Space exploration is important for scientific discovery, advancing technology, and the long-term survival of humanity. However, the impacts of microgravity and cosmic radiation during space travel on human physiology are not completely understood. While microgravity results in a loss of skeletal muscle mass and function, the effect on cardiac muscle, in particular the contractile elements, is not as clear. Here, we examine the effect of spaceflight on the myocardial contractile function of skinned cardiomyocytes from mice that traveled to the International Space Station (spaceflight, N = 5) and age-matched ground controls (ground control, N = 5, and vivarium, N = 3). These experiments allow for the characterization of the mechanical properties of the sarcomere, the fundamental unit of contraction. The functional experiments showed that ~38.5 days in space does not alter force-generating capacity (Tmax), calcium sensitivity (EC50), and cooperativity of the sarcomere. The passive force and cross-sectional area (CSA) were the same between the spaceflight and ground control groups. We next performed mass spectrometry (MS) analysis, and gene ontology analysis confirmed that pathways associated with sarcomere contractility remained unchanged. However, the MS data showed that the spaceflight group exhibited immune‑related proteomic changes compared to the ground controls. Together, these results suggest that ~38.5 days of space travel does not substantially affect the intrinsic contractile state of murine cardiomyocytes.

OBJECTIVE:Microvascular hyperpermeability and blood-brain barrier (BBB) dysfunction is a key consideration in neurological disorders, particularly tauopathies, a group of neurodegenerative disorders driven by misfolded and aggregated tau protein. Tau pathology has been shown to activate microglial NLRP3 inflammasome, an innate immune system sensor that responds to changes in the microenvironment, including cellular stress. Increases in reactive oxygen species (ROS), for example, activate NLRP3 inflammasome signaling, which provides a platform for the maturation of caspase-1 enzyme. Mature caspase-1 can cleave and release pro-inflammatory IL-1β cytokine. Both NLRP3 inflammasome and IL-1β may activate downstream MMP-9 enzyme, a known inducer of endothelial cell barrier hyperpermeability. Endothelial cells make up the innermost layer of the BBB and as such, largely govern BBB structural and functional integrity. Whether tau can activate NLRP3 inflammasome signaling in cerebral endothelial cells is unknown. The objective of this study is therefore to understand the role of tau, in various states of aggregation, on endothelial cell permeability and to investigate if NLRP3 inflammasome signaling occurs in this context. METHODS:Human brain microvascular endothelial cells (HBMECs) were grown as a monolayer in Transwell inserts and exposed to various tau polymorphs, including tau monomers, tau oligomers (oTau), and tau fibrils (fTau). Barrier permeability was measured using FITC-dextran fluorescent tracer (10 kDa) and Trans-Endothelial Electrical Resistance (TEER). Relative changes in gene expression were measured using RT-qPCR and normalized to GAPDH. Levels of NLRP3 sensor protein and IL-1β were measured by ELISA. Relative activity of caspase-1 and MMP-9 enzymes was calculated using fluorometry. Cell viability was reported using calcein AM, a measure of cell membrane integrity, and by measuring the redox potential of XTT. ROS formation, apoptosis, and necrosis were determined using commercially available kits. An NLRP3 inflammasome inhibitor, MCC950, was applied in blocking studies prior to tau treatments. RESULTS:Tau oligomers, but not monomers or fibrils, induced endothelial cell hyperpermeability in a dose-independent manner. At concentrations that compromised barrier function (100 nM; overnight), oTau did not alter cell viability and did not increase apoptosis or necrosis. Tau oligomers increased the formation of ROS and increased levels of both NLRP3 sensor protein and IL-1β cytokine. Enzymatic activity of caspase-1 and MMP-9 also increased in response to oTau, without changes in gene expression. These alterations were attenuated when NLRP3 inflammasome signaling was inhibited via MCC950, strongly suggesting that oTau activates NLRP3 inflammasome signaling in cerebral endothelial cells. CONCLUSIONS:Our study is the first to document a role for NLRP3 inflammasome signaling in cerebral endothelial cells following exposure to tau oligomers. Given the importance of endothelial cell functioning in BBB integrity, these data are significant in that they demonstrate a key role for endothelial cell signaling in tau pathogenicity and propose a mechanism by which tauopathies compromise BBB functional integrity. Taken together, these findings warrant future investigation into the therapeutic potential of NLRP3 inflammasome inhibition to ameliorate tauopathy-related barrier breach.

