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  • Destroying Existing Microglia is Necessary for Replacement Strategies to Work
  • A Two Part Interview with Greg Bailey of Juvenescence
  • Age-Related Downregulation of Rubicon Causes Excessive Autophagy in Adipocytes, Contributing to Metabolic Dysfunction
  • Aging as a Target is a New Therapeutic Frontier
  • Investigating the Cause of Irregular Heartbeat Following Cell Therapy for Regeneration of Heart Tissue
  • The Prospects for LANDO Upregulation as a Treatment for Alzheimer’s Disease
  • DOK7 Gene Therapy Regrows Neuromuscular Junctions to Improve Aged Muscle Function
  • Higher Body Mass Index Correlates with Reduced Cerebral Blood Flow
  • IL-6 and TGFβ1 Upregulation with Age Cause Detrimental Changes in Hematopoiesis
  • Cartilage Regrowth: Steering Microfracture to Provoke Regeneration of Fully Functional Cartilage
  • A Gene Therapy Approach to Clearing Persistent Herpesviruses
  • Increased Levels of Methylmalonic Acid May Raise the Risk of Metastasis in Older People
  • Evidence for Klotho to Act on Life Span in Part via Resistance to Hypertension
  • FOXO3a Suppresses Genomic Instability
  • Human Studies Link Calorie Restriction to Improved Cardiometabolic Status

Destroying Existing Microglia is Necessary for Replacement Strategies to Work

Today’s open access research is a demonstration in mice of approaches to replace near all microglia in the central nervous system. Microglia are innate immune cells of the brain, involved not just in destroying pathogens and errant cells, but also in ensuring the correct function of neural connections. With the progression of aging, their behavior shifts to become more harmful and inflammatory, and their numbers include ever more senescent cells. Senescent cells generate tissue dysfunction and chronic inflammation via the senescence-associated secretory phenotype, but beyond that microglia tend to adopt a more aggressive and inflammatory set of behaviors even when not senescent. This detrimental change is the consequence of some mix of persistent infection, protein aggregates, and other forms of the underlying molecular damage that drives aging.

Microglial dysfunction contributes meaningfully to age-related neurodegeneration, as illustrated by the benefits produced in animal models by the selective destruction of senescent microglia. That approach has turned back the tau pathology characteristic of Alzheimer’s disease in mice, for example. There is also evidence for inflammatory microglia to be involved in the progression of Parkinson’s disease.

More than just the senescent cells need to be replaced, or otherwise have their behavior changed for the better, however. Approaches involving clearance of a large fraction of microglia, and allowing them to regenerate thereafter, have seemed viable. Efforts to replace microglia with transplanted cells have proven challenging, however: even hematopoietic stem cell transplantation, such as via a bone marrow transplant, doesn’t replace more than a small fraction of the existing microglia. As researchers here demonstrate, it is necessary to first destroy near all microglia in order to leave an empty niche in the brain that will generate signals telling the body to replace these cells. Will replacement be necessary for the treatment of age-related microglial dysfunction, rather than genetic dysfunction? It seems plausible that hematopoietic stem cell replacement will be adopted as an approach to immune system rejuvenation, so why not pair it with clearance of cell populations that should be replaced?

Efficient Strategies for Microglia Replacement in the Central Nervous System

Microglia are important immune cells in the central nervous system (CNS). Dysfunctions of gene-deficient microglia contribute to the development and progression of multiple CNS diseases. Microglia replacement by nonself cells has been proposed to treat microglia-associated disorders. However, some attempts have failed due to low replacement efficiency, such as with the traditional bone marrow transplantation approach.

Engrafted cells in previous transplantation approaches do not extensively proliferate in the recipient brain, which explains the low efficiency of transplantation. Indeed, the proliferation-dependent turnover rate of microglia is rather slow in homeostatic conditions. In contrast, we have demonstrated that residual microglia exhibit an astonishing proliferation capacity after pharmacological depletion (~99%). This potentially suggests that microglial proliferation relies on an empty microglial niche. We therefore reasoned that the microglia-free niche is a vital prerequisite for successful engraftment of nonself microglia (or microglia-like cells). Colony-stimulating factor 1 receptor (CSF1R) is essential for microglia survival. PLX5622 is a CSF1R inhibitor with improved specificity compared to its analog, PLX3397. To create the microglia-free niche, we utilized PLX5622 to inhibit CSF1R.

We then developed highly efficient approaches for nonself microglia replacement that are effective in the adult normal mouse at the CNS-wide scale. First, microglia replacement by bone marrow transplantation (mrBMT) is capable of inducing allografted bone marrow cells (BMCs) to differentiate into microglia-like cells in the entire CNS, replacing 92.66% of resident microglia in the brain, 99.46% in the retina, and 92.61% in the spinal cord, respectively. Second, microglia replacement by peripheral blood (mrPB) is able to induce peripheral blood cells (PBCs) to microglia-like cells and replace 80.74% of resident microglia in the brain and 74.01% in the retina. Third, to precisely manipulate microglia in a specified brain region without affecting the rest of the brain, we further developed microglia replacement by microglia transplantation (mrMT). The engrafted microglia via mrMT resemble the natural characteristics of naive microglia.

When determining superiority of a strategy, replacement efficiency and source availability are the two most important dimensions to take into consideration. Among the three microglia replacement approaches, mrBMT achieves the highest replacement efficiency – 92.66% in the brain, 99.46% in the retina, and 92.61% in the spinal cord. However, mrBMT uses the BMC as the donor cell, which is clinically hard to acquire due to the invasive nature of the procedure and the aversive response from the donor. Such scarce availability of the source is likely to restrict its potential of becoming a widely used standard clinical method for microglial replacement. On the other hand, mrPB greatly broadens the donor source by using PBC, the largest donor cell pool, while maintaining high replacement efficiency CNS wide, just slightly inferior to mrBMT. Abundant availability of donor cells and the relatively high efficiency of cell replacement make mrPB an ideal approach to manipulate microglia at the whole-CNS scale.

