MicroRNA‐145 as one negative regulator of astrogliosis

Astrogliosis occurs at the lesion site within days to weeks after spinal cord injury (SCI) and involves the proliferation and hypertrophy of astrocytes, leading to glia scar formation. Changes in gene expression by deregulated microRNAs (miRNAs) are involved in the process of central nervous system neurodegeneration. Here, we report that mir‐145, a miRNA enriched in rat spinal neurons and astrocytes, was downregulated at 1 week and 1 month after SCI. Our in vitro studies using astrocytes prepared from neonatal spinal cord tissues indicated that potent inflammagen lipopolysaccharide downregulated mir‐145 expression in astrocytes, suggesting that SCI‐triggered inflammatory signaling pathways could play the inhibitory role in astrocytic mir‐145 expression. To induce overexpression of mir‐145 in astrocytes at the spinal cord lesion site, we developed a lentivirus‐mediated pre‐miRNA delivery system using the promoter of glial fibrillary acidic protein (GFAP), an astrocyte‐specific intermediate filament. The results indicated that astrocyte‐specific overexpression of mir‐145 reduced astrocytic cell density at the lesion border of the injured spinal cord. In parallel, overexpression of mir‐145 reduced the size of astrocytes and the number of related cell processes, as well as cell proliferation and migration. Through a luciferase reporter system, we found that GFAP and c‐myc were the two potential targets of mir‐145 in astrocytes. Together, the findings demonstrate the novel role of mir‐145 in the regulation of astrocytic dynamics, and reveal that the downregulation of mir‐145 in astrocytes is a critical factor inducing astrogliosis after SCI. GLIA 2015;63:194–205     

GFAP null astrocytes are a favorable substrate for neuronal survival and neurite growth

During the development of the CNS, astrocytes play a key role as a substrate for neuronal migration and axonal growth. These neuron-astrocyte interactions could be regulated, in part, by the astrocytic cytoskeleton. Nestin, vimentin, and glial fibrillary acidic protein (GFAP) are the three identified proteins constitutive of intermediate filaments present in astrocytes. In the present study, we used mice deficient in GFAP to define the influence of the major protein of the astrocytic cytoskeleton on neuron survival and axonal growth in a model of neuron-astrocyte coculture. We observed that GFAP null astrocytes are a better substrate for neuronal survival and neurite outgrowth than wild-type astrocytes. This may be correlated with the relatively late occurrence of GFAP expression in astrocyte maturation when the early steps of neurogenesis are completed.     

Upregulation in astrocytic connexin 43 gap junction levels may exacerbate generalized seizures in mesial temporal lobe epilepsy

Following brain injury, and during the process of neurodegeneration, a reactive astrocytic proliferation occurs. This is accompanied by an increase in the synthesis of neuropeptides, cytokines, growth factors and glial fibrillary acidic protein (GFAP), a cell-specific marker for reactive astrocytes. Astrocytes are extensively coupled by gap junctions of the Cx43 connexin subtype. Several studies have shown that in severe trauma, coupling between astrocytes may add to the spread of the damaged area. In this study we ask whether the astrocytosis which is a feature of other neurodegenerative diseases also occurs in mesial temporal lobe epilepsy (MTLE) and whether it is accompanied by an increase in astrocytic communication through an upregulation of Cx43 gap junction channel proteins. In order to examine the astrocytic response and the expression pattern of Cx43 protein, double immunohistochemical labeling studies were undertaken using antibodies against GFAP and Cx43 applied to human hippocampal tissue resected from patients with MTLE, and to normal human control hippocampal tissue. Immunofluorescent labeling of astrocytes and Cx43 was examined using confocal laser scanning microscopy. The images obtained were quantitatively analysed and reconstructed using three-dimensional volume rendering. The results of this study have established that not only is astrocytosis greater in MTLE-affected tissues than previously suggested, but it is accompanied by a highly significant increase in astrocytic Cx43 protein levels. We hypothesize that this surprisingly large upregulation in Cx43 may exacerbate generalized seizures in the progression of MTLE.     