Background Pediatric Acute-onset Neuropsychiatric Syndrome (PANS) is characterized by the abrupt onset of obsessive–compulsive symptoms and/or restrictive eating accompanied by disturbances in sleep, affect regulation, behavior, motor function, and sensory processing. Increasing evidence implicates systemic immune activation and circulating autoantibodies in basal ganglia dysfunction. Blood–brain barrier (BBB) impairment has been proposed as a mechanism by which peripheral inflammatory mediators access the central nervous system; however, the molecular pathways linking systemic inflammation to BBB disruption in PANS remain incompletely defined. Methods To determine whether circulating factors contribute to BBB dysfunction, human brain endothelial cell (BEC) monolayers were exposed to plasma from PANS patients during symptomatic flare and recovery, as well as from matched healthy controls. Barrier integrity was assessed by paracellular permeability assays; transcriptomic changes were analyzed using bulk RNA sequencing. Junctional organization and cytoskeletal architecture were examined by immunofluorescence microscopy. Circulating and endothelial-derived mediators associated with barrier disruption were quantified using multiplex bead–based immunoassays and enzyme-linked immunosorbent assays. Results Plasma from PANS flare (n = 15 samples) significantly increased BEC monolayer permeability compared to plasma from matched controls (~ 50% change in permeability; p < 0.001), corresponding with increased concentration of S100B, an in vivo biomarker of BBB permeability (~ 2 fold increase vs controls; p < 0.01). Transcriptomic profiling of flare samples demonstrated downregulation of genes essential for endothelial stability, including those encoding tight junction, adherens junction, and extracellular matrix components. Immunofluorescence of flare samples confirmed disruption of zonula occludens-1 (ZO-1) and vascular endothelial cadherin (VE-Cadherin), accompanied by increased actin stress fiber formation, consistent with enhanced cytoskeletal tension and junctional disassembly. Matrix metalloproteinase-9 (MMP-9) concentration was elevated in flare plasma vs controls (> 2.5 fold increase; p < 0.001) and was highly correlated with BEC monolayer permeability (r = 0.84; p < 0.001). Inhibition of MMP-9 in flare samples (n = 6) resulted in significant decrease in BEC monolayer permeability ( p < 0.01), approaching that of matched controls. Conclusions Circulating factors present during PANS flares induce brain endothelial dysfunction in vitro , correlating with in vivo biomarker findings. Elevated MMP-9 functions as a key downstream effector linking systemic inflammation to endothelial barrier disruption, providing mechanistic insight into how peripheral immune activation may facilitate neuroinflammation in PANS.

Background: HIV-associated neurocognitive impairment (HAND) is a common complication of HIV-1 infection, which can be exacerbated by exposure to cigarette smoke (CS). Tight junction proteins (TJPs) of the blood-brain barrier (BBB) play a crucial role in maintaining BBB integrity and preventing the entry of circulating toxic factors, including those resulting from HIV-1 infection, into the central nervous system. Both CS exposure and HIV-1 infection can independently disrupt TJPs and compromise BBB integrity; however, the combined or individual effects of these factors on BBB TJPs remain poorly understood. Methods: An in vitro BBB comprised of Sprague-Dawley rat brain microvascular endothelial cell (RBMVEC) transwell cultures was exposed to wild-type (WT) and HIV-1 transgenic (TG) rat sera, alone or in combination with cigarette smoke extract (CSE) and analyzed for trans-endothelial electrical resistance (TEER) and paracellular permeability to 10 kDa fluorescein isothiocyanate (FITC)-dextran. Immunofluorescence staining was performed to assess the effects of treatment on the cellular localization and expression of the TJPs, “zonula occludens-1 (ZO-1) and claudin-5. Results: Pretreatment TEER measures were significantly higher for cultures treated with WT serum alone compared to those treated with TG serum or with CSE. Compared to pretreatment, TEER measures were significantly reduced by treatment with WT serum alone, CSE alone, WT serum + CSE, and TG serum + CSE. TG serum alone or TG serum + CSE resulted in statistically significant increased permeability compared to WT serum. All treatments decreased TJP staining intensity, and, in some cases, altered TJP localization. These effects were most prominent following incubation with either CSE alone, TG serum alone, or TG serum + CSE. Conclusions: CSE and TG serum induced separate and additive toxic effects on BBB function and integrity, which may underlie mechanisms that are associated with more severe HAND among HIV+ cigarette smokers.

