摘要
目的研究线粒体乙醛脱氢酶2(ALDH2)缺失对体外培养的小鼠骨髓内皮祖细胞(EPCs)生长状态及功能的影响。方法分别取C57BL/6小鼠(WT)和相同背景来源的A/d/h2基因缺失(Aldh2-/-,KO)小鼠的股骨和胫骨,PBS冲洗获得骨髓细胞组份,密度梯度离心收集单个核细胞白膜层,并进行细胞计数,同时观察不同基因型来源的细胞在不同的培养体系及不同时间点的生长特点,采用乙酰化低密度脂蛋白(Dil-ac-LDL)吞噬实验和荆豆凝集素(FITC-UEA-1)结合实验验证EPC的状态和功能。结果 Aldh2-/-小鼠的骨髓单个核(BMNCs)数目与野生型小鼠相比显著降低(p<0.05,n=7),但是成集落能力两组之间无差异。在M199培养基中培养不同基因型来源的BMNCs时,细胞呈链条状结构,而在EGM-2培养基中培养时,细胞呈集落式生长。培养第十天时,WT小鼠来源EPCs的Dil-ac-LDL和FITC-UEA-1双阳性率达到80.23±3.98%,而A/dh2-/-小鼠EPCs的双阳性率只有52.66±10.54%。培养至第三周时,CD31流式检测表明,Aldh2基因缺失降低EPCs CD31的表达。结论A/dh2基因缺失影响EPCs的增殖效率及功能。这一作用可能会影响其在临床缺血性疾病中的推广应用。
Objective To study the effects of mitochondrial ALDH2 on the differentiation and function of endothelial progenitor cells in vitro. Methods The femur and tibia were isolated from wild type C57BL/6 mice and Aldh2-/-mice from the same gene background, and rinsed by PBS, then the bone marrow mononuclear cells was centrifuged in mice Ficoll-Hypaque solution and the buffy coat was harvested and counted, we observed the differentiation and function of EPCs with different gene background at time points, which were verified by abilities of uptake Dil-ac-LDL and binging FITC-UEA-1 lectin. Results Compared with WT-BMNCs, the number of KO-BMNCs significantly declined （p〈0.05, n=7）, while the cloning efficiency showed no difference. EPCs grew in the shape of chain when cultured in M199 medium, shape of colony in EGM-2 medium. At day 10, the Dil-ac-LDL and FITC-UEA-1 double positive EPCs was 80.23 ±3.98% and 52.66 ±10.54% in WT ＆ KO mice respectively. The analysis of cytometer flow at the third week demonstrated that AMh2 deficiency decreased the CD31 expression of EPCs compared with WT- EPCs. Conclusion ALDH2 deficiency compromised the proliferation and function of EPCs, which might affect the application of EPCs in clinic for isehemic diseases.
作者
孙晓垒
孙淑惠
朱洪
董震
马鑫
韩莎莎
孙爱军
SUN Xiao-lei SUN Shu-hui ZHU Hong DONG Zhen MA Xin HAN Sha-sha SUN Ai-jun.(Institute of Biomedical Science, Fudan University, Shanghai 200032, China Department of Medical Microbiology and Parasitology, Basic Medical School of Fudan University, Key Laboratory of Molecular Virology, Ministry of Education and Health, Shanghai 200032, China.)
出处
《中国分子心脏病学杂志》
CAS
2016年第5期1857-1860,共4页
Molecular Cardiology of China
基金
国家自然科学基金(81570224)
国家自然基金创新群体(81521001)
