Department of Emergency, Ningbo Medical Center Lihuili Hospital, Ningbo, China.
Eur Rev Med Pharmacol Sci. 2020 Jan;24(2):862-869. doi: 10.26355/eurrev_202001_20070.
To observe the effect of the micro ribonucleic acid (miR)-10a on sepsis-induced liver injury in rats through the transforming growth factor-b1 (TGF-b1)/Smad signaling pathway.
The rat model of sepsis was established via cecal ligation and puncture, in which miR-10a was overexpressed and silenced using liposome transfection. The rats were randomly divided into miR-10a mimics group (Mimics group, n=10) and miR-10a inhibitors group (Inhibitors group, n=10), and the sham operation group (Sham group, n=10) was also set up. The transfection efficiency of miR-10a in liver tissues in each group was detected via quantitative Real Time-Polymerase Chain Reaction (qRT-PCR), the serum liver function indexes aspartate aminotransferase (AST), alanine aminotransferase (ALT) and lactate dehydrogenase (LDH) were determined. Moreover, the content of the serum reactive oxygen species (ROS), glutathione (GSH), and GSH peroxidase (GSH-Px) was determined using enzyme-linked immunosorbent assay (ELISA). The content of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and myeloperoxidase (MPO) in liver tissues was detected, and the pathological changes in liver tissues were observed through hematoxylin-eosin (HE) staining. Finally, the expression levels of cytochrome P4502E1 (CYP2E1) and TGF-b1/Smad signaling pathway genes and proteins in liver tissues were detected via qRT-PCR and Western blotting.
The expression of miR-10a was significantly increased in Mimics group (p<0.05) and extremely low in the Inhibitors group (p<0.05). In Mimics group, the levels of serum AST, ALT, and LDH were significantly increased (p<0.05), the content of ROS, TNF-α, IL-6, and MPO was substantially increased (p<0.05), while that of GSH and GSH-Px notably declined (p<0.05). According to the HE staining results, the liver cells were orderly arranged in the Inhibitors group, and they were disorderly arranged with more inflammatory cells in the Mimics group. The results of the gene and protein assays showed that the expression levels of CYP2E1, TGF-b1, and Smad2 in Mimics group were markedly higher than those in the Sham group (p<0.05), while they displayed the opposite trends in the Inhibitors group (p<0.05).
Silencing miR-10a can inhibit the occurrence of sepsis-induced liver injury in rats by downregulating the TGF-β1/Smad pathway.
通过转化生长因子-β1（TGF-β1）/Smad 信号通路观察微小 RNA-10a（miR-10a）对脓毒症诱导的大鼠肝损伤的影响。
采用盲肠结扎穿孔法建立脓毒症大鼠模型，通过脂质体转染过表达和沉默 miR-10a。大鼠随机分为 miR-10a 模拟物组（Mimics 组，n=10）和 miR-10a 抑制剂组（Inhibitors 组，n=10），同时设置假手术组（Sham 组，n=10）。通过实时定量聚合酶链反应（qRT-PCR）检测各组肝组织中 miR-10a 的转染效率，测定血清肝功能指标天冬氨酸转氨酶（AST）、丙氨酸转氨酶（ALT）和乳酸脱氢酶（LDH）。此外，采用酶联免疫吸附试验（ELISA）测定血清中活性氧（ROS）、谷胱甘肽（GSH）和谷胱甘肽过氧化物酶（GSH-Px）的含量。检测肝组织中肿瘤坏死因子-α（TNF-α）、白细胞介素-6（IL-6）和髓过氧化物酶（MPO）的含量，通过苏木精-伊红（HE）染色观察肝组织的病理变化。最后，通过 qRT-PCR 和 Western blot 检测肝组织中细胞色素 P4502E1（CYP2E1）和 TGF-β1/Smad 信号通路基因和蛋白的表达水平。
Mimics 组 miR-10a 的表达明显升高（p<0.05），而 Inhibitors 组的表达极低（p<0.05）。在 Mimics 组，血清 AST、ALT 和 LDH 水平明显升高（p<0.05），ROS、TNF-α、IL-6 和 MPO 含量显著增加（p<0.05），而 GSH 和 GSH-Px 含量明显下降（p<0.05）。根据 HE 染色结果，Inhibitors 组的肝细胞排列整齐，而 Mimics 组的肝细胞排列紊乱，有较多的炎症细胞浸润。基因和蛋白检测结果显示，Mimics 组 CYP2E1、TGF-β1 和 Smad2 的表达水平明显高于 Sham 组（p<0.05），而 Inhibitors 组的表达趋势则相反（p<0.05）。
沉默 miR-10a 可通过下调 TGF-β1/Smad 通路抑制脓毒症诱导的大鼠肝损伤的发生。
