Inside the labs

Of the nearly 300 laboratories investigating different aspects of basic science and translational and clinical research at Yale School of Medicine (YSM), dozens are dedicated to inquiries related to the aging process. For a sampling of the research that is underway, Yale Medicine Magazine spoke with laboratory leaders who study the effects of aging on the immune system, metabolism, the brain, and endocrine function.

Albert Shaw

One might not expect to find aging among the interests of an infectious disease specialist—but it is, in fact, the focus of research in the laboratory of Albert Shaw, MD, PhD, professor of medicine (infectious diseases) and a member of the Yale Center for Research on Aging (Y-Age).

Older adults are at increased risk of morbidity and mortality due to such infectious diseases as pneumonia or sepsis, and their immune systems mount a weaker response post-vaccination, which leaves them less protected than their younger counterparts. These disparities were highlighted by the COVID-19 pandemic. Research shows that in 2023, adults aged 65 and older accounted for up to 63% of COVID-related hospitalizations and 90% of deaths. “COVID-19 had a devastating impact on older adults, particularly those in long-term care facilities and nursing homes, and continues to do so even with the availability of excellent vaccines,” Shaw says.

To better understand why older adults are more vulnerable to infectious diseases such as COVID-19, Shaw’s lab is investigating how the function of various immune system cells changes with age. The researchers are particularly interested in uncovering mechanisms linked to increased levels of inflammation that occur in older adults. “We think this chronic inflammation is the source of some of the altered immune functioning with age,” says Shaw.

Shaw’s team uses a variety of techniques to analyze the immune response in older versus younger adults. Flow cytometry, for example, is a method that uses antibodies recognizing specific proteins within or on the surface of cells that are linked to fluorescent compounds; researchers employ instruments with multiple lasers to detect and quantitate such proteins. Mass cytometry is a newer technique in which antibodies are linked to heavy metal tags detectable through mass spectroscopy, allowing the simultaneous assessment of over 40 cell parameters. Shaw also uses such methods as RNA sequencing to analyze gene expression, proteomics for protein expression, metabolomics for elucidation of key metabolites, and spatial transcriptomics methods pioneered by YSM experts to analyze gene expression in sections of human tissue from older versus younger adults. The researchers’ goal is to form a comprehensive picture of the mechanisms controlling the human immune response, as well as understanding how this response is affected by age.

Shaw’s team is zeroing in on a few potential key factors affected by aging, including receptors of the innate immune system and recent work on human platelets. The innate immune system is the part of the immune system that controls the earliest responses to infectious agents or vaccines, and primes the activation of the more specific immune responses controlled by antibodies and T cells. Platelets are best known as the cells that mediate blood clotting, but emerging research is revealing their prominent role in regulating the immune response. “Even though they’re very small cells, there are so many of them in the blood—millions in a teaspoon—that we think in aggregate they may contribute to dysfunction in the immune system in older adults,” Shaw says.

Their recent research—a collaboration with Steven Kleinstein, PhD, Anthony N. Brady Professor of Pathology; Ruth Montgomery, PhD, professor of medicine and professor of epidemiology (microbial diseases); Heather Allore, PhD, professor of medicine (geriatrics) and of biostatistics; and Thomas Gill, MD, Humana Foundation Professor of Medicine (Geriatrics)—shows that platelets have elevated levels of RNAs encoding genes critical for platelet signaling and activation in older compared to younger adults. Furthermore, the expression of these activation RNAs in platelets from older adults who met the criteria for frailty—a geriatric syndrome characterized by increased vulnerability to adverse health outcomes—was further increased compared to their expression in healthy older adults. The researchers believe that changes like these may play a role in driving age-associated chronic inflammation, as well as the higher rates of clotting-related diseases in older adults.

In addition, Shaw’s team is studying immunological differences between younger and older adults who have received various vaccinations. In 2022, Shaw was among a group of Yale researchers who received a $12 million award from the NIH as part of the Human Immunology Project Consortium (HIPC) to investigate vaccine responses in vulnerable groups. His lab is comparing the immune reactions of older adults in long-term care facilities to those of healthy community-dwelling older individuals and younger adults following administration of two different influenza vaccines approved for adults aged 65 and older. Through identifying immunological, gene expression, and proteomic differences, the team hopes to gain insights into designing better vaccines and therapeutics to strengthen the immune response in older adults.

Vishwa Deep Dixit

Scientists have long believed that the immune system is important only for protecting us from disease. But over the past several decades, researchers have found that immune cells are present within and important to nearly every organ in the body. “In addition to pathogen defense, they play an important role in the normal functioning of those organs,” says Vishwa Deep Dixit, DVM, PhD, Waldemar Von Zedtwitz Professor of Pathology and professor of immunobiology and of comparative medicine and the director Y-Age.