Protective role of melatonin in domoic acid-induced neuronal damage in the hippocampus of adult rats

Domoic acid (DA), a kainite-receptor agonist and potent inducer of neurotoxicity, has been administered intravenously in adult rats in the present study (0.75 mg/kg body weight) to demonstrate neuronal degeneration followed by glial activation and their involvement with inducible nitric oxide synthase (iNOS) in the hippocampus. An equal volume of normal saline was administered in control rats. The pineal hormone melatonin, which protects the neurons efficiently against excitotoxicity mediated by sensitive glutamate receptor, was administered intraperitoneally (10 mg/kg body weight), 20 min before, immediately after, and 1 h and 2 h after the DA administration, to demonstrate its role in therapeutic strategy. Histopathological analysis (Nissl staining) demonstrated extensive neuronal damage in the pyramidal neurons of CA1, CA3 subfields and hilus of the dentate gyrus (DG) in the hippocampus at 5 days after DA administration. Sparsely distributed glial fibrillary acidic protein (GFAP)-immunoreactive astrocytes were observed in the hippocampus at 4-24 h after DA administration and in the control rats. Astrogliosis was evidenced by increased GFAP immunoreactivity in the areas of severe neuronal degeneration at 5 days after DA administration. Along with this, microglial cells exhibited an intense immunoreaction with OX-42, indicating upregulation of complement type 3 receptors (CR3). Ultrastructural study revealed swollen or shrunken degenerating neurons in the CA1, CA3 subfields and hilus of the DG and hypertrophied astrocytes showing accumulation of intermediate filament bundles in the cytoplasm were observed after administration of DA. Although no significant change could be observed in the mRNA level of iNOS expression between the DA-treated rats and controls at 4-24 h and at 5-day time intervals, double immunofluorescense revealed co-expression of induced iNOS with GFAP immunoreactive astrocytes, but not in the microglial cells, and iNOS expression in the neurons of the hippocampal subfields at 5 days after DA administration. Expression of iNOS was not observed in the hippocampus of control rats. DA-induced neuronal death, glial activation, and iNOS protein expression were attenuated significantly by melatonin treatment and were comparable to the control groups. The results of the present study suggest that melatonin holds potential for the treatment of pathologies associated with DA-induced brain damage. It is speculated that astrogliosis and induction of iNOS protein expression in the neurons and astrocytes of the hippocampus may be in response to DA-induced neuronal degeneration.     

Experimental epidural hematoma causes cerebral infarction and activates neocortical glial and neuronal genesis in adult guinea pigs

Epidural hematoma (EDH) is a type of life‐threatening traumatic brain injury. Little is known about the extent to which EDH may cause neural damage and regenerative response in the cerebral cortex. Here we attempted to explore these issues by using guinea pigs as an experimental model. Unilateral EDH was induced by injection of 0.1 ml autologous blood into the extradural space, with experimental effects examined at 7, 14, 30, and 60 days postlesion. An infarct developed in the cortex deep to the EDH largely after 7 days postlesion, with neuronal death occurred from layers I to V in the central infarct region, as evidenced by loss of immunoreactivity (IR) for neuron‐specific nuclear antigen (NeuN). Glial fibrillary acidic protein (GFAP) IR appeared as a cellular band surrounding the infarct and extending into the periinfarct cortex along the pia. Doublecortin (DCX) IR emerged in these same areas, with labeled cells appearing as astrocytic and neuronal profiles. DCX/GFAP colocalization was found in these regions commonly at 7 and 14 days postlesion, whereas DCX/NeuN‐colabeled neurons were detectable at 30 and 60 days postlesion. Subpopulations of GFAP‐, DCX‐, or NeuN‐immunoreactive cells colocalized with the endogenous proliferative marker Ki‐67 or bromodeoxyuridine (BrdU) after pulse‐chase with this birth‐dating marker. The results suggest that experimental EDH can cause severe neuronal loss, induce significant glial activation, and promote a certain degree of local neuronal genesis in adult guinea pig neocortex. These findings point to potential therapeutic targets for improving neuronal recovery in clinical management of EDH. © 2012 Wiley Periodicals, Inc.     

