Fibroblasts repair blood-brain barrier damage and hemorrhagic brain injury via TIMP2.
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ABSTRACT:
SUBMITTER: Xu L
PROVIDER: S-EPMC9769996 | biostudies-literature | 2022 Nov
REPOSITORIES: biostudies-literature
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Fibroblasts repair blood-brain barrier damage and hemorrhagic brain injury via TIMP2.
Xu Lingling L Nirwane Abhijit A Xu Ting T Kang Minkyung M Devasani Karan K Yao Yao Y
Cell reports 20221101 8
The function of fibroblasts in intracerebral hemorrhage (ICH) remains elusive. By targeting Col1α1, a fibroblast-specific marker, we generate mice with ablated Col1α1<sup>+</sup> fibroblasts. These mutants show exacerbated blood-brain barrier (BBB) damage, enlarged injury volume, and worse neurological function, highlighting a beneficial role of Col1α1<sup>+</sup> fibroblasts in ICH. Echoing these findings, fibroblasts significantly decrease endothelial permeability in an in vitro ICH model. Nex ...[more]
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Project description:Background: Intracerebral hemorrhage (ICH) is a fatal subtype of stroke that lacks effective therapy. Blood-brain barrier (BBB) damage is a hallmark of ICH-induced brain injury that leads to edema formation, leukocytes infiltration, influx of blood components into the perihematomal (PHE) region, and eventually brain injury. Astrocytes are essential for the formation and maintenance of the BBB by providing secreted molecules that contribute to the association between these cells. Sonic hedgehog (SHH) derived from astrocytes promotes the maturity and integrity of the BBB by upregulating tight junctions (TJs) in brain capillary endothelial cells (ECs). However, the effect of SHH on BBB in ICH has not been investigated. Methods: Cyclopamine (CYC) is a potent, selective inhibitor that specifically blocks the SHH signaling pathway. Here, we used pharmacological inhibitions (CYC and its derivatives) to determine a critical role of the SHH signaling pathway in promoting BBB integrity after ICH by mechanisms of regulating the TJ proteins in vivo and in vitro. Results: The expression of astrocytic SHH was upregulated in mouse brains after ICH. Compared with the vehicle-treated group, inhibition of the SHH signaling pathway with CYC and its derivatives treatments aggravated neurological function deficits, brain edema, hematoma volume, and BBB impairment by downregulating TJs in ECs through the SHH-Gli-1 axis in vivo and in vitro. Conclusions: SHH signaling pathway at the level of the BBB provides a barrier-promoting effect, suggesting that the SHH signaling pathway may function as a potential therapeutic target for restoring BBB function in ICH.
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Project description:Blood-brain barrier (BBB) disruption is a serious pathological consequence of traumatic brain injury (TBI), for which there are limited therapeutic strategies. Tissue inhibitor of metalloproteinase-2 (TIMP2), a molecule with dual functions of inhibiting MMP activity and displaying cytokine-like activity through receptor binding, has been reported to inhibit VEGF-induced vascular hyperpermeability. Here, we investigate the ability of TIMP2 to ameliorate BBB disruption in TBI and the underlying molecular mechanisms. Both TIMP2 and AlaTIMP2, a TIMP2 mutant without MMP-inhibiting activity, attenuated neurological deficits and BBB leakage in TBI mice; they also inhibited junctional protein degradation and translocation to reduce paracellular permeability in human brain microvascular endothelial cells (ECs) exposed to hypoxic plus inflammatory insult. Mechanistic studies revealed that TIMP2 interacted with α3β1 integrin on ECs, inhibiting Src activation-dependent VE-cadherin phosphorylation, VE-cadherin/catenin complex destabilization, and subsequent VE-cadherin internalization. Notably, localization of VE-cadherin on the membrane was critical for TIMP2-mediated EC barrier integrity. Furthermore, TIMP2-mediated increased membrane localization of VE-cadherin enhanced the level of active Rac1, thereby inhibiting stress fiber formation. All together, our studies have identified an MMP-independent mechanism by which TIMP2 regulates EC barrier integrity after TBI. TIMP2 may be a therapeutic agent for TBI and other neurological disorders involving BBB breakdown.
