Astrocytes send out long processes that are terminated by endfeet at the vascular surface and regulate vascular functions as well as homeostasis at the vascular interface. To date, the astroglial mechanisms underlying these functions have been poorly addressed. Here we demonstrate that a subset of messenger RNAs is distributed in astrocyte endfeet. We identified, among this transcriptome, a pool of messenger RNAs bound to ribosomes, the endfeetome, that primarily encodes for secreted and membrane proteins. We detected nascent protein synthesis in astrocyte endfeet. Finally, we determined the presence of smooth and rough endoplasmic reticulum and the Golgi apparatus in astrocyte perivascular processes and endfeet, suggesting for local maturation of membrane and secreted proteins. These results demonstrate for the first time that protein synthesis occurs in astrocyte perivascular distal processes that may sustain their structural and functional polarization at the vascular interface.

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Brain mural cells form a heterogeneous family which significantly contributes to the maintenance of the blood-brain barrier and regulation of the cerebral blood flow. Current procedures to isolate them cannot specifically separate their distinct subtypes, in particular vascular smooth muscle cells (VSMCs) and mid-capillary pericytes (mcPCs), which differ among others by their expression of smooth muscle actin (SMA). We herein describe an innovative method allowing SMA+ VSMCs and SMA- mcPCs to be freshly isolated from the rat cerebral cortex. Using differential RNA-Seq analysis, we then reveal the specific gene expression profile of each subtype. Our results refine the current description of the role of VSMCs in parenchymal cortical arterioles at the molecular level and provide a unique platform to identify the molecular mechanisms underlying the specific functions of mcPCs in the brain vasculature.

Development of human resources for eye health (HReH) is a major focus of the Global Action Plan 2014 to 2019 to reduce the prevalence of avoidable visual impairment by 25% by the year 2019. The eye health workforce is thought to be much smaller in sub-Saharan Africa than in other regions of the world but data to support this for policy-making is scarce. We collected HReH and cataract surgeries data from 21 countries in sub-Sahara to estimate progress towards key suggested population-based VISION 2020 HReH indicators and cataract surgery rates (CSR) in 2011.

Streptococcus pneumoniae or pneumococcus (PN) is a major causative agent of bacterial meningitis with high mortality in young infants and elderly people worldwide. The mechanism underlying PN crossing of the blood brain barrier (BBB) and specifically, the role of non-endothelial cells of the neurovascular unit that control the BBB function, remains poorly understood. Here, we show that the astroglial connexin 43 (aCx43), a major gap junctional component expressed in astrocytes, plays a predominant role during PN meningitis. Following intravenous PN challenge, mice deficient for aCx43 developed milder symptoms and showed severely reduced bacterial counts in the brain. Immunofluorescence analysis of brain slices indicated that PN induces the aCx43-dependent destruction of the network of glial fibrillary acid protein (GFAP), an intermediate filament protein specifically expressed in astrocytes and up-regulated in response to brain injury. PN also induced nuclear shrinkage in astrocytes associated with the loss of BBB integrity, bacterial translocation across endothelial vessels and replication in the brain cortex. We found that aCx4-dependent astrocyte damages could be recapitulated using in vitro cultured cells upon challenge with wild-type PN but not with a ply mutant deficient for the pore-forming toxin pneumolysin (Ply). Consistently, we showed that purified Ply requires Cx43 to promote host cell plasma membrane permeabilization in a process involving the Cx43-dependent release of extracellular ATP and prolonged increase of cytosolic Ca2+ in host cells. These results point to a critical role for astrocytes during PN meningitis and suggest that the cytolytic activity of the major virulence factor Ply at concentrations relevant to bacterial infection requires co-opting of connexin plasma membrane channels.

Megalencephalic Leukoencephalopathy with subcortical Cysts (MLC) is a type of vacuolating leukodystrophy, which is mainly caused by mutations in MLC1 or GLIALCAM. The two MLC-causing genes encode for membrane proteins of yet unknown function that have been linked to the regulation of different chloride channels such as the ClC-2 and VRAC. To gain insight into the role of MLC proteins, we have determined the brain GlialCAM interacting proteome. The proteome includes different transporters and ion channels known to be involved in the regulation of brain homeostasis, proteins related to adhesion or signaling as several G protein-coupled receptors (GPCRs), including the orphan GPRC5B and the proposed prosaposin receptor GPR37L1. Focusing on these two GPCRs, we could validate that they interact directly with MLC proteins. The inactivation of Gpr37l1 in mice upregulated MLC proteins without altering their localization. Conversely, a reduction of GPRC5B levels in primary astrocytes downregulated MLC proteins, leading to an impaired activation of ClC-2 and VRAC. The interaction between the GPCRs and MLC1 was dynamically regulated upon changes in the osmolarity or potassium concentration. We propose that GlialCAM and MLC1 associate with different integral membrane proteins modulating their functions and acting as a recruitment site for various signaling components as the GPCRs identified here. We hypothesized that the GlialCAM/MLC1 complex is working as an adhesion molecule coupled to a tetraspanin-like molecule performing regulatory effects through direct binding or influencing signal transduction events.

