Department of Spinal Surgery, The Fifth Hospital of Harbin, Harbin City 150040, Heilongjiang Province, P.R. China.
Department of Orthopedic, The third affiliated hospital of Harbin Medical University, Harbin City 150040, Heilongjiang Province, P.R. China.
Biosci Rep. 2019 Sep 24;39(9). doi: 10.1042/BSR20191405. Print 2019 Sep 30.
MicroRNAs (miRNAs) are involved in the regulation of osteogenic differentiation and chondrification The purpose of the present study was to explore the potential mechanism of miR-128 in osteoporosis (OP).: Quantitative real-time PCR (qRT-PCR) was used to determine the expression of miR-128 in femoral neck trabecular bones of OP patients (=40) and non-OP patients (=40). C2C12 cells were transfected with miR-128 mimic or inhibitor to determine the effect of miR-128 on osteoblastic differentiation of C2C12 cells. Bioinformatics and luciferase reporter genes were used to determine the molecular mechanism of miR-128 in osteoblastic differentiation of C2C12 cells.: The qRT-PCR results showed that the expression level of miR-128 in bone samples of OP patients was significantly higher than that of non-OP patients, while miR-128 was significantly down-regulated during the osteogenic differentiation of C2C12 cells. In addition, the results showed that overexpression of miR-128 significantly inhibited the mRNA and protein expression levels of osteocalcin (OC), alkaline phosphatase (ALP) and collagen I type-α1 (COL1A1) in C2C12 cells, while miR-128 inhibitor could reverse this effect. Bioinformatics analysis and dual-luciferase reporter assay found that silencing information regulatory protein 6 (SIRT6) was a direct target of miR-128. The qRT-PCR and Western Blot results found that miR-128 significantly down-regulated the mRNA and protein expressions of SIRT6. Furthermore, silencing SIRT6 significantly inhibited the promoting effect of the miR-128 inhibitor on the expression of osteoblast markers.: The above results confirmed that miR-128 inhibited osteoblast differentiation in OP by down-regulating SIRT6 expression, thus accelerating the development of OP.
微小 RNA（miRNAs）参与成骨分化和软骨形成的调节。本研究旨在探讨 miR-128 在骨质疏松症（OP）中的潜在机制。采用定量实时 PCR（qRT-PCR）检测 40 例 OP 患者和 40 例非 OP 患者股骨颈小梁骨中 miR-128 的表达。用 miR-128 模拟物或抑制剂转染 C2C12 细胞，以确定 miR-128 对 C2C12 细胞成骨分化的影响。采用生物信息学和荧光素酶报告基因检测 miR-128 在 C2C12 细胞成骨分化中的分子机制。qRT-PCR 结果显示，OP 患者骨样本中 miR-128 的表达水平明显高于非 OP 患者，而 C2C12 细胞成骨分化时 miR-128 表达明显下调。此外，结果表明，miR-128 过表达显著抑制 C2C12 细胞中骨钙素（OC）、碱性磷酸酶（ALP）和 I 型胶原-α1（COL1A1）的 mRNA 和蛋白表达水平，而 miR-128 抑制剂可逆转此作用。生物信息学分析和双荧光素酶报告基因检测发现，沉默信息调节蛋白 6（SIRT6）是 miR-128 的直接靶标。qRT-PCR 和 Western blot 结果发现，miR-128 显著下调 SIRT6 的 mRNA 和蛋白表达。此外，沉默 SIRT6 显著抑制 miR-128 抑制剂对成骨标志物表达的促进作用。上述结果证实，miR-128 通过下调 SIRT6 表达抑制 OP 中成骨细胞分化，从而加速 OP 的发展。
