Pericyte abnormalities precede strial capillary basement membrane thickening in Alport mice
Pericyte abnormalities precede strial capillary basement membrane thickening in Alport mice
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DOI：
10.1016/j.heares.2020.107935
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发表时间：
2020-05-01
期刊：
影响因子：
2.8
通讯作者：
Cosgrove, Dominic
中科院分区：
文献类型：
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作者：
Dufek, Brianna;Meehan, Daniel T.;Cosgrove, Dominic
在129只Sv常染色体Alport小鼠中，毛细血管基底膜（SCBM）在5至9周龄之间逐渐老化，导致缺氧微环境，伴有代谢应激和促炎细胞因子和趋化因子的诱导。这些事件的发生伴随着耳蜗内电位的下降和在不会永久影响年龄/应变匹配的同窝仔的条件下对噪声诱导的听力损失的易感性。在这里，我们的目的是了解SCBM增厚发生前发生的事件。Alport纹具有正常的厚度，并且在SCBM中显示与野生型小鼠相当的细胞外基质（ECM）分子水平。3周Alport小鼠的听力阈值与野生型小鼠没有差异。我们使用来自从3周Alport小鼠和野生型同窝小鼠分离的血管纹的RNA进行RNAseq分析。使用Incidity Pathway Analysis软件处理数据，并使用手动程序进一步提取。RNAseq分析揭示了参与细胞粘附、细胞迁移、突起形成以及肌动蛋白和微管蛋白细胞骨架动力学的基因的显著失调。总的来说，这些数据表明纹的细胞结构的变化可能是明显的。为了验证这一观点，我们对这些动物的血管纹进行了双重免疫荧光分析，这些血管纹用抗同种凝集素gs-ib4（内皮细胞标记物）和抗结蛋白（周细胞标记物）抗体染色。结果显示，与野生型同窝小鼠相比，在Alport小鼠的z-堆叠共聚焦图像中，周细胞脱离和迁移以及周细胞上形成膜皱褶的证据。这通过TEM分析证实。我们实验室的早期工作表明，内皮素A受体阻断剂可以防止SCBM增厚和ECM在SCBM中的积累。用内皮素-1处理培养的周细胞诱导肌动蛋白细胞骨架重排，增加丝状肌动蛋白与球状肌动蛋白的比例。总的来说，这些发现表明Alport SCBM中IV型胶原组成的变化导致周细胞室的细胞损伤，激活分离和改变细胞骨架动力学。这些事件先于Alport小鼠的SCBM增厚和听力损失，因此构成了Alport神经病理学中迄今为止识别的最早事件。(C)2020爱思唯尔B.V.保留所有权利。
In 129 Sv autosomal Alport mice, the strial capillary basement membranes (SCBMs) progressively thicken between 5 and 9 weeks of age resulting in a hypoxic microenvironment with metabolic stress and induction of pro-inflammatory cytokines and chemokines. These events occur concomitant with a drop in endocochlear potential and a susceptibility to noise-induced hearing loss under conditions that do not permanently affect age/strain-matched littermates. Here we aimed to gain an understanding of events that occur before the onset of SCBM thickening. Alport stria has normal thickness and shows levels of extracellular matrix (ECM) molecules in the SCBMs commensurate with wild-type mice. Hearing thresholds in the 3-week Alport mice do not differ from those of wild-type mice. We performed RNAseq analysis using RNA from stria vascularis isolated from 3-week Alport mice and wild type littermates. Data was processed using Ingenuity Pathway Analysis software and further distilled using manual procedures. RNAseq analysis revealed significant dysregulation of genes involved in cell adhesion, cell migration, formation of protrusions, and both actin and tubulin cytoskeletal dynamics. Overall, the data suggested changes in the cellular architecture of the stria might be apparent. To test this notion, we performed dual immunofluorescence analysis on whole mounts of the stria vascularis from these same animals stained with anti-isolectin gs-ib4 (endothelial cell marker) and anti-desmin (pericyte marker) antibodies. The results showed evidence of pericyte detachment and migration as well as the formation of membrane ruffling on pericytes in z-stacked confocal images from Alport mice compared to wild type littermates. This was confirmed by TEM analysis. Earlier work from our lab showed that endothelin A receptor blockade prevents SCBM thickening and ECM accumulation in the SCBMs. Treating cultured pericytes with endothelin-1 induced actin cytoskeletal rearrangement, increasing the ratio of filamentous to globular actin. Collectively, these findings suggest that the change in type IV collagen composition in the Alport SCBMs results in cellular insult to the pericyte compartment, activating detachment and altered cytoskeletal dynamics. These events precede SCBM thickening and hearing loss in Alport mice, and thus constitute the earliest event so far recognized in Alport strial pathology. (C) 2020 Elsevier B.V. All rights reserved.