调节Lnk/干细胞因子-干细胞因子受体通路对糖尿病状态骨髓间充质干细胞成骨功能的影响

目的 探讨调节Lnk/干细胞因子（SCF）-干细胞因子受体（cKit）通路对糖尿病状态大鼠骨髓间充质干细胞（BMSCs）成骨功能的影响。方法 分离糖尿病大鼠BMSCs，免疫细胞化学染色鉴定后，将细胞分为空白对照组、阴性对照组、RNA干扰组，并用免疫印迹实验检测干扰效果。体外模拟糖尿病状态下诱导BMSCs成骨分化，并检测Lnk/SCF-cKit通路主要蛋白Lnk、SCF、cKit及成骨相关蛋白碱性磷酸酶（ALP）、骨钙素（OCN）、Ⅰ型胶原蛋白a1（ColⅠa1）的表达。结果 分离培养的细胞表达CD₄₄、CD₉₀，不表达CD_11b、CD₄₅，符合BMSCs表面标记物特征。与其他糖尿病状态BMSCs相比，RNA干扰组Lnk表达降低，SCF、cKit表达增加（P<0.05）。成骨诱导分化28 d，与其他糖尿病状态BMSCs相比，RNA干扰组细胞Lnk表达降低，SCF、cKit、ALP、OCN、ColⅠa1表达增加（P<0.05）。结论 抑制Lnk表达、激活SCF-cKit通路可改善糖尿病状态大鼠BMSCs的成骨功能。

Changes in bone marrow mesenchymal stem cells osteogenesis by the regulation of Lnk/stem cell factor-cKit signaling

Objective Changes in the osteogenesis of diabetic rat bone marrow mesenchymal stem cells (BMSCs) by the regulation of Lnk/stem cell factor (SCF)-cKit signaling were investigated. Methods BMSCs were isolated from diabetic rats and identified by immunocytochemical staining. These cells were divided into the control group (untransfected group), negative control group (transfected with control plasmid), and RNA interference group (transfected with Lnk-targeting RNA interference plasmid). Western blot was performed to analyze the effect of interference. The BMSCs were induced for osteogenic differentiation under diabetic conditions, and Western blot was used to examine the expressions of Lnk, SCF, and cKit in Lnk/SCFcKit signaling and osteogenic proteins alkaline phosphatase (ALP), osteocalcin (OCN), and collagen type Ⅰ a1 (ColIa1). Results Isolated cells expressed CD₄₄ and CD₉₀ but not CD_11b or CD₄₅. This phenomenon was characteristic of BMSCs. Compared with other diabetic BMSCs, cells in the RNA interference group expressed low Lnk but high SCF and cKit (P< 0.05). Thereafter, 28 days after induction of osteogenic differentiation, cells in the RNA interference group expressed low Lnk but high SCF, cKit, ALP, OCN, and ColIa1 compared with other diabetic BMSCs (P<0.05). Conclusion The inhibition of Lnk expression and activation of SCF-cKit pathway may improve the osteogenic differentiation of BMSCs under diabetic conditions.