Friends and foes: Extracellular vesicles in aging and rejuvenation.

Extracellular vesicles (EVs) are released by many different cell types throughout the body and play a role in a diverse range of biological processes. EVs circulating in blood as well as in other body fluids undergo dramatic alterations over an organism's lifespan that are only beginning to be elucidated. The exact nature of these changes is an area of active and intense investigation, but lacks clear consensus due to the substantial heterogeneity in EV subpopulations and insufficiencies in current technologies. Nonetheless, emerging evidence suggests that EVs regulate systemic aging as well as the pathophysiology of age-related diseases. Here, we review the current literature investigating EVs and aging with an emphasis on consequences for the maintenance of human healthspan. Intriguingly, the biological utility of EVs both in vitro and in vivo and across contexts depends on the states of the source cells or tissues. As such, EVs secreted by cells in an aged or pathological state may impose detrimental consequences on recipient cells, while EVs secreted by youthful or healthy cells may promote functional improvement. Thus, it is critical to understand both functions of EVs and tip the balance toward their beneficial effects as an antiaging intervention.

Endothelial ether lipids link the vasculature to blood pressure, behavior, and neurodegeneration.

Vascular disease contributes to neurodegeneration, which is associated with decreased blood pressure in older humans. Plasmalogens, ether phospholipids produced by peroxisomes, are decreased in Alzheimer's disease, Parkinson's disease, and other neurodegenerative disorders. However, the mechanistic links between ether phospholipids, blood pressure, and neurodegeneration are not fully understood. Here, we show that endothelium-derived ether phospholipids affect blood pressure, behavior, and neurodegeneration in mice. In young adult mice, inducible endothelial-specific disruption of PexRAP, a peroxisomal enzyme required for ether lipid synthesis, unexpectedly decreased circulating plasmalogens. PexRAP endothelial knockout (PEKO) mice responded normally to hindlimb ischemia but had lower blood pressure and increased plasma renin activity. In PEKO as compared with control mice, tyrosine hydroxylase was decreased in the locus coeruleus, which maintains blood pressure and arousal. PEKO mice moved less, slept more, and had impaired attention to and recall of environmental events as well as mild spatial memory deficits. In PEKO hippocampus, gliosis was increased, and a plasmalogen associated with memory was decreased. Despite lower blood pressure, PEKO mice had generally normal homotopic functional connectivity by optical neuroimaging of the cerebral cortex. Decreased glycogen synthase kinase-3 phosphorylation, a marker of neurodegeneration, was detected in PEKO cerebral cortex. In a co-culture system, PexRAP knockdown in brain endothelial cells decreased glycogen synthase kinase-3 phosphorylation in co-cultured astrocytes that was rescued by incubation with the ether lipid alkylglycerol. Taken together, our findings suggest that endothelium-derived ether lipids mediate several biological processes and may also confer neuroprotection in mice.

Chi3l1/YKL-40 is controlled by the astrocyte circadian clock and regulates neuroinflammation and Alzheimer's disease pathogenesis.

Regulation of glial activation and neuroinflammation are critical factors in the pathogenesis of Alzheimer's disease (AD). YKL-40, a primarily astrocytic protein encoded by the gene Chi3l1, is a widely studied cerebrospinal fluid biomarker that increases with aging and early in AD. However, the function of Chi3l1/YKL-40 in AD is unknown. In a cohort of patients with AD, we observed that a variant in the human CHI3L1 gene, which results in decreased CSF YKL-40 expression, was associated with slower AD progression. At baseline, Chi3l1 deletion in mice had no effect on astrocyte activation while modestly promoting microglial activation. In a mouse APP/PS1 model of AD, Chi3l1 deletion decreased amyloid plaque burden and increased periplaque expression of the microglial lysosomal marker CD68, suggesting that Chi3l1 may suppress glial phagocytic activation and promote amyloid accumulation. Accordingly, Chi3l1 knockdown increased phagocytosis of zymosan particles and of ß-amyloid peptide in both astrocytes and microglia in vitro. We further observed that expression of Chi3l1 is regulated by the circadian clock, as deletion of the core clock proteins BMAL1 or CLOCK/NPAS2 strongly suppresses basal Chi3l1 expression, whereas deletion of the negative clock regulators PER1/PER2 increased Chi3l1 expression. Basal Chi3l1 mRNA was nonrhythmic because of a long mRNA half-life in astrocytes. However, inflammatory induction of Chi3l1 was gated by the clock. Our findings reveal Chi3l1/YKL-40 as a modulator of glial phagocytic activation and AD pathogenesis in both mice and humans and suggest that the astrocyte circadian clock regulates inflammatory Chi3l1 induction.

The wrinkling of time: Aging, inflammation, oxidative stress, and the circadian clock in neurodegeneration.

A substantial body of research now implicates the circadian clock in the regulation of an array of diverse biological processes including glial function, metabolism, peripheral immune responses, and redox homeostasis. Sleep abnormalities and other forms of circadian disruption are common symptoms of aging and neurodegeneration. Circadian clock disruption may also influence the aging processes and the pathogenesis of neurodegenerative diseases. The specific mechanisms governing the interaction between circadian systems, aging, and the immune system are still being uncovered. Here, we review the evidence supporting a bidirectional relationship between aging and the circadian system. Further, we explore the hypothesis that age-related circadian deterioration may exacerbate multiple pathogenic processes, priming the brain for neurodegeneration.

Circadian regulation of astrocyte function: implications for Alzheimer's disease.