| S-EPMC10849766 | biostudies-literature
Project description:BackgroundTraumatic Brain Injury (TBI) is a major cause of disability and mortality, to which there is currently no comprehensive treatment. Blood Brain Barrier (BBB) dysfunction is well documented in human TBI patients, yet the molecular mechanisms that underlie this neurovascular unit (NVU) pathology remains unclear. The apolipoprotein-E (apoE) protein has been implicated in controlling BBB integrity in an isoform dependent manner, via suppression of Cyclophilin A (CypA)-Matrix metallopeptidase-9 (MMP-9) signaling cascades, however the contribution of this pathway in TBI-induced BBB permeability is not fully investigated.MethodsWe exposed C57Bl/6 mice to controlled cortical impact and assessed NVU and BBB permeability responses up to 21 days post-injury. We pharmacologically probed the role of the CypA-MMP-9 pathway in BBB permeability after TBI using Cyclosporin A (CsA, 20 mg/kg). Finally, as the apoE4 protein is known to be functionally deficient compared to the apoE3 protein, we used humanized APOE mice as a clinically relevant model to study the role of apoE on BBB injury and repair after TBI.ResultsIn C57Bl/6 mice there was an inverse relationship between soluble apoE and BBB permeability, such that damaged BBB stabilizes as apoE levels increase in the days following TBI. TBI mice displayed acute pericyte loss, increased MMP-9 production and activity, and reduced tight-junction expression. Treatment with the CypA antagonist CsA in C57Bl/6 mice attenuates MMP-9 responses and enhances BBB repair after injury, demonstrating that MMP-9 plays an important role in the timing of spontaneous BBB repair after TBI. We also show that apoe mRNA is present in both astrocytes and pericytes after TBI. We report that APOE3 and APOE4 mice have similar acute BBB responses to TBI, but APOE3 mice display faster spontaneous BBB repair than APOE4 mice. Isolated microvessel analysis reveals delayed pericyte repopulation, augmented and sustained MMP-9 expression at the NVU, and impaired stabilization of Zonula Occludens-1, Occludin and Claudin-5 expression at tight junctions in APOE4 mice after TBI compared to APOE3 mice.ConclusionsThese data confirm apoE as an important modulator of spontaneous BBB stabilization following TBI, and highlights the APOE4 allele as a risk factor for poor outcome after TBI.
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Project description:The level of microRNA-9-5p (miRNA-9-5p) in brain tissues is significantly changed after traumatic brain injury (TBI). However, the effect of miRNA-9-5p for brain function in TBI has not been elucidated. In this study, a controlled cortical impact model was used to induce TBI in Sprague-Dawley rats, and an oxygen glucose deprivation model was used to mimic the pathological state in vitro. Brain microvascular endothelial cells (BMECs) and astrocytes were extracted from immature Sprague-Dawley rats and cocultured to reconstruct blood-brain barrier (BBB) in vitro. The results show that the level of miRNA-9-5p was significantly increased in brain tissues after TBI, and up-regulation of miRNA9-5p contributed to the recovery of neurological function. Up-regulation of miRNA-9-5p with miRNA agomir may significantly alleviate apoptosis, neuroinflammation, and BBB damage in rats after TBI. Moreover, a dual luciferase reporter assay confirmed that miRNA-9-5p is a post-transcriptional modulator of Ptch-1. In in vitro experiments, the results confirmed that up-regulation of miRNA-9-5p with miRNA mimic alleviates cellular apoptosis, inflammatory response, and BBB damage mainly by inhibiting Ptch-1. In addition, we found that the activation of Hedgehog pathway was accompanied by inhibition of NF-κB/MMP-9 pathway in the BMECs treated with miRNA-9-5p mimic. Taken together, these results indicate that up-regulation of miRNA-9-5p alleviates BBB damage and neuroinflammatory responses by activating the Hedgehog pathway and inhibiting NF-κB/MMP-9 pathway, which promotes the recovery of neurological function after TBI.
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Project description:Traumatic brain injury (TBI) involves primary mechanical damage and delayed secondary damage caused by vascular dysfunction and neuroinflammation. Intracellular components released into the parenchyma and systemic circulation, termed danger-associated molecular patterns (DAMPs), are major drivers of vascular dysfunction and neuroinflammation. These DAMPs include cell-free RNAs (cfRNAs), which damage the blood-brain barrier (BBB), thereby promoting edema, procoagulatory processes, and infiltration of inflammatory cells. We tested the hypothesis that intraperitoneal injection of Ribonuclease-1 (RNase1, two doses of 20, 60, or 180 µg/kg) at 30 min and 12 h after controlled-cortical-impact (CCI) can reduce secondary lesion expansion compared to vehicle treatment 24 h and 120 h post-CCI. The lowest total dose (40 µg/kg) was most effective at reducing lesion volume (- 31% RNase 40 µg/kg vs. vehicle), brain water accumulation (- 5.5%), and loss of BBB integrity (- 21.6%) at 24 h post-CCI. RNase1 also reduced perilesional leukocyte recruitment (- 53.3%) and microglial activation (- 18.3%) at 120 h post-CCI, but there was no difference in lesion volume at this time and no functional benefit. Treatment with RNase1 in the early phase following TBI stabilizes the BBB and impedes leukocyte immigration, thereby suppressing neuroinflammation. RNase1-treatment may be a novel approach to delay brain injury to extend the window for treatment opportunities after TBI.