OBJECTIVE:The blood-brain barrier (BBB) is a semi-permeable microvascular barrier, composed of endothelial cells conjoined by tight junction proteins. Following pathological conditions, i.e., traumatic brain injury (TBI), BBB dysfunction occurs, leading to microvascular hyperpermeability, resulting in cerebral edema formation and elevated intracranial pressure. Recent evidence suggests that the activation of pro-inflammatory signaling pathways is critical to BBB dysfunction. The NLRP3 inflammasome has been implicated as a key component of pro-inflammatory signaling. The aim of this study was to determine the upstream regulators of NLRP3 inflammasome activation that cause subsequent BBB aberration and microvascular hyperpermeability. METHODS:Brain microvascular endothelial cells were exposed to benzoyl ATP (BzATP) with or without MCC950. We employed immunocytochemical localization of tight junction proteins, fluorometric enzymatic assays, total gene expression analyses of ZO-1, and monolayer permeability studies to assess the effect of BzATP-induced injury on NLRP3 inflammasome activation/inhibition. RESULTS:BzATP treatment induced monolayer hyperpermeability and increased caspase-1 and MMP-9 activities. NLRP3 inhibition decreased caspase-1 and MMP-9 activities and rescued BzATP-induced monolayer permeability significantly. CONCLUSIONS:NLRP3 inflammasome signaling is critical to BBB endothelial cell dysfunction. Extracellular ATP is an upstream promoter of BBB hyperpermeability. NLRP3 inflammasome activation leads to subsequent caspase-1 and MMP-9-mediated tight junction protein disarray.

OBJECTIVES:The amino acid homocysteine (HCY) has been implicated in the pathobiology of several conditions, including spaceflight-associated neuro-ocular syndrome (SANS)-a collection of symptoms affecting near vision in astronauts. Blood-retinal barrier (BRB) and blood-brain barrier (BBB) dysfunctions are implicated in the pathobiology of SANS. Our objective was to assess how HCY affects BRB/BBB permeability and the role of the NLRP3 inflammasome in the modulation of such effects. METHODS:Human brain and retinal microvascular endothelial cells (HBMECs and HRMECs) were treated with 100 μM HCY alone or in conjunction with NLRP3 inflammasome inhibitor MCC950 at 1 μM. The assays performed included fluorometric assays to measure cell viability, an enzyme assay for caspase-1, expression of BRB/BBB tight junction protein zonula occludens-1 (ZO-1) by RT-PCR, and barrier permeability using FITC-dextran. RESULTS:In HRMECs and HBMECs, HCY-induced endothelial monolayer hyperpermeability significantly (p < 0.05). In HBMECs, the effect was attenuated by MCC950 (p < 0.05). Increased Caspase-1 activity was observed in both cell types following the addition of HCY. Following HCY addition, gene expression results denoting barrier damage were observed, particularly that of ZO-1 (p < 0.05). CONCLUSIONS:HCY induces hyperpermeability in retinal and brain endothelial cells. NLRP3-mediation in HCY-induced microvascular permeability is prominent in brain endothelial cells compared to retinal endothelial cells.

