Wang Yong, Yang Tao, Liu Yadong, Zhao Wei, Zhang Zhen, Lu Ming, Zhang Weiguo
Department of Joint Surgery, The First Affiliated Hospital of Dalian Medical University, Dalian 116011, China.
The 4th Department of Orthopedic Surgery, The Central Hospital Affiliated to Shenyang Medical College, Shenyang 110024, China.
Int J Mol Sci. 2017 May 4;18(5):975. doi: 10.3390/ijms18050975.
Slow growth and rapid loss of chondrogenic phenotypes are the major problems affecting chronic cartilage lesions. The role of microRNA-195 (miR-195) and its detailed working mechanism in the fore-mentioned process remains unknown. Fibroblastic growth factor 18 (FGF-18) plays a key role in cartilage homeostasis; whether miR-195 could regulate FGF-18 and its downstream signal pathway in chondrocyte proliferation and maintenance of chondrogenic phenotypes still remains unclear. The present research shows elevated miR-195 but depressed FGF-18 expressed in joint fluid specimens of 20 patients with chronic cartilage lesions and in CH1M and CH3M chondrocytes when compared with that in joint fluid specimens without cartilage lesions and in CH1W and CH2W chondrocytes, respectively. The following loss of function test revealed that downregulation of miR-195 by transfection of miR-195 inhibitors promoted chondrocyte proliferation and expression of a type II collagen α I chain (Col2a1)/aggrecan. Through the online informatics analysis we theoretically predicted that miR-195 could bind to a FGF-18 3' untranslated region (3'UTR), also, we verified that a miR-195 could regulate the FGF-18 and its downstream pathway. The constructed dual luciferase assay further confirmed that FGF-18 was a direct target of miR-195. The executed anti-sense experiment displayed that miR-195 could regulate chondrocyte proliferation and Col2a1/aggrecan expression via the FGF-18 pathway. Finally, through an in vivo anterior cruciate ligament transection (ACLT) model, downregulation of miR-195 presented a significantly protective effect on chronic cartilage lesions. Evaluating all of the outcomes of the current research revealed that a decrease of miR-195 protected chronic cartilage lesions by promoting chondrocyte proliferation and maintenance of chondrogenic phenotypes via the targeting of the FGF-18 pathway and that the miR-195/FGF-18 axis could be a potential target in the treatment of cartilage lesions.
软骨生成表型的缓慢生长和快速丧失是影响慢性软骨损伤的主要问题。微小RNA - 195（miR - 195）在上述过程中的作用及其详细作用机制尚不清楚。成纤维细胞生长因子18（FGF - 18）在软骨稳态中起关键作用；miR - 195是否能调节FGF - 18及其下游信号通路在软骨细胞增殖和软骨生成表型维持中的作用仍不清楚。本研究表明，与无软骨损伤的关节液标本以及CH1W和CH2W软骨细胞相比，20例慢性软骨损伤患者的关节液标本以及CH1M和CH3M软骨细胞中miR - 195表达升高而FGF - 18表达降低。接下来的功能丧失试验表明，通过转染miR - 195抑制剂下调miR - 195可促进软骨细胞增殖以及II型胶原α1链（Col2a1）/聚集蛋白聚糖的表达。通过在线信息学分析，我们从理论上预测miR - 195可与FGF - 18的3'非翻译区（3'UTR）结合，此外，我们验证了miR - 195可调节FGF - 18及其下游通路。构建的双荧光素酶测定进一步证实FGF - 18是miR - 195的直接靶点。进行的反义实验表明，miR - 195可通过FGF - 18途径调节软骨细胞增殖和Col2a1/聚集蛋白聚糖表达。最后，通过体内前交叉韧带横断（ACLT）模型，下调miR - 195对慢性软骨损伤具有显著的保护作用。评估当前研究的所有结果表明，miR - 195的降低通过靶向FGF - 18途径促进软骨细胞增殖和维持软骨生成表型来保护慢性软骨损伤，并且miR - 195/FGF - 18轴可能是治疗软骨损伤的潜在靶点。
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