Abstract
ObjectiveTo investigate the expression of miR-195 and its target gene Bcl-2 in intervertebral disc degeneration (IVDD) and its effect on nucleus pulposus (NP) cell apoptosis.MethodsThe expressions of miR-195 and Bcl-2 in NP tissues of IVDD patients were quantified by qRT-PCR and western blotting, respectively. NP cells were divided into blank group, TNF-α group, TNF-α + miR-NC group, TNF-α + siBcl-2 group, and TNF-α + miR-195 inhibitors + siBcl-2 group. Cell proliferation was detected by MTT assay, cell apoptosis evaluated by flow cytometry, and mitochondrial membrane potential (MMP) tested by JC-1 staining. Moreover, the function of miR-195 on IVDD in vivo was investigated using a puncture-induced IVDD rat model.ResultsIVDD patients had significantly increased miR-195 expression and decreased Bcl-2 protein expression in NP tissues. The expression of miR-195 was negatively correlated with the expression of Bcl-2 in IVDD patients. Dual-luciferase reporter gene assay indicated that Bcl-2 was a target gene of miR-195. In comparison with blank group, TNF-α group showed decreased cell proliferation and MMP, increased cell apoptosis, upregulated expression of miR-195, Bax, and cleaved caspase 3, and downregulated Bcl-2 protein, while these changes were attenuated by miR-195 inhibitors. Additionally, siBcl-2 can reverse the protective effect of miR-195 inhibitors on TNF-α-induced NP cells. Besides, inhibition of miR-195 alleviated IVDD degeneration and NP cell apoptosis in the rat model.ConclusionMiR-195 was significantly upregulated in NP tissues of IVDD patients, and inhibition of miR-195 could protect human NP cells from TNF-α-induced apoptosis via upregulation of Bcl-2.
Highlights
Intervertebral disc degeneration (IVDD) is one of the major causes of low back pain, seriously endangering the public health and bringing huge economic burdens to the whole world [1, 2]
Inhibiting nucleus pulposus (NP) cell apoptosis may promote the synthesis of NP extracellular matrix (ECM) to slow down the IVDD progression, which is of great significance to improve the quality of life of IVDD patients [8]
The miR-195 and TNF-α expression in NP tissues were significantly higher from IVDD patients than those from the control participants
Summary
Intervertebral disc degeneration (IVDD) is one of the major causes of low back pain, seriously endangering the public health and bringing huge economic burdens to the whole world [1, 2]. MicroRNAs (miRNA) are a class of small single-strand non-coding RNA molecules about 18–25 nt, and play critical regulatory roles in cell differentiation, proliferation and survival [9]. MiRNA is shown to involve in the development and progression of IVDD, primarily via affecting cell apoptosis, inflammatory signal response, and ECM components [10, 11]. Xiaoming Cao et al found the increased miR-195 in osteoarthritis, which could affect the collagen synthesis in osteoarthritis progression via targeting PTHrP [13]. Relevant studies on the role of miR-195 in IVDD are still unclear. We hypothesized that miR195 may play its regulatory role in NP cell apoptosis of IVDD through the regulation of Bcl-2
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