大网膜包裹明胶-间充质干细胞复合物对马兜铃酸肾病保护作用的研究

doi:10.3877/cma.j.issn.2095-3216.2022.01.002

中华肾病研究电子杂志 ›› 2022, Vol. 11 ›› Issue (01) : 7 -14. doi: 10.3877/cma.j.issn.2095-3216.2022.01.002

论著

大网膜包裹明胶-间充质干细胞复合物对马兜铃酸肾病保护作用的研究

付章宁¹, 宋成成¹, 耿晓东², 迟坤¹, 洪权³, 吴镝³^,^†(

)

^1. 100853　北京，解放军医学院；100853　北京，解放军总医院第一医学中心肾脏病医学部、解放军肾脏病研究所、肾脏疾病国家重点实验室、国家慢性肾病临床医学研究中心、肾脏疾病研究北京市重点实验室
^2. 100853　北京，解放军总医院第一医学中心肾脏病医学部、解放军肾脏病研究所、肾脏疾病国家重点实验室、国家慢性肾病临床医学研究中心、肾脏疾病研究北京市重点实验室；066100　秦皇岛，联勤保障部队北戴河康复疗养中心
^3. 100853　北京，解放军总医院第一医学中心肾脏病医学部、解放军肾脏病研究所、肾脏疾病国家重点实验室、国家慢性肾病临床医学研究中心、肾脏疾病研究北京市重点实验室

收稿日期:2021-12-09 出版日期:2022-02-28
通信作者: 吴镝
基金资助:
国家自然科学基金面上项目(81870491，82070741); 解放军总医院"国家杰出青年科学基金"培育专项(2019-JQPY-002)

Protective effect of omentum-encapsulated gelatin-mesenchymal stem cells complex on aristolochic acid nephropathy

Zhangning Fu¹, Chengcheng Song¹, Xiaodong Geng², Kun Chi¹, Quan Hong³, Di Wu³^,^†()

^1. Chinese PLA Medical School, Beijing 100853; Department of Nephrology, First Medical Center of Chinese PLA General Hospital, Chinese PLA Institute of Nephrology, State Key Laboratory of Kidney Diseases, National Clinical Research Center for Kidney Diseases, Beijing Key Laboratory of Kidney Diseases, Beijing 100853
^2. Department of Nephrology, First Medical Center of Chinese PLA General Hospital, Chinese PLA Institute of Nephrology, State Key Laboratory of Kidney Diseases, National Clinical Research Center for Kidney Diseases, Beijing Key Laboratory of Kidney Diseases, Beijing 100853; Chinese PLA Joint Logistics Support Force Beidaihe Rehabilitation Center, Qinhuangdao 066100, Hebei Province; China
^3. Department of Nephrology, First Medical Center of Chinese PLA General Hospital, Chinese PLA Institute of Nephrology, State Key Laboratory of Kidney Diseases, National Clinical Research Center for Kidney Diseases, Beijing Key Laboratory of Kidney Diseases, Beijing 100853

Received:2021-12-09 Published:2022-02-28
Corresponding author: Di Wu

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引用本文：

付章宁, 宋成成, 耿晓东, 迟坤, 洪权, 吴镝. 大网膜包裹明胶-间充质干细胞复合物对马兜铃酸肾病保护作用的研究[J]. 中华肾病研究电子杂志, 2022, 11(01): 7-14.

Zhangning Fu, Chengcheng Song, Xiaodong Geng, Kun Chi, Quan Hong, Di Wu. Protective effect of omentum-encapsulated gelatin-mesenchymal stem cells complex on aristolochic acid nephropathy[J]. Chinese Journal of Kidney Disease Investigation(Electronic Edition), 2022, 11(01): 7-14.