Cerebral edema is a consequential outcome of traumatic brain injury (TBI) and may lead to intracranial hypertension, necessitating urgent medical attention. One of the primary causes of cerebral edema is microvascular hyperpermeability, characterized by excessive leakage of intravascular fluid and proteins via blood–brain barrier (BBB) dysregulation. Prolonged activation of reactive oxygen species (ROS) formation and inflammatory pathways due to BBB hyperpermeability results in poor patient outcomes. The primary goal of this study was to ascertain if quercetin, a bioflavonoid plant pigment, would protect against BBB breakdown and hyperpermeability in the acute context following TBI. We used a mixed in vitro and in vivo model to test the effects of quercetin pretreatment on endothelial cell tight junctions in murine models of TBI and stress-induced hyperpermeability. Hydrogen peroxide (H2O2), a key contributor of secondary injuries following TBI, was used as an inducer of oxidative stress in cerebral endothelial cells in vitro. BBB tight junction/cytoskeletal integrity was assessed using immunofluorescence of junctional proteins zonula occludens-1, β-catenin, and vascular endothelial–cadherin, alongside filamentous actin labeling and a monolayer permeability assay. Intracellular ROS and H2O2 levels were determined using fluorescent probes. In vivo experiments consisted of intravital microscopy of brain pial vasculature in a mouse model of TBI. The results demonstrate that quercetin (100 μM; 1 h) attenuated H2O2 (100 μM; 2 h)–induced monolayer hyperpermeability and ROS formation significantly and decreased the loss of tight junction and cytoskeletal integrity. Quercetin treatment (50 mg/kg) after injury decreased TBI-induced vascular hyperpermeability significantly compared to sham. These results indicate that quercetin provides BBB protection by decreasing oxidative stress–induced loss of tight junction/cytoskeletal integrity, ultimately resulting in decreased microvascular hyperpermeability. The data suggest that quercetin may be a viable therapeutic option for preventing or managing cerebral oedema acutely following TBI.

Background: Dopaminergic neurotransmission is critical to managing a variety of physiological activities, including sexual behavior. Erectile dysfunction is frequently related to low dopamine levels and hyperglycemia, both of which can be alleviated by Mucuna pruriens, a natural source of levodopa and other bioactive compounds. The existing hypothesis depicts that decreased dopaminergic neurotransmission and hyperglycemia lead to erectile dysfunction. . . Objectives: The primary objective of this study was to establish the hypoglycemic action and sexual stimulatory effects of Mucuna pruriens in rodents. The secondary objective was to evaluate the effect on general rodent behavior, which can validate the safety profile of Mucuna pruriens for clinical use. Methods: Standardized Mucuna pruriens extract was given orally to male rodents for a predetermined amount of time. Blood samples were withdrawn from the male rats to measure the glucose levels at predetermined intervals. During mating trials, sexual activity metrics such as mount frequency and delay, intromission frequency, and ejaculation frequency were recorded. Standard open-field and exploratory behavior tests were used to track general behavior and evaluate any unfavorable or unusual changes. Results: Mucuna pruriens significantly decreased blood glucose levels and increased male sexual activity and behaviors in rodents (mount frequency and latency, intromission frequency, and ejaculation frequency). Conclusion: Thus, Mucuna pruriens can be the alternative natural bioactive to prevent and treat sexual dysfunction.

Traumatic brain injury (TBI) is one of the leading causes of morbidity and mortality among young adults and the elderly. In the United States, TBI is responsible for around 30 percent of all injuries brought on by injuries in general. Vasogenic cerebral edema due to blood-brain barrier (BBB) dysfunction and the associated elevation of intracranial pressure (ICP) are some of the major causes of secondary injuries following traumatic brain injury. Matrix metalloproteinase-9 (MMP-9) is a therapeutic target for being an enzyme that degrades the proteins that make up a part of the microvascular basal lamina as well as inter-endothelial tight junctions of the blood-brain barrier. MMP-9-mediated BBB dysfunctions and the compromise of the BBB is a major pathway that leads the development of vasogenic cerebral edema, elevation of ICP, poor cerebral perfusion and brain herniation following traumatic brain injury. That makes MMP-9 an effective therapeutic target and endogenous or exogenous MMP-9 inhibitors as therapeutic drugs for preventing secondary brain damage after traumatic brain injury. Although our understanding of the mechanisms that underlie the primary and secondary stages of damage following a TBI has significantly improved in recent years, such information has not yet resulted in the successful development of novel pharmacological treatment options for traumatic brain injury. Recent pre-clinical and/or clinical studies have demonstrated that there are several compounds with specific or non-specific MMP-9 inhibitory properties either directly binding and inhibiting MMP-9 or by indirectly inhibiting MMP-9, with potential as therapeutic agents for traumatic brain injury. This article reviews the efficacy of several such medications and potential agents that include endogenous and exogeneous compounds that are at various levels of research and development. MMP-9-based therapeutic drug development has enormous potential in the pharmacological treatment of cerebral edema and/or neuronal injury resulting from traumatic brain injury.