Selective Oestrogen Receptor Agonists Rescued Hippocampus Parameters in Male Spontaneously Hypertensive Rats

Spontaneously hypertensive rats (SHR) show pronounced hippocampus alterations, including low brain‐derived neurotrophic factor (BDNF) expression, reduced neurogenesis, astrogliosis and increased aromatase expression. These changes are reverted by treatment with 17β‐oestradiol. To determine which oestradiol receptor (ER) type is involved in these neuroprotective effects, we used agonists of the ERα [propylpyrazole triol (PPT)] and the ERβ [diarylpropionitrite (DPN)] given over 2 weeks to 4‐month‐old male SHR. Wistar Kyoto normotensive rats served as controls. Using immunocytochemistry, we determined glial fibrillary protein (GFAP)+ astrocytes in the CA1, CA3 and hilus of the dentate gyrus of the hippocampus, aromatase immunostaining in the hilus, and doublecortin (DCX)+ neuronal progenitors in the inner granular zone of the dentate gyrus. Brain‐derived neurotrophic factor mRNA was also measured in the hippocampus by the quantitative polymerase chain reaction. In SHR, PPT had no effect on blood pressure, decreased astrogliosis, slightly increased BDNF mRNA, had no effect on the number of DCX+ progenitors, and increased aromatase staining. Treatment with DPN decreased blood pressure, decreased astrogliosis, increased BDNF mRNA and DCX+ progenitors, and did not modify aromatase staining. We hypothesise that, although both receptor types may participate in the previously reported beneficial effects of 17β‐oestradiol in SHR, receptor activation with DPN may preferentially facilitate BDNF mRNA expression and neurogenesis. The results of the present study may help in the design of ER‐based neuroprotection for the encephalopathy of hypertension.     

Differential regulation of ciliary neurotrophic factor and its receptor in the rat hippocampus following transient global ischemia

To investigate a potential role of ciliary neurotrophic factor (CNTF) in transient global ischemia, we have studied the postischemic regulatory changes in the expression of CNTF and its receptor, the ligand-binding alpha-subunit (CNTFRalpha). Immunoblot analysis demonstrated CNTF levels were slightly upregulated already during the first day after ischemia and then increased markedly by more than 10-fold until 2 weeks postischemia. Immunoreactivity for CNTF became detectable 1 day after ischemia and was localized in reactive astrocytes. The intensity of the immunolabeling was maximal in CA1 during the phase of neuronal cell death (days 3-7 postischemia) and in the deafferented inner molecular layer of the dentate gyrus. Upregulation of CNTF expression was less pronounced in CA3 and absent in the stratum lacunosum moleculare and the outer molecular layer of the dentate gyrus and thus did not simply correlate with astroliosis as represented by upregulation of glial fibrillary acidic protein (GFAP). As shown by in situ hybridization, expression of CNTFRalpha mRNA was restricted to neurons of the pyramidal cell and granule cell layers in control animals. Following ischemia, reactive astrocytes, identified by double labeling with antibodies to GFAP, transiently expressed CNTFRalpha mRNA with a maximum around postischemic day 3. This astrocytic response was most pronounced in CA1 and in the hilar part of CA3. These results show that CNTF and its receptor are differentially regulated in activated astrocytes of the postischemic hippocampus, indicating that they are involved in the regulation of astrocytic responses and the neuronal reorganizations occurring after an ischemic insult.     

Up-regulation of the extracellular matrix glycoprotein tenascin-R during axonal reorganization and astrogliosis in the adult rat hippocampus

Interactions between cells and extracellular matrix (ECM) molecules play a crucial role during brain development. The ECM glycoprotein tenascin-R (TN-R) has been implicated in the control of axon targeting, neural cell adhesion, migration and differentiation. Here, we have focused on the putative role of TN-R in chronic brain diseases involving increased neuronal excitability, as found in epilepsy. An episode of pilocarpine-induced status epilepticus (SE) led over a period of 3-30 days to neuron loss in the hippocampal hilus, CA3 and CA1 with reactive mossy fiber sprouting and astrogliosis in these regions. We found a focal up-regulation of granular TN-R immunoreactivity within the neuropil of segments of the CA3 pyramidal cell layer, the extent of this up-regulation paralleled the degree of pyramidal cell loss, mossy fiber sprouting and astrogliosis in these CA3 segments. In contrast, parvalbumin immunoreactivity and Wisteria floribundi agglutinin (WFA)-labeled perineuronal nets were reduced in CA3 segments with neuronal cell loss. The parallel development of increase in focal granular TN-R immunoreactivity, reactive mossy fiber sprouting and astrogliosis in CA3 implies a role for TN-R in axon targeting and synapse formation and/or in astrocytic targeting and interactions with the ECM during lesion-induced sprouting in the adult brain.     

