脂肪干细胞的分离提取、鉴定及与脂肪颗粒的混合移植研究

袁志坚+何文涓（等）

[摘要] 目的 分离提取脂肪干细胞（ADSCs），对其进行成脂、成骨的鉴定，并与脂肪颗粒混合移植，观察移植后的脂肪组织的存活情况。 方法 取大鼠腹股沟脂肪，磷酸盐缓冲液清洗，0.1%Ⅰ型胶原酶37℃消化1 h，离心，取沉淀，用含10%血清的DMEM培养基进行培养、传代，取第3代ADSCs用于成脂、成骨鉴定；另取第3代ADSCs用于移植，分为两组：ADSCs（密度为5×107/mL）0.9 mL+脂肪颗粒0.6 mL组、脂肪颗粒1.5 mL组，分别移植于大鼠背部两侧，共移植24只大鼠，分别于2周、1、3个月时脱颈椎处死8只大鼠，再取出背部脂肪组织。 结果 大鼠ADSCs成脂、成骨诱导后，经油红O、茜素红染色呈阳性，移植2周、1个月时，两组取出的脂肪组织的体积变化不大；但移植3个月后，取背部脂肪组织，称重：脂肪颗粒1.5 mL组平均重量为（0.4551±0.0024）g，而ADSCs（密度为5×107/mL）0.9 mL+脂肪颗粒0.6 mL组的平均重量为（0.7798±0.0033）g，两组脂肪组织的重量差异有统计学意义（P < 0.05）。 结论 原代分离、培养的ADSCs生长状况良好，具有向成脂、成骨分化的能力，与脂肪颗粒混合移植后组织的存活率高，并能够改善脂肪颗粒单独移植时的液化吸收情况。

[关键词] 脂肪干细胞；成脂诱导；成骨诱导；移植

[中图分类号] R329 [文献标识码] A [文章编号] 1673-7210（2014）02（a）-0009-05

Isolation， culture and transplantation of rat adipose-derived stem cells

YUAN Zhijian1 HE Wenjuan1 ZHOU Hong1 JIANG Meiling2

1.Wuxi Higher Health Vocational School， Jiangsu Province， Wuxi 214028， China； 2.China Pharmaceutical University， Jiangsu Province， Nanjing 210009， China

[Abstract] Objective To isolate and culture the rat adipose-derived stem cells （ADSCs） and to identificate its multipotent differentiation ability and to observe the survival of transplantation of ADSCs with adipose compared with adipose in the back of the rats. Methods Adipose tissue were obtained from inguinal region and washed in PBS， then digested with 0.1% collagenase type Ⅰ at 37℃ for 1 h followed by centrifugation and ADSCs pellet was resuspended in DMEM with 10% fetal bovine serum （FBS） and seeded in flasks， cells at passages 3 were used for adipogenic differentiation and osteogenic differentiation， both sides of twenty-four rats were received subcutaneously 5×107/mL ADSCs resuspended in 0.9 mL of DMEM medium+0.6 mL of adipose and 1.5 mL of adipose， took out the adipose tissue of eight rats after two weeks， one month and three months respectively. Results ADSCs stained positively for oil red-O and alizarin red， two weeks and one month later， the weight of adipose had no difference with each other， however， after three months， the average weight of 1.5 mL of adipose group and the 5×107/mL ADSCs resuspended in 0.9 mL of DMEM medium+0.6 mL of adipose group were （0.4551±0.0024） g and （0.7798±0.0033） g respectively， thus， there were significant differences between the two groups （P < 0.05）. Conclusion ADSCs is in good condition and has adipogenic and osteogenic differentiation ability， ADSCs with adipose can improve the survival and the liquefied of the adipose after transplantation when compared with adipose alone.