Department of Pediatrics, Renming Hospital, Wuhan University, Wuhan, Hubei 430060, P.R. China.
Mol Med Rep. 2017 Nov;16(5):6013-6019. doi: 10.3892/mmr.2017.7307. Epub 2017 Aug 22.
Systemic inflammatory response syndrome is a pathophysiological inflammatory response mediated largely by tumor necrosis factor‑α (TNF‑α), in response to infectious or non‑infectious stimuli. TNF‑α secretion in response to bacterial lipopolysaccharide (LPS) is regulated in part by disintegrin and metalloproteinase domain‑containing protein 17 (ADAM17). Therefore, the present study aimed to identify an effective inhibitor of ADAM17, in order to control inflammation and associated processes. In the present study, a lentiviral vector expressing short hairpin (sh)RNA targeting the ADAM17 gene was constructed. U937 cells were infected with the lentivirus and stimulated with LPS. ADAM17 expression was assessed by western blotting and TNF‑α secretion was assessed by ELISA analysis. The lentivirus was additionally tested in vivo in a mouse model of endotoxemia and sTNF‑α expression was assessed by flow cytometry in peritoneal macrophages. In vitro, the ADAM17 shRNA lentivirus reduced ADAM17 expression, and prevented TNF‑α maturation in U937 cells. In vivo, mice exposed to the ADAM17 shRNA lentivirus prior to LPS‑induced endotoxemia exhibited fewer signs of inflammation and less tissue damage compared with the control mice. In conclusion, the present study successfully constructed a shRNA lentiviral vector targeting the ADAM17 gene that exhibited apparent in vitro and in vivo effects on TNF‑α processing in response to an LPS challenge. The results of the present study may aid the design and improvement of drugs designed to inhibit the function of ADAM17, and suggested a novel means of controlling inflammation and associated processes.
全身炎症反应综合征是一种病理生理学炎症反应，主要由肿瘤坏死因子-α（TNF-α）介导，以响应感染或非感染性刺激。TNF-α对细菌脂多糖（LPS）的分泌部分受解整合素金属蛋白酶域蛋白 17（ADAM17）调节。因此，本研究旨在鉴定 ADAM17 的有效抑制剂，以控制炎症和相关过程。在本研究中，构建了表达靶向 ADAM17 基因的短发夹（sh）RNA 的慢病毒载体。U937 细胞用慢病毒感染并用 LPS 刺激。通过 Western blot 评估 ADAM17 表达，通过 ELISA 分析评估 TNF-α分泌。该慢病毒还在脂多糖诱导的内毒素血症的小鼠模型中进行了体内测试，并通过流式细胞术评估腹腔巨噬细胞中的可溶性 TNF-α表达。在体外，ADAM17 shRNA 慢病毒降低了 ADAM17 的表达，并阻止了 U937 细胞中 TNF-α的成熟。在体内，与对照小鼠相比，预先用 ADAM17 shRNA 慢病毒处理的 LPS 诱导的内毒素血症小鼠表现出较少的炎症迹象和较少的组织损伤。综上所述，本研究成功构建了靶向 ADAM17 基因的 shRNA 慢病毒载体，该载体在体外和体内对 LPS 刺激下 TNF-α处理均表现出明显的作用。本研究的结果可能有助于设计和改进旨在抑制 ADAM17 功能的药物，并提出了一种控制炎症和相关过程的新方法。