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Project description:Low-level blast exposure can result in neurological impairment for military personnel. Currently, there is a lack of experimental data using sex as a biological variable in neurovascular outcomes following blast exposure. To model mild blast traumatic brain injury (mbTBI), male and female rats were exposed to a single 11 psi static peak overpressure blast wave using the McMillan blast device and cohorts were then euthanized at 6 h, 24 h, 7 d and 14 d post-blast followed by isolation of the amygdala. After mbTBI, animals experience immediate bradycardia, although no changes in oxygen saturation levels or weight loss are observed. Male mbTBI animals displayed significantly higher levels of anxiety-like behavior (open field and elevated plus maze) compared to male sham groups; however, there was no anxiety phenotype in female mbTBI animals. Blast-induced neurovascular damage was explored by measuring expression of tight junction (TJ) proteins (zonula occludens-1 (ZO-1), occludin and claudin-5), glial fibrillary acidic protein (GFAP) and astrocyte end-feet coverage around the blood-brain barrier (BBB). Western blot analysis demonstrates that TJ protein levels were significantly decreased at 6 h and 24 h post-mbTBI in male rats, but not in female rats, compared to sham. Female animals have decreased GFAP at 6 h post-mbTBI while male animals display decreased GFAP expression at 24 h post-mbTBI. By 7 d post-mbTBI, there were no significant differences in TJ or GFAP levels between groups in either sex. At 24 h post-mbTBI, vascular integrity and astrocytic end-feet coverage around the BBB was significantly decreased in males following mbTBI. These results demonstrate that loss of GFAP expression may be due to astrocytic damage at the BBB. Our findings also demonstrate sex differences in acute vascular and behavioral outcomes after single mbTBI. Female animals display a lack of BBB pathology after mbTBI corresponding to improved acute neuropsychological outcomes as compared to male animals.
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Project description:Traumatic brain injury (TBI) could result in edema and cause an increase in intracranial pressure of the brain resulting in mortality and morbidity. Although there is hyperosmolarity therapy available for this pathophysiological event, it remains controversial. Recently, several groups have shown docosahexaenoic acid (DHA) to improve functional and histological outcomes following brain injury based on reduction of neuroinflammation and apoptosis. However, the effect of DHA on blood-brain barrier (BBB) dysfunction after brain injury has not been fully studied. Here, a controlled cortical impact rat model was used to test the effect of a single dose of DHA administered 30 min post injury. Modified neurological severity score (mNSS) and forelimb asymmetry were used to determine the functional outcomes. Neuroimaging and histology were used to characterize the edema and BBB dysfunction. The study showed that DHA-treated TBI rats had better mNSS and forelimb asymmetry score than vehicle-treated TBI rats. Temporal analysis of edema using MRI revealed a significant reduction in edema level with DHA treatment compared to vehicle in TBI rats. Histological analysis using immunoglobulin G (IgG) extravasation showed that there was less extravasation, which corresponded with a reduction in aquaporin 4 and astrocytic metalloprotease 9 expression, and greater endothelial occludin expression in the peri-contusional site of the TBI rat brain treated with DHA in comparison to vehicle treatment. In conclusion, the study shows that DHA can exert its functional improvement by prevention of the edema formation via prevention of BBB dysfunction after TBI.
| S-EPMC7503959 | biostudies-literature
Project description:Astrocytes undergo disease-specific transcriptomic changes upon brain injury. However, phenotypic changes of astrocytes and their functions remain unclear after hemorrhagic stroke. Here we reported hemorrhagic stroke induced a group of inflammatory reactive astrocytes with high expression of Gfap and Vimentin, as well as inflammation-related genes lipocalin-2 (Lcn2), Complement component 3 (C3), and Serpina3n. In addition, we demonstrated that depletion of microglia but not macrophages inhibited the expression of inflammation-related genes in inflammatory reactive astrocytes. RNA sequencing showed that blood-brain barrier (BBB) disruption-related gene matrix metalloproteinase-3 (MMP3) was highly upregulated in inflammatory reactive astrocytes. Pharmacological inhibition of MMP3 in astrocytes or specific deletion of astrocytic MMP3 reduced BBB disruption and improved neurological outcomes of hemorrhagic stroke mice. Our study demonstrated that hemorrhagic stroke induced a group of inflammatory reactive astrocytes that were actively involved in disrupting BBB through MMP3, highlighting a specific group of inflammatory reactive astrocytes as a critical driver for BBB disruption in neurological diseases.
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