To understand RS1 gene interaction networks in the X-linked retinoschisis (XLRS) mouse retina (Rs1^−/y), we analyzed the transcriptome by RNA sequencing before and after in vivo expression of exogenous retinoschisin (RS1) gene delivered by AAV8. RS1 is a secreted cell adhesion protein that is critical for maintaining structural lamination and synaptic integrity of the neural retina. RS1 loss-of-function mutations cause XLRS disease in young boys and men, with splitting (“schisis”) of retinal layers and synaptic dysfunction that cause progressive vision loss with age. Analysis of differential gene expression profiles and pathway enrichment analysis of Rs1-KO (Rs1^−/y) retina identified cell surface receptor signaling and positive regulation of cell adhesion as potential RS1 gene interaction networks. Most importantly, it also showed massive dysregulation of immune response genes at early age, with characteristics of a microglia-driven proinflammatory state. Delivery of AAV8-RS1 primed the Rs1-KO retina toward structural and functional recovery. The disease transcriptome transitioned toward a recovery phase with upregulation of genes implicated in wound healing, anatomical structure (camera type eye) development, metabolic pathways, and collagen IV networks that provide mechanical stability to basement membrane. AAV8-RS1 expression also attenuated the microglia gene signatures to low levels toward immune quiescence. This study is among the first to identify RS1 gene interaction networks that underlie retinal structural and functional recovery after RS1 gene therapy. Significantly, it also shows that providing wild-type RS1 gene function caused the retina immune status to transition from a degenerative inflammatory phenotype toward immune quiescence, even though the transgene is not directly linked to microglia function. This study indicates that inhibition of microglial proinflammatory responses is an integral part of therapeutic rescue in XLRS gene therapy, and gene therapy might realize its full potential if delivered before microglia activation and photoreceptor cell death. Clinical Trials. gov Identifier NTC 02317887.

中文翻译：

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X-连锁视网膜劈裂小鼠 (Rs1−/y) 视网膜的遗传拯救诱导 AAV8-RS1 基因转移后视网膜小胶质细胞炎症状态的静止并识别视网膜恢复的基因网络

为了了解X 连锁视网膜劈裂 (XLRS) 小鼠视网膜 ( Rs1 ^{-/y ) 中的}RS1基因相互作用网络，我们通过 RNA 测序分析了AAV8 递送的外源性视网膜劈裂 ( RS1 ) 基因在体内表达前后的转录组。RS1 是一种分泌的细胞粘附蛋白，对于维持神经视网膜的结构分层和突触完整性至关重要。RS1功能丧失突变导致年轻男孩和男性出现 XLRS 疾病，视网膜层分裂（“分裂”）和突触功能障碍，导致随着年龄的增长进行性视力丧失。Rs1 -KO的差异基因表达谱分析和通路富集分析（Rs1 ^-/y ) 视网膜将细胞表面受体信号传导和细胞粘附的正调节识别为潜在的RS1基因相互作用网络。最重要的是，它还表现出早期免疫反应基因的大量失调，具有小胶质细胞驱动的促炎状态的特征。AAV8- RS1的递送使Rs1 -KO 视网膜朝着结构和功能恢复的方向发展。疾病转录组向恢复阶段过渡，上调与伤口愈合、解剖结构（相机型眼）发育、代谢途径和为基底膜提供机械稳定性的胶原蛋白 IV 网络有关的基因上调。AAV8- RS1表达也将小胶质细胞基因特征减弱到低水平，朝向免疫静止。这项研究是最早确定RS1基因治疗后视网膜结构和功能恢复基础的RS1基因相互作用网络的研究之一。重要的是，它还表明提供野生型RS1基因功能导致视网膜免疫状态从退行性炎症表型转变为免疫静止，即使转基因与小胶质细胞功能没有直接联系。这项研究表明，抑制小胶质细胞促炎反应是 XLRS 基因治疗中治疗拯救的一个组成部分，如果在小胶质细胞激活和感光细胞死亡之前进行基因治疗，则可能会发挥其全部潜力。临床试验。政府标识符 NTC 02317887。