Inflammatory Macrophages are Found to Contribute to Harmful Ventricular Remodeling in Heart Failure

It is already well known that the immune cells called macrophages are involved in the mechanisms of heart failure, and in the research noted here the details of that role are further explored. Macrophages are important in processes of regeneration and tissue growth throughout the body, but also in the propagation of inflammation in response to damaging circumstances. A growing theme in the research of past years is the polarization of macrophages, meaning their division into several subtypes based on behavior. Some are inflammatory and aggressive, attacking pathogens but also hindering regeneration, while others are not inflammatory and undertake a variety of activities to directly aid tissue regeneration. A useful response to injury requires both behaviors in some proportion, and at different times, but later life and many age-related conditions are characterized by the presence of far too many inflammatory macrophages. Removing these macrophages or adjusting their state shows promise as a basis for therapy.

The researchers here find macrophages displaying a CCR2 receptor, which correlates fairly well with the inflammatory polarization, are necessary for much of the harmful growth of the heart that takes place in later life, as the cardiovascular system becomes damaged and dysfunctional. One of the more important components of heart failure is this hypertrophy of heart tissue. The muscle grows larger and weaker, initially in response to the failure of blood pressure feedback mechanisms that takes place alongside the development of hypertension, but later a range of other mechanisms are also involved. Clearly, inflammatory macrophages are doing their part to generate an unhelpful growth response - and so selectively removing them could be a useful form of therapy.

Prevention of blood pressure issues is probably a better first option for those not already old, however. If rejuvenation therapies can (a) prevent the processes that lead to the stiffening of blood vessels, such as cross-linking and calcification, and (b) prevent the atherosclerotic plaque that narrows blood vessels, such as by clearing out the harmful lipid compounds that cells cannot effectively break down, then hypertension and other blood pressure issues could be largely eliminated. Given a life-long normal blood pressure, the impact of inflammatory processes on the heart will be that much less severe. They must still be dealt with, as the secondary consequence of fibrosis remains an issue, but that can happen in a context of better overall health and physical robustness.

Immune cell target identified that may prevent or delay heart failure after pressure overload

Researchers have found that preventing the early infiltration of CCR2+ macrophages into the heart, after experimental pressure overload in a mouse model, significantly lessened the heart's enlargement and reduced pumping ability that leads to later heart failure. Thus, this infiltration is a required step in the path toward heart failure. Macrophages are immune cells that engulf and remove damaged or dead cells in response to tissue injury or infection. They also may present antigens to other immune cell types. The most common forms of pressure overload are aortic stenosis - a narrowing of the aortic valve of the heart that forces the heart muscle to overwork - and high blood pressure.

The researchers used two different methods to prevent early macrophage infiltration - an inhibitor of the macrophage cell-surface CCR2 chemokine receptor, and an antibody that selectively removes CCR2+ macrophages. Migrating macrophages use the CCR2 receptor to home in on damaged tissues in the body that are releasing chemokines. Preventing early macrophage infiltration may offer a therapeutic target in human disease. Researchers had previously known that pressure-overload heart failure is associated with inflammation caused by activated T-cells. The present study showed the link between infiltrating macrophages and the T-cell response during pressure overload of the heart.

One week after inducing pressure load, that the heart showed increased expression of three attractant chemokines that are able to bind to the CCR2 receptor on macrophages. The researchers also found an increased number of monocytes with the cell-surface markers Ly6C and CCR2 circulating in the blood, and they saw an eightfold increase in CCR2+ macrophages infiltrating into the heart. Those macrophages are derived from the circulating monocytes. Thus increased circulating monocytes might serve as an easily measurable biomarker that reflects cardiac tissue CCR2+ macrophage expansion. The circulating monocytes - along with other clinical, imaging and biochemical biomarkers - could guide patient selection for a prospective clinical trial to find out whether modulating CCR2 macrophages in humans with pressure-overload hypertrophy will delay or prevent later transition to heart failure.

CCR2+ Monocyte-Derived Infiltrating Macrophages Are Required for Adverse Cardiac Remodeling During Pressure Overload

Inflammation is a hallmark of chronic heart failure (HF) initially triggered by nonimmune modes of cardiac injury, such as myocardial infarction, genetic mutations, and mechanical stress (e.g., pressure overload). Moreover, the systemic and myocardial immune cell profiles underlying the inflammatory response in the various etiologies of HF are of considerable importance for disease progression. For example, in chronic ischemic HF, expanded populations of both innate immune cells (e.g., macrophages) and T cells in the heart promote tissue injury and pathological remodeling. Chronic nonischemic HF due to pressure overload is characterized by CD4+ T-cell activation, which has been shown to play a critical role in promoting adverse cardiac remodeling. We recently demonstrated that during cardiac pressure overload, proinflammatory macrophage expansion in the heart occurs early, before sustained systolic dysfunction, but resolves during the chronic stage.

Importantly, although pressure-overload HF is characterized by T-cell activation, prior work also indicates that such activation is dependent on antigen presentation, because the progression of HF is ameliorated upon blockade of T-cell costimulatory molecules on antigen presenting cells (APCs). The requirement for specific antigen recognition implies an essential pathogenetic role for macrophages and other APCs, although their specific function in the development of pressure-overload HF remains poorly defined. Recent studies have characterized cardiac macrophage populations in the heart with disparate functions, including tissue-resident, embryonically derived macrophages and infiltrating monocyte-derived macrophages. The normal heart is seeded with resident macrophages that are not replenished by circulating monocytes under steady-state conditions. Resident cardiac macrophages are minimally inflammatory and promote angiogenesis and tissue repair. However, cardiac injury and aging stimulate the infiltration of monocyte-derived macrophages that are proinflammatory, promote tissue injury, and the death and substitution of resident cells.

Monocyte-derived macrophages can be distinguished by the expression of C-C chemokine receptor 2 (CCR2). Although we and others have documented expansion of cardiac macrophages during the early phase of pressure overload, it is unknown whether the macrophages are monocyte-derived, and whether these cells play an important role in subsequent T-cell recruitment and activation, and associated long-term adverse cardiac remodeling. Accordingly, here we tested the hypothesis that CCR2+ monocyte-derived macrophages infiltrate the heart early following pressure-overload-induced hemodynamic stress, and that this macrophage population plays a critical role in the activation of T cells and the ensuing transition to failure.

An Interview with Vitalik Buterin, Patron of SENS Rejuvenation Research

Vitalik Buterin is the originator of Ethereum, but also a strong supporter of research and development aimed at bringing aging under medical control. He recently stepped up to make a $2.4 million donation to the SENS Research Foundation to support the scientific programs there, and thus help to hasten the advent of the first generation of working rejuvenation therapies. This is very welcome support at a critical juncture in the development of means of human rejuvenation, biotechnologies that will be based on periodic repair of the forms of cell and tissue damage that cause aging. The Life Extension Advocacy Foundation volunteers arranged this interview with Buterin, one of a number of articles resulting from the recent Undoing Aging conference that they hope to publish soon.

Wealthy people usually donate money towards research into and treatment of cancer, Alzheimer's disease, and other diseases. Why did you decide to donate Ethereum to the fight against aging?

The first reason is just because there are many other people who donate to fight against cancer and other specific diseases, which, of course, is very important and necessary. The second reason is that there is strong scientific evidence that aging is the root of the most serious diseases.

It turns out that if you slow down aging or even reverse it, you can save people from serious illnesses such as malignant tumors, stroke, and Alzheimer's disease.

Exactly. After all, if you do not prevent these diseases by eliminating aging, you will have to provide treatment to people who are already sick and suffering and whose quality of life is worsening, and the economy will be under enormous pressure because the treatment is often expensive, caregiving is needed, etc. These problems could be avoided. Studies of aging are very important right now, yet there are still very few people who invest money in this field, unfortunately.

Why do you think that is so?

Most people simply do not know or do not believe that aging can be successfully manipulated. However, I have read Ending Aging by Dr. Aubrey de Grey, I'm interested in scientific discoveries, and I see that this is plausible. Researchers can already extend the life of laboratory animals significantly, and it is necessary to refine these technologies in order to transfer them to humans. And research and full-scale clinical trials of anti-aging therapies in humans requires money.

