Fight Aging!

One of the research groups involved in developing biomarkers of aging based on characteristic epigenetic changes published a most interesting paper earlier this month, linked below, in which they use their tools to investigate cellular senescence and cell aging. Biomarkers to measure biological age, the degree to which an individual is damaged and their biology has become dysfunctional in response to that damage, are an important line of development. An effective biomarker might be used to quickly assess the overall benefits of a potential rejuvenation therapy in mammals. As an alternative to running full life span studies this would dramatically reduce the time and cost required for such research. Cheaper research and faster results are certainly good for the pace of progress where they can be achieved.

Individual cells age, accumulating metabolic waste and unrepaired damage in the case of long-lived cells, or marching towards the Hayflick limit placed on the number of divisions permitted them in the case of short-lived somatic cells, but the relationship between cellular aging and tissue aging is not a straightforward one. A living being is a a dynamic system in which the majority of cells that make up its tissues are at some point replaced, on a schedule of days or months in most cases. Only in the central nervous system and a few other places do we find individual cells lasting a very long time, even the whole lifetime, and in which the steady accumulation of damage and waste are more important factors. In most tissues the importance is the pace of turnover, the number of lingering senescent cells that behave badly and refuse to die, the level of waste in the environment outside cells, and the quality of the stem cells that are responsible for producing a supply of new somatic cells to keep tissue functional.

Cells become senescent, removing themselves from the cycle of division and replication in response to a range of circumstances: damage, a toxic local environment, short telomeres as a result of reaching the Hayflick limit, and so forth. This probably serves to reduce cancer risk, at least initially, but senescent cells generate harmful signals that degrade surrounding tissues and produce localized inflammation. As their numbers grow, this damaging behavior contributes meaningfully to degenerative aging. As illustrated by recent research linking mitochondrial dsyfunction and cellular senescence, cells are very complicated machines: senescence isn't a single uniform state, not all senescent states are similar, and nor is it the case that all of the states that can be created in the laboratory are known to occur to a significant degree in living tissues. So this is an interesting area of cellular biochemistry to explore, even as clinical development is moving ahead on the blunt and direct approach of clearing senescent cells from the body so as to remove their detrimental effects. This is absolutely the way things should be: taking the fast road to therapies that will effectively treat the causes of aging even in the absence of detailed understanding, and those researchers who can raise funds for more leisurely investigation and mapping can continue their work in the meanwhile. If only this were the case in other fields relevant to aging, but for the most part only the leisurely investigation is taking place there.

Epigenetic clock analyses of cellular senescence and ageing

One model of ageing posits that the failure of tissues to function properly is due to the depletion of stem cells. Stem cells, which are the reservoir cells of tissues, may have finite capacities of proliferation such as being limited by the lengths of their telomeres. Their eventual depletion leads to the deficit of properly functioning cells, causing phenotypic changes that constitute ageing. While this model is plausible and supported by strong circumstantial evidence, it is presently difficult to prove or refute directly, not least because the identification of specific tissue stem cells is difficult. Similarly, the association between telomere length and ageing, although widely reported, is not without inconsistencies.

There is however, another model of ageing which is based on the observation that the number of senescent cells in the body increases in function of organism age. While this could be interpreted to mean that senescent cells cause ageing, it could also equally mean that senescent cells are a consequence of ageing. In this regard, it is noteworthy that there is increasing evidence to demonstrate that senescent cells are not benign. Instead they secrete bio-chemicals that are detrimental to normal functioning of neighbouring cells. The senescence-associated secretory phenotype (SASP) proteins include cytokine, chemokines and proteases and their paracrine activities are very diverse and include oncogenic characteristics that stimulate cellular proliferation and epithelial-mesenchymal transition. Importantly, SASP proteins also promote chronic inflammation, which is the origin of almost all age-related pathologies. As such, SASP proteins, through their different effects on normal and cancer cells, induce deterioration of the tissue. Recently, it was demonstrated that removal of senescent cells in mice delays ageing-associated disorders, providing very strong support for the notion that senescent cells mediate the effects of ageing. Hence it follows that to understand ageing, it is necessary to understand cellular senescence. This model of active induction of ageing (via senescent cells) does not exclude the role of stem cell depletion described above, which could indeed be a result of stem cell senescence.

