Extracellular vesicles produced by bone marrow mesenchymal stem cells (BMSC-EVs) can play important roles in the repair of injured tissues. Though numerous studies have reported the effect of EVs on renal fibrosis, the underlying mechanisms remain unclear. We hypothesized that BMSC-EVs containing milk fat globule–epidermal growth factor–factor 8 (MFG-E8) could attenuate renal fibrosis by inhibiting the RhoA/ROCK pathway. We investigated whether BMSC-EVs have anti-fibrotic effects in a rat model of renal fibrosis, in which rats were subjected to unilateral ureteral obstruction (UUO), as well as in cultured HK2 cells. Extracellular vesicles from BMSCs were collected and co-cultured with HK2 cells during transforming growth factor-β1 (TGF-β1) treatment. HK2 cells co-cultured with TGF-β1 were also treated with the ROCK inhibitor, Y-27632. Compared with the Sham group, UUO rats displayed fibrotic abnormalities, accompanied by an increased expression of α-smooth muscle actin and Fibronectin and reduced expression of E-cadherin. These molecular and pathological changes suggested increased inflammation in damaged kidneys. Oxidative stress, as evidenced by an increased level of MDA and decreased levels of SOD1 and Catalase, was also observed in UUO kidneys. Additionally, activation of cleaved caspase-3 and PARP1 and increased apoptosis in the proximal tubules confirmed tubular cell apoptosis in the UUO group. All of these phenotypes exhibited by UUO rats were suppressed by treatment with BMSC-EVs. However, the protective effect of BMSC-EVs was completely abolished by the inhibition of MFG-E8. Consistent with the in vivo results, treatment with BMSC-EVs reduced inflammation, oxidative stress, apoptosis, and fibrosis in HK-2 cells stimulated with TGF-β1 in vitro. Interestingly, treatment with Y-27632 protected HK-2 cells against inflammation and fibrosis, although oxidative stress and apoptosis were unchanged. Our results show that BMSC-EVs containing MFG-E8 attenuate renal fibrosis in a rat model of renal fibrosis, partly through RhoA/ROCK pathway inhibition.

中文翻译：

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在UUO大鼠模型中，骨髓间充质干细胞产生的细胞外囊泡可部分抑制RhoA / ROCK通路，从而减轻肾纤维化。

骨髓间充质干细胞（BMSC-EV）产生的细胞外囊泡在受损组织的修复中起重要作用。尽管大量研究报告了电动汽车对肾纤维化的影响，但其潜在机制仍不清楚。我们假设含有乳脂球-表皮生长因子-8（MFG-E8）的BMSC-EV可以通过抑制RhoA / ROCK途径来减轻肾纤维化。我们研究了BMSC-EVs在大鼠肾纤维化模型中是否具有抗纤维化作用，在该模型中，大鼠遭受了单侧输尿管阻塞（UUO），并且在培养的HK2细胞中。收集来自骨髓间充质干细胞的细胞外囊泡，并在转化生长因子-β1（TGF-β1）处理期间与HK2细胞共培养。与TGF-β1共培养的HK2细胞也用ROCK抑制剂Y-27632处理。与假手术组相比，UUO大鼠出现纤维化异常，伴有α-平滑肌肌动蛋白和纤连蛋白的表达增加，而E-钙粘蛋白的表达减少。这些分子和病理学改变提示受损肾脏的炎症增加。在UUO肾脏中也观察到氧化应激，如MDA水平升高和SOD1和过氧化氢酶水平降低所证明。此外，裂解的caspase-3和PARP1的激活以及近端小管中凋亡的增加证实了UUO组中肾小管细胞的凋亡。用BMSC-EVs抑制UUO大鼠表现出的所有这些表型。然而，通过抑制MFG-E8，完全消除了BMSC-EV的保护作用。与体内结果一致，BMSC-EV的治疗可减轻炎症，TGF-β1体外刺激HK-2细胞的氧化应激，凋亡和纤维化。有趣的是，尽管氧化应激和细胞凋亡未改变，但用Y-27632处理可保护HK-2细胞免于炎症和纤维化。我们的研究结果表明，含有MFG-E8的BMSC-EV可以部分抑制RhoA / ROCK通路，从而减轻大鼠肾纤维化模型中的肾纤维化。