摘要
目的探讨人参皂苷对骨髓间充质干细胞(bone marrow derived mesenchymal stem cell,BMSC)通过旁分泌途径治疗心肌缺血的促进作用。方法从SD大鼠胫骨和股骨中分离培养BMSC,取第3代BMSC,流式细胞术检测细胞表面抗原CD44、CD45、CD34和CD29的表达。建立大鼠心肌缺血模型,将模型鼠随机分为模型组(每天常规喂食水)和人参皂苷组[每天常规喂食水及人参皂苷液(100 mg/100 ml),每日3次,100 ml/次],给药的同时,均经大鼠尾静脉缓慢注射BMSC悬液(1×105个/ml),1 ml/只,另设空白对照组(未建模的SD大鼠)。分别采集模型组建模后24、48、72 h及人参皂苷组建模后72 h的大鼠心脏血,分离血清,ELISA法检测血清中干细胞因子(stem cell factor,SCF)、干细胞衍生因子(stromal cell derived factor-1,SDF-1)、血管内皮生长因子(vascular endothelial growth factor,VEGF)及趋化因子受体4(chemokine receptor 4,CXCR4)的分泌水平。体外迁移试验检测不同浓度SDF-1(1、10、100 ng/ml)对BMSC迁移的影响;取各组大鼠建模后24、48、72 h的心肌组织提取液,检测其对BMSC及经SDF-1抑制剂AMD3100作用的BMSC迁移的影响。结果大鼠BMSC表面表达CD44、CD29,不表达CD34、CD45。模型组大鼠血清中细胞因子SCF、SDF-1、VEGF、CXCR4的分泌水平随建模时间延长显著上升,人参皂苷组大鼠建模后72 h血清中各细胞因子的表达水平均明显高于模型组(P<0.05)。SDF-1可显著增加BMSC的迁移数,且呈浓度依赖性(P<0.05);模型组和人参皂苷组大鼠心肌组织提取液作用BMSC的迁移数随建模时间的延长显著上升(P<0.05),两组相同时间点间差异有统计学意义(P<0.05);人参皂苷组72 h的大鼠心肌组织提取液作用的AMD3100处理的BMSC的迁移数明显低于模型组72 h(P<0.05)。结论人参皂苷能促进BMSC旁分泌作用,提高BMSC对心肌缺血性疾病的疗效,可作为治疗该疾病的辅助药物。
Objective To investigate the promoting effect of ginsenoside on treatment of myocardial ischemia with bone marrow mesenchymal stem cells （BMSCs） by paracrine effect. Methods BMSCs were isolated from shin-bone and thighbone of SD rats, of which the passage 3 were determined for surface antigens CD44, CD45, CD34 and CD29 by flow cytometry. Rat model of myocardial ischemia was established, and the model rats were divided into model control and test groups. The rats in model control group were fed by normal diet dialy, while those in test groups with ginsenoside （ 100 mg/100 ml） 3 times a day, and injected i.v. slowly with BMSC suspension （1 × 10^5cells/ml） at the same time, 1 ml for each, using normal SD rats as normal control. Heart blood samples were collected from the rats in model control group 24, 48 and 72 h and in test group 72 h after establishment of model, from which sera were extracted and determined for secretion levels of stem cell factor （SCF）, stromal cell derived factor-1 （SDF-1）, vascular endothelial growth factor （VEGF） and chemokine receptor 4 （CXCR4） by ELISA. The effects of SDF-1 at various concentrations （1, 10 and 100 ng/ml） on migration of BMSCs were determined by migration test in vitro. The cardiac muscle tissue extracts of rats in various groups were collected 24, 48 and 72 h after establishment of model, of which the effect on the migration of BMSCs untreated and those treated with SDF-1 inhibitor AMD3100 was determined. Results CD44 and CD29 were expressed, while CD34 and CD45 were unexpressed on the surface of BMSCs. The secretion levels of SCF, SDF-1, VEGF and CXCR4 in heart blood of rats in model control group increased significantly with the increasing hours after establishment of model. However, the secretion levels of these cytokines in test group 72 h after establishment of model were significantly higher than those in model control group （P 〈 0. 05）. SDF-1 showed dose-dependent effect on the migration of BMSCs. SDF-1 increased the counts of migrated BMSCs in a dose. dependent manner （P 〈 0. 05 ）. The counts of migrated BMSCs after treatment with cardiac muscle tissue extracts in model control and test groups increased significantly with the increasing hours （P 〈 0. 05 ）, which showed signifi- cant difference in two groups at the same time points（P 〈 0. 05）. However, the count of AMD3100-treated migrated BMSCs after treatment with cardiac muscle tissue extracts of rats in test group 72 h after establishment of model was significantly lower than that in model group （P 〈 0. 05 ）. Conclusion Ginsenoside promoted the paracrine of BMSCs and enhanced the curative effect of BMSCs on ischemic heart disease, which might be used as an accessory drug the disease.
出处
《中国生物制品学杂志》
CAS
CSCD
2013年第11期1617-1620,1624,共5页
Chinese Journal of Biologicals
基金
辽宁省科技厅计划项目(2011225015)
关键词
人参皂苷
骨髓间充质干细胞
旁分泌
心肌缺血
Ginsenoside
Bone marrow mesenchymal stem cells （BMSCs）
Paracrine
Myocardial ischemia
