A Circadian Clock in the Blood-Brain Barrier Regulates Xenobiotic Efflux.

Endogenous circadian rhythms are thought to modulate responses to external factors, but mechanisms that confer time-of-day differences in organismal responses to environmental insults/therapeutic treatments are poorly understood. Using a xenobiotic, we find that permeability of the Drosophila "blood"-brain barrier (BBB) is higher at night. The permeability rhythm is driven by circadian regulation of efflux and depends on a molecular clock in the perineurial glia of the BBB, although efflux transporters are restricted to subperineurial glia (SPG). We show that transmission of circadian signals across the layers requires cyclically expressed gap junctions. Specifically, during nighttime, gap junctions reduce intracellular magnesium ([Mg2+]i), a positive regulator of efflux, in SPG. Consistent with lower nighttime efflux, nighttime administration of the anti-epileptic phenytoin is more effective at treating a Drosophila seizure model. These findings identify a novel mechanism of circadian regulation and have therapeutic implications for drugs targeted to the central nervous system.

Senescent glia link mitochondrial dysfunction and lipid accumulation.

Senescence is a cellular state linked to ageing and age-onset disease across many mammalian species1,2. Acutely, senescent cells promote wound healing3,4 and prevent tumour formation5; but they are also pro-inflammatory, thus chronically exacerbate tissue decline. Whereas senescent cells are active targets for anti-ageing therapy6-11, why these cells form in vivo, how they affect tissue ageing and the effect of their elimination remain unclear12,13. Here we identify naturally occurring senescent glia in ageing Drosophila brains and decipher their origin and influence. Using Activator protein 1 (AP1) activity to screen for senescence14,15, we determine that senescent glia can appear in response to neuronal mitochondrial dysfunction. In turn, senescent glia promote lipid accumulation in non-senescent glia; similar effects are seen in senescent human fibroblasts in culture. Targeting AP1 activity in senescent glia mitigates senescence biomarkers, extends fly lifespan and health span, and prevents lipid accumulation. However, these benefits come at the cost of increased oxidative damage in the brain, and neuronal mitochondrial function remains poor. Altogether, our results map the trajectory of naturally occurring senescent glia in vivo and indicate that these cells link key ageing phenomena: mitochondrial dysfunction and lipid accumulation.

Hypermetabolic state is associated with circadian rhythm disruption in mouse and human cancer cells.

Crosstalk between metabolism and circadian rhythms is a fundamental building block of multicellular life, and disruption of this reciprocal communication could be relevant to disease. Here, we investigated whether maintenance of circadian rhythms depends on specific metabolic pathways, particularly in the context of cancer. We found that in adult mouse fibroblasts, ATP levels were a major contributor to signal from a clock gene luciferase reporter, although not necessarily to the strength of circadian cycling. In contrast, we identified significant metabolic control of circadian function across a series of pancreatic adenocarcinoma cell lines. Metabolic profiling of congenic tumor cell clones revealed substantial diversity among these lines that we used to identify clones to generate circadian reporter lines. We observed diverse circadian profiles among these lines that varied with their metabolic phenotype: The most hypometabolic line [exhibiting low levels of oxidative phosphorylation (OxPhos) and glycolysis] had the strongest rhythms, while the most hypermetabolic line had the weakest rhythms. Pharmacological enhancement of OxPhos decreased the amplitude of circadian oscillation in a subset of tumor cell lines. Strikingly, inhibition of OxPhos enhanced circadian rhythms only in the tumor cell line in which glycolysis was also low, thereby establishing a hypometabolic state. We further analyzed metabolic and circadian phenotypes across a panel of human patient-derived melanoma cell lines and observed a significant negative association between metabolic activity and circadian cycling strength. Together, these findings suggest that metabolic heterogeneity in cancer directly contributes to circadian function and that high levels of glycolysis or OxPhos independently disrupt circadian rhythms in these cells.

Circadian Rhythms of the Blood-Brain Barrier and Drug Delivery.

The blood-brain barrier (BBB) is a critical interface separating the central nervous system from the peripheral circulation, ensuring brain homeostasis and function. Recent research has unveiled a profound connection between the BBB and circadian rhythms, the endogenous oscillations synchronizing biological processes with the 24-hour light-dark cycle. This review explores the significance of circadian rhythms in the context of BBB functions, with an emphasis on substrate passage through the BBB. Our discussion includes efflux transporters and the molecular timing mechanisms that regulate their activities. A significant focus of this review is the potential implications of chronotherapy, leveraging our knowledge of circadian rhythms for improving drug delivery to the brain. Understanding the temporal changes in BBB can lead to optimized timing of drug administration, to enhance therapeutic efficacy for neurological disorders while reducing side effects. By elucidating the interplay between circadian rhythms and drug transport across the BBB, this review offers insights into innovative therapeutic interventions.