A vital function of immune cells includes regulating the metabolism of our organs. In Dixit’s laboratory, his team is investigating the interactions between the metabolic and immune systems. “The interaction between these is absolutely critical for maintaining function and homeostasis,” he says. The team’s work provides important insights into why we age and clues for potential new therapies for enhancing health span.

As we grow older, fat cells start to accumulate within some organs. This fat accumulation triggers the activation of the immune cells residing in the tissues, which in turn can lead to increased levels of inflammation. “That is now recognized to be one of the major pillars of mechanisms of aging,” says Dixit. “When the crosstalk between immune and metabolic systems goes awry, it leads to inflammation, immune and metabolic dysfunction, and the process of aging.”

Dixit’s team is using various transcriptomic technologies, including single-cell sequencing, RNA sequencing, proteomics, and metabolomics, to study the composition of fat tissues and identify potential targets that increase health span or delay the process of aging. The researchers’ goal is to identify the underlying immunometabolic mechanisms that drive aging-induced disease risk so that specific interventions can be designed to improve the health of older adults.

One finding is that a specific pathway that regulates inflammation, known as the NLRP3 inflammasome, is an important driver of the aging process. The Yale team’s research has shown that blocking the activity of this immunological complex not only reduces inflammation but also extends the health span of animal models. Other groups have shown that lowering NLRP3 delays aging of certain organs and enhances lifespan. Now, multiple ongoing clinical trials are investigating how drugs that inhibit NLRP3 impact disease. “Time will tell whether these interventions will have beneficial effects in human aging, as they have in animal models,” Dixit says.

Another avenue is caloric restriction, which may also offer insight into a mechanism to slow the aging process. In 2022, Dixit’s team analyzed genetic data collected from fat tissue that was examined as part of a National Institute on Aging study called the Comprehensive Assessment of the Long-term Effects of Reducing Intake of Energy (CALERIE), in which participants reduced their calorie intake by 14% for two years. Caloric restriction extends the lifespan and reduces inflammation in animal models, but how this dietary intervention affects the related process in human aging is not known. Dixit’s new study offered evidence that mild reduction of calories significantly improves immune function and reduces inflammation and thymic aging in humans. However, long-term calorie restriction is difficult to maintain. “So we want to identify the drivers that mediate and mimic the effects of calorie restriction on health and lifespan,” says Dixit.

This work has led to the identification of two molecules named PLA2G7 and SPARC, which are inhibited when people restrict their calorie intake. “These studies have identified that there are calorie restriction mimetics that can be harnessed,” says Dixit. “And mechanisms such as these could be potential future drug targets that enhance health span and may even increase quality of lifespan.”

Nenad Sestan

Our brains are continuously changing throughout our lifetimes. A newborn’s brain looks quite different from an adult’s. And the brain of a healthy older person will differ from that of someone living with an age-related chronic disease.

Nenad Sestan, MD, PhD, Harvey and Kate Cushing Professor of Neuroscience and professor of comparative medicine, of genetics and of psychiatry, studies how the human brain develops over time. Over the years, his team has created a research biobank comprised of postmortem brain specimens donated for scientific study. The tissues collected from each donor contain billions of neural cells. Each cell is “a bag of RNA,” says Sestan, produced when an enzyme transcribes a sequence of a gene’s DNA. Cell types are differentiated by the RNA they contain. However, the RNA repertoire will change over time—even within the same cell.

Emerging technologies are now giving Sestan’s team an unprecedented view inside these neural cells. Next-generation sequencing, for example, enables the team to study a large number of cells in parallel to see when and where genes are expressed in the brain tissue. These analyses are helping his laboratory to uncover biological differences between younger and older brains, which occur divergently in humans, nonhuman primates, mammals, and other species. “Instead of looking at only one molecule or one gene, as we did several years ago, we can profile every single gene, transcript, and protein to create a huge dataset,” Sestan says. “Before, we could only see a few continents, but now we have a very detailed map where we can see countries, counties, towns, cities, and streets.”

To make sense of these large datasets, and even to combine and integrate their features, Sestan’s team uses bioinformatics and artificial intelligence. The results have applications for understanding how our most important organ ages. “Now, we can tell how one’s brain cells change from age 3 to age 20, 40, 60, or even 90 and 100,” Sestan says. In doing so, the researchers are studying how healthy individuals age compared to those who are affected by age-related neurological or psychiatric disorders.

Eventually, large datasets are curated and sculpted into “brain atlases,” helping Sestan’s lab understand the molecular and cellular signatures of brain aging and identify various biomarkers involved in healthy and unhealthy aging. This process, in turn, brings insight into potential targets that scientists can test in model systems to see whether they can slow or reverse certain negative effects of aging. Furthermore, understanding how cells change with age can help scientists build biological models for predicting someone’s age based on signatures in the brain.

