MiR-370-3P inhibits proliferation of human articular chondrocytes by regulating JAK2/STAT5B signaling pathway

Abstract

Objective To investigate whether the non-coding single chain RNA miR-370-3P regulates the proliferation of human articular chondrocytes (HC-a) through the JAK2/STAT5B signaling pathway and to explore its potential mechanism. Methods The over- /under-expression of miR-370-3P models were constructed by transfecting HC-a cells, and then the cells were divided into 5 groups: miR-370-3P mimic group (over-expression group), miR-370-3P inhibitor group (under-expression group), mimic normal control group (mNC group), inhibitor normal control group (iNC group), and normal control group (NC group). The mRNA levels of miR-370-3P, JAK2 and STAT5B in each group were detected by qRT-PCR, and the protein levels of JAK2 and STAT5B were determined using Western blotting. In addition, MTT assay was adopted to measure the proliferation of HC-a cells in each group, and the binding of miR-370-3P with JAK2 and STAT5B was detected by dual luciferase assay. Results As compared with the NC group, the mRNA expression of miR-370-3P was upregulated in the mimic group and down-regulated in the inhibitor group (P < 0.05), indicating successful construction of miR-370-3P over-expression/under-expression models. By contrast, the mRNA expression of JAK2 and STAT5B was decreased in the mimic group and increased in the inhibitor group (P < 0.05), with no significant difference between the empty vector control groups (mNC and iNC groups) and the NC group. Western blotting results also confirmed the down-regulation of JAK2 and STAT5B in the mimic group and upregulation in the inhibitor group (P < 0.05). MTT assay showed that miR-370-3P overexpression reduced the survival rate of HC-a cells (P < 0.05), suggesting an inhibitory effect on HC-a cells proliferation. The results of dual luciferase assay displayed that after miR-370-3P was co-transfected with JAK2 (wild type) as well as STAT5B (wild type), the fluorescence intensity was significantly down-regulated compared with the control group (P < 0.05), indicating that miR-370-3P can directly bind to JAK2-3'UTR and STAT5B-3'UTR to function. Conclusion miR-370-3P can inhibit the proliferation of HC-a cells by direct regulation of JAK2/STAT5B signaling pathway.

Keywords

• mir-370-3p
• jak2/stat5b
• human articular chondrocytes
• cell proliferation