Hippocampal GABA and glutamate transporter immunoreactivity in patients with temporal lobe epilepsy

OBJECTIVE: Sodium-coupled transporters remove extracellular neurotransmitters and alterations in their function could enhance or suppress synaptic transmission and seizures. This study determined hippocampal gamma-aminobutyric acid (GABA) and glutamate transporter immunoreactivity (IR) in temporal lobe epilepsy (TLE) patients. METHODS: Hippocampal sclerosis (HS) patients (n = 25) and non-HS cases (mass lesion and cryptogenic; n = 20) were compared with nonseizure autopsies (n = 8). Hippocampal sections were studied for neuron densities along with IR for glutamate decarboxylase (GAD; presynaptic GABA terminals), GABA transporter-1 (GAT-1; presynaptic GABA transporter), GAT-3 (astrocytic GABA transporter), excitatory amino acid transporter 3 (EAAT3; postsynaptic glutamate transporter), and EAAT2-1 (glial glutamate transporters). RESULTS: Compared with autopsies, non-HS cases with similar neuron counts showed: 1) increased GAD IR gray values (GV) in the fascia dentata outer molecular layer (OML), hilus, and stratum radiatum; 2) increased GAT-1 OML GVs; 3) increased astrocytic GAT-3 GVs in the hilus and Ammon's horn; and 4) no IR differences for EAAT3-1. HS patients with decreased neuron densities demonstrated: 1) increased OML and inner molecular layer GAD puncta; 2) decreased GAT-1 puncta relative to GAD in the stratum granulosum and pyramidale; 3) increased GAT-1 OML GVs; 4) decreased GAT-3 GVs; 5) increased EAAT3 IR on remaining granule cells and pyramids; 6) decreased glial EAAT2 GVs in the hilus and CA1 stratum radiatum associated with neuron loss; and 7) increased glial EAAT1 GVs in CA2/3 stratum radiatum. CONCLUSIONS: Hippocampal GABA and glutamate transporter IR differ in TLE patients compared with autopsies. These data support the hypothesis that excitatory and inhibitory neurotransmission and seizure susceptibility could be altered by neuronal and glial transporters in TLE patients.     

Altered astrocytic response to activation in SOD1G93A mice and its implications on amyotrophic lateral sclerosis pathogenesis

Amyotrophic Lateral Sclerosis (ALS) is a fatal, rapidly progressive, neurodegenerative disease caused by motor neuron degeneration. Despite extensive efforts, the underlying cause of ALS and the path of neurodegeneration remain elusive. Astrocyte activation occurs in response to central nervous system (CNS) insult and is considered a double edged sword in many pathological conditions. We propose that reduced glutamatergic and trophic response of astrocytes to activation may, over time, lead to accumulative CNS damage, thus facilitating neurodegeneration. We found that astrocytes derived from the SOD1^G93A ALS mouse model exhibit a reduced glutamatergic and trophic response to specific activations compared to their wild‐type counterparts. Wild‐type astrocytes exhibited a robust response when activated with lipopolysaccharide (LPS), G5 or treated with ceftriaxone in many parameters evaluated. These parameters include increased expression of GLT‐1 and GLAST the two major astrocytic glutamate transporters, accompanied by a marked increase in the astrocytic glutamate clearance and up‐regulation of neurtrophic factor expression. However, not only do un‐treated SOD1^G93A astrocytes take up glutamate less efficiently, but in response to activation they show no further increase in any of the glutamatergic parameters evaluated. Furthermore, activation of wild‐type astrocytes, but not SOD1^G93A astrocytes, improved their ability to protect the motor neuron cell line NSC‐34 from glutamate induced excitotoxicity. Our data indicates that altered astrocyte activation may well be pivotal to the pathogenesis of ALS. © 2012 Wiley Periodicals, Inc.     