Do you have plans to continue supporting research projects on aging and life extension, or is your current contribution of 2.4 million dollars likely to be all?

Of course I'm ready to invest more into it. However, right now, I am mostly investigating what the scientists are working on, what the most promising directions are, and what else should be supported.

What, in your opinion, is the main problem currently hampering the fight against aging on Earth?

There is not enough public support. Huge resources, as I said, are invested in research and treatment of single diseases, but the problem is that if we focus only on specific diseases, this will only slightly improve the lives of people who are already chronically sick. Only a few years will be added to their lives.


Increased Elastin Production as a Therapy for Age-Related Arterial Stiffening

Elastin, as the name might suggest, is an important structural molecule in the extracellular matrix of elastic tissues, such as blood vessels. Elastin content in blood vessel walls falls with age, alongside the stiffening of those blood vessels, though it is an open question as to the degree to which that is secondary to various mechanisms such as chronic inflammation, presence of senescent cells, and so forth. A very interesting study in mice from a few years ago demonstrated improved elasticity in the lung tissue of mice resulting from clearance of senescent cells, for example.

It is also an open question as to whether the reduction in elastin is as important as the cross-linking of molecules in the extracellular matrix when it comes to stiffening of blood vessels - absent the ability to selectively fix just one of these problems, firm answers will remain elusive. And that is before we consider other mechanisms such as calcification, probably also due in large part to the presence of senescent cells, or disrupted signaling that hampers the ability of smooth muscle cells in blood vessels to coordinate vasoconstriction and vasodilation.

The line of evidence constructed in the research results noted here is somewhat tenuous, since it was carried out in animal models of a genetic condition in which elastin levels are abnormally low, and with a focus on young patients rather than older individuals. It doesn't necessarily follow that because a boost in elastin production helped to restore blood vessel elasticity in this situation, then the same result will occur in old patients. Old blood vessels may have reduced elastin to some degree, but also have the range of other problems mentioned above. If there is a suitable drug candidate or other means of increased elastin production ready to go, as appears to be the case, then it would seem cost-effective to try it and see - but I'd wager on better results from cross-link breaking if this turns out to be a matter of significant investment in further research first.

Arteries in young, healthy humans and other mammals stretch easily because they contain a protein called elastin. Elastin is produced only during development, however, and is slowly lost with aging. Stiff arteries contribute to development of high blood pressure and significantly increase the risk of sudden death, stroke, myocardial infarction, and cognitive decline. "We know that genetic conditions, such as Williams-Beuren Syndrome (WS) and supravalvar aortic stenosis (SVAS), lead to abnormally low levels of elastin in developing arteries. As a result, children with WS or SVAS have stiff, narrow arteries and high blood pressure. Like older adults, they are also at increased risk of sudden death and stroke. We therefore tested whether a medicine called minoxidil would not only reduce blood pressure but also would help relax arteries and increase their diameter, thus improving organ perfusion."

Minoxidil is perhaps best known for its potential to improve hair growth when applied to the skin. In a different formulation, minoxidil is sometimes prescribed orally for high blood pressure that has not responded to other medications. Earlier studies have suggested that minoxidil may increase elastin deposition even in mature tissues. The research team conducted the work in experimental models of hypertension and chronic vascular stiffness associated with WS and SVAS. They used ultrasound imaging and magnetic resonance imaging-based arterial spin labeling to gauge minoxidil's impact on vessel mechanics, carotid and cerebral blood flow, and gene expression.

"Minoxidil not only lowered blood pressure, but also increased arterial diameter and restored carotid and cerebral blood flow. Minoxidil also reduced functional arterial stiffness and increased arterial elastin content. Equally important, these beneficial changes persisted weeks after the drug was no longer in the bloodstream. The sustained improvements and the increased elastin gene expression suggest that minoxidil treatment may help remodel stiff arteries. Such remodeling may benefit humans whose elastin insufficiency is due to either advanced age or genetic conditions."


Artificial Cell Components and Membranes, the Start of a Fusion Between Biology and Biotechnology Inside the Body

There will be no bright dividing line between evolved cellular component and artificial molecular machinery in the future of medicine and human enhancement. It is already possible to produce programmable DNA machinery that can react to the environment in simple ways, or to adjust the programming of cells by altering the production or activities of specific proteins. As understanding of the cell improves, it will be possible to produce nanoscale structures that act in similar ways to cellular components. Researchers are starting down this road with the production of various forms of manufactory, artificial membranes that enclose anything from cells or bacteria to a minimal set of DNA or other molecular machinery that can produce specific proteins or other molecules in response to circumstances. The articles below look at the two ends of this scale: an entire cell wrapped in a membrane on the one hand, versus much smaller components designed to be taken up and used by cells, releasing molecules in response to internal signals.

For the future, it is possible to envisage all sorts of further possibilities. Tweaks to existing structures to make them better: enhanced lysosomes equipped with a better range of digestive enzymes, improving the ability of long-lived cells to break down unwanted molecular waste; mitochondria with a stripped down, best of breed mitochondrial genome, based on the most performant of those evolved in our species; protein production and protein clearance structures based upon those found in other species that are much more efficient than the human model; cultured gut bacteria that are designed from the ground up, with minimal genomes, to be entirely beneficial; and more. Or simple artificial cells that replace or augment some of the simpler functions of evolved cells, such as the production of a needed protein or removal of an unwanted protein. Or wholly new structures within a cell that trickle out signal molecules that permanently increase cellular stress responses. Or sophisticated manufactories capable of producing all of the known cancer suppression genes, delivered by the billion, taken up into all cells, where they lie dormant, waiting to triggered into activity in cancerous cells. There are so very many options for improvement.

Further down the line, machinery that looks very different from cells will start to become a viable proposition. Diamondoid nanotechnology, for example, coupled with molecular manufacturing to mass produce devices that look nothing like cells, but can be vastly more efficient than any cell at a specific task. Nanomachines that can store hundreds as times as much oxygen as a red blood cell; that can identify and destroy pathogens without flagging; that can assist in the repair and maintenance of the inner machinery of living cells. The fusion of machine and biology will become highly sophisticated and varied. The importance of the designation of biological or artificial will fade, and ultimately we will become just as designed and enhanced as any of of the countless component parts in our cells.

Artificial and biological cells work together as mini chemical factories

Researchers have fused living and non-living cells for the first time in a way that allows them to work together, paving the way for new applications. The system encapsulates biological cells within an artificial cell. Using this, researchers can harness the natural ability of biological cells to process chemicals while protecting them from the environment. This system could lead to applications such as cellular 'batteries' powered by photosynthesis, synthesis of drugs inside the body, and biological sensors that can withstand harsh conditions.

Previous artificial cell design has involved taking parts of biological cell 'machinery' - such as enzymes that support chemical reactions - and putting them into artificial casings. The new study goes one step further and encapsulates entire cells in artificial casings. The artificial cells also contain enzymes that work in concert with the biological cell to produce new chemicals. In the proof-of-concept experiment, the artificial cell systems produced a fluorescent chemical that allowed the researchers to confirm all was working as expected.

"Biological cells can perform extremely complex functions, but can be difficult to control when trying to harness one aspect. Artificial cells can be programmed more easily but we cannot yet build in much complexity. Our new system bridges the gap between these two approaches by fusing whole biological cells with artificial ones, so that the machinery of both works in concert to produce what we need. This is a paradigm shift in thinking about the way we design artificial cells, which will help accelerate research on applications in healthcare and beyond."

Tiny implants for cells are functional in vivo

In the cells of higher organisms, organelles such as the nucleus or mitochondria perform a range of complex functions necessary for life. Researchers are working to produce organelles of this kind in the laboratory, to introduce them into cells, and to control their activity in response to the presence of external factors (e.g. change in pH values or reductive conditions). These cellular implants could, for example, carry enzymes able to convert a pharmaceutical ingredient into the active substance and release it "on demand" under specific conditions. Administering drugs in this way could considerably reduce both the amounts used and the side effects. It would allow treatment to be delivered only when required by changes associated with pathological conditions (e.g., a tumor).

