How Would One Go About Building a Company to Bring Cheap Senolytics to the World?

Let us for a moment choose to believe that the dasatinib and quercetin combination is a senolytic treatment that does as well in humans as it does in mice. This is to say it kills about 25-50% of senescent cells in the tissues usually most affected by oral medications, meaning the kidney, liver, and cardiovascular system, and some unknown but lower fraction elsewhere. Whether or not this is the case has yet to be determined; the first pilot studies are still running at Betterhumans, and they likely won't tell us the size of the effect in terms of fraction of cells removed. Viable assays for cellular senescence that can be used in human medicine are in short supply - there is only the one that I know of that is ready to go, and even that has just reached the final stage of laboratory proof of concept. If it is the case, however, that treatment with dasatinib and quercetin works in much the same way in humans, then it should have a notably positive effect on the state of health for older individuals, given that the accumulation of senescent cells is one of the causes of aging.

The distinguishing feature of dasatinib and quercetin are that they are cheap. A senolytic therapy would be undergone perhaps once every few years at most; it kills the unwanted cells it can kill, and it is pointless to do it again before there has been enough time for new senescent cells to emerge at their slow pace. Quercetin is a widely used supplement, and enough of it for a single treatment costs less than a dollar. Dasatinib can be purchased from manufacturers for between $20 and $150 for a single dose suitable for senolytic therapy, depending on where the manufacturer is based. The FDA approved packaging of dasatinib, called Sprycel, costs $300-600 for the same amount, assuming you can find someone willing to break down a bottle of tablets to sell you the small amount needed. It is certainly possible to purchase Sprycel for less than this by ordering from outside the US.

If this pharmaceutical does work in humans as imagined above, then at these prices it is a therapy that would be affordable for a sizable portion of the world's population. It isn't the only candidate senolytic drug that is cheap enough to consider in this way. Once the first of these treatments are proven to be at least passably useful in human patients, what is the path to putting these low-cost rejuvenation therapies into the hands of hundreds of millions of people, the majority of which are not wealthy, as soon as possible? We should give this some thought, as it is a opportunity that will likely arrive much sooner than most of us expected it to. This is a big deal: early senolytics could provide a gain in health for much of humanity if the opportunity is managed correctly. That makes it worth consideration even prior to proof arriving in human studies.

There are always roadblocks. Like all such matters, the use of dasatinib is tied up by patents and regulation. No-one can build a large-scale business selling a pharmaceutical where the intellectual property and regulatory approval are owned by a large and influential concern - in this case Bristol-Myers Squibb (BMS). It is certainly the case that there is a healthy marketplace of scofflaws outside the US who sell directly from manufacturers, but they are not a single target, and it is hardly worth BMS's time to try to squash them while dasatinib is generating only the level of revenue possible for a cancer drug. That economic calculus may well change if it suddenly becomes a viable treatment for every older individual, and physicians show interest in off-label use - that is a vastly larger potential market. Certainly, BMS exerted their influence to block attempts to produce a cheaper generic version in India. That was associated with the Indian government and thus had a convenient single point of attack, unlike the manufacturer marketplace.

Dasatinib is still patent protected, at least until 2020, which means that any earnest effort to make dasatinib a household term in the near future would have to engage with BMS and gain at last tacit approval in order to grow. After 2020, no permission is needed. BMS will continue to tinker with their formula to extend patent protection on the versions of Sprycel that they sell, but they will no longer be able to directly make life difficult for those who wish to manufacture and sell dasatinib per the original formulation. The price will likely drop considerably at that point. So how could a group proceed if willing to found a company to work on distribution of low-cost senolytics?

The Non-Profit Approach

The most obvious option is to build a non-profit that focuses on education and partnership. The goal would be to deliver low-cost dasatinib and the understanding needed for widespread use to less wealthy regions of the world. The non-profit would focus on building relationships with physicians, medical organizations, manufacturers, and the product owner, BMS. There is considerable precedent for this sort of endeavor, and many larger pharmaceutical companies carry out in-house programs of this nature. It can benefit the pharmaceutical company considerably even if they make little to no revenue from the use of their product in those markets. It is usually the case that they wouldn't have been able to sell at profitable prices there anyway, and the program can be very good for their public image - something that Big Pharma entities are always in need of, for some strange reason.

The For-Profit Scofflaw Approach

Prior to 2020, one would require deep pockets and to be based outside the US, preferably in a country that doesn't regard the US with any great favor, in order to build something large that undercuts BMS, or even simply to sell into markets that BMS chooses not to serve. Being a small company that ships dasatinib at low cost from China to other parts of the world is probably viable, but growth to any significant size would bring a quick end to the endeavor. As mentioned, an attempt was made in India, where there is a history of threatening to break international intellectual property agreements in order to bring low-cost medications to that part of the world. That failed, and I'd say that India is probably the most likely region to successfully host a defensible patent breaking exercise.

The For-Profit BMS Enabler Approach

The enabler approach runs something like this: establish a path to obtain Sprycel in bulk at a workably low cost, and in an approved manner for the regulatory framework, and then build a revenue stream based on selling wrapped packages of services and Sprycel to physicians, nursing home operators, and other interested groups. Businesses and other organizations are better customers to start with in less wealthy regions, as there is a greater chance of being able to gain sufficient revenue to expand. Optionally, partner with BMS, though this is typically hard to do without connections.

The packages sold might include: educational materials and classes; professional services to assist with insurance and other regulatory concerns for prescribing off-label usage; membership of a network that helps bring in patients interested in the treatment and thus contributes to a physician's bottom line; tests and organization of testing services to evaluation results; and so forth. Everything is carried out in a such a way that it benefits BMS, such that the company has incentives to allow the business to grow. There are many possible variations on this theme, some of which are similar to the promotional activities carried out by the pharmaceutical companies themselves, while others look more like patient or physician associations or service organizations.

The Wait Until 2020 Approach

In either non-profit or for-profit models by which dasatinib might be distributed to the less wealthy regions of the world in volume, the prospects look a lot better once BMS is no longer the gatekeeper. The price of manufacture will fall precipitously, and an enterprising group with a good approach and competent execution might be able to do quite well in markets traditionally neglected by large pharmaceutical concerns. "Quite well" in this case would mean - under the assumptions at the top of this post - a significant number of people living incrementally longer in better health at a cost that is reasonable for them, considering the benefits achieved. That seems a worthwhile goal to aim for.

A Measure of Cerebrospinal Fluid Flow Suggests that Brain Aging Commences Early

There is a growing faction in the neurodegenerative research community whose members think it likely that rising levels of metabolic waste in brain, such as tau and amyloid aggregates, are due to failing drainage of cerebrospinal fluid. That drainage is a primary method of removal, and as it declines the wastes build up. The Methuselah Foundation is somewhat ahead of the game here, having incubated Leucadia Therapeutics to develop a possible solution. A number of other groups have turned their attention to this topic, and it has been interesting to see a flurry of papers in the last year or so. The work noted here is related, though the researchers are looking at circulation of cerebrospinal fluid within the brain, driven by cardiovascular activity, rather than drainage. The open access paper - worth looking at, but very dry - describes a low-cost way of assessing this flow and some exploration of the findings. Their measurements start to show changes at a comparatively early age, much earlier than one would expect for a process linked to cardiovascular function. This is quite interesting, though it is far too early to do more than speculate on why this might be the case.

