同种异体脂肪干细胞介导RhoA-YAP通路在慢性创面修复机制研究  

Study of repair mechanism in chronic wound with homogeneity adipose-derived stem cells via RhoA/YAP signaling pathway

作　　者：郑炜[1] 程天平[1] 李向云[1] 杨帆[1] 郭亚峰 张龙军 ZHENG Wei;CHENG Tian-ping;LI Xiang-yun;YANG Fan;GUO Ya-feng;ZHANG Long-jun(Department of Bum and Orthopedics,Xingtai People's Hospital,Xingtai 054000,Hebei,China)

机构地区：[1]邢台市人民医院烧伤整形科

出　　处：《生物医学工程与临床》2019年第6期640-646,共7页Biomedical Engineering and Clinical Medicine

基　　金：河北邢台市科技计划项目(2018ZC170)

摘　　要：目的分析同种异体脂肪干细胞(ADSC)移植促进慢性创面修复的效果及Ras同源基因家族成员A(RhoA)-Yes相关蛋白(YAP)信号通路活化的机制。方法选择2个月龄SD大鼠31只,雌雄不限,体质量112~120 g,平均体质量116.7 g。以其中1只提取ADSC并鉴定;30只大鼠随机分为5组:模型组(自身皮瓣移植),ADSC移植组,RhoA抑制剂C3干预组(简称C3干预组),慢病毒转染沉默YAP表达组(简称shYAP组),C3和慢病毒转染联合干预组(简称联合组)。分别在移植后7 d和21 d采用激光多普勒血流计测定创面血流量、皮瓣成活率,逆转录聚合酶链反应(RT-PCR)和Western blot法检测创面RhoA、YAP、结缔组织生长因子(CTGF)和血管内皮生长因子(VEGF)mRNA和蛋白的定量表达水平。结果体外成功分离并鉴定为ADSC。30只大鼠均成活至实验21 d。ADSC移植组7 d和21 d皮肤血流量[7 d(256.3±56.2) PU,21 d (421.5±82.8) PU]、皮瓣成活率[7 d (32.6±10.6)%,21 d (45.8±17.5)%]、RhoA mRNA(7 d0.36±0.07,21 d 0.58±0.12)、YAP mRNA(7 d 0.33±0.06,21 d 0.52±0.13)、CTGF mRNA(7 d 0.49±0.11,21 d 0.67±0.15)和VEGF mRNA(7 d 0.48±0.14,21 d 0.68±0.16)及蛋白定量表达水平(7 d 0.37±0.07,21 d 0.59±0.12;7 d 0.35±0.06,21 d 0.54±0.13;7 d 0.46±0.11,21 d 0.65±0.15;7 d 0.47±0.14,21 d 0.67±0.16)均显著高于其他各组,联合组显著低于C3干预组和shYAP组,差异有统计学意义(P <0.05)。结论同种异体ADSC可提高慢性创面移植皮瓣的存活率,促进创面愈合,增加创面组织CTGF和VEGF表达,可能通过介导RhoA-YAP信号通路的活化程度来实现。Objective To study the effect of homogeneity adipose-derived stem cells(ADSC) transplantation for promoting chronic wounds repairment and mechanism of RhoA/YAP signaling pathway activation. Methods Thirty-one male or female SD rats aged 2 months old, body mass of 112-120 g with mean body mass of 116.7 g were selected. One of the rats was extracted ADSC and identified. Thirty rats were randomly divided into 5 groups: model group(self flap graft), ADSC transplantation group,RhoA inhibitor C3 intervention group(C3 intervention group), lentiviral transfection silencing YAP expression group(shYAP group), C3 and lentiviral transfection combined intervention group(combined group). Blood flow and flap survival rate were measured by laser Doppler flowmeter at 7-day and 21-day after transplantation, the reverse transcription polymerase chain re-action(RT-PCR) and Western blot were used to detect RhoA, YAP, connective tissue growth factor(CTGF) and vascular en-dothelial growth factor(VEGF) mRNA and protein quantitative expression level. Results ADSC was successfully isolated and identified in vitro. Thirty rats survived until 21 th day of experiment. At 7 th day and 21 th day, blood flow in ADSC transplantation group were(256.3 ± 56.2) PU and(421.5 ± 82.8) PU, respectively;flap survival rate were(32.6 ± 10.6) % and(45.8 ±17.5) %, respectively;RhoA mRNA were 0.36 ± 0.07 and 0.58 ± 0.12, respectively;YAP mRNA were 0.33 ± 0.06 and 0.52 ±0.13, respectively;CTGF mRNA were 0.49 ± 0.11 and 0.67 ± 0.15, respectively;VEGF mRNA were 0.48 ± 0.14 and 0.68 ±0.16, respectively. Protein quantitative expression levels at 7 th day vs 21 th day were(0.37 ± 0.07 vs 0.59 ± 0.12;0.35 ± 0.06 vs0.54 ± 0.13;0.46 ± 0.11 vs 0.65 ± 0.15;0.47 ± 0.14 vs 0.67 ± 0.16). The above values were significantly higher than that in other groups. The tested values in combined group were significantly lower than that in the C3 intervention group and shYAP group, and the differences were statistically significant(P < 0.05). Conclusion It is demo

关 键 词：脂肪干细胞 慢性创面 RhoA-YAP信号雪路 结缔组织生长因子(CTGF) 血管内皮生长因子(VEGF) 慢病毒 鼠 

分 类 号：R318[医药卫生—生物医学工程]