整合素靶向肽促进SD大鼠骨髓间充质干细胞的增殖  

Integrin-targeted peptide promotes proliferation of bone marrow mesenchymal stem cells in SD rats

作　　者：贾麒钰 黄晓夏 郭建 黄金勇 郭晓斌 阿卜杜萨拉木·阿力木江 吴桐 马创[1] Jia Qiyu;Huang Xiaoxia;Guo Jian;Huang Jinyong;Guo Xiaobin;Abdussalam·Alimujiang;Wu Tong;Ma Chuang(Department of Microprosthetic Surgery,First Affiliated Hospital of Xinjiang Medical University,Urumqi 830054,Xinjiang Uygur Autonomous Region,China;Department of Traumatology and Orthopedics,First Affiliated Hospital of Xinjiang Medical University,Urumqi 830054,Xinjiang Uygur Autonomous Region,China;Department of Joint Surgery,Orthopedic Center,First Affiliated Hospital of Xinjiang Medical University,Urumqi 830054,Xinjiang Uygur Autonomous Region,China)

机构地区：[1]新疆医科大学第一附属医院骨科中心显微修复外科,新疆维吾尔自治区乌鲁木齐市830054 [2]新疆医科大学第一附属医院骨科中心创伤骨科,新疆维吾尔自治区乌鲁木齐市830054 [3]新疆医科大学第一附属医院骨科中心关节外科,新疆维吾尔自治区乌鲁木齐市830054

出　　处：《中国组织工程研究》2022年第30期4780-4786,共7页Chinese Journal of Tissue Engineering Research

基　　金：新疆维吾尔自治区自然科学基金(2020D01C263),项目负责人:郭晓斌;国家自然科学基金(81760397),项目负责人:马创。

摘　　要：背景:研究报道整合素靶向肽多巴胺-精氨酸-甘氨酸-天冬氨酸(dopamine-arginine-glycine-aspatic acid,DOPA-RGD)可以抑制破骨细胞活化和促进细胞黏附,然而对于整合素靶向肽干预骨髓间充质干细胞或改性钛基材料的最适浓度及最适时间尚不明确。目的:探究DOPA-RGD肽促进大鼠骨髓间充质干细胞增殖的最适干预质量浓度及最适干预时间。方法:体外分离培养SD大鼠骨髓间充质干细胞,选取纯度达标的细胞进行下一步干预实验。96孔培养板中接种第3代骨髓间充质干细胞,对照组细胞培养采用单纯α-MEM完全培养基,其余各实验组用含2.5,5,10,20,40,80 mg/L DOPA-RGD肽条件培养基培养,CCK-8法测定DOPA-RGD肽干预1,3,5,7 d的细胞增殖情况,筛选出DOPA-RGD肽干预骨髓间充质干细胞的最适质量浓度及干预时间,流式细胞仪测定细胞凋亡率进一步验证最适干预质量浓度,采取划痕实验验证最适质量浓度DOPA-RGD肽对骨髓间充质干细胞迁徙能力的影响。结果与结论:①重复测量方差分析结果显示,DOPA-RGD肽干预不同时间组间的吸光度值改变不同,干预至第5天时增殖效果最佳,存在显著性差异(F=3012.618,P<0.001),通过LSD法对分组因素行组间两两比较,10 mg/L DOPA-RGD肽组与质量浓度≥20 mg/L的3个DOPA-RGD肽组及对照组比较存在显著性差异(P<0.01);②细胞凋亡率测定显示,与其余各组相比,10 mg/L DOPA-RGD肽干预第5天可显著降低骨髓间充质干细胞的凋亡率(P<0.001);③与对照组相比,10 mg/L DOPA-RGD肽干预24 h后骨髓间充质干细胞呈现出一定迁徙能力,迁徙速度明显加快;④结果表明,10 mg/L DOPA-RGD肽干预第5天对骨髓间充质干细胞有显著的促增殖效应。BACKGROUND:Studies have reported that integrin-targeted peptide dopamine-arginine-glycine-aspatic acid(DOPA-RGD)could inhibit osteoclast activation and promote cell adhesion,while the optimal concentration and optimal time to apply DOPA-RGD peptide to intervene with bone marrow mesenchymal stem cells or modified titanium-based materials are unclear OBJECTIVE:To investigate the optimal mass concentration and the optimal intervention time of DOPA-RGD peptide to produce pro-proliferative effect when acting on bone marrow mesenchymal stem cells in rats.METHODS:SD rat bone marrow mesenchymal stem cells were isolated and cultured in vitro and the cells with the required purity were selected for the next step of intervention.The third-generation bone marrow mesenchymal stem cells were inoculated in 96-well culture plates.Cells in the control group were cultured with simpleα-MEM complete medium,while the remaining of the experimental groups were cultured with conditioned medium containing 2.5,5,10,20,40,and 80 mg/L DOPA-RGD peptide.Cell proliferation was measured by CCK-8 method after 1,3,5,and 7 days of DOPA-RGD peptide intervention.The optimal mass concentration and intervention time of DOPA-RGD peptide in bone marrow mesenchymal stem cells were selected,and the optimal mass concentration of DOPA-RGD peptide in bone marrow mesenchymal stem cells was further verified by apoptosis rate using flow cytometry.The effect of optimal mass concentration of DOPA-RGD peptide on the migration ability of bone marrow mesenchymal stem cells was verified by scratch assay.RESULTS AND CONCLUSION:(1)Using repeated measures analysis of variance,the absorbance values of the cells intervened by DOPA-RGD peptide changed differently between different time length groups,and the best proliferation effect was achieved by the 5th day of intervention,and the difference was significant(F=3012.618,P<0.001).Paired comparison of grouping factors by the LSD method showed that there was a significant difference between the 10 mg/L DOPA-RGD peptide grou

关 键 词：整合素 DOPA-RGD肽 整合素靶向肽 骨髓间充质干细胞 细胞增殖 细胞迁徙 牵张成骨 大鼠 

分 类 号：R459.9[医药卫生—治疗学] R318[医药卫生—临床医学]