Now, researchers have succeeded in integrating artificial organelles into the cells of living zebrafish embryos. The artificial organelles are based on tiny capsules that form spontaneously in solution from polymers and can enclose various macromolecules such as enzymes. The artificial organelles presented here contained a peroxidase enzyme that only begins to act when specific molecules penetrate the wall of the capsules and support the enzymatic reaction. To control the passage of substances, the researchers incorporated chemically modified natural membrane proteins into the wall of the capsules. These act as gates that open according to the glutathione concentration in the cell. At a low glutathione value, the pore of the membrane proteins are "closed" - that is, no substances can pass. If the glutathione concentration rises above a certain threshold, the protein gate opens and substances from outside can pass through the pore into the cavity of the capsule. There, they are converted by the enzyme inside and the product of the reaction can leave the capsule through the open gate.

The researchers chose zebrafish embryos because their transparent bodies allow excellent tracking of the cellular implants under a microscope when they are marked with a fluorescent dye. After the artificial organelles were injected, they were "eaten" by macrophages and therefore made their way into the organism. The researchers were then able to show that the peroxidase enzyme trapped inside the artificial organelle was activated when hydrogen peroxide produced by the macrophages entered through the protein gates.

A Tissue Engineered Retinal Patch Improves Vision in Macular Degeneration Patients

The trial results announced here represent a promising step forward in efforts to regenerate an age-damaged retina, particularly because the patients were in an advanced stage of their degenerative condition and nonetheless achieved a meaningful degree of restored sight. Macular degeneration has a number of different manifestations, and here the wet form was treated, which involves excessive growth of blood vessels in the retina and consequent death of the retinal cells necessary for vision. Researchers have established an approach involving the generation of a patch of engineered retinal cells that can be implanted to restore some of the lost retinal function. Given the details, it is interesting to speculate on the degree to which the transplanted cells are helping by integrating into the retina versus helping by issuing signals that spur local regeneration. In most cell therapies it is the latter, but here the transplanted cells are more organized into a tissue-like structure.

Human embryonic stem cells (ESCs) represent a promising source for cellular replacement therapies owing to their availability, pluripotency, and unlimited self-renewal capacity. However, they also carry risks of neoplastic change, uncontrolled proliferation, and differentiation to inappropriate cell types. The eye is advantageous in investigating hESC-based cell therapy as it is accessible and confined, and the transplanted cells can be monitored directly in vivo, with the possibility of being removed or destroyed if there is evidence of neoplastic change. Furthermore, long-term immunosuppression can be delivered locally.

Late age-related macular degeneration (AMD) is characterized by irreversible cell loss, initially of retinal pigment epithelium (RPE) cells and subsequently of neuroretinal and choroidal cells, and thus may be amenable to hESC-based cell therapy. Suspensions of hESC-derived RPE (hESC-RPE) cells have been transplanted in human subjects with dry AMD and Stargardt's disease, but the extent of cell survival and restoration of vision remains ambiguous.

We developed a therapeutic, biocompatible hESC-RPE monolayer on a coated synthetic membrane, herein termed a 'patch', for transplantation in wet and early-stage dry AMD. The choice of membrane material and its preparation, including the human vitronectin coating, has not been described previously to our knowledge. In contrast to RPE suspensions, cells on the patch are delivered fully differentiated, polarized, and with the tight junction barrier formed, that is, in a form close to their native configuration. The synthetic membrane allows the patch to be handled easily and robustly. The main disadvantage of the patch is that it requires a purpose-built delivery tool and a more complicated surgery compared to cell suspensions, and the use of hESCs may require immunosuppression, unlike an autologous cell source.

The clinical trial was designed as a phase 1, open-label, safety and feasibility study of implantation of an hESC-RPE patch in two subjects with acute wet AMD and recent rapid vision decline. For safety reasons and to obtain an early efficacy signal, the trial involved patients with severe wet AMD only, although we aim to study the RPE patch in early dry AMD in the future. We reported three serious adverse events to the regulator. These were exposure of the suture of the fluocinolone implant used for immunosuppression, a retinal detachment, and worsening of diabetes following oral prednisolone. All three incidents required readmission to the hospital, with the first two incidents requiring further surgery and the third being treated medically. The three incidents were treated successfully. Both patients achieved an improvement in best-corrected visual acuity of more than 15 letters at 12 months after transplantation.

Although 12 months is sufficient to begin to describe cell survival and clinical outcomes, it is early in terms of safety monitoring, especially for late teratoma formation. The patients will be followed for five years after surgery. These two early cases are also instructive as they show an encouraging outcome despite very advanced disease, which increases the complexity of surgery and involves more damaged neuroretina.


An Update on Immune System Recreation as a Treatment for Multiple Sclerosis

The destruction of near all immune cells followed by cell therapy to speed recreation of the immune system is a fairly harsh procedure, as the only way to clear a sufficiently high fraction of immune cells at the moment is essentially a form of chemotherapy. It is an effective treatment for autoimmune conditions, however, albeit with a significant risk of death, in line with that for many major surgeries. This makes it suitable in its current form only for more severe autoimmune disorders in which the patients tend to be younger and more robust, but with a very poor prognosis. In past years researchers have demonstrated considerable success with multiple sclerosis, and the article here provides an update on ongoing trials. The results continue to be impressive.

In the future, the chemotherapy approach will be replaced with more targeted, less harmful methods of selective cell destruction - consider the Oisin Biotechnologies cell destruction technology turned against immune system markers, for example. More gentle cell destruction methodologies will make immune system recreation viable as a way to rejuvenate aged immune systems, even in very old, frail individuals, clearing out all of the misconfigured, senescent, exhausted, or otherwise harmful immune cells. That is why it is worth keeping an eye on progress in this line of research.

Doctors say a stem cell transplant could be a "game changer" for many patients with multiple sclerosis (MS). Results from an international trial show that it was able to stop the disease and improve symptoms. It involves wiping out a patient's immune system using cancer drugs and then rebooting it with a stem cell transplant. Just over 100 patients took part in the trial, in hospitals in Chicago, Sheffield, Uppsala in Sweden and Sao Paolo in Brazil. They all had relapsing remitting MS - where attacks or relapses are followed by periods of remission. The interim results were released at the annual meeting of the European Society for Bone and Marrow Transplantation in Lisbon.

The patients received either haematopoietic stem cell transplantation (HSCT) or drug treatment. After one year, only one relapse occurred among the stem cell group compared with 39 in the drug group. After an average follow-up of three years, the transplants had failed in three out of 52 patients (6%), compared with 30 of 50 (60%) in the control group. Those in the transplant group experienced a reduction in disability, whereas symptoms worsened in the drug group. "The data is stunningly in favour of transplant against the best available drugs - the neurological community has been sceptical about this treatment, but these results will change that."

The treatment uses chemotherapy to destroy the faulty immune system. Stem cells taken from the patient's blood and bone marrow are then re-infused. These are unaffected by MS and they rebuild the immune system. "We are thrilled with the results - they are a game changer for patients with drug resistant and disabling multiple sclerosis. This is an interim analysis, but with that caveat, this is the best result I have seen in any trial for multiple sclerosis." The transplant costs around $40,000, about the same as the annual price of some MS drugs. Doctors stress it is not suitable for all MS patients and the process can be gruelling, involving chemotherapy and a few weeks in isolation in hospital.


Nicotinamide Supplementation Looks Little Better than Resveratrol in Mice

Hopefully the Fight Aging! audience recalls the years-long hype over resveratrol, driven by the self-serving processes that enabled investors in Sitris Pharmaceuticals to make a sizable profit at the expense of GSK, and supplement sellers to open up a new market for the credulous. The only meaningful results from all of that turned out to be an increased knowledge of the biochemistry of sirtuins, one very thin slice of the broad metabolic response to calorie restriction. Resveratrol and its ilk are not meaningful calorie restriction mimetics, and you are far better off cutting a few hundred calories from your daily intake or exercising a little more.