Absence of the astrocyte-specific membrane protein MLC1 is responsible for megalencephalic leukoencephalopathy with subcortical cysts (MLC), a rare type of leukodystrophy characterized by early-onset macrocephaly and progressive white matter vacuolation that lead to ataxia, spasticity, and cognitive decline. During postnatal development (from P5 to P15 in the mouse), MLC1 forms a membrane complex with GlialCAM (another astrocytic transmembrane protein) at the junctions between perivascular astrocytic processes. Perivascular astrocytic processes along with blood vessels form the gliovascular unit. It was not previously known how MLC1 influences the physiology of the gliovascular unit. Here, using the Mlc1 knock-out mouse model of MLC, we demonstrated that MLC1 controls the postnatal development and organization of perivascular astrocytic processes, vascular smooth muscle cell contractility, neurovascular coupling, and intraparenchymal interstitial fluid clearance. Our data suggest that MLC is a developmental disorder of the gliovascular unit, and perivascular astrocytic processes and vascular smooth muscle cell maturation defects are primary events in the pathogenesis of MLC and therapeutic targets for this disease.

Astrocytes are the most abundant glial cells of the central nervous system and have recently been recognized as crucial in the regulation of brain immunity. In most neuropathological conditions, astrocytes are prone to a radical phenotypical change called reactivity, which plays a key role in astrocyte contribution to neuroinflammation. However, how astrocytes regulate brain immunity in healthy conditions is an understudied question. One of the astroglial molecule involved in these regulations might be Connexin 43 (Cx43), a gap junction protein highly enriched in astrocyte perivascular endfeet-terminated processes forming the glia limitans. Indeed, Cx43 deletion in astrocytes (Cx43KO) promotes a continuous immune recruitment and an autoimmune response against an astrocyte protein, without inducing any brain lesion. To investigate the molecular basis of this unique immune response, we characterized the polysomal transcriptome of hippocampal astrocytes deleted for Cx43. Our results demonstrate that, in the absence of Cx43, astrocytes adopt an atypical reactive status with no change in most canonical astrogliosis markers, but with an upregulation of molecules promoting immune recruitment, complement activation as well as anti-inflammatory processes. Intriguingly, while several of these upregulated transcriptional events suggested an activation of the γ-interferon pathway, no increase in this cytokine or activation of related signaling pathways were found in Cx43KO. Finally, deletion of astroglial Cx43 was associated with the upregulation of several angiogenic factors, consistent with an increase in microvascular density in Cx43KO brains. Collectively, these results strongly suggest that Cx43 controls immunoregulatory and angiogenic properties of astrocytes.

Primary Sjögren syndrome (pSS) is a chronic systemic autoimmune disease characterized by xerophthalmia, xerostomia, and potential peripheral or central neurological involvement. In pSS, the prevalence of cognitive disorders is generally sparse across literature and the impact of pain on cognitive profile is unclear. The aim of this study was to determine the relation between pain, cognitive complaint, and impairment in a very homogenous population of 10 pSS patients with painful small fiber neuropathy (PSFN) and spontaneous cognitive complaint. Neurological exam, neuropsychological assessment, clinical evaluation measuring pain level, fatigue, anxiety, depression, and cognitive complaint were performed. Our results showed that 100% of patients had cognitive dysfunction especially in executive domain (80%). The most sensitive test was the Wisconsin Card Sorting Test (WCST), abnormal in 70% of our population. Moreover, we found clear cut significant correlations between pain levels and 3 measures of WCST: the number of errors (R = -0.768, P = .0062), perseverations (R = 0.831, P = .0042), and categories (R = 0.705, P = .02). In the literature review, the impact of pain is underexplored and results could be discordant. In a homogeneous cohort of pSS patients with PSFN, a cognitive complaint seems to be a valid reflection of cognitive dysfunction marked by a specific executive profile found with the WCST. In this preliminary study, this profile is linked to the level of pain and highlights that an appropriate management of pain control and a cognitive readaptation in patients could improve the quality of life.