A Two Part Interview with Greg Bailey of Juvenescence

Juvenescence and Life Biosciences are presently the two large business development companies in the growing longevity industry. They act much like venture funds, in that they create or take controlling positions in biotech startups, but are organized as companies in structure, with the ability to later go public. The resulting entity looks much like a Big Pharma company with many subsidiaries. It is quite possible to do this at a smaller scale and bootstrap towards much the same end goal – see Ichor Therapeutics and its portfolio companies, for example.

Greg Bailey is one of the cofounders of Juvenescence, alongside Jim Mellon and Declan Doogan. Today I’ll point out a recent two part interview, covering a mix of what the company is doing and the present state of investor interest in the longevity industry. Juvenescence is very visible as a company, since Jim Mellon does a great deal of work to spread his views on (a) the merits of working to extend the healthy human life span, and (b) the enormous returns on investment that will be generated by even early and limited success in treating aging as a medical condition. The longevity industry will become the largest and most beneficial industry in the world, given that every adult human being is a potential customer, and without health and life little else matters. The present medical industry, the business of treating ill people, will become a sidebar to the far more extensive provision of preventative treatments to control aging.

Perceptions among investors and the public at large as to how far along we are towards that goal will rise and fall with the year to year successes and failures of Juvenescence and similar companies. In a young industry, whether it is fair or not, the fortunes of all fellow travelers are affected by the performance of the household names.

Juvenesence (1): from strength to strength in R&D and investment firepower

Juvenescence seems to be moving from success to success; we started by asking Greg to bring us up-to-speed on news so far.

Well, the good news is that we closed morphoceuticals [spontaneous tissue regeneration therapy] this year; the bad news is that they need access to their labs. So COVID-19 has slowed the process on our development of spontaneous regeneration of a limb or an organ using bioelectrics – but we have an amazing team working on this and hopefully will be back in the lab soon. We recently closed a joint venture with a company called G3, and the new company is called Juvenomics. Basically they have 2500 people’s complete omics data – proteome, genome, etc., and they have another 4000 patients that they have partial omics data on. We have the use of all that data to try and generate new drugs and drug combinations for anti-aging and modifying aging. So it’s really exciting to get access to that data, and our extraordinary machine learning team, which has made great strides on the data science side, will be the group that will use this data to create drugs, drug combinations and repurpose drugs.

We are on track to launch our first product in the end of September, Metabolic Switch; it is a ketone ester that in mammals is geroprotective, neuroprotective, and cardioprotective. I couldn’t be more excited about that launch! And it should be relatively affordable, if you have a monthly subscription it’s quite reasonable. We signed our second product for Juvenescence Life, our non-RX division. This a product that increases autophagy. It will improve cognition, boost your immunity, help your bones, your cardiac health, your skin, hair … So I’m really excited about adding that one; hopefully we will be able to launch next year (and hopefully is the operative word there), which would be great news.

And how about research?

Our RX Division continues to move forward; it looks like we’re going to have two to four drugs in the clinic next year and we are starting human trials for obesity, cachexia, and immunometabolism, fibrosis, as well as combinations of those products – when products move into trials, that’s a time I find really exciting. In our regeneration division, LyGenesis starts its phase 1/2A clinic, to be able to regenerate a liver using lymph nodes, and that starts this year. We are on track to try and undertake thymus replacement using the lymph nodes, over the next few years. This is very exciting and would obviously be a great step forward for immuno-resilience, since it involutes at 3% a year every year after you turn 20. This is possibly why, unfortunately, 70 or 80-year-olds are dying from COVID-19.

When we spoke last we understood that you were fundraising – how’s that going?

We’re going out to raise 150m in our C round, which hopefully will be a prelude to an IPO, and, of course, we’re talking to banks, to make sure that we have a balanced syndicate for that initiative. So all in all, it’s been a little busy! There’s still this wild discrepancy between how much money is made available to social apps and media apps and one of the most powerful transformative scientific opportunities that humanity has ever had to modify aging. I think it’s like 10 times, maybe 20 times discrepancy in capital available from VCs, so, hopefully, the VCs and the sophisticated investors will understand that work to modify aging is happening now, not in 10 years’ time.

Juvenescence (2): “It’s all about prevention, and that’s incredibly disruptive”

Are there other issues that you need to educate investors about, or are they becoming better informed as time goes on?

It’s going in the right direction; there’s already a difference response from when I talked to people in March compared with now. Maybe I’m getting better at telling the story, or maybe the company is moving along, and obviously the fact that biotech is on fire is certainly very helpful. However, I think that what still needs an enormous amount of education about is the scale of things. Tackling aging is very different from manufacturing a drug for breast cancer, cardiac disease, or inflammatory bowel disease. The population for breast cancer in Europe is probably in the 400,000s, but 400 million Europeans are getting older – it’s just completely different metrics.

Most investors appear to prefer seed-to-early-stage investing; have you found this to be the case in your networks?

This is about talking to the banks again; this is not going to be biotech investors, because they’ll think it’s early stage and worth nothing. They don’t understand that the patient population is 7.8 billion people. So, it’s going to be thematic investors, ESG people [Environmental, Social, and Governance], sustainable, environmental investors; it’s going to be funds who understand the diversity of having non-RX, RX, and machine learning. It’s a retail event, so crowd-funding has an opportunity to do well. Most people are petrified of biotech, but it’s no different from mining: put some money in, if you’re lucky you find gold; if you’re not lucky, you don’t. The share price either goes up if you found gold, or drops if you didn’t. If we have a clinical trial, and it’s run by smart, educated people, then it has a better-than-average chance of it being positive and, if it’s right, there’s a 10x return; historically, this is what we saw at Medivation and Biohaven, predecessor companies with which I’ve been involved.

