切换至 "中华医学电子期刊资源库"
高级检索
中华肾病研究电子杂志

中华肾病研究电子杂志 ›› 2020, Vol. 09 ›› Issue (04) : 145 -151. doi: 10.3877/cma.j.issn.2095-3216.2020.04.001

所属专题： 文献；

论著

人羊水间充质干细胞的分离鉴定及其对抗Thy-1肾炎的治疗作用
张峻铭1, 张英杰1, 白雪源1, 陈香美1,†()   
  1. 1. 100853　北京，解放军总医院第一医学中心肾脏病科、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:2020-06-03 出版日期:2020-08-28
  • 通信作者: 陈香美
  • 基金资助:
    国家重点研发计划(2016YFA0101002)

Isolation and identification of human amniotic fluid mesenchymal stem cell and its therapeutic effect in anti-Thy-1 nephritis

Junming Zhang1, Yingjie Zhang1, Xueyuan Bai1, Xiangmei Chen1,†()   

  1. 1. 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, China
  • Received:2020-06-03 Published:2020-08-28
  • Corresponding author: Xiangmei Chen
  • About author:
    Corresponding author: Chen Xiangmei, Email:
全文 ( ) 下载PDF ( ) 导出引用
引用本文：

张峻铭, 张英杰, 白雪源, 陈香美. 人羊水间充质干细胞的分离鉴定及其对抗Thy-1肾炎的治疗作用[J]. 中华肾病研究电子杂志, 2020, 09(04): 145-151.

Junming Zhang, Yingjie Zhang, Xueyuan Bai, Xiangmei Chen. Isolation and identification of human amniotic fluid mesenchymal stem cell and its therapeutic effect in anti-Thy-1 nephritis[J]. Chinese Journal of Kidney Disease Investigation(Electronic Edition), 2020, 09(04): 145-151.

探讨羊水间充质干细胞(amniotic fluid mesenchymal stem cells，AF-MSCs)对抗Thy-1肾炎模型的治疗作用。

方法

采用差异性贴壁与机械性分离法从人孕中期羊水中分离出羊水间充质干细胞，流式细胞术鉴定其表面标志物，成脂、成骨诱导分化检测其分化能力。利用尾静脉注射抗Thy-1抗体建立SD大鼠抗Thy-1系膜增生性肾炎模型；将第5代羊水间充质干细胞通过尾静脉注射治疗抗Thy-1系膜增生性肾炎大鼠，检测大鼠尿蛋白的变化，PAS染色观察大鼠肾脏病理改变。

结果

羊水间充质干细胞顺利分离，流式结果显示其表达间充质干细胞标志物(CD29、CD44、CD73、CD90、CD105)和胚胎干细胞标志物(SSEA-4)而不表达造血干细胞标志物(CD34、CD45、CD133)，且在适当条件下能够被诱导分化为脂肪细胞和骨细胞。羊水间充质干细胞治疗后，与模型组相比，治疗组在第7、12天时24 h尿蛋白明显下降(P<0.01)，且肾脏病理结果表明系膜细胞增殖减轻，系膜区细胞外基质积聚减少。

结论

成功的从人孕中期羊水中分离出羊水间充质干细胞，羊水间充质干细胞治疗系膜增生性肾小球肾炎有效。

关键词: 羊水间充质干细胞, 系膜增生性肾小球肾炎, 干细胞治疗
Objective

To explore the therapeutic effect of amniotic fluid mesenchymal stem cells (AF-MSCs) in anti-Thy-1 mesangial proliferative glomerulonephritis.

Methods

AF-MSCs were isolated from human amniotic fluid during the second trimester by differential adherence and mechanical separation methods, and their surface markers were identified by flow cytometry, while their differentiation ability was detected by lipogenesis and osteogenesis induction methods. The anti-Thy-1 mesangial proliferative nephritis model in SD rats was established by injecting anti-Thy-1 antibody into tail vein. The rats with anti-Thy-1 mesangial proliferative nephritis were treated with the 5th-generation AF-MSCs through tail vein injection. Changes of urine protein of the rats were detected, and pathological changes of the rats kidneys were observed after PAS staining.

