流式分析人脂肪组织中血管周围干细胞含量的方法探究  

The method of sorting out perivascular stem cells from human adipose tissue through flow cytometry

作　　者：徐峰[1] 刘舒云[1] 王鑫[1,2] 彭江[1] 卢世璧[1] 袁玫[1] 许文静[1] 郭全义[1] 

机构地区：[1]解放军总医院骨科研究所,北京100853 [2]兰州军区乌鲁木齐总医院骨科,830000

出　　处：《中国医药生物技术》2015年第3期223-227,共5页Chinese Medicinal Biotechnology

基　　金：国家自然科学基金(81472092);国家高技术研究发展计划(863计划)(2012AA020502)

摘　　要：目的建立人脂肪组织中分离血管周围干细胞(PSCs)的方法,并研究其在脂肪组织细胞中所占的比例,为血管周围干细胞作为骨和软骨组织工程新的种子细胞奠定基础。方法取人的脂肪组织分别用Ⅰ型胶原酶和Ⅱ型胶原酶消化得到血管基质成分(SVF),用细胞计数仪及流式细胞仪检测SVF中细胞密度、活细胞比例和PSCs细胞所占的比例。结果用细胞计数仪分析得出用Ⅱ型胶原酶消化脂肪组织所得到的SVF中活细胞比例更高,且差异具有统计学意义(P<0.01),而细胞密度有增高趋势(P=0.18)。用流式细胞仪检测得出使用Ⅱ型胶原酶消化脂肪组织所得SVF中PSCs比例更高,且差异具有统计学意义(P<0.01)。使用Ⅰ型胶原酶消化每10 ml脂肪组织中可以分离得到PSCs活细胞为(7.98±2.38)×105个,使用Ⅱ型胶原酶消化则每10 ml脂肪组织中可以分离得到PSCs活细胞为(16.68±4.56)×105个。结论使用Ⅱ型胶原酶消化脂肪组织可以得到更多的血管周围干细胞PSCs,其在脂肪组织中的含量可以满足骨和软骨损伤后自体细胞移植修复的需要。Objective To establish the method of sorting out perivascular stem cells （PSCs） from human adipose tissue and study the proportion of these cells in adipose tissue cells. This research is to explore new seed cells for the bone and cartilage tissue engineering. Methods Stromal vascular fraction （SVF） was got from human adipose tissue that was digested by collagenase type I or collagenase type II. The cell density, proportion of living cells and proportion of PSCs in SVF were tested by the cell count and flow cytometry （FCM）. Results The proportion of living cells in SVF digested by collagenase type II was much higher through analyzing by the cell count and the difference was statistically significant （P〈0.01）. The cellular density in SVF digested by collagenase type II tend to increase （P=0.18）. The proportion of PSCs in SVF digested by collagenase type II was much higher through analyzing by the flow cytometry, and the difference was statistically significant （P〈0.01）. （7.98 ± 2.38） × 105 PSCs were got in 10 ml human adipose tissue digested by collagenase type I, while （16.68 ± 4.56） × 105 PSCs were got in 10 ml human adipose tissue digested by collagenase type II. Conclusion A higher amount of PSCs can be got from human adipose digested by collagenase type II, and the content of PSCs in the adipose tissue can satisfy the needs of autologous cell transplantation for the bone and cartilage repair.

关 键 词：血管周围干细胞 流式细胞术 种子细胞 

分 类 号：R329.2[医药卫生—人体解剖和组织胚胎学]