Age-Related Downregulation of Rubicon Causes Excessive Autophagy in Adipocytes, Contributing to Metabolic Dysfunction

Autophagy is a vital collection of cellular maintenance processes in which proteins and structures are broken down and recycled for their component parts. In short-lived laboratory species, dysfunctional autophagy shortens life span, while increased operation and efficiency of autophagy – as occurs in response to forms of stress such as heat, exercise, and calorie restriction – slows aging and extends life span.

The usual high level view of aging and autophagy is that autophagic activity declines with age, and that this loss of function contributes to cell and tissue dysfunction, and thus also to age-related disease and mortality. The picture is more complex, however. Different component mechanisms of autophagy decline in different ways and at different paces in different tissues, and this is a distinct issue from the question of whether or not autophagy is running at a given pace. The operation of autophagy is actually upregulated with age in at least some tissues. Too much autophagy can cause issues that are just as problematic as those resulting from too little autophagy, because it destroys necessary protein machinery in the cell, thus disrupting normal function.

Today’s research materials present an interesting example of the perils of too much autophagy. Here, this is specifically occurring in fat cells, and the researchers involved identify a protein that appears to regulate this excessive autophagy in older fat tissue. It is known that fat cells change their behavior for the worse with age, and the changes in autophagy noted here may be one of the more important mechanisms in this aspect of aging.

Is turning back the clock in aging fat cells a remedy for lifestyle diseases?

“Adipocytes produce hormones and cytokines that regulate the function of other metabolic organs. Age-related changes in adipose tissue result in metabolic disorders that are closely associated with life-threatening cardiovascular diseases. However, no one really knows what causes adipocyte dysfunction in aged organisms.” The research team decided to focus on autophagy, the process used by cells to eliminate unwanted or dysfunctional cellular components. Previous studies had shown that autophagy plays an important role in the prevention of various age-related disorders and is likely to be involved in the aging process. But most pertinent was the finding that autophagy is essential for the normal function and longevity of normal organs, such as liver or kidney.

“We previously showed that a protein called Rubicon, which inhibits autophagy, is upregulated in aging tissues. We therefore hypothesized that Rubicon likely accumulates in aged adipocytes, decreasing autophagic activity and contributing to the onset of metabolic disorders.” Surprisingly though, the researchers found that Rubicon levels were actually decreased in the adipose tissue of aged mice, resulting in increased autophagic activity. “As a result, the mice developed lifestyle diseases such as diabetes and fatty liver and had significantly higher cholesterol levels, despite being fed the same diet as control animals.” The researchers went on to identify the specific proteins affected by the increased levels of autophagy, showing that supplementation of these proteins restored adipocyte function.

Age-dependent loss of adipose Rubicon promotes metabolic disorders via excess autophagy

The systemic decline in autophagic activity with age impairs homeostasis in several tissues, leading to age-related diseases. A mechanistic understanding of adipocyte dysfunction with age could help to prevent age-related metabolic disorders, but the role of autophagy in aged adipocytes remains unclear. Here we show that, in contrast to other tissues, aged adipocytes upregulate autophagy due to a decline in the levels of Rubicon, a negative regulator of autophagy. Rubicon knockout in adipocytes causes fat atrophy and hepatic lipid accumulation due to reductions in the expression of adipogenic genes, which can be recovered by activation of PPARγ. SRC-1 and TIF2, coactivators of PPARγ, are degraded by autophagy in a manner that depends on their binding to GABARAP family proteins, and are significantly downregulated in Rubicon-ablated or aged adipocytes. Hence, we propose that age-dependent decline in adipose Rubicon exacerbates metabolic disorders by promoting excess autophagic degradation of SRC-1 and TIF2.

Aging as a Target is a New Therapeutic Frontier

My attention was recently drawn to an open access commentary on the present early stage of the scientific initiative to treat aging as a medical condition. It was published earlier this year, and slipped past my notice amidst all of the other interesting papers emerging at the time. It is illustrative of a number of similar commentaries, in scientific journals, at conferences, and so forth, and is reflective of the present tenor of discussion among researchers. The scientific community is largely optimistic about the potential to intervene in the aging process, even if opinions vary widely as to how hard it will be, from a technical perspective, how much of a benefit to health and life span we can expect to engineer over the next few decades, and how much of a roadblock to progress is presented present systems of regulation, none of which yet recognize aging as a legitimate target for medical development.

We live in interesting times, witnessing the emergence of a new and very important field of scientific endeavor, concurrent with the emergence of a new and energetic medical industry of longevity, a period in which previously small scientific and advocacy communities blossom into large and earnest movements that overtake and transform existing institutions. The last five years and the next five years are a tipping point in the slow, decades-long cycles of business and technology, the prior phase of ineffective approaches to the diseases of aging giving way to and era of new biotechnologies and new approaches that will ensure longer, healthier lives for all. To my eyes that transformation is best explained as a shift to targeting the causes of aging, to repair the underlying damage to cells and tissues, rather than trying to treat the symptoms of aging caused by that damage. The next hundred years of medical development will be the focused on the control and reversal of aging; longevity assurance will be the defining industry of the 21st century, ultimately larger than all of the rest of medicine put together.

Aging: therapeutics for a healthy future

Many consider aging as a purely chronological phenomenon; it is an immutable fact that we all get older. However, this is a simplification as individuals all age functionally in different ways and the concept of “biological aging” is more relevant than chronological aging. When we consider biological aging, we have a therapeutic target, not simply targeting getting old, rather treating physiological decline that is manifested by dysfunction and morbidity in late life.