Seasonal changes in astrocytes parallel neuronal plasticity in the song control area HVc of the canary

In the song control area HVc of the canary, intercellular dye‐coupling among astrocytes was studied by intracellular injection of neurobiotin into identified single astrocytes. Injection of individual astrocytes into acute slices resulted in dye spread to neighboring astrocytes, covering a sphere of up to 1 mm in diameter. The astrocytic nature of the dye‐coupled cells was verified by double labeling of neurobiotin‐filled cells with antisera for the astrocytic filament proteins GFAP or vimentin. The similarity in the number of dye‐coupled cells and the total number of astrocytes labeled by immunocytochemical markers indicate that dye‐coupling is specific for astrocytes and labels almost the entire local astrocytic population. Within the major nucleus for vocal control (HVc), approximately 25% more astroglial cells were present than in the surrounding forebrain tissue. There is no apparent hindrance of dye spread at the border of the HVc. The density of dye‐coupled astrocytes and the expression of cytoskeletal filament proteins differed markedly between the reproductive period in spring and the quiescent period in autumn. While vimentin is the major astroglial filament in autumn, GFAP is strongly expressed in spring. The density of dye‐coupled astrocytes reveals a marked increase in the reproductive period, followed by a reduction in autumn. The data indicate that the astrocytic population in the avian forebrain undergoes significant changes coincident with the known functional changes in the vocal control nuclei during periods of song production. GLIA 27:88–100, 1999. © 1999 Wiley‐Liss, Inc.     

Alteration of glial‐neuronal metabolic interactions in a mouse model of Alexander disease

Alexander disease is a rare and usually fatal neurological disorder characterized by the abundant presence of protein aggregates in astrocytes. Most cases result from dominant missense de novo mutations in the gene encoding glial fibrillary acidic protein (GFAP), but how these mutations lead to aggregate formation and compromise function is not known. A transgenic mouse line (Tg73.7) over‐expressing human GFAP produces astrocytic aggregates indistinguishable from those seen in the human disease, making them a model of this disorder. To investigate possible metabolic changes associated with Alexander disease Tg73.7 mice and controls were injected simultaneously with [1‐¹³C]glucose to analyze neuronal metabolism and [1,2‐¹³C]acetate to monitor astrocytic metabolism. Brain extracts were analyzed by ¹H magnetic resonance spectroscopy (MRS) to quantify amounts of several key metabolites, and by ¹³C MRS to analyze amino acid neurotransmitter metabolism. In the cerebral cortex, reduced utilization of [1,2‐¹³C]acetate was observed for synthesis of glutamine, glutamate, and GABA, and the concentration of the marker for neuronal mitochondrial metabolism, N‐acetylaspartate (NAA) was decreased. This indicates impaired astrocytic and neuronal metabolism and decreased transfer of glutamine from astrocytes to neurons compared with control mice. In the cerebellum, glutamine and GABA content and labeling from [1‐¹³C]glucose were increased. Evidence for brain edema was found in the increased amount of water and of the osmoregulators myo‐inositol and taurine. It can be concluded that astrocyte—neuronal interactions were altered differently in distinct regions. © 2010 Wiley‐Liss, Inc.     

Astrocytes modulate distribution and neuronal signaling of leptin in the hypothalamus of obese A vy mice