In light of this history I think it is entirely appropriate to be skeptical of the current hype surrounding the role of NAD+ in metabolism, and the various precursor molecules that can increase levels of NAD+ when taken as dietary supplements. When compared with sirtuins and resveratrol, the publicity here involves many of the same people, similar for-profit companies engineering the news cycle, and the same area of cellular biochemistry, which is to say aspects of calorie restriction closely related to sirtuins. My expectation is that, at the end of the day, this will result in nothing more than another increase in the knowledge of this portion of cellular biochemistry, while all the other claims regarding longevity and health are largely smoke and mirrors. Some people will make a lot of money, supplement sellers will prosper, and nothing will meaningfully change in human health as a result of all of this.

The first study in mice noted below is very similar in outcome to past studies of resveratrol, which is to say little in the way of gains in healthy mice, and some compensation for the detrimental effects of being overweight or obese. It is important to remember that mouse longevity is far more plastic than that of humans in response to calorie restriction and interventions that affect the same portions of cellular biochemistry as are involved in the calorie restriction response. Mice live 40% longer when calorie restricted; in humans the gain is unlikely to be larger than a few years, even though the observed health benefits are sizable. So an alleged calorie restriction mimetic that produces no gain in mouse longevity, or only helps to make overweight mice less metabolically abnormal, is not all that interesting. You might compare this with the second paper, which is a commentary from the usual suspects on how great the prospects are for supplementation related to NAD+ levels.

Nicotinamide Improves Aspects of Healthspan, but Not Lifespan, in Mice

The role in longevity and healthspan of nicotinamide (NAM), the physiological precursor of NAD+, is elusive. In the present study, we aimed to characterize the effects of chronic NAM supplementation on the longevity and healthspan characteristics of male C57BL/6J mice fed a synthetic low-fat diet (SD) and the corresponding high-fat diet (HFD). Because of the liver's importance in maintaining metabolic homeostasis, we carried out histological, biochemical, and untargeted metabolomics surveys to provide an unbiased view of the metabolic impact exerted by 62-week NAM supplementation on liver from SD- and HFD-fed mice.

Protein target validation combined with metabolic flux analysis enabled the identification of the underlying mechanisms of enhanced glucose disposal and reduced oxidative stress in response to NAM supplementation. Surprisingly, our data showed that NAM depresses NAD salvage and has complex effects on sirtuin expression and activity. NAM appears to have greater beneficial effects in mice subjected to HFD than SD, which might provide important clues about its therapeutic potential in the fight against obesity and associated comorbidities.

We report that chronic NAM supplementation improves healthspan measures in mice without extending lifespan. Analysis revealed NAM-mediated improvement in glucose homeostasis in mice on a high-fat diet (HFD) that was associated with reduced hepatic steatosis and inflammation concomitant with increased glycogen deposition and flux through the pentose phosphate and glycolytic pathways. Although neither hepatic NAD+ nor NADP+ was boosted by NAM, acetylation of some SIRT1 targets was enhanced by NAM supplementation in a diet- and NAM dose-dependent manner. Collectively, our results show health improvement in NAM-supplemented HFD-fed mice in the absence of survival effects.

Therapeutic Potential of NAD-Boosting Molecules: The In Vivo Evidence

Nicotinamide adenine dinucleotide (NAD) is one of the most important and interesting molecules in the body. It is required for over 500 enzymatic reactions and plays key roles in the regulation of almost all major biological processes. Above all, it may allow us to lead healthier and longer lives. Much of the renewed interest in NAD over the last decade can be attributed to the sirtuins, a family of NAD+-dependent protein deacetylases (SIRT1-7). Sirtuins have been shown to play a major regulatory role in almost all cellular functions. At the physiological level, sirtuins impact inflammation, cell growth, circadian rhythm, energy metabolism, neuronal function, and stress resistance.

By modulating NAD+-sensing enzymes, NAD+ controls hundreds of key processes from energy metabolism to cell survival, rising and falling depending on food intake, exercise, and the time of day. NAD+ levels steadily decline with age, resulting in altered metabolism and increased disease susceptibility. Restoration of NAD+ levels in old or diseased animals can promote health and extend lifespan, prompting a search for safe and efficacious NAD-boosting molecules that hold the promise of increasing the body's resilience, not just to one disease, but to many, thereby extending healthy human lifespan.

TREM2 as a Target to Enhance Immune Clearance of Amyloid in Alzheimer's Patients

Researchers have identified TREM2 as a target to potentially enhance the ability of immune cells in the brain to remove amyloid beta, solid deposits of misfolded proteins associated with the progression of Alzheimer's disease. Removal of amyloid beta remains the primary focus of the Alzheimer's research community, despite the continued lack of progress towards working therapies based on this approach. An increasing number of researchers are investigating alternatives to the existing approaches to amyloid immunotherapy, so far failing to achieve meaningful results in human trials.

The slow accumulation of amyloid beta and other metabolic waste in the brain looks a lot like the consequence of a slow failure of clearance mechanisms, as amyloid levels are actually quite dynamic from moment to moment. One candidate for this failure is the age-related deterioration of immune activity, in and of itself a very complex topic - which is one reason to think that therapies based on improved immune function might be helpful. Other candidates include failure of filtration of cerebrospinal fluid in the choroid plexus, or more recent views on the failure of cerebrospinal fluid drainage. Alzheimer's is a complex condition, and the brain is a complex organ.

Two new studies describe how TREM2, a receptor found on immune cells in the brain, interacts with toxic amyloid beta proteins to restore neurological function. The research, performed on mouse models of Alzheimer's disease, suggests boosting TREM2 levels in the brain may prevent or reduce the severity of neurodegenerative disorders including Alzheimer's disease. "Our first paper identifies how amyloid beta binds to TREM2, which activates neural immune cells called microglia to degrade amyloid beta, possibly slowing Alzheimer's disease pathogenesis. The second study shows that increasing TREM2 levels renders microglia more responsive and reduces Alzheimer's disease symptoms."

One of the hallmarks of Alzheimer's disease is the accumulation of amyloid plaques that form between neurons and interfere with brain function. Many drug companies have been working for years to reduce amyloid beta production to thwart Alzheimer's - but with minimal success. "TREM2 offers a potential new strategy. Researchers have known that mutations in TREM2 significantly increase Alzheimer's risk, indicating a fundamental role for this particular receptor in protecting the brain. This new research reveals specific details about how TREM2 works, and supports future therapeutic strategies to strengthen the link between amyloid beta and TREM2, as well as increasing TREM2 levels in the brain to protect against pathological features of the disease."

The first study showed that TREM2 binds quite specifically to amyloid beta. In particular, it connects with amyloid beta oligomers (proteins that bind together to form a polymer), which are the protein's most toxic configuration. Without TREM2, microglia were much less successful at binding to, and clearing out, amyloid beta. Further investigation showed that removing TREM2 downregulated microglial potassium ion channels, impairing the electrical currents associated with the activation of these immune cells. In addition, TREM2 turned on a number of mechanisms associated with the amyloid beta response in microglia.

In the second study researchers added TREM2 to a mouse model with aggressive Alzheimer's disease. They found that the added TREM2 signaling stopped disease progression and even restored cognitive function. As they learn more about how TREM2 modulates the amyloid signals that put microglia to work, the researchers have their work cut out for them. "It could be beneficial in early stages to activate microglia to eat up amyloid beta, but if you over-activate them, they may release an overabundance of cytokines (causing extensive inflammation) damaging healthy synaptic junctions as a side-effect from overactivation." Still, the ability to use the brain's existing immune mechanisms to clear amyloid offers intriguing possibilities.


Assessing Recent Changes in the Pace of Secondary Aging

Biological age, as opposed to chronological age, is driven by the intrinsic processes of primary aging, the accumulation of molecular damage outlined in the SENS rejuvenation research proposals, but also by the influence of the environment, secondary aging. The important contributions to secondary aging are excess visceral fat tissue as a consequence of diet, burden of infectious disease, lack of exercise, and smoking, acting through a range of mechanisms that overlap with the intrinsic processes of primary aging. There are others, but their effects are smaller and it is harder to see them in the data in comparison to the points above.

In the paper here, researchers make an effort to map recent changes in secondary aging, picking combinations of metrics from past data that might offer insight into the biological age of patients. I would say that there is little reason to expect primary aging to have altered significantly in the past few decades, given the landscape of medical technology, but it is certainly up for debate as to whether medications that control blood pressure and cholesterol levels might have some effect. They have certainly become more prevalent and effective over the time covered by the study data.