When biological aging becomes pathological, it can be considered as a failure of homeostasis. There is a progressive component, but ultimately a point is reached at which there is inability to counter the amassed toll on the body of time-dependent accumulation of cellular damage (DNA mutations, protein misfolding, oxidative stress, etc.), which occurs throughout life. With age, cellular processes (including stress response pathways invoked by damage) become less efficient, ultimately leading to cellular death and irreversible consequences. At younger ages, the body is able to mount compensatory responses and life is healthy. During middle age, the body’s ability to maintain homeostasis declines, resulting in chronic diseases that accelerate the gradual degradation of life quality, resulting in severe detriments, frailty and, eventually, mortality. There are many ways that a homeostatic balance can be maintained, giving many opportunities for development of therapeutics.

Development of drugs in recent years has focused almost entirely on selectivity and specificity with a primary goal to reduce the risks of any side effects. Such an approach is highly valid when considering specific indications targeting selected organs. Aging is different; the body ages systemically, although not necessarily uniformly, and it may indeed be preferential to have broad, systemic anti-aging effects. Consideration of systemic therapies, broadly distributed targets or organ systems that can have wide impacts may be strong and viable strategies to take. Approaches of potential broad value are to modulate processes that are ubiquitous, systemic and that potentially have multiple impacts such as targeting the plasma proteome, cellular senescence, proteostasis, and metabolic processes.

Is a golden bullet of a single drug to impact aging biology a realistic scenario? The diversity of age-related disorders, the multitude of potential endpoints, the complexities of genetic risk factors and environmental challenges accumulating over a lifetime all make a single therapeutic unlikely. However, if there are fundamental underlying mechanisms, such as cellular senescence or failure of proteostatic maintenance, or a natural mixture, such as plasma or a fraction of plasma able to modulate multiple mechanisms, then potentially a single therapeutic could halt, or at least delay, most age-related disorders. It is still early for us to ascertain this, but the prospect will be tested in the clinic.

Regulatory issues pose some hurdles to the development of anti-aging therapeutics, but with a common goal in mind these can be navigated. Firstly, the FDA requires clinical trial endpoints be related to specifically impacting health or quality of life – survival, function, or feeling, not biomarkers. This must be kept in mind as we develop drugs, although biomarkers are going to be critical in assessing efficacy especially over extended time periods, they will not by themselves be sufficient for approval. Secondly, payers require a specific disease code for patient reimbursement, these will need much consideration as we move concepts from targeting specific indications to generalizing age-related diseases. These areas, which can be resolved by working together, should not be left too late for consideration.

We are at an exciting juncture where the realities of anti-aging therapies are upon us, and discussing how we can practically advance such approaches is a necessity. Even though a majority of research and therapeutic development focuses on individual domains such as neuroscience or behavior alone, thinking in the context of a systemic impact as we age provides wholly new opportunities, not only to tackle neurological disorders, but a spectrum of age-related ailments. The involvement of multiple disciplines, perspectives, and constituents in the field will be needed to be successful. This collaborative approach must be triggered so that quality of life for all can be improved in the near future.

Investigating the Cause of Irregular Heartbeat Following Cell Therapy for Regeneration of Heart Tissue

The promise of cell therapies is twofold. Firstly, the ability to regenerate from injuries that do not normally heal, such as severed nerves, or large loss of tissue mass. Secondly, the ability to restore more youthful function to aged tissues that suffer from a lack of replacement somatic cells due to the decline of stem cell activity. This decline is in part due to a loss of viable cells, and in part due to changes in the signaling environment or damage to stem cell niches that cause stem cells to become less active in response. Which of these processes is the dominant cause of loss of activity with age most likely differs from cell population to cell population.

As noted in the research materials below, there is considerable enthusiasm for the use of cell therapies to regenerate damage to the heart, particularly that occurring as a consequence of a heart attack. However, the heart is a highly structured organ with an electrical component to its activity, and regenerative strategies must avoid disruption of that structure and behavior. Do so and the heartbeat becomes irregular, or perhaps worse than that. This makes the heart a more challenging target than organs such as the kidney or liver, which are less stringent in their requirements for a very specific structure and balance of cell populations.

Heart regeneration using stem cells: Why irregular heartbeats occur after transplantation

Once a part of a heart tissue is injured due to restricted blood flow during a heart attack, treatment options are dire to fix the function of the heart to previous capacity. Stem cells are promising because they can be manipulated to generate healthy cells to replace diseased cells. No other cells hold this promise. There are a few issues to clear before stem cell treatments can be implemented clinically for heart regeneration and one major obstacle is to understand why irregular, abnormal heartbeats occur two to four weeks after induced pluripotent stem cell-derived heart muscle cells are transplanted to the heart. The heartbeat stabilizes on its own after 12 weeks but researchers set out to find out why the arrhythmia occurs.

It was thought that the arrhythmia occurs from the activity of the transplanted cells. Arrhythmia during a heart attacks is often noted as “re-entry” or when the electricity inside the heart goes haywire and loops around inside the heart. Two previous groups who studied arrhythmia in hearts of transplanted cells thought it was not caused by re-entry, but that it is the activity of the transplanted cells. Therefore, this team set out to find the cause through observing the properties of the various cells according to time points.

They created embryonic stem cell-derived cardiomyocyte cells and observed their electrical properties. There are two types of heart muscle cells made from induced pluripotent stem cells. “Working-type”, which like the name implies, contracts and relaxes to produce exertion. The other is called “nodal-like”, which acts like an electric pacemaker. After the twelfth week in vivo, the graft starts to grow, but immediately after transplantation it is very small. At the twelfth week the small graft has grown and consists mostly of working-type cells. The nodal-like cells has decreased significantly by then. The researchers believe that the arrhythmia decreases then, because the number of and activity of the nodal-like cells have decreased, causing extra electrical activity to decrease. So why does the population nodal-like cells decrease in vivo? Two weeks after transplantation, it was observed that nodal-like cells don’t multiply after transplantation, while the working-type cells increase significantly after transplantation. “Perhaps if doctors could remove the nodal-like cells before transplantation, arrhythmia would not occur during future transplantation of heart cell grafts.”