Ligand-free mitochondria-localized mutant AR-induced cytotoxicity in spinal bulbar muscular atrophy.

Spinal bulbar muscular atrophy (SBMA), the first identified CAG-repeat expansion disorder, is an X-linked neuromuscular disorder involving CAG-repeat-expansion mutations in the androgen receptor (AR) gene. We utilized CRISPR-Cas9 gene editing to engineer novel isogenic human induced pluripotent stem cell (hiPSC) models, consisting of isogenic AR knockout, control and disease lines expressing mutant AR with distinct repeat lengths, as well as control and disease lines expressing FLAG-tagged wild-type and mutant AR, respectively. Adapting a small-molecule cocktail-directed approach, we differentiate the isogenic hiPSC models into motor neuron-like cells with a highly enriched population to uncover cell-type-specific mechanisms underlying SBMA and to distinguish gain- from loss-of-function properties of mutant AR in disease motor neurons. We demonstrate that ligand-free mutant AR causes drastic mitochondrial dysfunction in neurites of differentiated disease motor neurons due to gain-of-function mechanisms and such cytotoxicity can be amplified upon ligand (androgens) treatment. We further show that aberrant interaction between ligand-free, mitochondria-localized mutant AR and F-ATP synthase is associated with compromised mitochondrial respiration and multiple other mitochondrial impairments. These findings counter the established notion that androgens are requisite for mutant AR-induced cytotoxicity in SBMA, reveal a compelling mechanistic link between ligand-free mutant AR, F-ATP synthase and mitochondrial dysfunction, and provide innovative insights into motor neuron-specific therapeutic interventions for SBMA.

G1/S cell cycle regulators mediate effects of circadian dysregulation on tumor growth and provide targets for timed anticancer treatment.

Circadian disruption has multiple pathological consequences, but the underlying mechanisms are largely unknown. To address such mechanisms, we subjected transformed cultured cells to chronic circadian desynchrony (CCD), mimicking a chronic jet-lag scheme, and assayed a range of cellular functions. The results indicated a specific circadian clock-dependent increase in cell proliferation. Transcriptome analysis revealed up-regulation of G1/S phase transition genes (myelocytomatosis oncogene cellular homolog [Myc], cyclin D1/3, chromatin licensing and DNA replication factor 1 [Cdt1]), concomitant with increased phosphorylation of the retinoblastoma (RB) protein by cyclin-dependent kinase (CDK) 4/6 and increased G1-S progression. Phospho-RB (Ser807/811) was found to oscillate in a circadian fashion and exhibit phase-shifted rhythms in circadian desynchronized cells. Consistent with circadian regulation, a CDK4/6 inhibitor approved for cancer treatment reduced growth of cultured cells and mouse tumors in a time-of-day-specific manner. Our study identifies a mechanism that underlies effects of circadian disruption on tumor growth and underscores the use of treatment timed to endogenous circadian rhythms.

Losing TREC with age.

In this issue of Immunity, den Braber et al. (2012) highlight differences in naive T cell lifespan between mice and humans. Their data suggest that mechanisms of naive T cell maintenance may differ between mice and men.

Characterization of COVID-19 in Assisted Living Facilities - 39 States, October 2020.

The coronavirus disease 2019 (COVID-19) pandemic has highlighted the vulnerability of residents and staff members in long-term care facilities (LTCFs) (1). Although skilled nursing facilities (SNFs) certified by the Centers for Medicare & Medicaid Services (CMS) have federal COVID-19 reporting requirements, national surveillance data are less readily available for other types of LTCFs, such as assisted living facilities (ALFs) and those providing similar residential care. However, many state and territorial health departments publicly report COVID-19 surveillance data across various types of LTCFs. These data were systematically retrieved from health department websites to characterize COVID-19 cases and deaths in ALF residents and staff members. Limited ALF COVID-19 data were available for 39 states, although reporting varied. By October 15, 2020, among 28,623 ALFs, 6,440 (22%) had at least one COVID-19 case among residents or staff members. Among the states with available data, the proportion of COVID-19 cases that were fatal was 21.2% for ALF residents, 0.3% for ALF staff members, and 2.5% overall for the general population of these states. To prevent the introduction and spread of SARS-CoV-2, the virus that causes COVID-19, in their facilities, ALFs should 1) identify a point of contact at the local health department; 2) educate residents, families, and staff members about COVID-19; 3) have a plan for visitor and staff member restrictions; 4) encourage social (physical) distancing and the use of masks, as appropriate; 5) implement recommended infection prevention and control practices and provide access to supplies; 6) rapidly identify and properly respond to suspected or confirmed COVID-19 cases in residents and staff members; and 7) conduct surveillance of COVID-19 cases and deaths, facility staffing, and supply information (2).