Blood-brain barrier (BBB) dysfunction and hyperpermeability have been implicated in a myriad of brain pathologies. The Evans Blue assay is one of the most popular methods for studying BBB integrity and permeability in rodent models of brain disorders. Under normal physiological conditions, the BBB is impermeable to albumin, so Evans Blue when injected intravenously binds to serum albumin and remains restricted within blood vessels. In traumatic and ischemic injuries, and other brain pathologies that result in BBB hyperpermeability, neighboring endothelial cells partially lose their close contacts to each other, and the BBB becomes permeable to proteins such as albumin. This paracellular leak of Evans blue-bound albumin is considered a reliable indicator of BBB dysfunction and hyperpermeability. Here, we describe the procedures for the evaluation of BBB integrity and hyperpermeability using Evans Blue extravasation assay in a mouse model of traumatic brain injury. The method described here focuses on intravenous injection of Evans Blue followed by Evans Blue dye extraction. This is followed by the measurement of fluorescence intensity of Evans Blue to determine the dye extravasation as a direct indicator of BBB hyperpermeability.

Blood-brain barrier (BBB) dysfunction and hyperpermeability that occurs following traumatic and ischemic insults lead to various downstream ill effects such as cerebral edema and elevation of intracranial pressure. The inter-endothelial tight junctions that consist of tight junction proteins are critical regulators of BBB dysfunctions and hyperpermeability. The major tight junction-associated proteins of the BBB are occludin, claudins, and junctional adhesion molecules that are intracellularly linked to the adaptor protein zonula occludens-1 (ZO-1). Quantitative measurement of tight junction-associated proteins provides valuable insight into barrier integrity and mechanisms that regulate microvascular hyperpermeability. Western blot analysis is a commonly used method to separate and identify proteins in a mixture using gel electrophoresis. Understanding the changes in the expression of one or more of these proteins is critical to evaluating barrier integrity and permeability in health and disease. Furthermore, studying them will provide insight into the associated downstream signaling pathways and evaluation of therapeutic approaches for regulating BBB permeability. Herein, we have described the protocol for immunoblot analysis of ZO-1 as an indicator of tight junction integrity in brain microvascular endothelial cells.

The blood vessels that vascularize the central nervous system (CNS) exhibit unique properties, termed the blood-brain barrier (BBB). The BBB allows these blood vessels to tightly regulate the movement of ions, molecules, and cells between the blood and the brain. The BBB is held together by tight junctions of the neighboring endothelial cells of the barrier, more specifically by tight junction proteins (TJPs) which can take the form of either integral transmembrane proteins or accessory cytoplasmic membrane proteins. BBB permeability can furthermore be affected by various factors, including but not limited to TJP expression, size, shape, charge, and type of extravascular molecules, as well as the nature of the vascular beds. The BBB is essential for the proper maintenance of CNS function, and its structural integrity has been implicated in several disorders and conditions. For instance, it has been shown that in the cases of traumatic brain injury (TBI), TBI-associated edema, and increased intracranial pressure are primarily caused by cases of hyperpermeability seen because of BBB dysfunction. Intravital microscopy is one of the most reliable methods for measuring BBB hyperpermeability in rodent models of BBB dysfunction in vivo. Here, we describe the surgical and imaging methods to determine the changes in BBB permeability at the level of the pial microvasculature in a mouse model of TBI using intravital microscopy.

