小鼠肾脏足细胞的原代培养和鉴定

发布时间：2018-07-10 02:01

本文选题：肾脏 + 足细胞　；参考：《吉林大学学报(医学版)》2017年01期

【摘要】：目的:建立可操作性好、简单易行且效率较高的小鼠肾脏足细胞分离及原代培养模式,为其进一步研究奠定基础。方法:取C57/BL6J小鼠肾脏,通过差异过筛法获取300目筛的肾小球,用制备好的KI-3T3培养基重悬肾小球后静置于铺被鼠尾胶原的培养皿中,4d后开始换液,7d后开始胰蛋白酶消化肾小球,并开始足细胞传代培养。5~7d传代1次,传代2~3次后收集细胞鉴定。采用倒置扭转显微镜观察小鼠肾脏足细胞形态表现,采用PCR法检测小鼠肾脏足细胞中nephrin,podocin和P-cadherin的表达。结果:肾小球种植3d后可见足细胞从组织中爬出,足细胞传代培养7d后在倒置相差显微镜下可观察到细胞胞体有突起生长。PCR法检测,分离培养的足细胞表达nephrin、podocin和P-cadherin。结论:差异过筛结合消化酶技术能够成功分离培养C57/BL6J小鼠肾脏足细胞。
[Abstract]:Objective: to establish a simple and efficient model of mouse kidney podocyte isolation and primary culture. Methods: three hundred mesh glomeruli of C57 / BL6J mouse kidney were obtained by differential sieve method. The glomeruli were suspended in the prepared KI-3T3 medium and put into a culture dish coated with rat tail collagen for 4 days. The glomeruli were digested by trypsin for 7 days. The podocytes were subcultured once for 7 days and collected for identification after 3 passages. The morphology of mouse renal podocytes was observed by inverted torsion microscope and the expression of nephrinpodocin and P-cadherin was detected by PCR. Results: podocytes could be seen crawling out of the tissue after 3 days of glomerular implantation. After 7 days of podocyte culture, the neurite growth of the cell body could be observed under inverted phase contrast microscope. The cultured podocin expressed nephrininpodocin and P-cadherin. Conclusion: differential screening combined with digestive enzyme technique can successfully isolate and culture renal podocytes of C57 / BL6J mice.
【作者单位】：甘肃省人民医院儿科;甘肃省中医药大学研究生处;宁夏医科大学研究生学院;
【基金】：国家自然科学基金地区项目资助课题(81360114) 甘肃省科技厅技术研究与开发专项计划资助课题(1305TCYA035) 甘肃省卫计委行业项目资助课题(GSWSKY-2015-12) 甘肃省委组织部陇原青年创新人才扶持计划资助课题[甘组通字(2014)4号]
【分类号】：R329.2

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