Sestan’s research has shown that not all cell types age in the same way, and that some cell types are more vulnerable to the aging process than others. This finding may explain why some cognitive skills, such as spatial awareness and memory, decline with age, while others like verbal abilities may actually improve. “Now we’re trying to understand why some cell types have different aging signatures than others,” says Sestan.

Sestan hopes that studying human brain tissue will provide answers that will help people attain a higher quality of life in their old age. There is, however, much to learn from studying other species, as well. There are certain animals, and even mammals, that have impressive lifespans. The Greenland shark, for example, can live for 300 to 500 years, and bowhead whales can live for over 200 years.

By comparing the composition of cell types and their regulatory capabilities with age in these animal brains versus our own, Sestan hopes to better understand the evolutionary differences between them as well as to decode secrets to other species’ remarkable longevity. “The notion that sharks have a complex brain and can live this long tells us that technically neurons can live 500 years,” he says. “We would like [to] learn from other animals how to help humans live longer and have a healthy brain.”

Hattie Chung

In humans, the ovary is among the first organs to age—a transition marked by menopause. Scientists have traditionally believed that ovarian aging is simply the result of egg depletion. Hattie Chung, PhD, assistant professor of medicine, who joined Yale School of Medicine as a member of the Yale Cardiovascular Research Center this year, is studying how cells in the ovary interact with one another and how these interactions change with age. She is also a member of Y-Age and has secondary appointments in the Department of Molecular, Cellular, and Developmental Biology and the Department of Obstetrics, Gynecology, and Reproductive Sciences. Her lab’s research is adding to the growing evidence that the ovaries play an important role beyond fertility—particularly in endocrine function—and that the ovarian aging process is much more intricate than previously thought.

All female organisms are born with a finite reserve of eggs, which are released monthly in humans through the menstrual cycle. “This is a very dramatic process that completely changes the [ovarian] tissue,” Chung says. “And it’s a very stressful process for the organ.”

First, ovarian cells must incubate eggs in multicellular units called follicles that are spatially organized. These follicles mature with the egg inside until they are ready for ovulation. A number of different cellular interactions occur during this maturation process. But these interactions can go awry—in fact, only 1% of follicles reach the ovulation stage.

When the follicle does survive, the egg is released via ovulation—a process that requires it to physically rupture the surface of the ovary. “This is a very inflammatory process,” says Chung. Ovulation occurs hundreds of times throughout a woman’s lifetime, which over time leads to a buildup of cellular debris and chronic inflammation in the ovary.

After the egg is released, the remaining follicular cells that supported its growth take on a second life and become a transient structure called the corpus luteum. This is the structure that produces most of a woman’s progesterone. “This change is also highly dynamic,” says Chung, as a variety of cells together infiltrate the ovary. “All these coordinated changes are occurring in a tiny organ all at once.”

At any given moment, the entire ovarian structure is undergoing change. Immune cells come and go to clear old structures. But like the rest of the body, the ovary is not a perfect machine. During each cycle, minor accidents occur that can contribute to inflammatory and fibrotic processes within the organ. “It’s a miracle that our [ovarian] tissues are able to achieve hundreds of remodelings to begin with,” says Chung. “Aging is highlighting all the ways that these processes can break.”

As a systems and computational biologist, Chung believes that the numerous coordinated changes that occur repeatedly make the ovary a fascinating model system. Her new lab uses spatial transcriptomics and advanced computational methods to study ovarian tissue, and their early findings are already revealing new insights into the aging process.

For instance, through a collaboration with the Broad Institute of MIT and Harvard, where she trained as a postdoc, and the Buck Institute for Research on Aging in Novato, California, Chung and her team have learned that there are actually two types of corpora lutea—one that makes progesterone and another that breaks it down. “The ratio of these two types of corpora lutea, we suspect, is key to regulating how we might have spikes in progesterone levels versus a decay across a cycle,” says Chung.

In older mice, the researchers found, the ratio of these two types of corpora lutea begins to change, which could hinder the regulation of progesterone production. Furthermore, as in other organ systems affected by aging, there is an extensive accumulation of immune cells in the aged ovary that have inflammatory functions. Understanding how and why immune cells linger in the ovary could uncover general principles of inflammaging (chronic, low-grade inflammation related to aging). Ovaries are like the canary in the coal mine for general aging. Hormones have a protective effect on a woman’s health. After menopause, women experience increased rates of cardiovascular disease, osteoporosis, and Alzheimer’s disease. Researchers believe that the decline of the ovary’s endocrine function triggers physiological changes in the body that accelerate the aging process. Among Chung’s goals is to determine whether scientists can reverse or slow the loss of endocrine function in the ovary, which in turn could boost women’s overall health. “We have big plans in the pipeline,” she says. “We’re taking it one small step at a time.”