The blood-brain barrier (BBB) is a highly complex and dynamic microvascular barrier that protects the brain parenchyma from the entry of pathogens, toxins, and other macromolecules and is a critical structure that helps to maintain brain homeostasis. The BBB is formed mainly by brain capillary endothelial cells and perivascular astrocytes and pericytes. One of the primary properties of the BBB is a tight regulation of paracellular permeability due to the presence of tight junctional complexes (also, adherens and gap junctions) between the neighboring microvascular endothelial cells. Alterations in the assembly of the tight junctions impair BBB properties, particularly influenced barrier integrity and permeability. The tight junctions of the BBB are mainly composed of proteins including claudins, occludin, and zonula occludens-1 (ZO-1). Zonula occludens-1 binds to the actin cytoskeleton, and its localization provides valuable information on the status of BBB integrity and permeability. Immunofluorescence localization of ZO-1 and/or other tight junction proteins is a reliable indicator of barrier integrity and permeability in microvascular endothelial cells. In microvascular endothelial cells, f-actin stress fiber formation significantly influences the rate and size of the inter-endothelial cell gap that form as cells retract from their borders. Rhodamine phalloidin is a popular conjugate used as a fluorescent label for f-actin. Herein, we describe the procedures for ZO-1 immunofluorescence and f-actin labeling followed by confocal microscopic imaging to determine barrier integrity and tight junction organization in brain microvascular endothelial cells in vitro.

The integrity of the blood-brain barrier (BBB), the protective barrier of the brain, is key to maintaining normal microvascular permeability and brain homeostasis. Brain microvascular endothelial cells are primary components of the blood-brain barrier. Transendothelial electrical resistance (TEER) is the electrical resistance across a cellular monolayer such as the brain microvascular endothelial cell monolayers. Measurement of TEER is considered a sensitive, reliable, and noninvasive method for evaluating barrier integrity and permeability of an endothelial cell monolayer under in vitro conditions. Measurement of TEER is also helpful for studying various cellular and molecular changes and signaling events that regulate barrier functions in endothelial monolayers. One of the in vitro endothelial cell barrier models that have been commonly used for measuring TEER is the BBB model using human or rodent brain microvascular endothelial cells grown as a monolayer. In this protocol, we describe how TEER is measured in brain microvascular endothelial cell monolayers, to determine blood-brain barrier integrity under in vitro conditions.

Traumatic brain injury, chronic traumatic encephalopathy, and Alzheimer's disease are three distinct neurological disorders that share common pathophysiological mechanisms involving neuroinflammation. One sequela of neuroinflammation includes the pathologic hyperphosphorylation of tau protein, an endogenous microtubule-associated protein that protects the integrity of neuronal cytoskeletons. Tau hyperphosphorylation results in protein misfolding and subsequent accumulation of tau tangles forming neurotoxic aggregates. These misfolded proteins are characteristic of traumatic brain injury, chronic traumatic encephalopathy, and Alzheimer's disease and can lead to downstream neuroinflammatory processes, including assembly and activation of the inflammasome complex. Inflammasomes refer to a family of multimeric protein units that, upon activation, release a cascade of signaling molecules resulting in caspase-induced cell death and inflammation mediated by the release of interleukin-1β cytokine. One specific inflammasome, the NOD-like receptor protein 3, has been proposed to be a key regulator of tau phosphorylation where it has been shown that prolonged NOD-like receptor protein 3 activation acts as a causal factor in pathological tau accumulation and spreading. This review begins by describing the epidemiology and pathophysiology of traumatic brain injury, chronic traumatic encephalopathy, and Alzheimer's disease. Next, we highlight neuroinflammation as an overriding theme and discuss the role of the NOD-like receptor protein 3 inflammasome in the formation of tau deposits and how such tauopathic entities spread throughout the brain. We then propose a novel framework linking traumatic brain injury, chronic traumatic encephalopathy, and Alzheimer's disease as inflammasomedependent pathologies that exist along a temporal continuum. Finally, we discuss potential therapeutic targets that may intercept this pathway and ultimately minimize long-term neurological decline.

