increased resistance to pathogenic bacterial infection (Garsin et al., 2003), suggesting an evolutionarily-conserved link between immunity and lifespan.

One of the unique and far-sighted contributions made by Roy within the general field of immune system aging was his adaptation of the well-studied fibroblast replicative senescence model system to human T cells. Indeed, it was the confluence of my own background in T cell immunity to viruses with Roy's interest in this novel aspect of human immunosenescence that led to my joining his research group within the Department of Pathology and Laboratory Medicine at UCLA in 1981. At the time, like most immunologists, I was unfamiliar with research on fibroblast replicative senescence, despite its prominent role within the field gerontology. Twenty years and many experiments later, it has become clear that replicative senescence is highly relevant to the human immune system. Indeed, a large body of research suggests that the immune system may constitute one of the clearest examples of the impact of the Hayflick Limit on human aging.

It is instructive to review the process by which Roy first became interested in the possible role of replicative senescence within the immune system, since it illustrates his characteristically broad, creative approach to science in general, and his unique ability to see the big picture. During the early 1980s, Roy made the astute observation that within the scientific community, there existed two major groups of investigators—immunologists and cell biologists—whose views on an important aspect of cell behavior with important ramifications for biology were mutually incompatible. Each group appeared to be almost totally unaware of the other, in that for at least 7 years both disciplines developed in parallel with very little manifest cognizance of a major conflict of theoretical tenets (Effros and Walford, 1984). The field of replicative (or cellular) senescence was based on the classic studies of Hayflick (Hayflick, 1965) and Hayflick and Moohead (Hayflick and Moohead, 1961), which had suggested that normal vertebrate diploid cells cultured in vitro invariably undergo a finite and fairly predictable number of cell divisions. Numerous theoretical explanations for this so-called ‘Hayflick Limit’ and its role in aging have been proposed, but the dogma itself has become a cornerstone of cell biologists' and gerontologists' thinking. During the same period when replicative senescence research was flourishing, it happened that a novel cytokine which promoted T cell proliferation in cell culture was discovered, an event that soon led to a flurry of immunology papers describing the seemingly unlimited growth of normal human T cells (Gillis and Smith, 1977, Morgan et al., 1976).

It was this apparent conflict between immunologists and cell biologists that captured Roy's attention and motivated us to determine in a systematic fashion whether or not human lymphocytes cultured in vitro were, in fact, restricted by the Hayflick Limit (Effros and Walford, 1984). The issue was not merely theoretical, but rather, a question with important implications to both cell biology and immunology: if normal T cells did not have a limited replicative potential, as had been suggested by numerous reports on putatively ‘permanent’ or ‘immortal’ T cells, the notion that replicative senescence was a general biological phenomenon would require revision. Alternatively, if normal T cells were restricted by the Hayflick Limit, the immortal T cell cultures described in the literature most likely consisted of cells that were abnormal, and therefore probably not appropriate models for analysis of normal immune function. In the event, extensive work in our own laboratory and by others has conclusively demonstrated that normal human T cells are, in fact, similar to all other mitotically-competent human cells and undergo replicative senescence in cell culture. These studies, which will be summarized below, not only elucidate the mechanism of T cell senescence itself, but also highlight the importance of analyzing replicative senescence in a variety of cell types, since there are unique aspects of the senescence program that emerged from the T cell experiments.

Section snippets

T cell replicative senescence

Immune responses to foreign pathogens require extensive proliferation, suggesting that a limited replicative potential might have a significant impact on the efficient function of lymphocytes. This requirement for massive clonal expansion is due to the intricate genetic mechanism by which T lymphocyte antigen receptors are generated, which allows a limited number of T cell receptor genes to create an immune system with an enormous range of specificities. Briefly, during a lymphocyte's

In vivo relevance of the in vitro model

The Holy Grail of replicative senescence studies has been to demonstrate that aging in vivo is accompanied by the accumulation of cells showing characteristics of replicative senescence identified in cell culture. Our identification of loss of CD28 gene expression as a marker of T cell senescence in vitro, therefore, provided an unparalleled opportunity to test whether similar cells accumulate during aging in vivo. Flow cytometric analysis of T cells from cohorts of different ages has clearly

Concluding remarks

Research over the past few decades has confirmed the highly original and insightful predictions made by Roy Walford regarding the role of the immune system as well as the process of T cell replicative senescence in various pathologies of human aging. Recent longitudinal studies on the elderly have provided clinical validation of many of Roy's theoretical predictions. The extensive analysis of the Swedish geriatric population over several decades has documented a cluster of immune parameters

Acknowledgments

The author acknowledges the support for the research described in this review, which include the National Institutes of Health, the UCLA Center on Aging, and the Plott Endowment (RBE holds that Plott Endowed Chair in Gerontology). I also thank Dr Graham Pawelec for critically reviewing the manuscript.

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