Overall this is an interesting exercise, but of little relevance to the future of aging. Gains from here on out will increasingly arise from the development of rejuvenation therapies that can repair the damage of primary aging, rather than from lifestyle improvements such as reduction in smoking or obesity. Greater potential gains in health and life span might be achieved through addressing primary aging; the scope of increased longevity through better lifestyle choices is far more limited. Our remaining healthy life span will be determined ever more by progress in rejuvenation biotechnology as time passes.

A new study suggests that at least part of the gains in life expectancy over recent decades may be due to a change in the rate of biological aging, rather than simply keeping ailing people alive. "This is the first evidence we have of delayed aging among a national sample of Americans. A deceleration of the human aging process, whether accomplished through environment or biomedical intervention, would push the timing of aging-related disease and disability incidence closer to the end of life. Life extension without changing the aging rate will have detrimental implications. Medical care costs will rise, as people spend a higher proportion of their lives with disease and disability. However, lifespan extension accomplished through a deceleration of the aging process will lead to lower healthcare expenditures, higher productivity, and greater well-being."

Using data from the National Health and Nutrition Examination Survey (NHANES) III (1988-19994) and NHANES IV (2007-2010), the researchers examined how biological age, relative to chronological age, changed in the U.S. while considering the contributions of health behaviors. Biological age was calculated using several indicators for metabolism, inflammation, and organ function, including levels of hemoglobin, total cholesterol, creatinine, alkaline phosphatase, albumin, and C-reactive protein in blood as well as blood pressure and breath capacity data.

While all age groups experienced some decrease in biological age, the results suggest that not all people may be faring the same. Older adults experienced the greatest decreases in biological age, and men experienced greater declines in biological age than females; these differences were partially explained by changes in smoking, obesity, and medication use. Slowing the pace of aging, along with increasing life expectancy, has important social and economic implications. The study suggests that modifying health behaviors and using prescription medications does indeed have significant impact on the health of the population.


Undoing Aging: An Interview with Aubrey de Grey

The Undoing Aging conference in Berlin is presently underway, a gathering of everyone who is anyone in the rejuvenation research community. It is hosted jointly by the SENS Research Foundation and the Forever Healthy Foundation, and is a unification of the varied themes of the past fifteen years of SENS conferences: the science of aging and its treatment from the earlier SENS conference series mixed with the industry, startup, and commercial development focus of the Rejuvenation Biotechnology series of recent years.

The first rejuvenation therapies to be implemented and shown to work, those based on clearance of senescent cells, are presently entering human trials and being carried forward to the clinic by startups. The next set of rejuvenation therapies, targeted at different mechanisms of aging such as cross-links and mitochondrial damage, are still in the laboratory, working their way towards completion. This is a time of transition, the birth of a new field of applied biotechnology, one that will grow to subsume most of the present medical community and provide services and products to every adult human being.

To commemorate the occasion, the Life Extension Advocacy Foundation volunteers have published a three-part interview with Aubrey de Grey of the SENS Research Foundation, the person who did the most to start this ball rolling back at the turn of the century. Later joined by a range of allies within and outside the scientific community, and then by a community of advocates and supporters, a bootstrapping process of growth towards the industry needed to bring an end to aging has been underway since. These are fairly lengthy interviews, and I'm not quoting more than a fraction of the whole here - you'll certainly find further interesting comments if you read the whole thing.

Undoing Aging With Aubrey de Grey Part One

Why did you choose Berlin and not California or elsewhere in the USA for the event?

Basically, because the suggestion came from our main German donor, Michael Greve, who is also the conference's main sponsor. Hard to argue with that!

Is SRF planning to make Undoing Aging into a recurring event, much like the Rejuvenation Biotechnology conferences in America?

We'll certainly be continuing to do both more science-centered events like Undoing Aging and the SENS Conferences, as well as more rejuvenation biotechnology industry-oriented events like the Rejuvenation Biotechnology series, but we haven't yet decided on the sequence and orientation of future meetings. We certainly want to maintain a strong conference presence in California, but it may be best to do that with smaller, more frequent events, such as the one we did with the California Life Sciences Association.

Recently, SRF has received significant donations amounting to over 7 million dollars. What priorities does SRF plan to address with this money?

First and foremost, we will be gearing up our existing programs in mitochondrial gene replacement, scaling up glucosepane research, rejuvenation biotechnology against cytosolic aggregates, and so on. We will also be initiating new ones; those are still being discussed with potential extramural collaborators, but you can expect some announcements later this year. They will all be within the same seven-strand framework that has defined SENS since the beginning. And after having sometimes in the past allocated nearly all of our available research budget at the beginning of the fiscal year and thereby limiting our ability to take advantage of new opportunities that arose later in the year, we will be maintaining a research reserve fund so that we are always poised to get good work funded year-round.

For anyone reading this who is thinking about doing the same as our recent donors, I will just say that we are a very long way from running out of productive ways to invest more money.

Undoing Aging With Aubrey de Grey Part Two

Regarding the use of senolytics, are you concerned about their potential to remove highly specialized cells like cardiomyocytes, which do not divide or do so very slowly?

Cells that don't divide (like cardiomyocytes and neurons) are far less likely to become senescent in the first place than cell types that divide; many of the main drivers of senescence are related to cell division. In the specific case of cardiomyocytes, there's already significant evidence in rodents that senolytics improve cardiac function overall. However, there is some reason for concern here, which is why we're already working to develop the next generation of senescent cell ablation therapies. The selectivity of senolytic drugs for senescent cells comes from the fact that they target the activity or expression of genes involved in cell survival, on which senescent cells are much more reliant than healthy cells under normal, unstressed conditions. But during times in which the cell is under stress, normal cells also rely on those same pathways to carry them through and give them time to recover. Future therapies can target truly senescent cells more selectively, and SENS Research Foundation is helping to advance those next-generation senescent cell therapies, even as UNITY Biotechnology prepares for human testing, through our investment in Oisín Biotechnologies.

Senolytic drugs gave mice increased healthy lifespan in experiments. Given that every living organism produces senescent cells the same way, could this mean that it may translate to humans?

Interventions that lead to gains in median lifespan only in laboratory mice, with no corresponding effect on a robust maximum lifespan (tenth-decile survivorship), still need to be heavily discounted when speculating on effects in humans. Interventions that only affect median lifespan primarily affect deaths in the first half of the lifespan - and here there is a critical difference between mice in a lab and modern humans, for whom medicine has already eliminated many causes of such early deaths, from vaccines (which also impact late-life mortality by reducing lifelong inflammatory burden), to surgery, to antibiotics, to drugs that more obviously affect middle-aged people.

The force of this reasoning is somewhat attenuated in the case of interventions like senescent cell clearance, which actually repair aging damage, than with interventions affecting environmental or metabolic risk factors driving "premature" disease (obesity, inflammation, cardiovascular risk factors, environmental toxins, etc). Still, you have to assume that the effect on lifespan of any single damage-repair intervention in isolation will be modest, based on the principle of the "weakest link in the chain": all the links are weakening over time, and shoring up only one of them still leaves the rest of the links damaged and ready to shear, whereupon the whole chain is broken. To move the needle on lifespan in modern humans, we have to push back on all of the cellular and molecular damage of aging, not just one form.

Regarding the breakdown of extracellular aggregates, what will you do if the first wave of treatments using antibodies is unable to repair the whole system?

Certainly, it's guaranteed that no first-generation SENS therapy will be able to repair every single contributor to any given category of aging damage - and it doesn't have to. All we have to do to reach "longevity escape velocity" is to remove or repair the specific forms of cellular and molecular aging damage within each category that meaningfully restrict our lives to the extremes of current lifespans. During the extra decades of healthy life that we'll then enjoy, scientists can then work to identify the constraints that limit life- and healthspan to those newly-expanded horizons. Accordingly, all SENS therapies will need to be iteratively improved; we will want safer and more effective ways to repair the damage targeted by earlier iterations of rejuvenation biotechnologies and also to repair additional specific targets within each category. It's only once those first therapies are developed and in use that we'll know what their specific limitations will be

Have you reviewed you position that nuclear mutations matter only in cancer in light of recent research results suggesting that certain ominous mutations in hematopoietic stem cells increase the risk of developing not only blood cancers (50 fold) but dying of all causes by 40%?