Physicists have devised a new method of investigating brain function. This new non-invasive technique could potentially be used for any diagnosis based on cardiovascular and metabolic-related diseases of the brain. The researchers deciphered oscillations in the cerebrospinal fluid which lies between the scalp and skull; a device for non-invasive recordings of this translucent fluid was developed and recordings on healthy subjects were made.

It has been shown that the circulation throughout the brain of this fluid is highly fluctuating, and that these fluctuations are slow but interconnected by the rhythms of breathing and the heart rate. Researchers found that some of these oscillations are linked with blood pressure, but are generally slower, occurring at lower frequencies, which have been shown in previous studies to be related to oscillations in vascular motion and blood oxygenation.

Preliminary results showed evidence of a decline in the coherence between these oscillations in participants over the age of 25, indicating that brain ageing may begin earlier than expected. "Combining the technique to noninvasively record the fluctuation corresponding to cerebrospinal fluid and our sophisticated methods to analyse oscillations which are not clock-like but rather vary in time around their natural values, we have come to an interesting and non-invasive method that can be used to study ageing."

Link: http://www.lancaster.ac.uk/news/articles/2018/brain-ageing-may-begin-earlier-than-expected/

Arguing for Tau to be More Important than Amyloid-β in Alzheimer's Disease

This isn't the first paper I've seen to argue the point that there should be a greater focus on tau aggregation in Alzheimer's disease, and that tau may be more important to the progression of the condition. As I'm sure the readers here are aware, Alzheimer's is characterized by the buildup of both amyloid-β and tau in the brain. Forms of these normally soluble proteins precipitate into solid deposits that are accompanied by a complex halo of biochemistry that degrades the function of neurons and ultimately kills these cells. The primary focus for development of therapies has long been the removal of amyloid-β, but despite enormous effort there is no light at the end of the tunnel yet. The history of clinical trials for amyloid-β clearance is one of unremitting failure, even recently in trials that produced evidence for amyloid-β to be removed to some degree in patients.

It is much debated as to whether trials are failing because amyloid-β is the wrong target, despite being harmful in and of itself, or because Alzheimer's is a hard problem. Alzheimer's research has proceeded in parallel with mapping the brain at the necessary level to talk about how exactly it is damaged by protein aggregates, and also in parallel with the development of immunotherapy technologies, both of which are challenging areas of research and development. The biochemistry of the brain, its operation, and its failure modes are all enormously complex. We seem to be reaching a tipping point, however, in which discontent with the focus on amyloid-β is spilling over into greater emphasis and funding for alternatives. Rightly or wrongly in this specific case, I think that diversity in approaches is almost always better in the long term.

The hallmarks of Alzheimer's disease (AD) pathology are marked by accumulation of extracellular amyloid-β (Aβ) plaques in the brain followed by intracellular neurofibrillary tangle (NFT) growth. Aβ upregulates the generation of NFTs by increasing glycogen synthase kinase-3 (GSK-3) activity, leading to the phosphorylation of tau. Phosphorylated tau (pTau) begins to self-assemble to form NFTs. Aβ plaques, soluble Aβ oligomers, and NFTs interfere with normal neuronal cell function by disrupting synaptic signaling. Each protein's accumulation leads to neuron damage, eliciting diminished brain mass and cognitive function.

The removal of Aβ plaques does not influence elimination of NFTs after NFTs have been established in the brain, but early intervention can prevent pTau development. Therefore, targeted late stage treatments may specifically eliminate Aβ without impacting pTau levels that have already accumulated, which enables NFTs to continue amplifying cognitive deficits. Comparison of differences in pTau and Aβ levels in treated mice illuminate differences between the proteins' impact on cognitive function. For example, pTau levels were reduced by chemical treatment as Aβ levels continued to increase, yet cognitive function improved. This result implies that there is a quantitative difference between how the two proteins effect cognitive deterioration, and moreover, that decreasing pTau may ultimately be more important than reducing Aβ in the quest to successfully treat AD.

The Amyloid Cascade Hypothesis states that Aβ is the center piece in AD pathology leading to hyperphosphorylation of tau and numerous neurotoxic pathways causing cell death. Treatments targeting Aβ and Aβ precursors have failed to pass clinical trials to improve patient outcomes. The presence of Aβ is associated with a decrease in cognitive performance; however, the quantitative level of Aβ inconsistently predicts the amount of cognitive decline. Instead, it is suggested that other contributors, such as the hyperphosphorylation of tau, are the functional cause of degeneration after the initial onset of AD.

The present study compares the effects of Aβ and pTau levels on cognitive performance in the Morris water maze (MWM) and Novel Object Recognition (NOR) through a large-scale meta-analysis of 3xTg-AD mouse model experiments. The triple-transgenic mouse model (3xTg-AD) of AD expresses tangle and plaque pathology as well as synaptic dysfunction. Multiple linear regression confirmed pTau is a stronger predictor of MWM performance than Aβ. Despite pTau's lower physical concentration than Aβ, pTau levels more directly and quantitatively correlate with 3xTg-AD cognitive decline.

Link: https://doi.org/10.3233/JAD-170490

An Energetic Exploration of the Biochemistry of Cellular Senescence is Underway

In 2011 a research group published the results from an animal study that demonstrated, in a way that couldn't be ignored, that the accumulation of senescent cells is a significant cause of aging and age-related disease. In fact, the evidence for this to be the case had been compelling for a very long time - this demonstration came nearly a decade after Aubrey de Grey, on the basis of the existing evidence at the time, included cellular senescence as one of the causes of aging in the first published version of his SENS research proposals. Yet nothing had been done to move ahead and achieve something with this knowledge. That did not change until researchers obtained sufficient philanthropic funding to run the 2011 animal study, using a sophisticated genetic mechanism that eliminated senescent cells as they formed.

From that point on, a slow-moving avalanche of interest and funding fell into this part of the field of aging research. All of the groups with an existing interest in cellular senescence, and that had previously struggled to raise sufficient resources to make progress, could now move rapidly. With the aim of selectively destroying senescent cells to reverse aspects of aging, small molecule senolytic pharmaceuticals and then other methods such as gene therapies and immunotherapies were discovered or constructed. Today there are at least a dozen such small molecule drugs, published and in the works, and a handful of increasingly well-funded startup biotech companies bringing these therapies to human trials and the clinic.

That is the practical side that will lead to rejuvenation treatments in the near future. But the pure scientific impulse isn't to build new technology, it is to learn how our biochemistry works. Much of the funding for further work on cellular senescence goes towards mapping and understanding its details. Now that it is inarguable that this phenomenon is important in the progression of degenerative aging, scores of research groups are picking apart the biochemistry of senescent cells. They are categorizing, trying to understand whether all senescence is essentially the same, or whether there are significant differences in different cell types. They are attempting to better grasp all of the relevant mechanisms that operate inside cells as senescence occurs, and how the triggering change works - or, indeed, whether or not it is a single trigger. They are exploring the details of the senescence-associated secretory phenotype (SASP), the means by which these cells cause harm to tissues.

