Epigenetic Clocks Continue to Tick Over Multiple Lifetimes

From fruit flies that survive a mere two weeks to the giant tortoises of the tropics that can live for nearly 200 years, every species has an allotted life expectancy. But while an organism’s lifespan may be constrained by its biology, not all its cells appear to play by the same rules.

In 2023, University of Minnesota immunologist David Masopust and his team published findings that the same population of T cells could be transferred from mouse to mouse to mouse, resulting in cells that survived for up to 10 years, approximately four mouse lifespans.¹ These comparatively ancient immune cells not only survived but remained functional: They proliferated in response to the appropriate antigen, displaying neither senescence nor uncontrolled growth.

In a recent paper published in Nature Aging, Masopust teamed up with immunologists Benjamin Youngblood and Caitlin Zebley at St. Jude Children's Research Hospital to study the epigenetic clocks in these mouse cells that seemed to defy aging.² Unlike most cells, whose clocks tick along with the passage of time, the T cell clocks seemed to record proliferation events, and they continued to tick forward well beyond the natural lifespan of the organism. This research could provide important insights into immunological phenomena like age-related immune senescence and T cell exhaustion, which can develop in the context of cancer or chronic infections.³

“It’s an absolutely remarkable mouse model and a great tool for exploring the relations between replicative history, epigenetic age, and chronological age,” wrote Andrew Yates, a quantitative immunologist at Columbia University who was not involved in the study, in an email to The Scientist.

Compared to most other cells in the human (and mouse) body, T cells have very different patterns of proliferation. Before an organism encounters a particular antigen—such as a protein on a specific strain of influenza virus, or sugar molecules on Streptococcus bacteria—it possesses only a few T cells with the cellular machinery needed to respond to this threat. “[Initially], the cells are super rare and not abundant enough to be really functionally relevant,” said Masopust. “But when you get an infection, these cells will become the fastest dividing cells in your body, dividing every six hours… [so they] can become numerically relevant before you die.” After the infection is cleared, this T cell population shrinks but remains much larger than before exposure; the cells are ready to spring into action with another burst of proliferation if they encounter the same antigen again.

Masopust wanted to test the limits of these proliferative capabilities. “There's an idea that every cell in your body only has so many population doublings before it basically becomes permanently undividable—a state that most people call senescence.”

There is plenty of evidence that T cells can become senescent during aging, chronic infections, and cancer.⁴ However, the mechanisms that drive senescence in these cells have not been conclusively established. “We felt that this was contextual, and not purely a function of proliferation or stimulation history,” said Masopust. “And we were willing to bet on that.”

To test this hypothesis, Masopust and his team needed a system that allowed them to separate cell-intrinsic and cell-extrinsic factors. In an initial group of mice, the researchers generated an army of memory T cells specific for a certain antigen by using a carefully timed protocol of vaccinations and booster shots. Then, they transferred some of these memory T cells to new mice, administered the vaccines to induce proliferation, and started the whole process over again.

“We just kept going and going and going until it kind of got absurd,” Masopust recalled. Ultimately, they transferred the T cells to new mice up to 17 times over the course of the following decade. As Masopust’s team demonstrated in their 2023 paper, these ancient cells remained functional, fulfilling their duties as needed without descending into either senescence or uncontrolled growth.

In the present study, researchers examined the inner workings of these cells. They found that the cells’ epigenetic clocks kept ticking well beyond a normal mouse lifetime. As the T cells aged over multiple passages, genome-wide methylation gradually declined and the researchers observed changes in the epigenetic profiles of several genetic loci, including genes that regulate tumor suppression and oncogenesis. These findings suggest a potential connection between the clocks and the mechanisms that help aging T cells avoid replicative senescence, but the extent to which these epigenetic changes simply record cell history—or whether they actively promote longevity by altering gene expression—is still murky.

The researchers were also curious about what the T cell clocks were recording. Traditional epigenetic clocks keep track of chronological time, but the T cell clocks appeared to function differently, suggesting that this clock kept track of the cell’s history of proliferation rather than its age.

But did this clock tick according to similar rules in human T cells? In healthy humans, naïve T cells, which have not undergone an antigen-induced proliferation event, appeared young regardless of the donor’s actual age. However, pediatric patients with T cell acute lymphoblastic leukemia, in which the cells undergo rapid, uncontrolled proliferation, had T cells that, according to this epigenetic clock, were between 100 and 200 years old, even though none of the patients were older than 15 years of age.

Yates, however, mentioned an important caveat, which the researchers also acknowledged in the paper. “The iterative transplantation process may—at least in part—be selecting for established epigenetic states that confer fitness, rather than changes in methylation being acquired progressively and within cell lineages over time. These alternatives are difficult to untangle,” wrote Yates. In other words, it may be difficult to determine the extent to which these epigenetic changes are markers of cell history or whether cells that acquired particular epigenetic changes were the only cells able to survive and proliferate.

Regardless, Masopust believes there are many potential future directions for this research. Understanding the T cell aging process and the “rules” that govern whether an aging T cell remains functional or becomes senescent could inform new strategies to fight age-related immunological decline.

Similarly, said Masopust, “We could learn about the regulation of [T cell] exhaustion in a different way; maybe we could apply this to adoptive cell therapies, which are newer, somewhat boutique ways of curing cancer with some very remarkable clinical results.” He added, “But one problem with those cells is generating enough that then have durability and won't become exhausted.” By studying the multilifetime T cells, which can be easily expanded and appear resistant to exhaustion, Masopust hopes to identify strategies that could evoke these same properties in cancer-fighting chimeric antigen receptor (CAR) T cells for patients.

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