The anti-inflammatory TIPE2 is an inhibitor of the oncogenic Ras.

The connection between cancer and inflammation is widely recognized, yet the underlying molecular mechanisms are poorly understood. We report here that TIPE2 provides a molecular bridge from inflammation to cancer by targeting the Ras signaling pathway. TIPE2 binds the Ras-interacting domain of the RalGDS family of proteins, which are essential effectors of activated Ras. This binding prevented Ras from forming an active complex, thereby inhibiting the activation of the downstream signaling molecules Ral and AKT. Consequently, TIPE2 deficiency led to heightened activation of Ral and AKT, resistance to cell death, increased migration, and dysregulation of exocyst complex formation. Conversely, TIPE2 overexpression induced cell death and significantly inhibited Ras-induced tumorigenesis in mice. Importantly, TIPE2 expression was either completely lost or significantly downregulated in human hepatic cancer. Thus, TIPE2 is an inhibitor of both inflammation and cancer, and a potential drug target for inflammatory and neoplastic diseases.

Human and rat gut microbiome composition is maintained following sleep restriction.

Insufficient sleep increasingly characterizes modern society, contributing to a host of serious medical problems. Loss of sleep is associated with metabolic diseases such as obesity and diabetes, cardiovascular disorders, and neurological and cognitive impairments. Shifts in gut microbiome composition have also been associated with the same pathologies; therefore, we hypothesized that sleep restriction may perturb the gut microbiome to contribute to a disease state. In this study, we examined the fecal microbiome by using a cross-species approach in both rat and human studies of sleep restriction. We used DNA from hypervariable regions (V1-V2) of 16S bacteria rRNA to define operational taxonomic units (OTUs) of the microbiome. Although the OTU richness of the microbiome is decreased by sleep restriction in rats, major microbial populations are not altered. Only a single OTU, TM7-3a, was found to increase with sleep restriction of rats. In the human microbiome, we find no overt changes in the richness or composition induced by sleep restriction. Together, these results suggest that the microbiome is largely resistant to changes during sleep restriction.

Chemokine treatment rescues profound T-lineage progenitor homing defect after bone marrow transplant conditioning in mice.

Development of T cells in the thymus requires continuous importation of T-lineage progenitors from the bone marrow via the circulation. Following bone marrow transplant, recovery of a normal peripheral T-cell pool depends on production of naïve T cells in the thymus; however, delivery of progenitors to the thymus limits T-lineage reconstitution. Here, we examine homing of intravenously delivered progenitors to the thymus following irradiation and bone marrow reconstitution. Surprisingly, following host conditioning by irradiation, we find that homing of lymphoid-primed multipotent progenitors and common lymphoid progenitors to the thymus decreases more than 10-fold relative to unirradiated mice. The reduction in thymic homing in irradiated mice is accompanied by a significant reduction in CCL25, an important chemokine ligand for thymic homing. We show that pretreatment of bone marrow progenitors with CCL25 and CCL21 corrects the defect in thymic homing after irradiation and promotes thymic reconstitution. These data suggest new therapeutic approaches to promote T-cell regeneration.

Both retention and recirculation contribute to long-lived regulatory T-cell accumulation in the thymus.

Natural Treg cells acquire their lineage-determining transcription factor Foxp3 during development in the thymus and are important in maintaining immunologic tolerance. Here, we analyzed the composition of the thymic Treg-cell pool using RAG2-GFP/FoxP3-RFP dual reporter mice and found that a population of long-lived GFP(-) Treg cells exists in the thymus. These long-lived Treg cells substantially increased with age, to a point where they represent >90% of the total thymic Treg-cell pool at 6 months of age. In contrast, long-lived conventional T cells remained at ∼ 15% of the total thymic pool at 6 months of age. Consistent with these studies, we noticed that host-derived Treg cells represented a large fraction (∼ 10%) of the total thymic Treg-cell pool in bone marrow chimeras, suggesting that long-lived Treg cells also reside in the thymus of these mice. The pool of long-lived Treg cells in the thymus was sustained by retention of Treg cells in the thymus and by recirculation of peripheral Treg cells back into the thymus. These long-lived thymic Treg cells suppressed T-cell proliferation to an equivalent extent to splenic Treg cells. Together, these data demonstrate that long-lived Treg cells accumulate in the thymus by both retention and recirculation.