Increased predominance of the matured ventricular subtype in embryonic stem cell-derived cardiomyocytes in vivo

Accumulating evidence suggests that human pluripotent stem cell-derived cardiomyocytes can affect “heart regeneration”, replacing injured cardiac scar tissue with concomitant electrical integration. However, electrically coupled graft cardiomyocytes were found to innately induce transient post-transplant ventricular tachycardia in recent large animal model transplantation studies. We hypothesised that these phenomena were derived from alterations in the grafted cardiomyocyte characteristics.

In vitro experiments showed that human embryonic stem cell-derived cardiomyocytes (hESC-CMs) contain nodal-like cardiomyocytes that spontaneously contract faster than working-type cardiomyocytes. When transplanted into athymic rat hearts, proliferative capacity was lower for nodal-like than working-type cardiomyocytes with grafted cardiomyocytes eventually comprising only relatively matured ventricular cardiomyocytes. RNA-sequencing of engrafted hESC-CMs confirmed the increased expression of matured ventricular cardiomyocyte-related genes, and simultaneous decreased expression of nodal cardiomyocyte-related genes. Temporal engraftment of electrical excitable nodal-like cardiomyocytes may thus explain the transient incidence of post-transplant ventricular tachycardia, although further large animal model studies will be required to control post-transplant arrhythmia.

The Prospects for LANDO Upregulation as a Treatment for Alzheimer’s Disease

Of late, researchers have identified a process known as LC3-associated endocytosis (LANDO) by which cells can ingest and then break down the amyloid-β deposits associated with Alzheimer’s disease. This raises the idea that perhaps some form of upregulation of LANDO would at least slow the progression of Alzheimer’s disease, though the balance of evidence to date is beginning to suggest that amyloid-β is the wrong target, at least in later stages of the condition. Researchers here show that, in animal models, LANDO can reduce the inflammation of brain tissue associated with neurodegenerative conditions, a finding that makes this approach perhaps more interesting. The chronic inflammation of aging is strongly implicated in the progression of neurodegeneration, and may be the primary mechanism linking various forms of early pathology and environment exposure to the much more harmful later stages of Alzheimer’s disease.

The researchers previously discovered the LANDO pathway in microglial cells, the primary immune cells of the brain and central nervous system. Scientists found that when genes required for this pathway are deleted, Alzheimer’s disease progression accelerates in a mouse model. The investigators also showed that LANDO protects against neuroinflammation, one of the hallmarks of Alzheimer’s disease. While continuing to investigate LANDO, the researchers identified a novel function of the protein ATG16L. This protein is critical for autophagy, the normal process by which a cell recycles its components during periods of stress or energy deprivation. While ATG16L is important for autophagy, it can also play a role in LANDO. The investigators found that if a region of ATG16L called the WD domain is deleted, LANDO is inhibited while autophagy continues.

Most mouse models used in Alzheimer’s disease research rely on making genetic changes to recreate the disease. For this work, researchers used a new model with a specific deficiency of just the WD domain of ATG16L. This means the model carries out autophagy normally but lacks the LANDO pathway. By the time the mice are 2 years old, they exhibit symptoms and pathology that mimic human Alzheimer’s disease. This spontaneous age-associated model of Alzheimer’s disease is the first created by deleting a single protein domain (WD on ATG16L) not previously associated with Alzheimer’s disease. The researchers also analyzed human Alzheimer’s disease tissue samples, looking at the expression of proteins that regulate LANDO, including ATG16L. Expression of these proteins is decreased by more than 50% in people with Alzheimer’s disease. This finding shows a correlation between how deficiency in LANDO combined with aging may lead to Alzheimer’s disease in the mouse model and in humans.

Reducing neuroinflammation has been proposed as a potential way to treat Alzheimer’s disease. To treat their new mouse model, researchers used a compound that inhibits the inflammasome – a complex of proteins that activates pro-inflammatory immune reactions. Researchers profiled the model’s behavior and found evidence of improved cognition and memory in addition to a decrease in neuroinflammation. “This work solidifies LC3-associated endocytosis as a pathway that prevents inflammation and inflammatory cytokine production in the central nervous system. While much of the data on LANDO suggests a significant role in neuroinflammatory and neurodegenerative diseases, there is also a strong possibility that it could be targeted as a therapy against cancer or even infectious diseases that rely on similar processes for survival.”

DOK7 Gene Therapy Regrows Neuromuscular Junctions to Improve Aged Muscle Function

One of the numerous possible contributing causes to sarcopenia, the name given to the characteristic age-related decline in muscle mass and strength, is the dysfunction and loss of neuromuscular junctions. These structures link muscles and nerves, but how much of the lost strength of sarcopenia is due to this cause versus, say, declining muscle stem cell activity. The best way to assess the contribution of any given form of damage to any specific age-related condition is to repair that damage, and only that damage, and then observe the results. With that in mind, researchers here report on their implementation of a gene therapy approach to force the regrowth of neuromuscular junctions in aged mice. The treated mice exhibit increased strength in comparison to their untreated peers, which is a promising step towards an eventual therapy for humans.

Age-related decline in motor function has a major impact on quality of human life. The motor impairment involves age-related changes at least in the nerve and muscle systems, including a pathogenic loss of skeletal muscle mass and strength, known as sarcopenia. Accumulating evidence raises the possibility that the age-related decline in motor function is caused, at least in part, by functional impairment of the neuromuscular junction (NMJ), a cholinergic synapse essential for motoneural control of skeletal muscle contraction. Many studies with rodents have shown age-related denervation at NMJs in addition to degeneration of the presynaptic motor nerve terminals, where the neurotransmitter acetylcholine is released, and the postsynaptic endplate, where acetylcholine receptors (AChRs) densely cluster, suggesting an impaired neuromuscular transmission with aging.

