摘要
目的探讨脂肪间充质干细胞(adipose-derived mesenchymal stem cells,ADSCs)在抑制博来霉素诱导裸鼠皮肤纤维化方面的作用及机制。方法选用5~6周龄雌性BALB/c裸鼠24只,完全随机分为空白对照组、移植对照组、模型组和移植组4组,每组6只。空白对照组:每天于裸鼠背部皮下注射PBS溶液2次,每次1ml,间隔1h注射第2次。移植对照组:先注射1×10^6/mlADSCs1ml,间隔1h再注射1mlPBS溶液。模型组:先注射1mg/ml博来霉素1ml,间隔1h再注射1mlPBS。移植组:先注射1ml博莱霉素,间隔1h再注射1mlADSCs。注射4周后取材,大体观察各组裸鼠皮肤质地;蛋白质印迹法检测纤维化相关蛋白的表达;RT-PCR检测与纤维化形成相关基因的表达;免疫组织化学法检测皮肤纤维化程度指标α-平滑肌肌动蛋白(α-smoothmuscleactin,α-SMA)的表达。实验数据以Graphpad7.0软件进行分析,2组间比较采用t检验,多组样本均数比较应用One-WayANOVA,P<0.05为差异具有统计学意义。结果蛋白质印迹法实验结果示,TGF-β1、Notch1及α-SMA表达在ADSCs移植组较模型组明显降低,而空白对照组与移植对照组比较无明显差别。Real-timePCR检测结果:模型组中TGF-β1、Ⅰ型胶原、Notch1、Smad3表达量分别为0.90±0.11、1.20±0.05、1.00±0.06和0.90±0.05,ADSC移植组中分别为0.43±0.05、0.63±0.04、0.60±0.05和0.20±0.11,比模型组明显降低,差异有统计学意义(t=5.29、8.05、5.50、13.44,P=0.006、0.001、0.005、0.000)。空白对照组、移植对照组、模型组和移植组皮肤组织中α-SMAmRNA表达分别为0.37±0.01、0.35±0.05、1.43±0.10、0.83±0.03,与空白对照组及移植对照组相比,模型组α-SMA表达量明显升高,差异有统计学意义(F=352.3,P=0.000),而ADSCs移植组相对模型组α-SMA表达量明显下降,差异有统计学意义(t=10.39,P=0.009)。结论ADSCs局部注射后可能通过旁分泌功能下调TGF-β1、Smad3及Notch1的表达,重塑细胞外基质、抑制成纤维�
Objective To investigate the role and the mechanism of adipose-derived mesenchymal stem cells (ADSCs) in inhibiting bleomycin-induced skin fibrosis in nude mice, in order to provide experimental basis for the prevention and treatment of dermatitis, keloid, scleroderma, and other skin fibrosis disorders. Methods Twenty-four female BALB/c nude mice aged 5-6 weeks were randomly divided into blank control group, transplantation control group, model group, and transplantation group. There were 6 mice in each group. In blank control group, PBS solution was injected subcutaneously on the back of nude mice twice a day, at intervals of 1 h. The transplanted control group was injected with 1×10^6/ml ADSCs 1 ml and then injected with 1 ml PBS solution at intervals of 1 h. In the model group, 1 mg/ml bleomycin was injected firstly, and then 1 ml PBS was injected at intervals of 1 h. In the transplantation group, 1 ml of bleomycin was injected firstly, and 1 ml of ADSCs were injected at intervals of 1 h. After 4 weeks injection, the skin texture of each group nude mice was grossly observed. The expression of fibrosis-related proteins was detected by Western blot. The expression of fibrosis-associated genes was detected by RT-PCR. The expression of α-smooth muscle actin (α-SMA), an indicator marker of skin fibrosis, was detected by immunohistochemistry staining. The experimental data was analyzed with GraphPad 7.0 software. Two groups were compared by t test, and multiple groups of samples were compared using one-way ANOVA. The P<0.05 was regarded as statistically significant. Results The results of Western blot showed that the expression of TGF-β1, Notch1 and α-SMA in ADSCs transplantation group was significantly lower than that in the model group, but there was no significant difference between the blank control group and the transplantation control group. Real-time PCR results showed that the expression levels of TGF-β1, type Ⅰ collagen, Notch1, and Smad3 were significantly higher than those in the ADSCs group. The
作者
韩兵
范金财
Han Bing;Fan Jincai(Ninth Department of Plastic Surgery, Plastic Surgery Hospital of Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100144, China)
出处
《中华整形外科杂志》
CAS
CSCD
北大核心
2019年第2期182-188,共7页
Chinese Journal of Plastic Surgery
关键词
脂肪间充质干细胞
皮肤
纤维化
细胞外基质
成纤维细胞
Adipose derived mesenchymal stem cells
Skin
Fibrosis
Extracellular matrix
Fibroblasts