Trafficking to the thymus.

The continuous production of T lymphocytes requires that hematopoietic progenitors developing in the bone marrow migrate to the thymus. Rare progenitors egress from the bone marrow into the circulation, then traffic via the blood to the thymus. It is now evident that thymic settling is tightly regulated by selectin ligands, chemokine receptors, and integrins, among other factors. Identification of these signals has enabled progress in identifying specific populations of hematopoietic progenitors that can settle the thymus. Understanding the nature of progenitor cells and the molecular mechanisms involved in thymic settling may allow for therapeutic manipulation of this process, and improve regeneration of the T lineage in patients with impaired T cell numbers.

Expression of functional P-selectin glycoprotein ligand 1 on hematopoietic progenitors is developmentally regulated.

T cell development requires periodic importation of hematopoietic progenitors into the thymus. The receptor-ligand pair P-selectin and P-selectin glycoprotein ligand 1 (PSGL-1) are critically involved in this process. In this study, we examined the expression of functional PSGL-1 on bone marrow hematopoietic progenitors. We demonstrate that functional PSGL-1 is expressed at low levels on hematopoietic stem cells, but upregulated on the cell surface of progenitors that bear other homing molecules known to be important for thymic settling. We found that progenitors able to home to the thymus expressed high levels of PSGL-1 transcripts compared with hematopoietic stem cells. We further demonstrate that hematopoietic progenitors lacking fucosyltransferase 4 and 7 do not express functional PSGL-1, and do not home efficiently to the thymus. These studies provide insight into the developmentally regulated expression of a critical determinant involved in progenitor homing to the thymus.

A circadian clock regulates the blood-brain barrier across phylogeny.

As the central regulatory system of an organism, the brain is responsible for overseeing a wide variety of physiological processes essential for an organism's survival. To maintain the environment necessary for neurons to function, the brain requires highly selective uptake and elimination of specific molecules through the blood-brain barrier (BBB). As an organism's activities vary throughout the day, how does the BBB adapt to meet the changing needs of the brain? A mechanism is through temporal regulation of BBB permeability via its circadian clock, which will be the focal point of this chapter. To comprehend the circadian clock's role within the BBB, we will first examine the anatomy of the BBB and the transport mechanisms enabling it to fulfill its role as a restrictive barrier. Next, we will define the circadian clock, and the discussion will encompass an introduction to circadian rhythms, the Transcription-Translation Feedback Loop (TTFL) as the mechanistic basis of circadian timekeeping, and the organization of tissue clocks found in organisms. Then, we will cover the role of the circadian rhythms in regulating the cellular mechanisms and functions of the BBB. We discuss the implications of this regulation in influencing sleep behavior, the progression of neurodegenerative diseases, and finally drug delivery for treatment of neurological diseases.

Differences in the content and coherence of autobiographical memories between younger and older adults: Insights from text analysis.

Several studies have shown that older adults generate autobiographical memories with fewer specific details than younger adults, a pattern typically attributed to age-relate declines in episodic memory. A relatively unexplored question is how aging affects the content used to represent and recall these memories. We recently proposed that older adults may predominately represent and recall autobiographical memories at the gist level. Emerging from this proposal is the hypothesis that older adults represent memories with a wider array of content topics and recall memories with a distinct narrative style when compared to younger adults. We tested this hypothesis by applying natural language processing approaches to a data set of autobiographical memories described by healthy younger and older adults. We used topic modeling to estimate the distribution (i.e., diversity) of content topics used to represent a memory, and sentence embedding to derive an internal similarity score to estimate the shifts in content when narrating a memory. First, we found that older adults referenced a wider array of content topics (higher content diversity) than younger adults when recalling their autobiographical memories. Second, we found older adults were included more content shifts when narrating their memories than younger adults, suggesting a reduced reliance on choronology to form a coherent memory. Third, we found that the content diversity measures were positively related to specific detail generation for older adults, potentially reflecting age-related compensation for episodic memory difficulties. We discuss how our results shed light on how younger and older adults differ in the way they remember and describe the past. (PsycInfo Database Record (c) 2024 APA, all rights reserved).