The research on this "clonal hematopoiesis" phenomenon is certainly provocative but doesn't ultimately change our view on this question. Remember first that it has never been our position that nuclear mutations matter only in causing cancer; at a minimum, they also matter in causing apoptosis ("cellular suicide," which denudes the body of functional cells with age, most importantly stem cells) and cellular senescence (ditto, plus the baleful effects of the senescence-associated secretory phenotype). And then remember that SENS is fundamentally an engineering approach to aging, focused on practical solutions rather than acquiring a full understanding of mechanistic details. Our position has been, therefore, that all the effects of nuclear mutations that meaningfully constrain current human lifespan/healthspan can be obviated by removing, repairing, or obviating the effects of mutations that are relevant to our health over the course of currently-normal lifespans: clearing senescent cells, replacing cells lost to apoptosis and senescence and other causes, and making the body impervious to cancer.

In clonal hematopoiesis, blood stem cells with one of a small number of mutations gain a selective advantage over blood stem cells with other genotypes, which allows them to "take over" the stem cell compartment. This isn't exactly what an oncologist would call "cancer," but it is a clear case of "too many cells" caused by nuclear mutations proliferating at the expense of their neighbors, which fits the operational criteria for the oncoSENS category. And the periodic purging of all native bone marrow stem cells and their wholesale replacement with fresh, mutation-free, cancer-proof ones - which would immediately eliminate clonal hematopoiesis - is already planned to be the very first clinical phase of the whole body interdiction of lengthening of telomeres (WILT) plan to pre-emptively shut down cancer.

Undoing Aging With Aubrey de Grey Part Three

Has your position on the relevance of telomere attrition changed since you first devised SENS, especially in the light of the recent results with fibrosis and your involvement with AgeX Therapeutics?

No. Let's start with the big picture. Neither I nor anyone sensible has ever suggested that telomere attrition has no functional effects in aging: telomere attrition causes cells to become senescent and runs down the proliferative capacity of stem cells, amongst other things. Nor have I suggested that there wouldn't be some short-term health benefits to activating telomerase or telomerase gene therapy in aging animals or animal models of age-related disease. The issue is rather that those short-term benefits come with the longer-term (and sometimes not so long-term) risk of increased rates of cancer.

So, why don't we see a plague of excess cancers in animal studies that show the benefits of telomerase-based treatments? Depending on the study, it's one or more of several reasons. The most common one is that such studies are usually too short-term. A related issue is that many of these studies involving animal models of age-related disease are actually done in quite young animals that have been damaged in some way that simulates aspects of an age-related disease. Because such animals are still quite young, they haven't yet lived long enough to have accumulated a high burden of the kinds of mutations that predispose cells to become cancerous. A third reason why many animal studies of telomerase treatments don't result in high reported rates of cancer is that the animals may actually be deficient in telomerase to begin with, such that telomerase gene therapies actually just restore the normal activity of telomerase in the animals.

The solution to problems caused by age-related attrition of telomeres is not to juice up telomerase to lengthen them again in often-damaged stem cells, but to take telomerase out of the picture, purge those defective stem cells, and replenish stem cell pools periodically with cancer-proofed, pristine replacement cells that are unable to replicate out of control.

You have been engineering glucosepane-eating bacteria that use enzymes effectively 'gifted' to them. Have the enzymes you identified demonstrated specificity to glucosepane?

We can say that Dr. David Spiegel's SRF-funded lab at Yale has identified some candidates, but we can't go into the details at this time. Still, expect some news on the commercialization front in the glucosepane space in coming months.

Given the state of immunotherapy, and taking into account the rate of progress in the field, how confident are you that OncoSENS may be unnecessary?

The recent progress in cancer immunotherapy has certainly made me much more optimistic than I was five years ago that new cancer therapies might hold off cancer for more than a very small number of years - but not that it might make whole body interdiction of lengthening of telomeres (WILT) redundant. If we had all the other components of a comprehensive panel of rejuvenation biotechnologies assembled and deployed, ongoing progress with these therapies might well give us a slightly longer runway along the path to "longevity escape velocity" than I had expected at the time. But only slightly; within an all-too-short few additional years, I expect that without WILT, the surging rocket of "longevity escape velocity" will still run headlong into a wall of cancer until we have a way to definitively defeat its evolutionary engine of selection and replication. At present, WILT is the sole foreseeable approach to doing that.

Which rejuvenation treatments can we reasonably expect to reach the clinic first?

If you don't count stem cell therapies (some of which are in clinical use, but not as rejuvenation biotechnology), it's a race between ablating senescent cells with senolytics (with UNITY Biotechnology expected to perform their first-in-human trials early next year) and one of the many immunotherapies targeting the intracellular or extracellular aggregates that drive the neurodegenerative diseases of aging.

Choose Wisely: Practical Applications of Philosophy in the Age of Cryopreservation

There are many people who subscribe to the idea that accurately preserving the fine structure of the brain on death, having that brain scanned and discarded, and the data of that scan later used to run a whole brain emulation is essentially no different from cold water drowning followed by successful resuscitation. There is a stop, and then a start. That the same pattern is running in a completely different system, and the original is destroyed, is immaterial: the pattern is the self. The rest of us would say that this individual died permanently with the destruction of the preserved brain, and the emulation is a copy - and possibly not even a continuous, surviving, single entity, depending on the implementation.

Which of these views you or I hold is entirely unimportant right up to the point at which it is possible to preserve the brain on death and have some choice about what happens next. Since we do presently live in the era of brain preservation by vitrification or, recently, vitrifixation, whether one holds a pattern identity view (the self is the pattern) or a continuity view (the self is the pattern as embodied in this particular set of matter) can turn out to be important. The former will kill you, if you let it steer your choices. Clearly I'm not the only one who feels that pattern identity beliefs have the potential to be dangerous to those who subscribe to them, as illustrated by this article on the options for near future development of improved methods of brain preservation.

As someone who is fully supportive of the ultimate goals of the cryonics enterprise, but still views the current state of the practice with some degree of skepticism, I make a point of acquainting myself with the latest evidence regarding the quality of cryonics procedures and their ability to preserve the foundations of a person's identity through time. Over the past two years or so, I have increasingly seen a recent achievement by 21st-Century Medicine (21CM) cited by some cryonics supporters as demonstrating the scientific validity of those procedures: namely 21CM's research on aldehyde-stabilized cryopreservation (ASC). This new technique has allowed them to win the Technology Prize awarded by the Brain Preservation Foundation (BPF) by demonstrating excellent preservation of brain ultrastructure. Were I to follow this line of reasoning, I could happily set aside my concerns about the adequacy of today's cryopreservation procedures, which had now been verified by scientific experts; the proper focus would now need to be on how to responsibly introduce those procedures into a clinical setting, for patients at the end of their lives who might request them.

It turns out, however, that things are not so simple. ASC is no doubt a step forward for the field of brain banking, and as its name indicates, it it is indeed a form of cryopreservation, since it involves vitrification of the brain at -135°C. Nonetheless, ASC does not count as cryonics, insofar as it uses a fixative solution prior to vitrification and cooling, which could potentially preclude revival of the original biological brain (an essential part of cryonics as traditionally understood). And indeed, biological revival with the help of future technology is not a priority for the Brain Preservation Foundation (BPF)'s president, Dr. Kenneth Hayworth. Rather, he envisages brain preservation as conducive to life extension via mind uploading: a process that would involve cutting the preserved brain into thin slices, scanning each slice, and feeding the resulting data to an advanced computer that would thereby be able to map out the entire network of neural connections in the person's original brain, and ultimately to emulate that person's mind. This is quite different from cryonics.

The BPF's commitment to holding brain preservation research to the highest standards of scientific rigour is laudable, and worth emulating. Nonetheless, for those interested in brain preservation with a view to enabling life extension, supporting cryonics-specific research remains the safer bet. We should not simply rely on the BPF's approach if our goal is to try and save those whom medicine in its current state cannot restore to life and health.

