UTX转染对骨髓间充质干细胞影响的体外研究△

Effect of UTX transfection on bone marrow mesenchymal stem cells in vitro

摘要:

[目的] 探索广泛转录 X 染色体三角形四肽重复蛋白 (ubiquitously transcribed tetratricopeptide repeat on chromosome X, UTX) 基因调控骨髓间充质干细胞 (bone marrow mesenchyml stem cells, BMSCs) 的作用。[方法] 自 C57 小鼠中分离提取 BMSCs，行形态与流式细胞学表面抗原鉴定。分别采用慢病毒转染，建立 UTX-down 组、UTX-NC 组和 UTX-up 组。行 QRTPCR 检测 UTXmRNA 表达，检测诱导成骨和成软骨情况，II 型胶原的表达。[结果]所获得的第三代细胞符合 BMSCs 的典型形态特征和表面抗原特征。QRT-PCR 检测 UTX 的 mRNA 相对表达量由低至高依次为：UTX-down 组＜ UTX-NC 组＜ UTX-up 组， 差异均有统计学意义(P＜0.05)。诱导成骨培养茜素红染着色细胞数、诱导成软骨后阿利新兰染色细胞数以及 II 型胶原免疫荧光染色 OD 值由低至高依次均为：UTX-down 组＜ UTX-NC 组＜ UTX-up 组，差异均有统计学意义(P＜0.05)。[结论]BMSCs 过表达 UTX 可促使成骨与成软骨的发生，促使 II 型胶原产生。有可能用于促使椎间盘纤维环缺损的修复。

Abstract:

[Objective] To explore the effect of ubiquitously transcribe tetropeptide repeat on chromosome X (UTX) genes transfection on bone marrow mesenchymal stem cells (BMSCs) in vitro. [Methods] BMSCs were isolated from C57 mice and identified by morphology and flow cytometry. Lentivirus transfection was used to establish UTX-down group, UTX-NC group and UTX-up group of cells. RT-PCR was performed to detect UTX mRNA expression. In addition, osteogenesis, chondrogenesis and type II collagen expression were assayed and compared. [Results] The third-generation cells obtained in this study proved consistent with the typical morphological characteristics and surface antigen characteristics of BMSCs. The relative expression levels of UTX mRNA detected by QRT-PCR were ranked down-up as follows: UTX-down group ＜ UTX-NC group ＜ UTX-up group, with statistically significant differences among the 3 groups (P＜0.05) . Furthermore, the number of alizarin red staining cells in osteogenic culture, the number of Alcian blue staining cells after chondrogenic cul- ture, and the OD values of type II collagen immunofluorescence staining all were ranked from low to high as follows: UTX-down group ＜ UTX-NC group ＜ UTX-up group, which all were statistically significantly different (P＜0.05) . [Conclusion] BMSCs overexpression of UTX do promote the occurrence of osteogenesis and chondrogenesis, and production of type II collagen. It may be used to promote the repair of annulus fibrosus defect of intervertebral disc.