目的

研究大网膜包裹明胶-间充质干细胞(MSCs)复合物对马兜铃酸肾病(AAN)小鼠肾脏的修复和保护作用。

方法

使用明胶与间充质干细胞共培养构建间充质干细胞-明胶复合物。用CCK-8法评价MSCs与明胶的生物相容性以及MSCs的增殖能力。将体重为20~25 g的健康雄性C57BL/6小鼠24只随机分为4组：正常对照组(Control组)、马兜铃酸肾病模型组(AAN组，腹腔注射马兜铃酸溶液)、大网膜包裹MSCs治疗组(MSCs组)、大网膜包裹MSCs-明胶复合物治疗组(MSCs-Gel组)。利用小动物活体成像系统观察MSCs在小鼠体内的分布和存活情况；检测各组小鼠的血肌酐、尿素氮水平变化，观察肾脏组织学变化及相关纤维化指标如α-SMA、TGF-β和胶原变化情况。

结果

明胶对MSCs的增殖能力无明显影响。与单纯MSCs组相比，MSC-Gel组的MSCs在小鼠肾脏内存活时间更长，且血肌酐、尿素氮明显降低，肾组织病理改变纤维化程度明显减轻，α-SMA、TGF-β、Col-Ⅰ表达亦显著降低(均P<0.05)。

结论

大网膜包裹明胶-MSCs复合物对AAN小鼠肾脏具有良好的修复和保护作用。

关键词: 大网膜, 明胶, 间充质干细胞, 马兜铃酸肾病

Objective

To investigate the repair and protective effects of omentum-encapsulated gelatin-mesenchymal stem cells (MSCs) complex on the kidneys of mice with aristolochic acid nephropathy (AAN).

Methods

The MSCs-gelatin complex was made by co-culture of gelatin and MSCs. The biocompatibility between MSCs and gelatin as well as the proliferation ability of MSCs were evaluated with the CCK-8. Twenty-four healthy male C57BL/6 mice weighing 20-25 g were randomly divided into 4 groups: control group (normal mice), AAN model group (intraperitoneal injection of aristolochic acid solution), MSCs group (greater omentum-encapsulated MSCs), and MSCs-Gel group (greater omentum-encapsulated MSCs plus gelatin complex). An in vivo fluorescence imaging system was used to observe the distribution and survival of MSCs in mice. The levels of serum creatinine and blood urea nitrogen of the mice were measured. The renal histological changes and fibrosis indicators such as α-SMA, TGF-β, and collagen were also evaluated.

Results

The gelatin had no significant influence on the proliferation ability of MSCs. Compared with the MSCs group, the MSC-Gel group showed significantly longer survival of the MSCs in the mice kidneys, lower levels of serum creatinine and blood urea nitrogen, less expression of α-SMA, TGF-β, and collagen Ⅰ, and lower level of renal fibrosis (P<0.05).

Conclusion

The omentum-encapsulated gelatin-MSCs complex had good repair and protective effects on the kidneys of AAN mice.

Key words: Greater omentum, Gelatin, Mesenchymal stem cells, Aristolochic acid nephropathy

图1 CCK-8法检测MSCs的细胞增殖能力注：MSCs-Gel组与MSCs组相比，^aP>0.05

图2 各组小鼠小动物活体荧光成像观察结果注：MSC组与MSC-Gel组相比，^aP<0.05

图3 各组小鼠体重变化情况及肾功能指标检测结果注：A：各组小鼠体重变化情况；B：各组小鼠血肌酐检测结果；C：各组小鼠尿素氮检测结果；与AAN组相比，^aP<0.05；与MSC组相比，^bP>0.05；与MSC组相比，^cP<0.05

图4 各组肾脏组织病理PAS染色(×200)注：与AAN组相比，^aP<0.05；与MSC组相比，^bP<0.05

图5 各组肾脏组织病理Masson染色(×200)注：与AAN组相比，^aP<0.05；与MSC组相比，^bP<0.05

图6 各组肾脏组织病理PSR染色(×200)注：与AAN组相比，^aP<0.05；与MSC组相比，^bP<0.05

图7 Western印迹检测各组肾脏中α-SMA、TGF-β和Col-Ⅰ蛋白的表达注：α-SMA：抗平滑肌抗体；TGF-β：转化生长因子β；和Col-Ⅰ：胶原蛋白Ⅰ；与MSC组相比，^aP<0.05

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