Results

The AF-MSCs were successfully isolated, and flow cytometry results showed that they expressed mesenchymal stem cell markers (CD29, CD44, CD73, CD90, CD105) and embryonic stem cell markers (SSEA-4) but not hematopoietic stem cell markers (CD34, CD45, CD133). The AF-MSCs could be induced to differentiate into adipocytes and bone cells under appropriate conditions. The urinary protein level of the AF-MSCs treatment group was significantly lower than that of the model group at 24 h, on day 7, and on day 12 (P<0.01). And renal pathology examination showed that the mesangial cell proliferation and mesangial extracellular matrix accumulation were both reduced.

Conclusion

AF-MSCs were successfully isolated from human amniotic fluid during the second trimester, and were effective in treating the anti-Thy-1 mesangial proliferative glomerulonephritis of rats.

Key words: Amniotic fluid mesenchymal stem cells, Mesangial proliferative glomerulonephritis, Stem cell therapy
图1 羊水细胞的形态(200×)
图2 羊水间充质干细胞的分离(100×)
图3 流式检测羊水间充质干细胞表面标志物的表达
图4 羊水间充质干细胞的诱导分化
图5 不同时间点24 h尿蛋白定量的变化
图6 不同时间点肾脏病理PAS染色(400×)
[1]
Zhou Q, Yang X, Wang M, et al. Changes in the diagnosis of glomerular diseases in East China: a 15-year renal biopsy study [J]. Ren Fail, 2018, 40(1): 657-664.
[2]
Jin M, Yin Z, Wei K, et al. Metanephric mesenchyme-derived Foxd1＋ mesangial precursor cells alleviate mesangial proliferative glomerulonephritis [J]. J Mol Med, 2019, 97(4): 553-561.
[3]
De Coppi P, Bartsch G Jr, Siddiqui MM, et al. Isolation of amniotic stem cell lines with potential for therapy [J]. Nat Biotechnol, 2007, 25(1): 100-106.
[4]
Hauser PV, De Fazio R, Bruno S, et al. Stem cells derived from human amniotic fluid contribute to acute kidney injury recovery [J]. Am J Pathol, 2010, 177(4): 2011-2021.
[5]
Sedrakyan S, Da Sacco S, Milanesi A, et al. Injection of amniotic fluid stem cells delays progression of renal fibrosis [J]. J Am Soc Nephrol, 2012, 23(4): 661-673.
[6]
Geng W, Wei R, Liu S, et al. Shenhua tablet inhibits mesangial cell proliferation in rats with chronic anti-Thy-1 nephritis [J]. Biol Res, 2016, 49: 17.
[7]
Kunter U, Rong S, Djuric Z, et al. Transplanted mesenchymal stem cells accelerate glomerular healing in experimental glomerulonephritis [J]. J Am Soc Nephrol, 2006, 17(8): 2202-2012.
[8]
Sakr S, Rashed L, Zarouk W, et al. Effect of mesenchymal stem cells on anti-Thy1.1 induced kidney injury in albino rats [J]. Asian Pac J Trop Biomed, 2013, 3(3): 174-181.
[9]
Kim J, Lee Y, Kim H, et al. Human amniotic fluid-derived stem cells have characteristics of multipotent stem cells [J]. Cell Prolif, 2007, 40(1): 75-90.
[10]
Dziadosz M, Basch RS, Young BK. Human amniotic fluid: a source of stem cells for possible therapeutic use [J]. Am J Obstet Gynecol, 2016, 214(3): 321-327.
[1] 杨蕴钊, 周诚, 石美涵, 赵静, 白雪源. 人羊水间充质干细胞对膜性肾病大鼠的治疗作用[J]. 中华肾病研究电子杂志, 2023, 12(04): 181-186.
[2] 项永生, 徐如祥. 骨髓基质干细胞在脑缺血疾病治疗的新进展[J]. 中华神经创伤外科电子杂志, 2017, 03(05): 300-304.
[3] 江学良. 重视溃疡性结肠炎的限时达标治疗[J]. 中华消化病与影像杂志(电子版), 2017, 07(06): 241-242.
[4] 王天仁, 张巧霞, 石瑶, 汤雯珺, 马捷. 神经干细胞在精神分裂症患者治疗中的研究进展[J]. 中华临床医师杂志(电子版), 2020, 14(12): 1002-1008.
阅读次数
全文


摘要

版权所有 中华医学会 中华医学电子音像出版社有限责任公司  京ICP备14006079号-1  网络出版服务许可证：(署)网出证(京)字第075号