摘要
目的通过大鼠肝星状细胞(hepatic stellate cells,HSCs)与大鼠脂肪间充质干细胞(rat adipose tissue-derived mesenchymal stem cells,r ADSCs)在内皮细胞分化诱导培养基中的共培养,观察HSCs对r ADSCs向内皮细胞(endothelial cells,ECs)分化的影响。方法共培养实验设置四组,阳性对照组、实验组、阴性对照组和空白对照组。诱导培养2周后,观察检测各组r ADSCs向ECs的分化情况。结果诱导2周后,三组r ADSCs细胞体积逐渐变小变圆,部分细胞出现典型的内皮细胞鹅卵石样改变;四组细胞v WF、VE-Cadherin和表达均阳性;Western blotting检测结果与免疫荧光结果一致,实验组e NOS蛋白表达明显高于阴性对照组和空白对照组(P<0.05);内皮细胞迁移试验显示,实验组每视野下细胞迁移数目明显高于阴性对照组和空白对照组(P<0.01);低密度脂蛋白吞噬试验显示,实验组细胞吞噬Di IAc-LDL的能力明显高于空白对照组(P<0.05);体外成血管试验显示实验组细胞体外成血管的能力明显高于其他三组细胞—阳性对照组(P<0.05)、阴性对照组和空白对照组(P<0.01)。结论 HSCs与r ADSCs的间接共培养可以促进大鼠血管内皮细胞生长因子(rat vascular endothelial growth factor,r VEGF)介导的r ADSCs向ECs的分化,并增强分化后ECs的功能。利用共培养体系对多种细胞共培养时进行生长分化现象的观察及其相互影响机制的研究,可以为组织工程功能性血管化组织器官及其生物模拟物的设计和创造提供更加可靠的理论指导和更加广阔的研究策略。
Objective To observe the effect of hepatic stellate cells on promoting rat adipose tissue-derived mesenchymal stem cells to differentiate into endothelial cells. Methods Four groups were designed： HSCs coculture group, the positive control, the negative control and the blank control. The characteristics of ECs induced from r ADSCs of the groups were detected with immunofluorescence, Western blotting, EC migration assay,acetylated low-density lipoprotein uptake assay and in vitro matrigel assay. Results During 2 weeks inducing,the r ADSCs got smaller and some of them developed cobblestone morphology. The ECs of the four groups stained positive for v WF and VE-cadherin, and the Western blotting showed the consistent results with immunofluorescence that the e NOS had a higher expression in HSCs co-culture group and the positive control compared with the negative control and the blank control（P〈0.05）. ECs of HSCs co-culture group got a stronger ability of migration compared with the negative control and the blank control（P〈0.01）, of uptaking Di I-Ac-LDL compared with the blank control（P〈0.05）, of forming vascular network-like structures compared with the positive control（P〈0.05）,compared with the negative control and the blank control（P〈0.01）. Conclusion In the co-culture system, HSC scan enhance the effect of VEGF on promoting r ADSCs differentiate into ECs and enhance the endothelial function of ECs. It will provide a more reliable theoretical guidance and research strategy for the design and manufacture of functional vascularized tissues and their biological mimics through the study of the growth and differentiation mechanism in co-culture system.
出处
《肝胆胰外科杂志》
CAS
2015年第2期125-132,共8页
Journal of Hepatopancreatobiliary Surgery
基金
浙江省外科学重中之重学科资助项目(2008-255)
浙江省自然科学基金资助项目(LY13H030009)
温州市科技局资助项目(Y20120170)
