Polyphyllin I suppressed the apoptosis of intervertebral disc nucleus pulposus cells Induced By IL-1 β by miR-503-5p/Bcl-2 axis

Abstract

AbstractBackground:no studies have shown the role and underlying mechanism of PPI-mediated anti- apoptosis activity in NP cells. We aimed to evaluate the effects of PPI in IL-1β-induced apoptosisin vitro.Methods:CCK-8 assay was used to detect the cell viability, cell apoptosis evaluated by double-stained flow cytometry (FITC Annexin V/PI), the expression of miRNA- 503-5p was quantified by qRT-PCR, the expression of Bcl-2, Bax and Cleaved caspase-3 were quantified by Western blot, Dual-luciferase reporter gene assay was used to detect the targeting relationship between miR-503-5p and Bcl-2.Results:PPI at 40 μg·mL-1markedly promoted the activity of NP cells (P＜0.01). Also, PPI reduced the level of apoptosis and enhanced activity induced by interleukin-1β( IL-1β) in NP cells (P＜0.001，0.01). PPI treatment significantly inhibited the expression of apoptosis-related protein Bax, Cleaved Caspase-3 (P＜0.05, 0.01), and enhanced the level of anti-apoptotic protein Bcl-2 (P＜0.01). The activity of NP cells was significantly decreased and the apoptosis rate of NP cells was increased under IL-1β treatment (P＜0.01， 0.001). Moreover, miR-503-5p was highly expressed in IL-1β-injured NP cells (P＜0.001). Furthermore, the effect of PPI on NP cell activity and apoptosis in IL-1β treatment was dramatically reversed by the overexpression of miR-503-5p (P＜0.01，0.01). The targeted binding of miR-503-5p to the 3'UTR of Bcl-2 mRNA was confirmed by dual luciferase reporter gene assays (P＜0.05). In further experiments, compared with miR-503-5p mimics, the effects of PPI on IL-1β-injured NP cell activity and apoptosis were greatly reversed by the co-overexpression of miR-503-5p and Bcl-2 (P＜0.05，0.05).Conclusion:PPI suppressed the apoptosis of intervertebral disc nucleus pulposus cells induced by IL-1β via miR-503-5p/Bcl-2 molecular axis.