The four open access papers noted here are recent examples of this sort of thing. There is a great deal to learn, and while the work is largely irrelevant to the senolytic therapies currently in development, there will no doubt be discoveries that steer and inform development of the second generation of more subtle and sophisticated therapies. Those will likely commence development five to ten years from now, and be mature and in widespread use by the early 2030s.

TNFα-senescence initiates a STAT-dependent positive feedback loop, leading to a sustained interferon signature, DNA damage, and cytokine secretion

Cellular senescence is a cell fate program that entails essentially irreversible proliferative arrest in response to damage signals. Tumor necrosis factor-alpha (TNFα), an important pro-inflammatory cytokine secreted by some types of senescent cells, can induce senescence in mouse and human cells. However, downstream signaling pathways linking TNFα-related inflammation to senescence are not fully characterized. Using human umbilical vein endothelial cells (HUVECs) as a model, we show that TNFα induces permanent growth arrest and increases p21CIP1, p16INK4A, and SA-β-gal, accompanied by persistent DNA damage and ROS production. By gene expression profiling, we identified the crucial involvement of inflammatory and JAK/STAT pathways in TNFα-mediated senescence. We found that TNFα activates a STAT-dependent autocrine loop that sustains cytokine secretion and an interferon signature to lock cells into senescence.

3′ UTR lengthening as a novel mechanism in regulating cellular senescence

Cellular senescence has been viewed as a tumor suppression mechanism and also as a contributor to individual aging. Widespread shortening of 3′ untranslated regions (3′ UTRs) in messenger RNAs (mRNAs) by alternative polyadenylation (APA) has recently been discovered in cancer cells. However, the role of APA in the process of cellular senescence remains elusive. Here, we found that hundreds of genes in senescent cells tended to use distal poly(A) (pA) sites, leading to a global lengthening of 3′ UTRs and reduced gene expression. Genes that harbor longer 3′ UTRs in senescent cells were enriched in senescence-related pathways. Rras2, a member of the Ras superfamily that participates in multiple signal transduction pathways, preferred longer 3′ UTR usage and exhibited decreased expression in senescent cells. Depletion of Rras2 promoted senescence, while rescue of Rras2 reversed senescence-associated phenotypes.

The SCN9A channel and plasma membrane depolarization promote cellular senescence through Rb pathway

Oncogenic signals lead to premature senescence in normal human cells causing a proliferation arrest and the elimination of these defective cells by immune cells. Oncogene-induced senescence (OIS) prevents aberrant cell division and tumor initiation. In order to identify new regulators of OIS, we performed a loss-of-function genetic screen and identified that the loss of SCN9A allowed cells to escape from OIS. The expression of this sodium channel increased in senescent cells during OIS. This upregulation was mediated by NF-κB transcription factors, which are well-known regulators of senescence. Importantly, the induction of SCN9A by an oncogenic signal or by p53 activation led to plasma membrane depolarization, which in turn, was able to induce premature senescence. Computational and experimental analyses revealed that SCN9A and plasma membrane depolarization mediated the repression of mitotic genes through a calcium/Rb/E2F pathway to promote senescence.

Mitochondrial (Dys) Function in Inflammaging: Do MitomiRs Influence the Energetic, Oxidative, and Inflammatory Status of Senescent Cells?

A relevant feature of aging is chronic low-grade inflammation, termed inflammaging, a key process promoting the development of all major age-related diseases. Senescent cells can acquire the senescence-associated (SA) secretory phenotype (SASP), characterized by the secretion of proinflammatory factors fuelling inflammaging. Cellular senescence is also accompanied by a deep reshaping of microRNA expression and by the modulation of mitochondrial activity, both master regulators of the SASP. Here, we synthesize novel findings regarding the role of mitochondria in the SASP and in the inflammaging process and propose a network linking nuclear-encoded SA-miRNAs to mitochondrial gene regulation and function in aging cells. In this conceptual structure, SA-miRNAs can translocate to mitochondria (SA-mitomiRs) and may affect the energetic, oxidative, and inflammatory status of senescent cells.

Assembling Cells and Scaffolds into a Suitable Trachea Replacement

Researchers here report on their efforts to build a suitable structure to replace a trachea, starting with patient cells and artificial scaffolds. Since the trachea is a thin-walled pipe, engineered tissue can be constructed in this way without the need for complex blood vessel networks, as at no point is the tissue so thick as to prevent direct perfusion of nutrients and oxygen to the inner cells. Unfortunately, it remains the case that decellularized donor tissue is the only reliable solution for the production of capillaries to support thicker tissues, scores of such vessels passing through every square millimeter. This is why most of the more ambitious work, closer to clinical application, involves thin tissues and tubular structures - larger blood vessels, skin, and so forth - while everyone else is working with the tiny sections of engineered tissue known as organoids.

Biomedical engineers are growing tracheas by coaxing cells to form three distinct tissue types after assembling them into a tube structure - without relying on scaffolding strategies currently being investigated by other groups. "The unique approach we are taking to this problem of trachea damage or loss is forming tissue modules using a patient's cells and assembling them into a more complex tissue." Recent tissue engineering approaches using synthetic or natural materials as scaffolding for cells have met with challenges. Difficulties have included uniformly seeding cells on the scaffolding, recreating the multiple different tissue types found in the native trachea, tailoring the scaffolding degradation rate to equal the rate of new tissue formation, and recreating important contacts between cells because of the intervening scaffold.

The trachea engineering strategy now being pursued, however, wouldn't have those problems because it doesn't rely on a separate scaffold structure. A new trachea replacement must do three critical things to function properly: (1) maintain rigidity to prevent airway collapse when the patient breathes; (2) contain immunoprotective respiratory epithelium, the tissue lining the respiratory tract, which moistens and protects the airway and functions as a barrier to potential pathogens and foreign particles; and (3) integrate with the host vasculature, or system of blood vessels, to support epithelium viability.

The self-assembling rings developed by researchers meet all three of those requirements because they can fuse together to form tubes of both cartilage and "prevascular" tissue types. Prevascular refers to tissues potentially ready to participate in the formation of blood vessels, though not yet functional in that way. The cartilage rings are formed by aggregating marrow-derived-stem cells in ring-shaped wells. Polymer microspheres containing a protein that induces the stem cells to become "chondrocytes," or cells that form cartilage, are also incorporated into the cell aggregates. These prevascular rings are comprised of both these marrow-derived stem cells and endothelial cells, the thin layer of cells that line the interior of blood vessels.

The researchers then coat the tubes with epithelial cells to form multi-tissue constructs that satisfy all of those requirements: cartilage provides rigidity, epithelium serves the role of immunoprotection and the vascular network would ultimately permit blood flow to feed and integrate the new trachea tissue. Using this method, the team has been able to engineer highly elastic "neo-tracheas" of various sizes, including tissues similar to human trachea. When these tracheas were implanted under the skin in mice, there was evidence the prevascular structures could join up with the host vascular supply.