At present, the causes of cellular senescence in vivo are not known for certain but in vitro, cells can become senescent through (i) telomere shortening via exhaustive replication (replicative senescence), (ii) over-expression of oncogene or (iii) DNA damage. While it is easy to perceive replicative senescence (RS) as part of a bona fide mechanism of ageing, it is more challenging to consider oncogene-induced senescence (OIS) as a significant contributor to natural ageing. Instead OIS has been proposed to function as a tumour suppressor mechanism. The only obvious common factor between RS and OIS is the co-opting of the DNA damage signalling mechanism to usher cells into arrest.

Recently, we developed a multivariate estimator of chronological age, referred to as epigenetic clock, based on methylation levels. The following features of this clock demonstrates that its age estimates capture several aspects of biological age: (a) it can accurately measure the age of cells regardless of tissue types including brain, liver, kidney, breast and lung (b) its accuracy is substantially higher than that of other molecular markers such as telomere length (c) it is able to predict mortality independent of health, life-style or genetic factors (d) its measurements correlate with cognitive and physical fitness amongst the elderly and (e) it is able to detect accelerated ageing induced by various factors including obesity, Down syndrome and HIV infection. Here, we apply this epigenetic clock to study the relationship between ageing and senescence of isogenic cells induced by exhaustive replication, ectopic oncogene over-expression or radiation-induced DNA damage.

We show that induction of replicative senescence (RS) and oncogene-induced senescence (OIS) are accompanied by ageing of the cell. However, senescence induced by DNA damage is not, even though RS and OIS activate the cellular DNA damage response pathway. Collectively, these two sets of observation make an effective case for the uncoupling of senescence from cellular ageing. This however, appears at first sight to be inconsistent with the fact that senescent cells contribute to the physical manifestation of organism ageing, as demonstrated elegantly by studies in which removal of senescent cells slowed down ageing. In the light of our observations however, it is proposed that cellular senescence is a state that cells are forced into as a result of external pressures such as DNA damage, ectopic oncogene expression and exhaustive proliferation of cells to replenish those eliminated by external/environmental factors. These senescent cells, in sufficient numbers, will undoubtedly cause the deterioration of tissues, which is interpreted as organism ageing. However, at the cellular level, ageing, as measured by the epigenetic clock, is distinct from senescence. It is an intrinsic mechanism that exists from the birth of the cell and continues. This implies that if cells are not shunted into senescence by the external pressures described above, they would still continue to age. This is consistent with the fact that mice with naturally long telomeres still age and eventually die even though their telomere lengths are far longer than the critical limit, and they age prematurely when their telomeres are forcibly shortened, due to replicative senescence. Hence senescence is a route by which cells exit prematurely from the natural course of cellular ageing.

Finally, it is necessary to address specifically the role of telomeres as it is easy to confound them with cellular ageing because at first view, they appear to share similar features. Since critical telomere length is attained after many rounds of proliferation, which takes a long time and hence occurs later in life, it is easy to mistake this for a functional link with age even though telomere length has only a modest correlation with chronological age, while cellular ageing as measured by the epigenetic clock has a far higher degree of association with biological ageing. The fact that maintenance of telomere length by telomerase did not prevent cellular ageing defines the singular role of telomeres as that of a means by which cells restrict their proliferation to a certain number; which was the function originally ascribed to it. Cellular ageing on the other hand proceeds regardless of telomere length.

Although the characteristics of cellular ageing are still not well known, the remarkable precision with which the epigenetic clock can measure it and correlate it to biological ageing remove any doubt of its existence, distinctiveness and importance. This inevitably raises the question of what is the nature of this cellular ageing, and what are its eventual physical consequences. Admittedly, the observations above do not purport to provide the answer, but they have however, cleared the path to its discovery by unshackling cellular ageing from senescence, telomeres and DNA damage response, hence inviting fresh perspectives into its possible mechanism. In summary, the results from these experiments, while apparently simple in their presentation, untangles a conceptual knot that hitherto tied senescence, DNA damage signalling, ageing and telomeres together in an incomprehensible way. Here we propose that cellular ageing, as measured by the epigenetic clock, is an intrinsic property of cells, and while independent, its speed can be affected by some factors; a feature that would undoubtedly be exploited to characterise and elucidate its mechanism.