To see why this is so, let us begin by noting the two main philosophical theories of personal identity through time that are relevant when discussing the respective merits of cryonics and mind uploading in this context. The first one, which we can call the "Physical Continuity" (PhyCon) theory, asserts that a person is identical with the physical substratum from which her mind emerges: that is to say, her brain, with its intricate web of neurons and synaptic connections. The second relevant theory can be referred to as the "Psychological Continuity" (PsyCon) theory. Roughly speaking, it says that you are to identical with the set of psychological features (memories, beliefs, desires, personality traits, etc.) that constitutes your mind. On this view, preserving you after you have been pronounced dead requires ensuring the persistence of enough of those psychological features, in an embodied mind of some sort (but one that need not be embodied in your current biological brain).

If that is the case, what is the prudent choice to make for those who wish to promote life extension through brain preservation? I submit that traditional cryonics is the more prudent option to pursue. This can be demonstrated using a simple argument that considers what the implications are if we assume that PhyCon and, respectively, PsyCon are true. Suppose first that PhyCon is true. If so, a cryonics procedure carried out properly will save a person's life, whereas using a technique like ASC that compromises the brain's potential for viability, followed by destructive scanning and uploading, will kill that person. If PsyCon is true, on the other hand, both methods can ensure survival. Indeed, adequate cryonic preservation of a person's brain would also preserve the ultrastructure grounding the various psychological features that defined that person.

None of this is meant to imply that the work of the BPF is without merit. On the contrary, the Foundation's approach demonstrates a number of virtues that can provide a model for the cryonics movement to follow. These include a commitment to rigorously and impartially evaluating the quality of brain preservation procedures, in accordance with the standards of scientific peer-review. Another example is the BPF's successful effort at crowdfunding its incentive prizes for brain preservation research, such as the two prizes won by 21CM.


Lysosomal Aggregates Linked to the Age-Related Decline of Neural Stem Cells

Reinforcing the SENS rejuvenation biotechnology view of the importance of lysosomal aggregates in aging, researchers here demonstrate a link between lyososomal function and the ability of neural stem cells to support brain tissue. Lysosomes inside cells are recycling machines, packed with enzymes capable of breaking down near everything they will encounter. They are the ultimate destination for damaged proteins and other broken cellular structures. Unfortunately, lysosomes do encounter molecular waste that they cannot handle, and long-lived cells become ever more burdened by damage as their lysosomes falter and become bloated. The processes of recycling and cellular maintenance back up and run down, and the cells become dysfunctional.

The solution envisaged by the SENS Research Foundation is to build therapies capable of safely breaking down the unwanted contents of lysosomes. The most promising way forward appears to be mining the bacterial world for enzymes that might serve as a starting point. The known resilient lysosomal wastes do not accumulate in graveyards, so we know those bacteria and their useful molecular tools are out there, waiting to be discovered. The first SENS program to work along these lines successfully discovered a number of candidate enzymes that proceeded to further development, and are currently at various stages in that process.

Young, resting neural stem cells in the brains of mice store large clumps of proteins in specialized cellular trash compartments known as lysosomes. As the cells age, they become less proficient at disposing of these protein aggregates, and their ability to respond readily to "make new neurons" signals wanes. Restoring the ability of the lysosomes to function normally rejuvenates the cells' ability to activate, the researchers found. "We were surprised by this finding because resting, or quiescent, neural stem cells have been thought to be a really pristine cell type just waiting for activation. But now we've learned they have more protein aggregates than activated stem cells, and that these aggregates continue to accumulate as the cells age. If we remove these aggregates, we can improve the cells' ability to activate and make new neurons. So if one were able to restore this protein-processing function, it could be very important to bringing older, more dormant neural stem cells 'back to life.'"

Researchers isolated several populations of cells for study from the brains of both young and old mice, including resting neural stem cells, activated neural stem cells, and the neural cell progenitors that arise from activated stem cells. They found that resting stem cells expressed many lysosome-associated genes, while activated stem cells expressed genes associated with a protein complex involved in protein destruction called a proteasome. Strict control of production and disposal allows cells to maintain the necessary protein inventory to carry out needed cellular functions.

"The fact that these young, pristine resting stem cells accumulate protein aggregates makes us wonder whether they actually serve an important function, perhaps by serving as a source of nutrients or energy upon degradation." Old resting stem cells express fewer lysosome-associated genes and begin to accumulate even higher levels of protein aggregates. "It's almost as if these older cells lose the ability to store, or park, these aggregates. We found that artificially clearing them by either activating lysosomes in older cells or subjecting them to starvation conditions to limit their protein production actually restored the ability of these older resting stem cells to activate. We'd like to know whether the aggregated proteins are the same in the young and old cells. What do they do? Are they good or bad? Are they storing factors important for activation? If so, can we help elderly resting stem cells activate more quickly by harnessing these factors? Their existence in young cells suggests they may be serving an important function."


A Start on Mapping Biomarkers of Cellular Senescence by Tissue and Age

Cellular senescence is one of the root causes of aging. Cells enter a senescent state in response to damage or the end of their replicative life span, and near all quickly self-destruct or are destroyed by the immune system. Others enter senescence to assist in regenerative processes following wounding, again being destroyed soon afterwards. Senescent cells that linger are a real problem, however. They generate harmful signaling that produces chronic inflammation, destructively remodels tissue structures, and changes the behavior of nearby cells for the worse. The accumulation of senescent cells over the years directly contributes to the progression of age-related dysfunction, disease, and risk of death.

Just how many senescent cells is any given individual burdened with, however? What should we expect from this cause of aging at a specific age? Is it negligible at 40 or 50, with a sudden leap to significant levels at 60? Does the answer vary by tissue type? How do the usual health-associated lifestyle choices affect these numbers? Are senescent cells significantly different from tissue to tissue in terms of the signals they generate and the harm done?

The answers to these questions are not yet established in any robust way, but the development of therapies capable of destroying senescent cells is proceeding regardless - there is plenty of evidence to show that removing these cells is beneficial, even without the greater insight into the fine details. This more detailed information is important, however, when it comes to the energy with which any particular individual should pursue access to the first generation of senolytic therapies capable of destroying senescent cells, and where those groups involved in therapeutic development should focus most of their attention.

One of the paths to a better understanding of how the burden of cellular senescence progresses with age, and how that progress varies by tissue type, is the production of a more detailed mapping of biomarkers of senescence. The open access paper here is an example of this sort of work, initially focused on mice. Better and more discerning markers of cellular senescence and the harms it creates will help to validate existing senolytic therapies and steer the development of new and better approaches.

Age- and Tissue-Specific Expression of Senescence Biomarkers in Mice

Cellular senescence plays a complex role, both beneficial and deleterious, in biological processes such as embryonic development, wound healing, tissue regeneration, and tumor suppression, as well as age-related disorders. Senescent cells accumulate within aged tissues and at sites of age-related pathology in vivo, and potentially contribute to the age-related decline of tissue function by affecting the growth, migration and differentiation, of neighboring cells, impacting overall tissue architecture, and promoting chronic inflammation. Indeed, studies on both progeroid and naturally aged mice showed that selective elimination of p16Ink4a-expressing senescent cells increased healthspan and lifespan. Thus, the selective elimination of senescent cells (senolytics) or the disruptions of the senescence-associated secretory phenotype (SASP) program have been developed as potential therapeutic strategies against aging.

However, while p16Ink4a expression has been used as a classical senescence biomarker, no biomarker of senescence identified thus far is entirely specific to the senescent state. Thus, due to the lack of robust biomarkers of cellular senescence in vivo, the precise extent of senescent cell accumulation in aged animals and the functional outcome of such an accumulation, along with the exact target cells of, and removal by, senolytics, remain unclear. Surprisingly, a systematic multi-tissue in vivo study of senescence markers during aging has not been conducted in wild-type animals.

In the era of senolytics, it becomes imperative to develop robust biomarkers of senescence in vivo for preclinical trials, especially with several senolytics now nearing human clinical studies. As a first step, in this study we profiled the expression of a panel of known molecular markers of senescence in multiple tissues in mice at multiple ages, ranging from young (4 months) to very old (30 months). The results demonstrate that the secretory profiles and classical hallmarks of cellular senescence in aged tissues are highly variable and complex, suggesting that a systematic and concerted effort is needed to develop robust biomarkers of senescence for the identification, quantification, and monitoring of senescent cells in vivo.