Link: http://thedaily.case.edu/living-human-tracheas/

Astrocytes Become Inflammatory in the Aging Brain

Astrocytes are one of the common types of support cell in the brain, performing a wide variety of tasks that range from repair to maintaining the balance of various signal and electrolyte molecules. Researchers find evidence to suggest that astrocytes shift into an inflammatory mode in large numbers with advancing age. Chronic inflammation is a feature of most neurodegenerative conditions, and of aging in the broader sense. It disrupts the complex relationships between cell types that are needed for most sophisticated behavior in tissues, such as regeneration, or any number of cell communication processes required for correct function of the brain.

This is particularly interesting in the context of recent findings regarding cellular senescence in astrocytes. A large fraction of these cells show some signs of senescence in older individuals, and one of the characteristic bad behaviors of senescent cells is the generation of chronic inflammation through the senescence-associated secretory phenotype. Researchers have pinned down astrocyte senescence as a contributing factor in Parkinson's disease, for example. It is also worth noting that this business of cells shifting into an inflammatory mode in greater numbers with advancing age is also observed in macrophages, where it disrupts regenerative processes, and in microglia, another of the support cells of the brain. They also generate chronic inflammation in brain tissue, which contributes to the complicated breakdown of the normal operation of the brain.

The decline of cognitive function occurs with aging, but the mechanisms responsible are unknown. Astrocytes instruct the formation, maturation, and elimination of synapses, and impairment of these functions has been implicated in many diseases. These findings raise the question of whether astrocyte dysfunction could contribute to cognitive decline in aging. We performed RNA sequencing of astrocytes from different brain regions across the lifespan of the mouse. We found that astrocytes have region-specific transcriptional identities that change with age in a region-dependent manner.

Detailed analysis of the differentially expressed genes in aging revealed that aged astrocytes take on a reactive phenotype of neuroinflammatory A1-like reactive astrocytes. Hippocampal and striatal astrocytes up-regulated a greater number of reactive astrocyte genes compared with cortical astrocytes. Moreover, aged brains formed many more A1 reactive astrocytes in response to the neuroinflammation inducer lipopolysaccharide.

We found that the aging-induced up-regulation of reactive astrocyte genes was significantly reduced in mice lacking the microglial-secreted cytokines (IL-1α, TNF, and C1q) known to induce A1 reactive astrocyte formation, indicating that microglia promote astrocyte activation in aging. Since A1 reactive astrocytes lose the ability to carry out their normal functions, produce complement components, and release a toxic factor which kills neurons and oligodendrocytes, the aging-induced up-regulation of reactive genes by astrocytes could contribute to the cognitive decline in vulnerable brain regions in normal aging and contribute to the greater vulnerability of the aged brain to injury.

Link: https://doi.org/10.1073/pnas.1800165115

Recent Genetic Studies Claiming a Slowing of Aging may be Largely Incorrect

It is fair to ignore most studies showing extension of life span in laboratory species conducted much prior to the turn of the century. A majority failed to control for calorie restriction, and thus the (usually small) effects evaporate when more rigorously tested. The way this works is that an intervention makes mice nauseous or otherwise uncomfortable, they eat less as a consequence, and thus live longer solely due to lowered calorie intake. This is on top of the usual estimate that most of all published research results are flawed in some way. That includes animal studies that use too few animals, and thus tend to be prone to statistical happenstance, for example. Small studies with few animals are distressingly common in the study of aging, where funding is typically very restricted. Matters did improve once it was no longer possible to be ignorant of the size of the calorie restriction effect on longevity in short-lived species, as that research gained increasing popularity and interest after the 1990s. But as the open access paper I'll point out here suggests, not improved enough.

I think that part of the problem is that too many people were - and still are - trying to evaluate marginal effects on aging. It is hard to accurately detect and quantify small effects in animal studies. A 10% life span extension observed in a group of twenty mice, as compared to a control group of twenty mice, tells us just about nothing other than perhaps it would be good to seek corroboration in a group five times that size - and this example is around the size of effect for most reported interventions based on adjusting the operation of metabolism to slow aging.

One thing I wish was better understood and discussed in our community of advocates, supporters, and researchers is that size of effect and reliability of effect matter enormously. They are the point of the exercise, and the future of our health depends upon them. Everything shown to result in either small or only intermittently apparent outcomes should be rapidly dropped in favor of the continuing search for truly useful approaches to aging. Senescent cell clearance is a shining example of reliability: it always works; it works on many different aspects of aging; it works to treat many different age-related diseases; in fact it puts just about everything else tried to date to shame. The only item from the camp of metabolic manipulation that is as reliable in animal studies is the use of mTOR inhibitors such as rapamycin - and they are notably less effective when it comes to impact on specific age-related diseases. All in all, far too much time and effort is wasted on hoping that unreliable approaches with small effects are magically hiding something useful.

A Reassessment of Genes Modulating Aging in Mice Using Demographic Measurements of the Rate of Aging

The discovery that single gene manipulations can significantly modulate longevity is arguably the major breakthrough in biogerontology thus far. Genetic manipulations of aging in mice are crucial to gather insights into the underlying mechanisms of aging, to discover pathways modulating longevity and to identify candidate genes for drug discovery. Moreover, the manipulation of the aging process in mammalian models (particularly mice) via genetic manipulation (gene knockouts, overexpression, etc.) is crucial to test mechanistic hypotheses of aging. However, determining if such genetic interventions actually affect the aging process and not some others factor of health is not always straightforward.

For example, should a genetic intervention reduce an organism's resistance to disease, this could conceivably reduce the lifespan of the organism, although the rate of aging would not have been affected. Differentiating between genetic interventions that affect the lifespan of an organism through altered health as opposed to changes in the rate of aging is therefore essential to gain insights on aging, and determine interventions with wide ranging effects.

There are two fundamental methods to determine if a life-extending genetic intervention has altered the rate of aging rather than general health. One can track the onset and progression of age-related ailments and physiological degeneration to determine if there is a shift in the onset and on progression of the ailment. In addition, efforts have been made to quantify aging rates with mathematical models such as the Gompertz law of mortality. From the Gompertz parameters, the mortality rate doubling time (MRDT) can be calculated. The MRDT is the amount of time it takes for the mortality rate to double for a given cohort.

A change in MRDT indicates a change in the demographic rate of aging, which is not a perfect reflection of biological aging but a metric that correlates with physiological deterioration and health. Although some studies have investigated MRDT, many authors still often assume that changes in the lifespan of mice following a genetic intervention directly equates to changes in the rate of aging, leading to the misrepresentation of certain genes as having a causal role in aging, when in reality they do not.

Many studies have reported altered median and/or maximum lifespan as a result of an intervention but lifespan alterations may have a number of causes, including altered age at onset of senescence and age-independent mortality. To address this lack of distinction, we previously used linear regression to fit the Gompertz model to longevity data from published mouse studies, and statistically compared the rates of aging in these cohorts. For example, we showed that caloric restriction increases the MRDT and thus retards the demographic rate of aging. Here, the same methodology was employed to reassess mouse longevity data published since 2005 and to identify which genes are more important in determining the demographic rate of aging.