Arsenic impairs Drosophila neural stem cell mitotic progression and sleep behavior in a tauopathy model.

Despite established exposure limits, arsenic remains the most significant environmental risk factor detrimental to human health and is associated with carcinogenesis and neurotoxicity. Arsenic compromises neurodevelopment, and it is associated with peripheral neuropathy in adults. Exposure to heavy metals, such as arsenic, may also increase the risk of neurodegenerative disorders. Nevertheless, the molecular mechanisms underlying arsenic-induced neurotoxicity remain poorly understood. Elucidating how arsenic contributes to neurotoxicity may mitigate some of the risks associated with chronic sublethal exposure and inform future interventions. In this study, we examine the effects of arsenic exposure on Drosophila larval neurodevelopment and adult neurologic function. Consistent with prior work, we identify significant developmental delays and heightened mortality in response to arsenic. Within the developing larval brain, we identify a dose-dependent increase in brain volume. This aberrant brain growth is coupled with impaired mitotic progression of the neural stem cells (NSCs), progenitors of the neurons and glia of the central nervous system. Live imaging of cycling NSCs reveals significant delays in cell cycle progression upon arsenic treatment, leading to genomic instability. In adults, chronic arsenic exposure reduces neurologic function, such as locomotion. Finally, we show arsenic selectively impairs circadian rhythms in a humanized tauopathy model. These findings inform mechanisms of arsenic neurotoxicity and reveal sex-specific and genetic vulnerabilities to sublethal exposure.

Age-regulated cycling metabolites are relevant for behavior.

Circadian cycles of sleep:wake and gene expression change with age in all organisms examined. Metabolism is also under robust circadian regulation, but little is known about how metabolic cycles change with age and whether these contribute to the regulation of behavioral cycles. To address this gap, we compared cycling of metabolites in young and old Drosophila and found major age-related variations. A significant model separated the young metabolic profiles by circadian timepoint, but could not be defined for the old metabolic profiles due to the greater variation in this dataset. Of the 159 metabolites measured in fly heads, we found 17 that cycle by JTK analysis in young flies and 17 in aged. Only four metabolites overlapped in the two groups, suggesting that cycling metabolites are distinct in young and old animals. Among our top cyclers exclusive to young flies were components of the pentose phosphate pathway (PPP). As the PPP is important for buffering reactive oxygen species, and overexpression of glucose-6-phosphate dehydrogenase (G6PD), a key component of the PPP, was previously shown to extend lifespan in Drosophila, we asked if this manipulation also affects sleep:wake cycles. We found that overexpression in circadian clock neurons decreases sleep in association with an increase in cellular calcium and mitochondrial oxidation, suggesting that altering PPP activity affects neuronal activity. Our findings elucidate the importance of metabolic regulation in maintaining patterns of neural activity, and thereby sleep:wake cycles.

A pathologically expanded, clonal lineage of IL-21 producing CD4+ T cells drives Inflammatory neuropathy.

Inflammatory neuropathies, which include CIDP (chronic inflammatory demyelinating polyneuropathy) and GBS (Guillain Barre Syndrome), result from autoimmune destruction of the peripheral nervous system (PNS) and are characterized by progressive weakness and sensory loss. CD4+ T cells play a key role in the autoimmune destruction of the PNS. Yet, key properties of pathogenic CD4+ T cells remain incompletely understood. Here, we use paired scRNAseq and scTCRseq of peripheral nerves from an inflammatory neuropathy mouse model to identify IL-21 expressing CD4+ T cells that are clonally expanded and multifunctional. These IL-21-expressing CD4+ T cells are comprised of two transcriptionally distinct expanded populations, which express genes associated with Tfh and Tph subsets. Remarkably, TCR clonotypes are shared between these two IL-21-expressing populations, suggesting a common lineage differentiation pathway. Finally, we demonstrate that IL-21 signaling is required for neuropathy development and pathogenic T cell infiltration into peripheral nerves. IL-21 signaling upregulates CXCR6, a chemokine receptor that promotes CD4+ T cell localization in peripheral nerves. Together, these findings point to IL-21 signaling, Tfh/Tph differentiation, and CXCR6-mediated cellular localization as potential therapeutic targets in inflammatory neuropathies.