Prolonged exposure to microgravity during long- and short-duration spaceflight has been found to produce many pathophysiological changes. Of these, a set constellation of changes called “Spaceflight Associated Neuro-ocular Syndrome” (SANS) induces neuro-ophthalmic changes. Recent evidence suggests astronauts who experienced SANS generally had higher concentrations of one-carbon pathway intermediate homocysteine (Hcy) prior to spaceflight. Hcy is a known inducer of inflammatory mediators and vascular endothelial dysfunction, leading to blood-brain barrier (BBB) breakdown. Mechanisms behind this dysfunction/hyperpermeability are not clearly known, but recent studies show evidence for loss of BBB integrity following microgravity/simulated microgravity. Recent studies from our lab suggest that activation of the nucleotide-binding domain leucine-rich repeat and pyrin domain containing receptor 3 (NLRP3) inflammasome plays a role in mediating BBB dysfunction and related microvascular hyperpermeability. Our main objective was to understand the mechanism(s) by which Hcy could induce BBB dysfunction/hyperpermeability, and potential association with SANS. We hypothesized that Hcy-mediated dysfunction/BBB hyperpermeability occurs via NLRP3 inflammasome activation and subsequent proteolytic breakdown of tight junction proteins. Understanding this pathway is critical to devise protection against SANS at a microvascular level. To study this relationship, human brain microvascular endothelial cells (HBMECs) were treated with various concentrations of Hcy. The monolayer permeability was evaluated using Transwell permeability assays with fluorescence tracer FITC-dextran (10-kDa). Matrixmetalloproteinase (MMP-9) and calpain activities were measured fluorometrically. Hcy induced endothelial monolayer hyperpermeability dose-dependently (p<0.05). NLRP3 inhibitor MCC950 decreased Hcy-induced hyperpermeability (p<0.05). Hcy treatment showed no significant effect on cell viability at lower concentration but decreased cell viability at higher concentrations. MCC950 inhibited Hcy-induced MMP-9 and calpain 1 activity. These results suggest a possible mechanism of BBB dysfunction/hyperpermeability by Hcy via activation of the NLRP3 inflammasome pathway. The proteolytic enzymes MMP-9 and calpain 1 are potential downstream mediators of Hcy-induced BBB dysfunction/hyperpermeability. Seeing that the blood-retinal barrier (BRB) shares structural-functional similarity with the BBB, human retinal microvascular endothelial cells (HRMEC) were examined for comparisons of barrier permeability. The results suggest Hcy-induced hyperpermeability in HRMEC monolayers and decreased expression of tight junction protein zonula occludens-1. Expression and effects were attenuated by NLRP3 inhibition. Our results suggest that NLRP3 inflammasome signaling may play an important role in promoting Hcy-induced barrier dysfunction and hyperpermeability in the blood-brain and blood-retinal barriers. Morehouse School of Medicine, Grady Hospital Surgery Dept, NASA This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Traumatic brain injury (TBI) is a major cause of death and disability in the United States, exacting a debilitating physical, social, and financial strain. Therefore, it is crucial to examine the impact of TBI on medically underserved communities in the U.S. The purpose of the current study was to review the literature on TBI for evidence of racial/ethnic differences in the U.S. Results of the review showed significant racial/ethnic disparities in TBI outcome and several notable differences in other TBI variables. American Indian/Alaska Natives have the highest rate and number of TBI-related deaths compared with all other racial/ethnic groups; Blacks/African Americans are significantly more likely to incur a TBI from violence when compared with Non-Hispanic Whites; and minorities are significantly more likely to have worse functional outcome compared with Non-Hispanic Whites, particularly among measures of community integration. We were unable to identify any studies that looked directly at underlying racial/ethnic biological variations associated with different TBI outcomes. In the absence of studies on racial/ethnic differences in TBI pathobiology, taking an indirect approach, we looked for studies examining racial/ethnic differences in oxidative stress and inflammation outside the scope of TBI as they are known to heavily influence TBI pathobiology. The literature indicates that Blacks/African Americans have greater inflammation and oxidative stress compared with Non-Hispanic Whites. We propose that future studies investigate the possibility of racial/ethnic differences in inflammation and oxidative stress within the context of TBI to determine whether there is any relationship or impact on TBI outcome.