Overall, only 7 of 54 genes were found to have a statistically significant effect on the demographic rate of aging as expected from longevity manipulations. These results suggest that only a relatively small proportion of interventions reported to affect longevity in mice do so through directly influencing the demographic rate of aging. Surprisingly, 20 of 54 genes had a statistically significant impact on the demographic rate of aging in the opposite direction than would be expected for the published longevity effects. One possible explanation is that many mutations impacted on various parameters affecting longevity in non-linear ways, and indeed we observed that increases in aging independent mortality correlated with a slower demographic aging rate. For instance, Sirt1 deficiency extended lifespan but increased the demographic rate of aging; its effect appeared to be exerted instead by delaying the age of onset of mortality rate escalation. This highlights the complex relationship between lifespan and the demographic rate of aging.

Another caveat of our approach concerns the number of mice used in some of the original studies, which ranged from 10 to 146 animals per cohort. Whilst research reported here has attempted to compensate for this by using the Gompertz equation which allows for small sample sizes, one cannot escape the low statistical power that accompanies such small sample sizes. Interestingly, caloric restriction has been shown to significantly retard the demographic rate of aging, but this was a large study with over 200 animals in total. Therefore, caution must be taken when interpreting some of the results detailed here from studies with small sample sizes. Indeed, we observed that in smaller experimental cohorts subjective decisions in estimating Gompertz parameters can significantly affect the results.

Our main conclusions are: 1) most genetic manipulations of longevity in mice do so by modulating aging-independent mortality; 2) there is substantial variation in the lifespan of controls of the same strain across experiments; 3) studies in which the lifespan of the controls is short have a greater lifespan increase, emphasizing the importance of having adequate control groups; 4) mouse lifespan studies employing small cohorts can yield unreliable results; 5) lifespan-reducing experiments tend to be noisier and more difficult to analyze for demographic parameters than life-extending experiments; 6) a greater aging-independent mortality is usually accompanied by a slower demographic aging rate.

How Does Age Affect Induced Pluripotency for Regenerative Medicine?

One of the more intriguing discoveries relating to the cell reprogramming used to produce induced pluripotent stem cells is that this process appears to reverse some aspects of cell aging. It perhaps triggers some fraction of the mechanisms at work in early embryonic development, those that ensure that children are born young, with nowhere near the load of persistent damage present in the adult parents. This is not a well-explored topic, unfortunately - it is still too recent for much to be said in certainty, and a sizable fraction of the evidence is conflicting. Related to all of this is the question of how exactly the age of the donor affects the reprogramming of donated cells. Near all potential uses of regenerative medicine based on reprogrammed cells involve age-related disease and older individuals. It is important to understand whether it is safe to proceed, how effective approaches might be in practice, and where the problems lie, so that they can be addressed.

Induced pluripotent stem cells (iPSCs) avoid many of the restrictions that hamper the application of human embryonic stem cells, and the donor's clinical phenotype is often known when working with iPSCs. Therefore, iPSCs seem ideal to tackle the two biggest tasks of regenerative medicine: degenerative diseases with genetic cause (e.g., Duchenne's muscular dystrophy) and organ replacement in age-related diseases (e.g., end-stage heart or renal failure), especially in combination with recently developed gene-editing tools.

In the setting of autologous transplantation in elderly patients, donor age becomes a potentially relevant factor that needs to be assessed. Here, we review and critically discuss available data pertinent to the questions: How does donor age influence the reprogramming process and iPSC functionality? Would it even be possible to reprogram senescent somatic cells? How does donor age affect iPSC differentiation into specialised cells and their functionality? We also identify research needs, which might help resolve current unknowns.

Until recently, most hallmarks of ageing were attributed to an accumulation of DNA damage over time, and it was thus expected that DNA damage from a somatic cell would accumulate in iPSCs and the cells derived from them. In line with this, a decreased lifespan of cloned organisms compared with the donor was also observed in early cloning experiments. Therefore, it was questioned for a time whether iPSC derived from an old individual's somatic cells would suffer from early senescence and, thus, may not be a viable option either for disease modelling nor future clinical applications. Instead, typical signs of cellular ageing are reverted in the process of iPSC reprogramming, and iPSCs from older donors do not show diminished differentiation potential nor do iPSC-derived cells from older donors suffer early senescence or show functional impairments when compared with those from younger donors.

Thus, the data would suggest that donor age does not limit iPSC application for modelling genetic diseases nor regenerative therapies. However, open questions remain, e.g., regarding the potential tumourigenicity of iPSC-derived cells and the impact of epigenetic pattern retention.

Link: https://doi.org/10.3389/fcvm.2018.00004

Towards Lasting Therapeutic Manufactories that Operate Inside the Body

Gene therapies involve delivering instructions into cells to ensure that specific proteins are manufactured, either temporarily or permanently. This is effectively a hijacking or programming of cellular mechanisms. There is another approach, which is to deliver suitable DNA machinery into the body, capable of manufacturing the desired proteins outside cells. This isn't helpful for all types of protein, but in many cases it is. That machinery needs protection, however: naked, it would be quickly removed by the immune system or otherwise broken down. One possibility is to employ engineered bacteria, which removes the need to introduce changes into a patient's cells, but adds a sizable set of other complications. Another approach is to build a suitable structure from scratch, such as a membrane that will not alert the immune system, containing a carefully limited set of DNA machinery that will turn out the desired proteins for a lengthy period of time, but is incapable of any other activity. These constructs would in many ways resemble extracellular vesicles more than cells, and the research community has been capable of building such things for a few years now.

Researchers have successfully treated a cancerous tumor using a "nanofactory" - a synthetic cell that produces anti-cancer proteins within the tumor tissue. The research combines synthetic biology, to artificially produce proteins, and targeted drug delivery, to direct the synthetic cell to abnormal tissues. The synthetic cells are artificial systems with capacities similar to, and, at times, superior to those of natural cells. Just as human cells can generate a variety of biological molecules, the synthetic cell can produce a wide range of proteins. Such systems bear vast potential in the tissue engineering discipline, in production of artificial organs and in studying the origins of life. Design of artificial cells is a considerably complex engineering challenge being pursued by many research groups across the globe.

The researchers integrated molecular machines within lipid-based particles resembling the natural membrane of biological cells. They engineered the particles such that when they "sense" the biological tissue, they are activated and produce therapeutic proteins, dictated by an integrated synthetic DNA template. The particles recruit the energy sources and building blocks necessary for their continued activity, from the external microenvironment (e.g., the tumor tissue).

After experiments in cell cultures in the lab, the novel technology was also tested in mice. When the engineered particles reached the tumor, they produced a protein that eradicated the cancer cells. The particles and their activity were monitored using a green fluorescent protein (GFP), generated by the particles. This protein can be viewed in real-time, using a fluorescence microscope. "By coding the integrated DNA template, the particles we developed can produce a variety of protein medicines. They are modular, meaning they allow for activation of protein production in accordance with the environmental conditions. Therefore, the artificial cells we've developed may take an important part in the personalized medicine trend - adjustment of treatment to the genetic and medical profile of a specific patient."