In humans, electrophysiological and muscle fiber-type studies suggested age-related denervation at NMJs. Indeed, it is reported that the denervation rate at NMJs increases upon aging, although age-related morphological changes at NMJs remain controversial. Moreover, a recent study suggests that the increased rate of NMJ denervation contributes to the reduction in muscle strength in patients with sarcopenia, supporting the idea that the NMJ is a possible therapeutic target for treating age-related motor dysfunction.

We previously generated AAV-D7, a recombinant muscle-tropic adeno-associated virus (AAV) serotype 9 vector carrying the human DOK7 gene under the control of the cytomegalovirus promoter, and demonstrated that therapeutic administration of AAV-D7 – DOK7 gene therapy – enlarges NMJs and improves impaired motor activity in a mouse model of familial amyotrophic lateral sclerosis (ALS). Given that NMJ denervation appears to play a crucial role in age-related decline in motor function, DOK7 gene therapy might also ameliorate age-related motor impairment by suppressing denervation at NMJs. Thus, in the present study, we examined whether DOK7 gene therapy improves the motor function in aged mice.

Here, we show that DOK7 gene therapy significantly enhances motor function and muscle strength together with NMJ innervation in aged mice. Furthermore, the treated mice showed greatly increased compound muscle action potential (CMAP) amplitudes compared with the controls, suggesting enhanced neuromuscular transmission. Thus, therapies aimed at enhancing NMJ innervation have potential for treating age-related motor impairment.

Higher Body Mass Index Correlates with Reduced Cerebral Blood Flow

Vascular aging is an important contribution to neurodegeneration. The brain is an energy-hungry organ, and reductions in blood flow with age have a negative impact on brain tissue. These reductions can occur for obvious reasons such as the weakening of the heart in cases of heart failure, but there are other, more subtle processes at work to reduce the delivery of nutrients to the brain, such as the progressive stiffening of blood vessels and reductions in capillary density. Researchers here note that greater excess fat tissue, as measured by body mass index, correlates with reduced blood flow in the brain. It is plausible that this is mediated by the higher levels of chronic inflammation generated in people with larger than needed visceral fat deposits, as inflammation accelerates dysfunction in the vascular system, as well as dysfunction in tissue maintenance in general.

As a person’s weight goes up, all regions of the brain go down in activity and blood flow, according to a new brain imaging study. One of the largest studies linking obesity with brain dysfunction, scientists analyzed over 35,000 functional neuroimaging scans using single-photon emission computerized tomography from more than 17,000 individuals to measure blood flow and brain activity. Low cerebral blood flow is the primary brain imaging predictor that a person will develop Alzheimer’s disease. It is also associated with depression, ADHD, bipolar disorder, schizophrenia, traumatic brain injury, addiction, suicide, and other conditions.

Striking patterns of progressively reduced blood flow were found in virtually all regions of the brain across categories of underweight, normal weight, overweight, obesity, and morbid obesity. These were noted while participants were in a resting state as well as while performing a concentration task. In particular, brain areas noted to be vulnerable to Alzheimer’s disease, the temporal and parietal lobes, hippocampus, posterior cingulate gyrus, and precuneus, were found to have reduced blood flow along the spectrum of weight classification from normal weight to overweight, obese, and morbidly obese.

IL-6 and TGFβ1 Upregulation with Age Cause Detrimental Changes in Hematopoiesis

Researchers here show that blocking the age-related upregulation of inflammatory signal molecules IL-6 and TGFβ1 can reverse some of the deterimental changes in the function of hematopoiesis. Hematopoiesis is the process by which hematopoietic stem cells and related progenitor cell populations generate immune cells and red blood cells. With age, the production of lymphoid immune cells declines, and this is an important component of the aging of the immune system.

It is interesting to note that both IL-6 and TGFβ1 are generated by senescent cells as a part of the senescence-associated secretory phenotype. Senescent cells accumulate with age throughout the body, and contribute to chronic inflammation, as well as to the progression of near all age-related conditions. Given this, we should perhaps expect senolytic treatments that selectively destroy senescent cells in old tissues to be capable of reversing those aspects of immune aging investigated here.

Hematopoietic ageing involves declining erythropoiesis and lymphopoiesis, leading to frequent anaemia and decreased adaptive immunity. How intrinsic changes to the hematopoietic stem cells (HSCs), an altered microenvironment and systemic factors contribute to this process is not fully understood. Here we use bone marrow stromal cells as sensors of age-associated changes to the bone marrow microenvironment, and observe up-regulation of IL-6 and TGFβ signalling-induced gene expression in aged bone marrow stroma.

Inhibition of TGFβ signalling leads to reversal of age-associated HSC platelet lineage bias, increased generation of lymphoid progenitors and rebalanced HSC lineage output in transplantation assays. In contrast, decreased erythropoiesis is not an intrinsic property of aged HSCs, but associated with decreased levels and functionality of erythroid progenitor populations, defects ameliorated by TGFβ-receptor and IL-6 inhibition, respectively. These results show that both HSC-intrinsic and HSC-extrinsic mechanisms are involved in age-associated hematopoietic decline, and identify therapeutic targets that promote their reversal.

Cartilage Regrowth: Steering Microfracture to Provoke Regeneration of Fully Functional Cartilage

Microfracture surgery is a poor approach to producing the regrowth of cartilage. It is a procedure that causes minor damage, and that damage in turn provokes a more extensive regeneration of joint tissue. The tissue is unfortunately not the same as normal cartilage, but is better than nothing in cases of serious damage or wear. Researchers here asked how this response to damage works, and whether it can be steered to produce fully functional cartilage instead of the present less optimal tissue.

Damaged cartilage can be treated through a technique called microfracture, in which tiny holes are drilled in the surface of a joint. The microfracture technique prompts the body to create new tissue in the joint, but the new tissue is not much like cartilage. Microfracture results in what is called fibrocartilage, which is really more like scar tissue than natural cartilage. It covers the bone and is better than nothing, but it doesn’t have the bounce and elasticity of natural cartilage, and it tends to degrade relatively quickly.

