Jin Rilong, Shen Miaoda, Yu Liedao, Wang Xuanwei, Lin Xiangjin
Department of Orthopedics Surgery, The First Affiliated Hospital, College of Medical Zhejiang University, Hangzhou, 310003 China.
Mol Cells. 2017 Mar;40(3):222-229. doi: 10.14348/molcells.2017.2314. Epub 2017 Mar 27.
Adipose-derived stem cells (ADSCs) were previously considered to have an anti-inflammatory effect, and Interleukin-1β (IL-1β) was found to be a pro-inflammatory factor in chondrocytes, but the mechanism underlying ADSCs and IL-1β is unclear. In this study, we investigate whether P2X7 receptor (P2X7R) signalling, regulated by microRNA 373 (miR-373), was involved in the ADSCs and IL-1β mediated inflammation in osteoarthritis (OA). Chondrocytes were collected from 20 OA patients and 20 control participants, and ADSCs were collected from patients who had undergone abdominal surgery. The typical surface molecules of ASDCs were detected by flow cytometry. The level of nitric oxide (NO) was determined by Griess reagent. Concentrations of prostaglandin E2 (PGE2), interleukin 6 (IL-6), matrix metallopeptidase 3 (MMP-3) were detected by enzyme-linked immunosorbent assay (ELISA). The expressions of IL-6, MMP-3, miR-373 and P2X7R were determined by real-time polymerase chain reaction (PCR), and Western blot was used to detect the protein expression of P2X7R. The typical potential characters of ADSCs were verified. In chondrocytes or OA tissues, the miR-373 expression level was decreased, but the P2X7R expression was increased. IL-1β stimulation increased the level of inflammatory factors in OA chondrocytes, and ADSCs co-cultured with IL-1β-stimulated chondrocytes decreased the inflammation. OA chondrocytes transfected with the miR-373 inhibitor increased the inflammation level. The miR-373 mimic suppressed the inflammation by targeting P2X7R and regulated its expression, while its effect was reversed by overexpression of P2X7R. IL-1β induced inflammation in OA chondrocytes, while ADSCs seemed to inhibit the expression of P2X7R that was regulated by miR-373 and involved in the anti-inflammatory process in OA.
脂肪来源干细胞（ADSCs）此前被认为具有抗炎作用，白细胞介素-1β（IL-1β）被发现是软骨细胞中的一种促炎因子，但ADSCs与IL-1β之间的潜在机制尚不清楚。在本研究中，我们探究由微小RNA 373（miR-373）调控的P2X7受体（P2X7R）信号通路是否参与了ADSCs和IL-1β介导的骨关节炎（OA）炎症反应。从20例OA患者和20例对照参与者中收集软骨细胞，并从接受腹部手术的患者中收集ADSCs。通过流式细胞术检测ADSCs的典型表面分子。用格里斯试剂测定一氧化氮（NO）水平。通过酶联免疫吸附测定（ELISA）检测前列腺素E2（PGE2）、白细胞介素6（IL-6）、基质金属蛋白酶3（MMP-3）的浓度。通过实时聚合酶链反应（PCR）测定IL-6、MMP-3、miR-373和P2X7R的表达，并使用蛋白质印迹法检测P2X7R的蛋白表达。验证了ADSCs的典型潜能特性。在软骨细胞或OA组织中，miR-373表达水平降低，但P2X7R表达增加。IL-1β刺激增加了OA软骨细胞中炎症因子的水平，与IL-1β刺激的软骨细胞共培养的ADSCs减轻了炎症。用miR-373抑制剂转染的OA软骨细胞炎症水平升高。miR-373模拟物通过靶向P2X7R抑制炎症并调节其表达，而P2X7R的过表达逆转了其作用。IL-1β诱导OA软骨细胞发生炎症，而ADSCs似乎抑制了受miR-373调控并参与OA抗炎过程的P2X7R的表达。