Link: http://ats.org/news/synthetic-cell-produces-anti-cancer-drugs-within-a-tumor/

HDAC3 Knockout Mice Exhibit Greatly Reduced Loss of Memory Function with Age

Work on the decline of memory formation with aging was presented at a recent conference and is doing the rounds in the press. The core of it was published and presented last year, so the overall topic isn't particularly new, but I didn't notice it at the time. The scientific group in question is interested in the role of histone deacetylases (HDACs) in memory. This is a long-running thread of research. Looking back in the Fight Aging! archives, inhibition of HDACs in the context of improved neural function was mentioned in 2012, and a trail of publications exists prior and since.

The processes of acetylation and deacetylation of histones are important to gene regulation, a core part of the machinery that controls the packaged state of nuclear DNA in the cell nucleus. Genes must be accessible to the machinery of the cell in order to begin transcription, the first step in the complex operations involved in constructing proteins from their genetic blueprints. Whether a specific gene is accessible or inaccessible is determined by the state of various different histones, among other mechanisms. What does this have to do with memory? The formation of memory requires reliable access to certain genes, and the production of their proteins, but it is apparently the case that access becomes less available with age. One of the histones, HDAC3, becomes overactive, keeping DNA more tightly packaged than was the case in younger individuals.

Researchers have now demonstrated that mice lacking HDAC3 do not seem to suffer much in the way of side-effects, and also do not suffer age-related loss of memory - though this effect differs in detail for the various types of memory tested to date. It is worth considering that these mechanisms are a snapshot of some middle layer of the long chain of cause and effect that stretches between the root causes of aging and the ultimate consequences of age-related disease and organ failure. Why does HDAC3 become more active in older individuals? What underlying process is taking place, and what other harms is that process causing? Leaping to interventions has a way of short-cutting the conversation about deeper causes that should be taking place, especially when the interventions are comparatively easy to implement - there are plenty of approaches to HDAC3 inhibition that might be taken at low cost and in the near future, even given the need to bypass the blood-brain barrier. But what is shut out by taking that path as the primary focus?

Research cracks code to restoring memory creation in older or damaged brains

Aging or impaired brains can once again form lasting memories if an enzyme that applies the brakes too hard on a key gene is lifted. "What we've discovered is that if we free up that DNA again, now the aging brain can form long-term memories normally. In order to form a long-term memory, you have to turn specific genes on. In most young brains, that happens easily, but as we get older and our brain gets older, we have trouble with that." That's because the 6 feet of DNA spooled tightly into every cell in our bodies has a harder time releasing itself as needed. Like many body parts, "it's no longer as flexible as it used to be." The stiffness in this case is due to a molecular brake pad called histone deacetylase 3, or HDAC3, that has become "overeager" in the aged brain and is compacting the material too hard, blocking the release of a gene called Period1. Removing HDAC3 restores flexibility and allows internal cell machinery to access Period1 to begin forming new memories.

Researchers had previously theorized that the loss of transcription and encoding functions in older brains was due to deteriorating core circadian clocks. But it was found that the ability to create lasting memories was linked to a different process - the overly aggressive enzyme blocking the release of Period1 - in the same hippocampus region of the brain. That's potentially good news for developing treatments. "New drugs targeting HDAC3 could provide an exciting avenue to allow older people to improve memory formation."

NIH Summit Examines What Makes a Healthy Aging Brain

Histone deacetylase HDAC3 is expressed predominantly in the brain and represses gene expression. Researchers knocked out the HDAC3 gene in the dorsal hippocampi of mice, then trained them at young or old ages on a novel object-location task. Young mice performed equally well, regardless of whether they expressed HDAC3. In older animals the story was different. Wild-type animals became forgetful, whereas HDAC3-deficient mice remembered just as well as did young mice. This suggests HDAC3 hampers memory as mice age. In keeping with this, long-term potentiation weakened with age in wild-type but not HDAC3 knockout mice. Since knocking out HDAC3 restored hippocampal expression of Period1 (Per1), a master regulator of the cellular circadian clock, HDAC3 might function to help regulate circadian genes in the hippocampus.

Distinct roles for the deacetylase domain of HDAC3 in the hippocampus and medial prefrontal cortex in the formation and extinction of memory

Histone deacetylases (HDACs) are chromatin modifying enzymes that have been implicated as powerful negative regulators of memory processes. HDAC3 has been shown to play a pivotal role in long-term memory for object location as well as the extinction of cocaine-associated memory, but it is unclear whether this function depends on the deacetylase domain of HDAC3. Here, we tested whether the deacetylase domain of HDAC3 has a role in object location memory formation as well as the formation and extinction of cocaine-associated memories. Using a deacetylase-dead point mutant of HDAC3, we found that selectively blocking HDAC3 deacetylase activity in the dorsal hippocampus enhanced long-term memory for object location, but had no effect on the formation of cocaine-associated memory.

When this same point mutant virus of HDAC3 was infused into the prelimbic cortex, it failed to affect cocaine-associated memory formation. With regards to extinction, impairing the HDAC3 deacetylase domain in the infralimbic cortex had no effect on extinction, but a facilitated extinction effect was observed when the point mutant virus was delivered to the dorsal hippocampus. These results suggest that the deacetylase domain of HDAC3 plays a selective role in specific brain regions underlying long-term memory formation of object location as well as cocaine-associated memory formation and extinction.

Further Investigation of the Role of Osteopontin in Hematopoietic Stem Cell Aging

The Hematopoietic stem cell population resident in bone marrow is responsible for generating blood cells and immune cells. Like all stem cell populations, their activity alters and declines with aging. This is one of the causes of the progressive disarray of the immune system in older individuals. If we want to rejuvenate the immune system, then restoring the youthful activity of hematopoietic stem cells is one of the items on the to-do list, alongside regrowth of the thymus, and clearing out the accumulation of exhausted, senescent, and misconfigured immune cells.

The protein osteopontin appears to have a sizable role in maintaining the hematopoietic stem cell population, but levels fall in older individuals. Researchers have demonstrated, in mice, that restoring high levels osteopontin can also restore a significant degree of hematopoietic stem cell activity. This is promising because it is comparatively simple to achieve and package as a therapy, but equally it isn't addressing whatever root causes underlie this narrow view of the picture. The open access paper here continues the investigation of osteopontin in the context of hematopoietic aging in mice, adding further evidence for its relevance.

In mammalian tissues that undergo high cell turnover, such as the hematopoietic system, a small population of stem cells maintains organ regeneration throughout the animal's life span. However, the functionality of stem cells declines during aging and can contribute to aging-associated impairments in tissue regeneration. Accumulating evidence indicates that aged hematopoietic stem cells (HSCs) increase in number due to a higher rate of self-renewal cell divisions while displaying reduced ability to reconstitute the immune system.

The phosphorylated glycoprotein osteopontin (OPN) is an extracellular matrix component of the bone marrow with important roles in tissue homeostasis, inflammatory responses, and tumor metastasis. The expression of OPN within the bone marrow is highly restricted to the endosteal surface, a location where HSCs have been found to reside preferentially. OPN binds to cells through integrins or the CD44 receptor, subsequently activating multiple signaling pathways. When HSCs are transplanted into wild-type (WT) or OPN knockout mice, they exhibit aberrant attachment and engraftment, suggesting the dependence of HSCs on OPN in these processes. Moreover, OPN deficiency within the bone marrow microenvironment results in an increase in primitive HSC numbers. More recently it has been reported that osteopontin exposure to aged HSC can attenuate their aging-associated phenotype.