For a long time, people assumed that adult cartilage did not regenerate after injury because the tissue did not have many skeletal stem cells that could be activated. Working in a mouse model, researchers documented that microfracture did activate skeletal stem cells. Left to their own devices, however, those activated skeletal stem cells regenerated fibrocartilage in the joint. But what if the healing process after microfracture could be steered toward development of cartilage and away from fibrocartilage? The researchers knew that as bone develops, cells must first go through a cartilage stage before turning into bone. They had the idea that they might encourage the skeletal stem cells in the joint to start along a path toward becoming bone, but stop the process at the cartilage stage.

The researchers used a powerful molecule called bone morphogenetic protein 2 (BMP2) to initiate bone formation after microfracture, but then stopped the process midway with a molecule that blocked another signaling molecule important in bone formation, called vascular endothelial growth factor (VEGF). “What we ended up with was cartilage that is made of the same sort of cells as natural cartilage with comparable mechanical properties, unlike the fibrocartilage that we usually get. It also restored mobility to osteoarthritic mice and significantly reduced their pain.”

A Gene Therapy Approach to Clearing Persistent Herpesviruses

Approaches that might effectively clear herpesviruses from the body are of considerable interest, as there is good evidence for the burden of persistent infection to have a meaningful impact on the pace of aging, largely via detrimental effects on the operation of the immune system over the course of years and decades. This is particularly true for cytomegalovirus, which may be a major cause of immunosenescence in near all people, but one might also look at the (presently disputed) evidence for HSV-1 to be a primary contributing cause of Alzheimer’s disease.

Infectious disease researchers have used a gene editing approach to remove latent herpes simplex virus 1, or HSV-1, also known as oral herpes. In animal models, the findings show at least a 90 percent decrease in the latent virus, enough researchers expect that it will keep the infection from coming back. The study used two sets of genetic scissors to damage the virus’s DNA, fine-tuned the delivery vehicle to the infected cells, and targeted the nerve pathways that connect the neck with the face and reach the tissue where the virus lies dormant in individuals with the infection.

In the study, the researchers used two types of genetic scissors to cut the DNA of the herpes virus. They found that when using just one pair of the scissors the virus DNA can be repaired in the infected cell. But by combining two scissors – two sets of gene-cutting proteins called meganucleases that zero in on and cut a segment of herpes DNA – the virus fell apart. The dual genetic scissors are introduced into the target cells by delivering the gene coding for the gene-cutting proteins with a vector, which is a harmless deactivated virus that can slip into infected cells. The researchers injected the delivery vector into a mouse model of HSV-1 infection, and it finds its way to the target cells after entering the nerve pathways. The researchers found a 92% reduction in the virus DNA present in the superior cervical ganglia, the nerve tissue where the virus lies dormant. The reductions remained for at least a month after the treatment.

“This is the first time that scientists have been able to go in and actually eliminate most of the herpes in a body. We are targeting the root cause of the infection: the infected cells where the virus lies dormant and are the seeds that give rise to repeat infections. Most research on herpes has focused on suppressing the recurrence of painful symptoms, and the team is taking a completely different approach by focusing on how to cure the disease. The big jump here is from doing this in test tubes to doing this in an animal. I hope this study changes the dialog around herpes research and opens up the idea that we can start thinking about cure, rather than just control of the virus.”

Increased Levels of Methylmalonic Acid May Raise the Risk of Metastasis in Older People

The article here discusses the interesting possibility that comparatively simple differences in circulating factors may be at the root of the higher risk of cancer metastasis in older people. Metastasis is the process by which cancer cells migrate from the primary tumor to form new tumors elsewhere. It is what makes cancers in much of the body hard to treat and ultimately fatal rather than merely harmful, problematic, but manageable. Thus targets that might potentially interfere in metastasis are of interest.

As we get older, the risk that we will develop cancer increases, because we accumulate genetic mutations and are continually exposed to cancer-causing substances. Most cancer-causing agents are found in the environment, but some are produced by our own bodies. Researchers now report that methylmalonic acid (MMA) – a by-product of protein and fat digestion – can accumulate in the blood with age, and might promote the spread of tumours. Methylmalonic acid is produced in cells in very small amounts. Usually, it becomes linked to the molecule coenzyme A to form methylmalonyl-CoA, and is converted to succinyl-CoA in a reaction that involves vitamin B12 as a cofactor. Succinyl-CoA subsequently enters the TCA cycle – a series of chemical reactions that are a key part of energy production in the cell.

Researchers report that MMA levels are significantly higher in the blood of healthy people over the age of 60 than in those under 30. The elevated level of MMA had not caused ill health in the individuals studied. However, the authors found that treating human cancer cells with serum from the blood of the older group, or with high concentrations of MMA, led them to adopt characteristics of metastatic cancer cells – those that can spread from a primary tumour to seed cancers elsewhere in the body. These characteristics include a loss of cell-cell attachment and an increase in mobility. When injected into mice, the cells formed metastatic tumours in the lungs.

The authors examined the gene-expression profiles of cells treated with MMA, and compared them with those of untreated cells. One of the genes most highly upregulated in response to MMA was SOX4, which encodes a transcription factor involved in the regulation of embryonic development and cancer progression. The authors demonstrated that repressing SOX4 expression blocked the cancer-cell response to MMA, and prevented the formation of metastatic tumours in mice that received injections of cancer cells treated with old serum. Thus, MMA indirectly induces an increase in the expression of SOX4, which in turn elicits broad reprogramming of gene expression and subsequent transformation of cells into a metastatic state.

