摘要目的 探讨利用脂肪间充质干细胞微泡(adipose tissue mesenchymal stem cell-derived microvesicles,ADSC-MV)辅助脂肪移植,提高移植脂肪体积保留率的可行性.方法 取华中科技大学同济医学院附属协和医院于2016年12月至2017年4月收治的5例20～30岁健康女性来院行腹部吸脂手术所得脂肪组织,酶解贴壁法分离ADSC,传代培养获取其上清液后梯度离心获取ADSC-MV.使用透射电镜观察微泡形态,PKH26染色观察微泡性质.将16只BCLA裸鼠采用随机数表法分2组,每组8只,每只裸鼠背部皮下左右两侧分别注射等量脂肪移植物:空白组每个部位注射0.35 ml脂肪颗粒;微泡组每个部位注射0.35 ml脂肪颗粒+7μg ADSC-MV(混合物终浓度为20μg/ml).于术后1、3个月取材,测量移植物重量及体积,HE染色切片观察脂肪细胞形态、组织纤维化及空洞形成情况,CD31免疫组织化学染色计量血管形成数目.使用SPSS 13.0进行统计学处理,2组间比较采用t检验,P<0.05为差异有统计学意义.结果 术后1个月及3个月,空白组移植物体积较微泡组小,肉眼可见明显脂肪液化部分,微泡组移植物体积较大,色泽均一,未见明显坏死、液化组织.术后3个月空白组移植物重量(53.23±9.64)mg,显著低于MV组的(106.50±14.46)mg(t=8.117,P<0.001).术后3个月空白组移植物体积(0.057±0.009)ml,微泡组(0.103±0.015)ml(t=8.117,P<0.001).HE染色显示,术后1、3个月空白组移植物内中心区域可见脂肪细胞被大量增生的纤维结缔组织及空洞分隔,微泡组移植物内形成的纤维结缔组织及空洞较少,且脂肪细胞大小较为均一.CD31免疫组织化学血管计数结果显示,术后3个月空白组每个视野下血管数为23.0±2.4,微泡组为29.6±4.0,移植物内血管密度高于空白组(t=2.825,P=0.022),且形成的纤维化及空洞均低于空白组.结论 脂肪间充质干细胞微泡可提高脂肪移植物体积保留率.

abstractsObjective To investigate the possibility of adipose tissue mesenchymal stem cell-derived microvesicles ( ADSC-MVs) to improve the retention volume of fat transplantation. Methods Human adipose tissue was obtained from 5 healthy female patients aged from 20 to 30 years, who came to the hospital for abdominal liposuction. Adipose tissue mesenchymal stem cells ( ADSCs) were acquired by collagenase enzymatic hydrolysis. ADSC-MVs were isolated from the supernatant of cultured ADSCs through ultra-centrifugation, and characterized by transmission electron microscope and PKH26 staining. Sixteen BALB/c-nu nude mice were randomly divided into 2 groups ( n=8, mice/group) by random number table method. In EV group, the mice were subcutaneously injected 0. 35 ml human fat, with 7 μg ADSC-EVs ( final concentration is 20 μg/ml) ,while 0.35 ml human fat were in blank group. The mice were sacrificed and the grafts were harvested at 1 month and 3 months after transplantation. Weight and volume of transplanted fat were tested. Morphology of adipocytes, fibrosis and necrosis of fat tissue were confirmed by HE staining. Blood vessel density were accessed by CD31 staining. SPSS 13. 0 was used for statistical analysis and Student-t test was applied to compare the data from two groups. Results One month after fat transplantation, the volume of fat grafts in blank group was lower than that in MV group. Besides, lipoliquefaction can be observed in fat grafts of blank group. Fat grafts in MV group had a higher volume and littler necrosis and lipolique faction. Three months after grafting, the volume of fat grafts in blank group [(0. 057±0. 009) ml] was significantly lower than that in MV group [(0. 103±0. 015) ml], t=8. 117, P<0. 001. MV group had an increased number of vessels (29. 6±4. 0/field) compared with grafts from the Blank group (23. 0±2. 4/field), t=2. 825, P=0. 022. Histological analysis revealed that the fat grafts in MV group consisted less fat necrosis and fibrosis than control group. Conclusions ADSC-MVs can improve the retention volume of fat transplantation.