Here, we study the impact of OPN on HSC function during aging using an OPN-knockout mouse model. We show that during aging OPN deficiency is associated with an increase in lymphocytes and a decline in erythrocytes in peripheral blood. In a bone marrow transplantation setting, aged OPN-deficient stem cells show reduced ability to reconstitute the immune system likely due to insufficient differentiation of HSCs into more mature cells. In serial bone marrow transplantation, aged OPN knockout bone marrow cells fail to adequately reconstitute red blood cells and platelets, resulting in severe anemia and thrombocytopenia as well as premature deaths of recipient mice. Thus, OPN has different effects on HSCs in aged and young animals and is particularly important to maintain stem cell function in aging mice.

Link: https://doi.org/10.1038/s41598-018-21324-x

Bacteria Engineered to Deliver CXCL12 Accelerate Wound Healing in Mice

Researchers here report on an interesting approach used to deliver a therapeutic molecule into wounds, and thereby accelerate regeneration. They engineered a common bacterial species to produce the molecule of interest, CXCL12, which is implicated in the processes of wound healing. Those processes are an intricate dance between various types of immune cell, stem cell, senescent cell and somatic cells in the injured tissue. In recent years researchers have gained an increased understanding of the scope of involvement of immune cells known as macrophages; the participation of the immune system has turned out to be much more important to the quality and pace of regeneration than was thought a few decades ago. Macrophages can adopt different states, or polarizations. Of the two commonly observed polarizations, one is inflammatory and harmful to regeneration, while the other assists regeneration. There appears to be some potential in therapies that adjust the proportions of a macrophage population in injured tissue to favor the second type - and this is one of the goals that the researchers here aimed to achieve in their study.

During the inflammation phase of wound healing, immune cells accumulate in response to alarm signals, cytokines, and chemokines released by injured or activated cells. The chemokine CXCL12 (Stromal cell-Derived Factor 1α) is associated with beneficial effects in models of cutaneous wounds and binds CXCR4 expressed by immune cells and keratinocytes. Macrophages and neutrophils represent the major immune cell populations at the wound site, where they are essential for keeping invading microorganisms at bay and also for fueling the healing process by secreting additional chemokines, growth factors, and matrix digesting enzymes. During the course of healing, macrophages shift phenotype toward an anti-inflammatory one and subsequently promote tissue restitution. This shift is induced by macrophage phagocytosis of cell debris and by microenvironmental signals such as CXCL12.

Chronic wounds are often associated with underlying pathologic processes that increase susceptibility for acquiring wounds (e.g., peripheral neuropathies) and/or reduced healing abilities as seen in persons with arterial or venous insufficiencies. Several experimental and clinical trials have investigated the effects of local application of growth factors alone or coupled to different biomaterials on different types of chronic wounds, but with modest results so far.

This study aimed to accelerate wound healing by targeting the function of immune cells through local bioengineering of the wound microenvironment. To achieve this, a technology optimized to deliver chemokines directly to wounded skin was developed, whereby lactic acid bacteria were used as vectors. Lactobacillus reuteri bacteria were transformed with a plasmid encoding the chemokine CXCL12 previously associated with beneficial effects in models of healing and blood-flow restoration. Bacteria-produced lactic acid reduced the pH in the wound and thereby potentiated the effects of the produced CXCL12 by prolonging its bioavailability. The overall result of topical wound treatment with this on-site chemokine delivery system was strongly accelerated wound closure to an extent not reported before.

Importantly, treatment with CXCL12-delivering Lactobacilli also improved wound closure in mice with hyperglycemia or peripheral ischemia, conditions associated with chronic wounds, and in a human skin wound model. Further, initial safety studies demonstrated that the topically applied transformed bacteria exerted effects restricted to the wound, as neither bacteria nor the chemokine produced could be detected in systemic circulation.

Link: https://doi.org/10.1073/pnas.1716580115

Development of Exosome Delivery as a Regenerative Therapy Continues Apace

If many stem cell therapies produce their benefits largely through the signaling generated by the transplanted cells, in a brief window of time before these cells die, unable to integrate into the local tissue, then why not skip the cells entirely and just deliver the signals? This is made an easier prospect by the fact that a great deal of cell to cell signaling takes the form of extracellular vesicles such as exosomes, tiny membrane-bound packages of various molecules. Thus researchers don't need to completely map and understand the entire set of signals used in order to recreate most of the signaling effects of stem cells. Given a cultured stem cell population, the exosomes that the cells produce can be harvested and then employed as a therapy. Further down the line, after the mapping and the understanding is complete, then manufacture of exosomes from scratch will probably become the standard approach. For now, cells are required for that much, at least.

The research noted here is an illustrative example of present work on exosome-based regenerative therapies; a fair number of research groups are working towards treatments for various tissue types and age-related conditions. As a class, exosome therapies seem about as promising as early stem cell therapies, based on the results to date in animal models, and are arguably more easily controlled and managed than cells. Just considering the logistics of manufacture and storage, the costs should be significantly lower. Scientists are working their way up from mice to larger animal models, and the first human clinical trials for various conditions are on the near horizon. It is a significant shift in focus for the stem cell research community, and it will be interesting to see where this leads in the next few years.

Stem-cell based stroke treatment repairs brain tissue

A team of researchers and ArunA Biomedical, a startup company, have developed a new treatment for stroke that reduces brain damage and accelerates the brain's natural healing tendencies in animal models. The research team created a treatment called AB126 using extracellular vesicles (EV), fluid-filled structures known as exosomes, which are generated from human neural stem cells. Fully able to cloak itself within the bloodstream, this type of regenerative EV therapy appears to be the most promising in overcoming the limitations of many cell therapies ­- with the ability for exosomes to carry and deliver multiple doses - as well as the ability to store and administer treatment. Small in size, the tiny tubular shape of an exosome allows EV therapy to cross barriers that cells cannot.

Following the administration of AB126, the researchers used MRI scans to measure brain atrophy rates in preclinical, age-matched stroke models, which showed an approximately 35 percent decrease in the size of injury and 50 percent reduction in brain tissue loss - something not observed acutely in previous studies of exosome treatment for stroke. Outside of rodents, the results were replicated using a porcine model of stroke. Based on these pre-clinical results, ArunA Biomedical plans to begin human studies in 2019. The company has plans to expand this initiative beyond stroke for preclinical studies in epilepsy, traumatic brain and spinal cord injuries later this year.

Human Neural Stem Cell Extracellular Vesicles Improve Tissue and Functional Recovery in the Murine Thromboembolic Stroke Model

Over 700 drugs have failed in stroke clinical trials, an unprecedented rate thought to be attributed in part to limited and isolated testing often solely in "young" rodent models and focusing on a single secondary injury mechanism. Here, extracellular vesicles (EVs), nanometer-sized cell signaling particles, were tested in a mouse thromboembolic (TE) stroke model. Neural stem cell (NSC) and mesenchymal stem cell (MSC) EVs derived from the same pluripotent stem cell (PSC) line were evaluated for changes in infarct volume as well as sensorimotor function.