We tested the hypothesis that astrocytic activity modulates neuronal uptake and signaling of leptin in the adult-onset obese agouti viable yellow (A ^vy ) mouse. In the immunohistochemical study, A ^vy mice were pretreated with the astrocyte metabolic inhibitor fluorocitrate or phosphate-buffered saline (PBS) vehicle intracerebroventricularly (icv) followed 1 h later by Alexa568-leptin. Confocal microscopy showed that fluorocitrate pretreatment reduced astrocytic uptake of Alexa568-leptin 30 min after icv while increasing neuronal uptake in the arcuate nucleus and dorsomedial hypothalamus. Fluorocitrate also induced mild astrogliosis and moderately increased pSTAT3 immunopositive neurons in response to Alexa568-leptin in the dorsomedial hypothalamus. In the Western blotting study, A ^vy mice were pretreated with either PBS or fluorocitrate, and received PBS or leptin 1 h later followed by determination of pSTAT3 and GFAP expression an additional 30 min afterward. The results show that fluorocitrate induced a mild pSTAT3 activation but attenuated leptin-induced pSTAT3 activation and decreased GFAP expression independently of leptin treatment. We conclude that inhibition of astrocytic activity resulted in enhanced neuronal leptin uptake and signaling. This suggests opposite roles of astrocytes and neurons in leptin’s actions in the A ^vy mouse with adult-onset obesity.     

Generation of GFAP::GFP astrocyte reporter lines from human adult fibroblast‐derived iPS cells using zinc‐finger nuclease technology

Astrocytes are instrumental to major brain functions, including metabolic support, extracellular ion regulation, the shaping of excitatory signaling events and maintenance of synaptic glutamate homeostasis. Astrocyte dysfunction contributes to numerous developmental, psychiatric and neurodegenerative disorders. The generation of adult human fibroblast‐derived induced pluripotent stem cells (iPSCs) has provided novel opportunities to study mechanisms of astrocyte dysfunction in human‐derived cells. To overcome the difficulties of cell type heterogeneity during the differentiation process from iPSCs to astroglial cells (iPS astrocytes), we generated homogenous populations of iPS astrocytes using zinc‐finger nuclease (ZFN) technology. Enhanced green fluorescent protein (eGFP) driven by the astrocyte‐specific glial fibrillary acidic protein (GFAP) promoter was inserted into the safe harbor adeno‐associated virus integration site 1 (AAVS1) locus in disease and control‐derived iPSCs. Astrocyte populations were enriched using Fluorescence Activated Cell Sorting (FACS) and after enrichment more than 99% of iPS astrocytes expressed mature astrocyte markers including GFAP, S100β, NFIA and ALDH1L1. In addition, mature pure GFP‐iPS astrocytes exhibited a well‐described functional astrocytic activity in vitro characterized by neuron‐dependent regulation of glutamate transporters to regulate extracellular glutamate concentrations. Engraftment of GFP‐iPS astrocytes into rat spinal cord grey matter confirmed in vivo cell survival and continued astrocytic maturation. In conclusion, the generation of GFAP::GFP‐iPS astrocytes provides a powerful in vitro and in vivo tool for studying astrocyte biology and astrocyte‐driven disease pathogenesis and therapy. GLIA 2016;64:63–75     

Astrocytic modulation of potassium under seizures

The contribution of an impaired astrocytic K^+ regulation system to epileptic neuronal hyperexcitability has been increasingly recognized in the last decade.A defective K^+ regulation leads to an elevated extracellular K^+ concentration([K^+]o).When[K^+]o reaches peaks of 10-12 mM,it is strongly associated with seizure initiation during hypersynchronous neuronal activities.On the other hand,reactive astrocytes during a seizure attack restrict influx of K^+ across the membrane both passively and actively.In addition to decreased K^+ buffering,aberrant Ca^2+ signaling and declined glutamate transport have also been observed in astrogliosis in epileptic specimens,precipitating an increased neuronal discharge and induction of seizures.This review aims to provide an overview of experimental findings that implicated astrocytic modulation of extracellular K^+ in the mechanism of epileptogenesis.     

Reaction of astrocytes in the gerbil hippocampus following transient ischemia: immunohistochemical observations with antibodies against glial fibrillary acidic protein, glutamine synthetase, and S-100 protein.

