Adipose-derived mesenchymal stem cell transplantation combined with cordyceps polysaccharide improves renal function and hypercoagulable state of rats with nephrotic syndrome

doi:10.3969/j.issn.2095-4344.2016.50.015

Abstract

Abstract:

BACKGROUND: Cordyceps polysaccharide is a commonly used adjuvant drug for clinical treatment of nephrotic syndrome. As a classic model of nephrotic syndrome induced by adriamycin, the Sprague-Dawley rat model of nephrotic syndrome exhibits similar clinical manifestations and pathological changes to minimal-change nephropathy in humans.
OBJECTIVE: To observe the effect of adipose-derived mesenchymal stem cells (ADMSCs) transplantation combined with cordyceps polysaccharide on renal function and hypercoagulable state in rats with nephrotic syndrome.
METHODS: ADMSCs suspension was made in vitro and labeled using PKH26. Fifty Sprague-Dawley rats were randomized into normal (no intervention), model, ADMSCs, cordyceps polysaccharide and combined treatment groups (n=10/group). Adriamycin administration was performed to make rat models of nephrotic syndrome in the latter three groups. After modeling, model rats were respectively given no treatment, ADMSCs intravenously for 3 days, cordyceps polysaccharide intragastrically for 12 weeks, or their combined use. Then, 24-hour urinary protein, serum total cholesterol, triglyceride, total protein, albumin, high-density lipoprotein, low-density lipoprotein, creatinine, blood urea nitrogen levels and coagulation changes were detected at 12 weeks. Meanwhile, histopathological changes of renal tissues were observed under light microscope; survival and distribution of PKH26-labeled ADMSCs were observed by fluorescence microscopy; and expression of Hpa gene in renal tissue was detected by RT-PCR.
RESULTS AND CONCLUSION: Compared with the model group, the 24-hour urinary protein, serum total cholesterol, triglyceride, low-density lipoprotein, creatinine, and blood urea nitrogen levels were significantly lower, while the serum total protein, albumin and high-density lipoprotein levels were significantly higher in the three treatment groups (P < 0.05). These indicators showed significant differences between the combined group and ADMSCs and cordyceps polysaccharide groups (P < 0.05). Both ADMSCs transplantation and cordyceps polysaccharide significantly relieved the hypercoagulable state of rats with nephrotic syndrome, and their combined effects were stronger (P < 0.05). After treatment, the pathological improvement in the kidney tissues was found in the three treatment groups; moreover, it was better in the combined treatment group than the ADMSCs and cordyceps polysaccharide groups. Better improvement in the number of PKH26-labeled ADMSCs and the expression of Hpa mRNA was observed in the combined treatment group compared with the ADMSCs and cordyceps polysaccharide groups. In conclusion, the combination of ADMSCs transplantation and cordyceps polysaccharide can improve kidney function and hypercoagulable state in rats with nephrotic syndrome, reducing pathological damage to the kidney tissue.

中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程

Key words: Mesenchymal Stem Cells, Stem Cell Transplantation, Nephrosis, Lipoid, Tissue Engineering

CLC Number:

R394.2

Guo Zhi-bo, Zhang Chen-jie, Ma Li-na, Gao Da-wei, Song Ju-xing, Li Xiao-jing. Adipose-derived mesenchymal stem cell transplantation combined with cordyceps polysaccharide improves renal function and hypercoagulable state of rats with nephrotic syndrome[J]. Chinese Journal of Tissue Engineering Research, 2016, 20(50): 7549-7556.

Figures/Tables 2

2.1 实验动物数量分析动物分组实验纳入50只SD大鼠，随机分为5组，全部进入结果分析，无脱失。 2.2 脂肪间充质干细胞的形态观察体外培养第3代的脂肪间充质干细胞两至三天后即可铺满瓶底达80%以上；1周后可见脂肪间充质干细胞成集落样生长，核仁明显，核染色质细，胞浆丰富，结构清晰，形态具有典型的成纤维样细胞的形态，细胞表面光滑，分布较为均匀，可见明显漩涡状的细胞克隆，杂细胞基本消失。传第3代的脂肪间充质干细胞形态逐渐均一，呈平行排列生长或旋涡状生长，见图1。 2.3 各组大鼠24 h尿蛋白及各血清指标检测结果与正常组比较，模型组的24 h尿蛋白定量及血清总胆固醇、三酰甘油、低密度脂蛋白、肌酐、尿素氮水平均明显升高(P < 0.05)，血清总蛋白、白蛋白、高密度脂蛋白水平则明显降低(P < 0.01)。脂肪间充质干细胞组、人工虫草多糖组及联合组的24 h尿蛋白定量及血清总胆固醇、三酰甘油、低密度脂蛋白、肌酐、尿素氮水平均明显低于模型组(P < 0.05)，而血清总蛋白、白蛋白和高密度脂蛋白水平则明显高于模型组(P < 0.05)。联合组各项指标与脂肪间充质干细胞组及人工虫草多糖组比较，差异有显著性意义(P < 0.05)。各组大鼠24 h尿蛋白及血清各指标检测结果见表1，2。 2.4 各组大鼠凝血指标变化各组大鼠的凝血指标检测结果显示，脂肪间充质干细胞组、人工虫草多糖组大鼠的血液高凝状态较模型组明显缓解，联合组进一步缓解，各组间差异有显著性意义(P < 0.05)。各组大鼠的凝血指标变化比较见表3及图2。 2.5 各组大鼠的肾组织病理学变化联合组的病理改善情况明显优于脂肪间充质干细胞组和人工虫草多糖组，脂肪间充质干细胞组和人工虫草多糖组较模型组稍有改善，但不及联合组。各组大鼠的病理学变化见图3。 2.6 PKH26标记结果荧光显微镜观测PKH26标记后，在551 nm激光作用下，脂肪间充质干细胞膜完整，发出红色荧光；培养24 h后，呈梭形或多角形红色标记的脂肪间充质干细胞，联合组明显多于脂肪间充质干细胞组及人工虫草多糖组，组间比较差异均有显著性意义(P < 0.05)，见图4。 2.7 各组大鼠Hpa基因表达情况模型组Hpa基因表达较正常组明显增加，脂肪间充质干细胞组、人工虫草多糖组Hpa基因表达较模型组减少，联合组Hpa基因表达量明显减少，组间比较差异均有显著性意义(P < 0.05)。各组大鼠Hpa基因表达见图5。"

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