NSC EVs improved cellular, tissue, and functional outcomes in middle-aged rodents, whereas MSC EVs were less effective. Acute differences in lesion volume following NSC EV treatment were corroborated by MRI in 18-month-old aged rodents. NSC EV treatment has a positive effect on motor function in the aged rodent as indicated by beam walk, instances of foot faults, and strength evaluated by hanging wire test. Increased time with a novel object also indicated that NSC EVs improved episodic memory formation in the rodent. The therapeutic effect of NSC EVs appears to be mediated by altering the systemic immune response. These data strongly support further preclinical development of a NSC EV-based stroke therapy and warrant their testing in combination with FDA-approved stroke therapies.

Macrophages Make a Significant Contribution to Heart Failure

Researchers here implicate the immune cells known as macrophages in the progression of a particularly problematic form of heart failure. Macrophages are very important to the processes of tissue maintenance and regeneration, but they have several different characteristic states, or polarizations: one is inflammatory and aggressive, hindering regeneration, while the other is actively beneficial for regeneration. Researchers are finding that adjusting the proportion of these two states can be beneficial. The situation in heart failure - and a number of other age-related conditions - may well be made worse due to the balance in macrophage populations tipping away from assisting regeneration and towards chronic inflammation. In support of that view, stem cell therapies that have the primary outcome of reducing inflammation have been shown to be helpful in treating the form of heart failure examined here.

Researchers have discovered that the immune cells called macrophages contribute to a type of heart failure for which there currently is no effective treatment, heart failure with preserved ejection fraction (HFpEF). The concept of heart failure traditionally referred to a loss of the organ's pumping capacity, which is called systolic heart failure. But in HFpEF the heart retains the ability to pump or eject blood into the circulation. What is compromised is the ability of the heart muscle to relax and allow blood to flow into the left ventricle, reducing the amount of blood available to pump into the aorta. Symptoms of HFpEF are similar to those of heart failure in general, but since factors contributing to the condition are not well understood, it has been difficult to find promising therapies.

Interactions among cells within the heart - including macrophages - are essential to normal cardiac function but can also contribute to problems. For example, after the heart muscle is damaged by a heart attack, macrophages induce the cells called fibroblasts to generate the connective tissues that help reinforce damaged tissue. But excessive fibroblast activation can lead to the distortion and stiffening of tissues, further reducing cardiac function.

To explore a potential role for macrophages in HFpEF, the team examined cardiac macrophages in two mouse models that develop the sort of diastolic dysfunction - impaired relaxation of the heart muscle - that characterizes HFpEF. Those animals were found to have increased macrophage density in the left ventricle and exhibited elevated levels of a factor called IL-10, which is known to contribute to fibroblast activation. Deletion of IL-10 from cardiac macrophages in one model, in which the development of hypertension is induced, prevented the upregulation of macrophages and reduced the numbers and activation of cardiac fibroblasts. Levels of cardiac macrophages were also elevated in tissue biopsies from human patients with HFpEF, as were levels of circulating monocytes, which are precursors of macrophages.

"Not only were numbers of inflammatory cardiac macrophages increased in both the mice and in humans with HFpEF, but their characteristics and functions were also different from those in a healthy heart. Through their participation in the remodeling of heart tissue, these macrophages increase the production of extracellular matrix, which reduces diastolic relaxation. Our findings regarding the cell-specific knockout of IL-10 are the first to support the contribution of macrophages to HFpEF. Heart muscle cells and fibroblasts have been considered the major contributors to HFpEF. Our identification of the central involvement of macrophages should give us a new focus for drug development."

Link: http://www.massgeneral.org/News/pressrelease.aspx?id=2212

BACE1 Deletion Eliminates Amyloid Deposits in a Mouse Model of Alzheimer's Disease

BACE1 is one of the proteins involved in early stages of the production of amyloid-β, a form of metabolic waste that aggregates into solid deposits in the aging brain, and is characteristic of Alzheimer's disease. Inhibition of BACE1 so as to reduce levels of amyloid-β is a strategy pursued by a number of research groups, though it has to be said that disenchantment with the years of failure in the dominant strategy of clearing amyloid-β appears to be reaching a tipping point these days. While it is clear that amyloid-β is harmful, it may not be the most effective point of intervention. Or perhaps earlier efforts to remove amyloid-β were not going about it in the right way, and different approaches would work. It is very hard to say, as the aging brain is a complex mix of many different, interacting forms of damage and dysfunction.

The research here can be read as strong support for the BACE1 inhibition approach to Alzheimer's disease, given the size of the effect, though the same questions remain as in any other success in reducing amyloid from the mouse models of Alzheimer's disease. If none of the others successfully translated to human therapies, and failed in trials, how confident or hopeful should we be here? A great many people are asking themselves exactly that these days, which is why we can observe the growth of support for the impaired cerebrospinal fluid drainage model of Alzheimer's disease, or the microbial model of the condition, and a range of further theorizing on different causes and different priorities in research and development.

With a large swath of the population entering its senior years, the number of Alzheimer's disease (AD) cases are expected to skyrocket, placing a tremendous burden on the healthcare system. Yet, a glimmer of hope may have just emerged as investigators report that gradually depleting an enzyme called BACE1 completely reverses the formation of amyloid plaques in the brains of mice with AD, subsequently improving the animals' cognitive function. "To our knowledge, this is the first observation of such a dramatic reversal of amyloid deposition in any study of AD mouse models."

One of the earliest events in AD is an abnormal buildup of the beta-amyloid (Aß) peptide, which can form large, amyloid plaques in the brain and disrupt the function of neuronal synapses. Also known as beta-secretase, BACE1 helps produce the Aß peptide by cleaving the amyloid precursor protein (APP). Drugs that inhibit BACE1 are therefore being developed as potential AD treatments but, because BACE1 controls many important processes by cleaving proteins other than APP, these drugs could have serious side effects.

Mice completely lacking BACE1 suffer severe neurodevelopmental defects. To investigate whether inhibiting BACE1 in adults might be less harmful, the research team generated mice that gradually lose this enzyme as they grow older. These mice developed normally and appeared to remain perfectly healthy over time. "To mimic BACE1 inhibition in adults, we generated BACE1 conditional knockout (BACE1fl/fl) mice to induce deletion of BACE1 after passing early developmental stages. Strikingly, sequential and increased deletion of BACE1 in an adult AD mouse model was capable of completely reversing amyloid deposition. This reversal in amyloid deposition also resulted in significant improvement in gliosis and neuritic dystrophy. Moreover, synaptic functions, as determined by long-term potentiation and contextual fear conditioning experiments, were significantly improved, correlating with the reversal of amyloid plaques."

Remarkably, the loss of BACE1 also improved the learning and memory of mice with AD. However, when the researchers made electrophysiological recordings of neurons from these animals, they found that depletion of BACE1 only partially restored synaptic function, suggesting that BACE1 may be required for optimal synaptic activity and cognition.

Link: https://www.genengnews.com/gen-news-highlights/alzheimers-disease-reversed-in-mouse-model/81255493