An immunohistochemical method was used to study the distribution and changes with time of the astrocytic reaction in the gerbil hippocampus following transient ischemia. Three markers were investigated with specific antibodies to glial fibrillary acidic protein (GFAP), glutamine synthetase (GS), and S-100 protein. On Day 2 after ischemia, and more prominently on Day 3, reactive astrocytes were intensely stained for GFAP in the hippocampal formation, especially in the CA1 region and dentate gyrus. This response by astrocytes preceded CA1 pyramidal cell degeneration, which became apparent on Day 5. On Day 5, immunoreactive cells were not stained as intensely as on Day 3, but cells in the CA1 region and dentate gyrus were still more intensely stained than those in normal animals. GS and S-100 showed similar changes in distribution after ischemia, although the change in GS was less prominent: the hilus of the dentate gyrus was most intensely stained. Both immunoreactivities seemed to increase rather transiently on Day 2 or 3 and to decrease to the initial level on Day 5. The fact that reactive astrocytes appeared in CA1 before the onset of visible neural degeneration indicates that signals from indisposed neurons may be transmitted to astrocytes for their quick functioning. It is also suggested that degenerative changes occur in the dentate gyrus and may be involved in the delayed neural death of CA1 pyramidal cells. These observations indicate that astrocytes play a role in the neural degeneration induced by ischemia and that several types of astrocytes seem to react differently.     

Reactive astrocytes in multiple sclerosis impair neuronal outgrowth through TRPM7-mediated chondroitin sulfate proteoglycan production

Multiple sclerosis (MS) is a chronic inflammatory disorder of the central nervous system (CNS), characterized by inflammation-mediated demyelination, axonal injury and neurodegeneration. The mechanisms underlying impaired neuronal function are not fully understood, but evidence is accumulating that the presence of the gliotic scar produced by reactive astrocytes play a critical role in these detrimental processes. Here, we identified astrocytic Transient Receptor Potential cation channel, subfamily M, member 7 (TRPM7), a Ca²⁺-permeable nonselective cation channel, as a novel player in the formation of a gliotic scar. TRPM7 was found to be highly expressed in reactive astrocytes within well-characterized MS lesions and upregulated in primary astrocytes under chronic inflammatory conditions. TRPM7 overexpressing astrocytes impaired neuronal outgrowth in vitro by increasing the production of chondroitin sulfate proteoglycans, a key component of the gliotic scar. These findings indicate that astrocytic TRPM7 is a critical regulator of the formation of a gliotic scar and provide a novel mechanism by which reactive astrocytes affect neuronal outgrowth.     

体外培养大鼠星形胶质细胞受损后的激活与反应性胶质化

用鼠脑星形胶质细胞（ＡＳ）原代培养技术建立体外ＡＳ机械性损伤模型。以ｃ—Ｆｏｘｓ、ｂＦＧＦ、ＰＣＮＡ、ＧＦＡＰ—ｍＲＮＡ、ＧＦＡＰ和Ｍｙｏｓｉｎ作为观察指标研究反应性星形胶质化的形成机制。结果显示：１．ｃ—Ｆｏｓ蛋白于损伤后４５ｍｉｎ即有阳性表达，伤后２ｈ消失；２．损伤后２ｈ，损伤边缘的ＡＳ开始表达ｂＦＧＦ，１２ｈ达高峰，２ｄ后表达强度开始回落；３．损伤边缘的部分ＡＳ于损伤后２ｈ开始表达ＰＣＮＡ，伤后１ｄ，ＰＣＮＡ阳性的ＡＳ沿损伤边缘呈列兵式整齐排列，２ｄ后ＰＣＮＡ阳性的ＡＳ分布于损伤周围区域；４．损伤后４ｈ，损伤边缘的ＡＳ开始表达Ｍｙｏｓｉｎ，并逐渐增加，而且朝向损伤区胞浆突起的阳性表达强于背向损伤区的突起；５．损伤边缘的ＡＳ于损伤后６ｈ开始表达ＧＦＡＰ—ｍＲＮＡ，１ｄ达高峰，２ｄ开始回落，３ｄ则只在少数ＡＳ中可检出ＧＦＡＰ—ｍＲＮＡ；６．损伤后１ｄ，ＧＦＡＰ表达明显增强，胞体肥大并向损伤区伸出粗大突起，２ｄＧＦＡＰ达高峰，３ｄ肥大ＡＳ的胞体和突起覆盖损伤区；７．在体外ＡＳ机械性损伤模型上，在没有神经元和其它复杂因素影响的条件下，ＡＳ对损伤的主要反应是胞体的肥大、突起的粗大，并能独立形成反应性星形胶质化。