The wide diversity in tissue-specific profiles we observed was striking. Nevertheless, the matrix metalloproteinase Mmp12 represents a robust SASP factor that showed consistent age-dependent increases in expression across all tissues analyzed in this study. It has been demonstrated that mice lacking Mmp12 are protected from vascular injury, M2 macrophage accumulation, and perivascular heart fibrosis. Together with our data, this finding suggests that Mmp12 upregulation with age has a deleterious impact on heart function.

In this study, we did not observe significant age-dependent upregulation of the prominent SASP cytokine Il6 in any tissue, although an upward trend was observed that was consistent in magnitude with previous observations in the heart and kidney. This modest age-related upward trend could be explained by a previous report which demonstrated that senescent cell-secreted IL-6 acts in an autocrine manner, reinforcing the senescent state, rather than inducing senescence or promoting dysfunction in neighboring cells.

The decreased expression of Il6 with age we observed in the hypothalamus could be indicative of a lack or loss of senescent cells in that tissue with age. In support of this interpretation, p16Ink4a expression was non-detectable in the hypothalamus at any age. Taken together, these results suggest that some other age-related process results in the increased expression of the pro-inflammatory factors Il1b, Mmp12, Cxcl1, and Cxcl2 observed in the aged hypothalamus. Conversely, p16Ink4a expression was upregulated with age in all other tissues analyzed, consistent with previous reports, and thus reinforcing the importance of p16Ink4a as a biomarker of tissue aging.

Questions still remain, however, regarding the ultimate identity of the cells targeted for senolytic elimination in previous studies, as it has been demonstrated repeatedly that p16Ink4a expression is not exclusive to senescent cells, and thus does not represent an unequivocal target for senolytic therapies. Interestingly, however, CDKN2A (the gene that encodes p16Ink4a) was one of the top human genes that exhibited elevated expression with age, in 6 out of 9 tissues, including subcutaneous adipose, tibial artery, lung, skeletal muscle, tibial nerve, and whole blood, as detected by RNA-seq analysis. Thus, utilizing p16Ink4a-expressing cells as a biomarker of tissue aging and a target of senolytic therapies could still prove to be an effective strategy in the future treatment of age-related diseases in humans.

Stem Cell Signaling from Gums Might be Used to Accelerate Healing in Other Tissues

Why do gums heal more rapidly than skin? These research results follow that question down into the cellular biochemistry of regeneration and stem cell activity, in search of the important differences between gums and skin. The authors have uncovered a potentially interesting mechanism in the signaling of stem cells present in gum tissue, one that might be exploited to speed up healing of wounds elsewhere in the body. Investigations of stem cell signaling and its role in regeneration are a growing focus in the research community. Many classes of future regenerative therapies may well do away with cell transplants in favor of delivering only the signals generated by those cells.

Ever notice how a cut inside the mouth heals much faster than a cut to the skin? Gum tissue repairs itself roughly twice as fast as skin and with reduced scar formation. One reason might be because of the characteristics of gingival mesenchymal stem cells, or GMSCs, which can give rise to a variety of cell types. "This study represents the convergence of a few different paths we've been exploring. First, we know as dentists that the healing process is different in the mouth; it's much faster than in the skin. Second, we discovered in 2009 that the gingiva contains mesenchymal stem cells and that they can do a lot of good therapeutically. And, third, we know that mesenchymal stem cells release a lot of proteins. So here we asked, how are the gingival mesenchymal stem cells releasing all of these materials, and are they accelerating wound healing in the mucosal tissues?"

From earlier work it was clear that mesenchymal stem cells perform many of their functions by releasing signaling molecules in extracellular vesicles. So to understand what distinguishes mesenchymal stem cells in the gingiva from those in the skin, researchers began by comparing these extracellular vesicles between the two types. They found that the GMSCs contained more proteins overall, including the inflammation-dampening IL-1RA, which blocks a proinflammatory cytokine.

Next the team zoomed in to look at what might be controlling the release of IL-1RA and other cytokines. They had a suspect in the protein Fas, which they had earlier connected to immune regulation. They found that in gingival MSCs had more Fas than skin MSCs, and that mice deficient in Fas had reduced IL-1RA as well as reduced secretion of IL-1RA. Further molecular probing revealed that Fas formed a protein complex with Fap-1 and Cav-1 to trigger the release of small extracellular vesicles. To identify the connection with wound healing, they examined wound tissue and found that IL-1RA was increased in GMSCs around the margins of wounds. Mice lacking IL-1RA or in which the protein was inhibited took longer to heal gingival wounds. In contrast, when the researchers isolated IL-1RA that had been secreted from GMSCs and injected it into wounds, it significantly accelerated wound healing.

These findings may have special significance for people with diabetes, a major complication of which is delayed wound healing. In the study, the researchers found that GMSCs in mice with diabetes were less able to secrete extracellular vesicles compared to GMSCs in healthy mice, and their GMSCs also had less IL-1RA secretion. Introducing extracellular vesicles secreted from the GMSCs of healthy mice reduced wound healing time in diabetic mice. "Our paper is just part of the mechanism of how these stem cells affect wound healing, but I think we can build on this and use these cells or the extracellular vesicles to target a lot of different diseases, including the delayed wound healing seen in diabetic patients."


An Interview with a Buck Institute Neuroscientist

This short, interesting interview is with one of the Buck Institute neuroscience researchers with an interest in cellular senescence as a component of degenerative aging. Exhibited here is perhaps the most optimism that I recall seeing in public comments from any of the Buck Institute faculty - but if I were involved in cellular senescence research, I'd be fairly optimistic as well. This part of the field is progressing rapidly, producing solid evidence of the association of cellular senescence with the development of age-related disease, and of the benefits that can be obtained by removing these unwanted cells.

The field seems to have agreed upon nine hallmarks of aging, do you believe it is feasible for us to one day be able to treat all of them?

Ten years ago I would have wondered how feasible this was, but based on the progress that has been made in the last few years I do think it is plausible that we will be able to address each of those pillars of aging and that by addressing these underlying mechanisms that drive aging we are going to be able to treat age-related disease. I think we have a tendency to view age-related diseases in silos but many of these disorders have a lot in common. I think we are on the brink of solutions to these problems, not in the next decades, but within years.

Could you briefly describe senescence and its impact on neurodegenerative diseases?

Senescence is a process in cells that stops cells from dividing, it gets activated when certain types of damage occurs. From an evolutionary perspective, cellular senescence is there to prevent damaged cells from undergoing the kind of rapid division that leads to tumors. This is great in the short term, but if they persist they give off toxic pro-inflammatory factors which can damage neighboring tissues. A lot of research in the field goes into understanding this process and what we can do to prevent this toxic effect.

For a long time the field of neuroscience ignored senescence because everyone just looked at the neurons, which don't divide. However we also started looking at the other cell types in the brain that do divide, namely astrocytes. They are a major support cell in the brain that also secrete growth factors that help neurons grow and communicate, they are also much more abundant than neurons. We then discovered that these cells do undergo senescence by looking at post-mortem tissue from Parkinson's disease brains and found astrocytes that had become senescent. We showed in animal models that if we could remove aggregations of these cells we could slow some of the disease process. This is very exciting because it means we can push this strategy forward into human clinical trials as it is a possible therapeutic strategy that has not been explored before. We were one of the first labs to look into this but now a lot of other labs around the world are jumping into cellular senescence to try and tackle age-related disorders.

You also explore the protein TFEB to boost lysosomal function and autophagy?

We were looking at a young-onset model of Parkinson's disease that has a mutation in the Parkin gene which marks damaged mitochondria for disposal via autophagy. We then learned that one of the major factors in that process is this transcription factor called TFEB, which is a master regulator of autophagy. This has now become a potential target for treating Parkinson's and Alzheimer's because these diseases are the result of protein build ups and dysfunctional mitochondria. It is thought that if we boost TFEB then cells will be able to better dispose of these protein build ups. We screened a number of compounds that boosts TFEB in the brains of these animals are now trying to move this forward to clinical trial.