Evidence for Klotho to Act on Life Span in Part via Resistance to Hypertension

Klotho is a longevity-associated protein. More of it in mice extends life, less of it shortens life. In humans, a number of studies have shown klotho levels to correlate with longevity. Beyond life span, a higher level of klotho also positively influences cognitive function, but the evidence to date shows the protein acting in the kidney. Researchers here demonstrate a link to hypertension, which is quite interesting, as the raised blood pressure of hypertension is strongly linked to both age-related mortality and cognitive decline. Increased blood pressure accelerates the progression of vascular conditions such as hypertension, leads to heart failure, and causes pressure damage to delicate tissues such as the brain. Sustained control of blood pressure is well demonstrated to reduce mortality in older people.

It has been known that high salt intake causes hypertension, but its exact mechanism was not understood until this study which found for the first time that Klotho deficiency, an anti-aging factor produced in the kidneys, causes aging-associated hypertension through high salt intake. Klotho acts as a hormone and is secreted into the blood from the kidneys. Its presence decreases with age causing the vascular and arterial system to stiffen. A recent study had shown the inverse relationship between the Klotho concentration and BP salt sensitivity. Hypertension is caused by excessive intake of salt, but the sensitivity of blood pressure to salt varies from individual to individual, and highly sensitive people are more likely to have high blood pressure.

In general, young people are less sensitive and are unlikely to develop hypertension, whereas older people are more sensitive to salt and are likely to develop hypertension. However, the mechanism of increased salt sensitivity with aging was unknown. Therefore, the research group first confirmed that salt sensitivity increased in aged mice, and revealed that the cause is that the blood concentration of the anti-aging factor Klotho protein decreases with age. Furthermore, the group clarified the molecular mechanism Wnt5a-RhoA pathway for the first time. In experiments using aged mice and cells, abnormal activation of this pathway could be reversed by supplementation with Klotho protein. As a result, it was possible to establish that the cause of salt-sensitive hypertension due to aging is Klotho protein decline.

The results of this experiment showed that Klotho supplementation could prevent the development of hypertension in the elderly and that Klotho levels could be a predictive marker for the development of hypertension. Trials for human verification is currently underway. Aging, a universal phenomenon, causes not only hypertension but dementia and frailty, and impairs the healthy life expectancy of individuals. The aging-related phenomenon of Klotho protein deficiency may be related to the onset of dementia and sarcopenia, or the loss of muscle-mass and usage associated with aging. Its onset mechanism is currently under investigation.

FOXO3a Suppresses Genomic Instability

FOXO3a is one of the very few genes for which an association with longevity has been identified in multiple human studies – though one should bear in mind that even though it shows up fairly reliably, the effect size is small. Still, near all such associations between human genetic variants and longevity cannot be replicated. Given this, there is an interest in understanding exactly how FOXO3a acts to influence life span. Here, researchers provide evidence that is suggestive of an effect on the burden of mutations in nuclear DNA, particularly double strand breaks. This is interesting in the context of recent work that links DNA repair activity for double strand breaks with the progressive detrimental shift in gene expression that takes place with age.

Genomic instability is one of the hallmarks of aging, and both DNA damage and mutations have been found to accumulate with age in different species. Certain gene families, such as sirtuins and the FoxO family of transcription factors, have been shown to play a role in lifespan extension. However, the mechanism(s) underlying the increased longevity associated with these genes remains largely unknown and may involve the regulation of responses to cellular stressors, such as DNA damage.

Here, we report that FOXO3a reduces genomic instability in cultured mouse embryonic fibroblasts (MEFs) treated with agents that induce DNA double-strand breaks (DSBs), that is, clastogens. We show that DSB treatment of both primary human and mouse fibroblasts upregulates FOXO3a expression. FOXO3a ablation in MEFs harboring the mutational reporter gene lacZ resulted in an increase in genome rearrangements after bleomycin treatment; conversely, overexpression of human FOXO3a was found to suppress mutation accumulation in response to bleomycin. We also show that overexpression of FOXO3a in human primary fibroblasts decreases DSB-induced γH2AX foci. Knocking out FOXO3a in mES cells increased the frequency of homologous recombination and non-homologous end-joining events. These results provide the first direct evidence that FOXO3a plays a role in suppressing genome instability, possibly by suppressing genome rearrangements.

Human Studies Link Calorie Restriction to Improved Cardiometabolic Status

The data noted here is not news to anyone who has followed calorie restriction research. It is well understood that the practice of calorie restriction is beneficial to long term health, reducing the impact of aging over time. It extends life by up to 40% in mice, but is nowhere near as effective as that in longer-lived mammals, such as our own species, even given similar short-term effects on metabolism. That said, it is interesting to note that enough robust studies of calorie restriction in humans have taken place over the past few decades to justify review papers on the topic.

Uncertainty remains about the risk/benefit balance of calorie restriction (CR) and its transferability to the current medical practice. Although during the last years different reviews and systematic reviews were published related effect of some type of CR on health, there are still no systematic reviews quantitatively summarizing the potential association between CR and multiple dimensions of health status. In fact, different systematic reviews have explored the association between CR and asthma, hypercholesterolemia, cardiovascular health, or bone health. Some systematic reviews have examined the general effects of diet or intermittent energy restriction on health, while others took in consideration specific populations such as intensive care units patients, athletes, or animal models. Hence, the aim of this study was to assess the effects of CR on dimensions of the WHO health concept, with a systematic review and meta-analyses of randomized controlled trials performed on this topic.

A total of 29 articles were retrieved including data from eight randomized controlled trials. All included trials were at low risk for performance bias related to objective outcomes. Collectively, articles included 704 subjects. Among the 334 subjects subjected to CR, the compliance with the intervention appeared generally high. Meta-analyses proved benefit of CR on reduction of body weight, BMI, fat mass, total cholesterol, while a minor impact was shown for LDL, fasting glucose, and insulin levels. No effect emerged for HDL and blood pressure after CR. Data were insufficient for other hormone variables in relation to meta-analysis of CR effects. Our conclusion is that CR is a nutritional pattern linked to improved cardiometabolic status. However, evidence is limited on the multidimensional aspects of health and requires more studies of high quality to identify the precise impact of CR on health status and longevity.