The circadian clock regulates rhythms in gene transcription that have a profound impact on cellular function, behavior, and disease. Circadian dysfunction is a symptom of aging and neurodegenerative diseases, and recent studies suggest a bidirectional relationship between impaired clock function and neurodegeneration. Glial cells possess functional circadian clocks which may serve to control glial responses to daily oscillations in brain activity, cellular stress, and metabolism. Astrocytes directly support brain function through synaptic interactions, neuronal metabolic support, neuroinflammatory regulation, and control of neurovascular coupling at blood and CSF barriers. Emerging evidence suggests that the astrocyte circadian clock may be involved in many of these processes, and that clock disruption could influence neurodegeneration by disrupting several aspects of astrocyte function. Here we review the literature surrounding circadian control of astrocyte function in health and disease, and discuss the potential implications of astrocyte clocks for neurodegeneration.

Circadian clock protein Rev-erbα regulates neuroinflammation.

Circadian dysfunction is a common attribute of many neurodegenerative diseases, most of which are associated with neuroinflammation. Circadian rhythm dysfunction has been associated with inflammation in the periphery, but the role of the core clock in neuroinflammation remains poorly understood. Here we demonstrate that Rev-erbα, a nuclear receptor and circadian clock component, is a mediator of microglial activation and neuroinflammation. We observed time-of-day oscillation in microglial immunoreactivity in the hippocampus, which was disrupted in Rev-erbα-/- mice. Rev-erbα deletion caused spontaneous microglial activation in the hippocampus and increased expression of proinflammatory transcripts, as well as secondary astrogliosis. Transcriptomic analysis of hippocampus from Rev-erbα-/- mice revealed a predominant inflammatory phenotype and suggested dysregulated NF-κB signaling. Primary Rev-erbα-/- microglia exhibited proinflammatory phenotypes and increased basal NF-κB activation. Chromatin immunoprecipitation revealed that Rev-erbα physically interacts with the promoter regions of several NF-κB-related genes in primary microglia. Loss of Rev-erbα in primary astrocytes had no effect on basal activation but did potentiate the inflammatory response to lipopolysaccharide (LPS). In vivo, Rev-erbα-/- mice exhibited enhanced hippocampal neuroinflammatory responses to peripheral LPS injection, while pharmacologic activation of Rev-erbs with the small molecule agonist SR9009 suppressed LPS-induced hippocampal neuroinflammation. Rev-erbα deletion influenced neuronal health, as conditioned media from Rev-erbα-deficient primary glial cultures exacerbated oxidative damage in cultured neurons. Rev-erbα-/- mice also exhibited significantly altered cortical resting-state functional connectivity, similar to that observed in neurodegenerative models. Our results reveal Rev-erbα as a pharmacologically accessible link between the circadian clock and neuroinflammation.

Cell-Autonomous Regulation of Astrocyte Activation by the Circadian Clock Protein BMAL1.

Circadian clock dysfunction is a common symptom of aging and neurodegenerative diseases, though its impact on brain health is poorly understood. Astrocyte activation occurs in response to diverse insults and plays a critical role in brain health and disease. We report that the core circadian clock protein BMAL1 regulates astrogliosis in a synergistic manner via a cell-autonomous mechanism and a lesser non-cell-autonomous signal from neurons. Astrocyte-specific Bmal1 deletion induces astrocyte activation and inflammatory gene expression in vitro and in vivo, mediated in part by suppression of glutathione-S-transferase signaling. Functionally, loss of Bmal1 in astrocytes promotes neuronal death in vitro. Our results demonstrate that the core clock protein BMAL1 regulates astrocyte activation and function in vivo, elucidating a mechanism by which the circadian clock could influence many aspects of brain function and neurological disease.

Genome-independent hypoxic repression of estrogen receptor alpha in breast cancer cells.

BACKGROUND: About 75-80% of breast tumors express the estrogen receptor alpha (ER-α) and are treated with endocrine-target therapeutics, making this the premier therapeutic modality in the breast cancer clinic. However, acquired resistance is common and about 20% of resistant tumors loose ER-α expression via unknown mechanisms. Inhibition of ER-α loss could improve endocrine therapeutic efficacy, benefiting a significant number of patients. Here we test whether tumor hypoxia might commonly produce ER-α loss. METHODS: Using standard molecular and cellular biological assays and a work station/incubator with controllable oxygen levels, we analyze the effects of hypoxia on ER-α protein, mRNA, and transcriptional activity in a panel of independently-derived ER-α positive cell lines. These lines were chosen to represent the diverse genetic backgrounds and mutations commonly present in ER-α positive tumors. Using shRNA-mediated knockdown and overexpression studies we also elucidate the role of hypoxia-inducible factor 1-alpha (HIF-1α) in the hypoxia-induced decrease in ER-α abundance. RESULTS: We present the first comprehensive overview of the effects of bona fide low environmental oxygen (hypoxia) and HIF-1α activity on ER-α abundance and transcriptional activity. We find that stabilized HIF-1α induces rapid loss of ER-α protein in all members of our diverse panel of breast cancer cell lines, which involves proteolysis rather than transcriptional repression. Reduced ER-α severely attenuates ER-α directed transcription, and inhibits cell proliferation without overt signs of cell death in the cell lines tested, despite their varying genomic backgrounds. CONCLUSIONS: These studies reveal a common hypoxia response that produces reduced ER-α expression and cell cycle stalling, and demonstrate a common role for HIF-1α in ER-α loss. We hypothesize that inhibitors of HIF-1α or the proteasome might stabilize ER-α expression in breast tumors in vivo, and work in combination with endocrine therapies to reduce resistance. Our data also suggests that disease re-occurrence in patients with ER-α positive tumors may arise from tumor cells chronically resident in hypoxic environments. We hypothesize that these non-proliferating cells may survive undetected until conditions change to oxygenate the environment, or cells eventually switch to proliferation via other signaling pathways.