CAJ | 학술논문

目的探讨Notch信号在大鼠骨髓间充质干细胞(MSCs)向内皮细胞分化中的作用及其对诱导分化后细胞功能的影响.方法分离、培养大鼠MSCs,用含血管内皮生长因子(VEGF165)和碱性成纤维细胞生长因子(bFGF)的细胞培养液培养大鼠MSCs 2周,诱导MSCs向内皮细胞分化;采用免疫荧光技术鉴定细胞;用逆转录-聚合酶链反应(RT-PCR)检测诱导分化前后细胞上Notch信号受体和配体的表达.用γ-内分泌酶抑制剂阻断细胞Notch信号通路的转导,应用划痕实验检测细胞迁移能力;将细胞接种在半固体培养基上,观察其形成毛细血管样结构的能力.结果诱导MSCs向内皮细胞分化后细胞表达CD31和Flk1,说明其具备内皮细胞的特性.MSCs上表达有Notch信号的受体Notchl和配体Jaggedl的mRNA;但诱导前后细胞上Notch信号受体Notchl mRNA表达差异无统计学意义(0.59±0.01比0.59±0.01,P>0.05),其配体Jaggedl mRNA表达稍有上升趋势(1.01±0.02比0.99±0.03,P>0.05).阻断Notch信号通路转导可增强诱导后内皮细胞的迁移能力[划痕空白处细胞数(个):44.61±4.34比40.06±2.43,P<0.053及形成毛细血管样结构的能力(细胞分级:3.67±0.82比2.00±0.89,P<0.01).结论 Notch信号在大鼠MSCs向内皮细胞分化过程中可能具有重要作用,阻断Notch信号通路转导可增强诱导后内皮细胞的功能.
목적 탐토Notch신호재대서골수간충질간세포(MSCs)향내피세포분화중적작용급기대유도분화후세포공능적영향.방법 분리、배양대서MSCs,용함혈관내피생장인자(VEGF165)화감성성섬유세포생장인자(bFGF)적세포배양액배양대서MSCs 2주,유도MSCs향내피세포분화;채용면역형광기술감정세포;용역전록-취합매련반응(RT-PCR)검측유도분화전후세포상Notch신호수체화배체적표체.용γ-내분비매억제제조단세포Notch신호통로적전도,응용화흔실험검측세포천이능력;장세포접충재반고체배양기상,관찰기형성모세혈관양결구적능력.결과 유도MSCs향내피세포분화후세포표체CD31화Flk1,설명기구비내피세포적특성.MSCs상표체유Notch신호적수체Notchl화배체Jaggedl적mRNA;단유도전후세포상Notch신호수체Notchl mRNA표체차이무통계학의의(0.59±0.01비0.59±0.01,P>0.05),기배체Jaggedl mRNA표체초유상승추세(1.01±0.02비0.99±0.03,P>0.05).조단Notch신호통로전도가증강유도후내피세포적천이능력[화흔공백처세포수(개):44.61±4.34비40.06±2.43,P<0.053급형성모세혈관양결구적능력(세포분급:3.67±0.82비2.00±0.89,P<0.01).결론 Notch신호재대서MSCs향내피세포분화과정중가능구유중요작용,조단Notch신호통로전도가증강유도후내피세포적공능.
Objective To research the role of Notch signaling during the differentiation of bone marrow mesenchymal stem cells (MSCs) into endothelial cells and its effect on the functions of the differentiated cells.Methods Rat bone marrow MSCs were isolated and cultured in vitro.then the cells were treated with vascular endothelial growth factor (VEGF165)and basic fibroblast growth factor(bFGF) for 2 weeks to induce it to differentiate into endothelial cells.The differentiated cells were identified by fluorescence immunoassay.The receptors and ligands of the Notch signaling were detected by reverse transcription-polymerase chain reaction(RT-PCR)before and after the differentiation.γ-secretase inhibitor was used to block Notch pathway.Migration ability of cells were assessed by scarification test.Cells were inoculated on semisolid gel to study their ability of forming the capillary-like structure.Results After inducing MSCs to differentiate into endothelial cells by VEGF165 and bFGF,MSCs gained the characteristics of the endothelial cells with expression of CD31 and Flk1.There were Notchl mRNA and Jaggedl mRNA expressions in rat bone marrow MSCs.The expression changes in the receptor Notchl were not statistically significant on the differentiated cells(0.59±0.01 vs.0.59±0.01,P>0.05),but there was a trend towards an increase of Jaggedl mRNA(1.01±0.02 vs.0.99±0.03,P>0.05).When Notch pathway was blocked,the differentiated cells' migration ability was increased (number of cells on the scratched area:44.61±4.34 vs.40.06±2.43,P<0.05),and the ability of forming capillary-like structure was also increased(cells classification:3.67±0.82 vs.2.00±0.89,P<0.01).Conclusion Notch signaling may have an important role during the differentiation of MSCs into endothelial cells.The function of differentiated cells were strengthened when Notch pathway was blocked.