Isolation and epithelial co-culture of mouse renal peritubular endothelial cells

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• 作者：Ye Zhao (1)
  Hong Zhao (1) (2)
  Yun Zhang (1) (3)
  Tania Tsatralis (1)
  Qi Cao (1)
  Ya Wang (1)
  Yiping Wang (1)
  Yuan Min Wang (4)
  Steve I Alexander (4)
  David C Harris (1)
  Guoping Zheng (1)
  
  1. Centre for Transplant and Renal Research ; Westmead Millennium Institute ; The University of Sydney ; Sydney ; NSW ; Australia
  2. Department of Biochemistry and Molecular Biology ; Shanxi Medical University ; Taiyuan ; PR China
  3. Experimental Centre of Science and Research ; The First Clinical Hospital of Shanxi Medical University ; Taiyuan ; PR China
  4. Centre for Kidney Research ; Children鈥檚 Hospital at Westmead ; Sydney ; NSW ; Australia
  
• 关键词：Peritubular endothelial cells ; Tubular epithelial cells ; CD146 ; Co ; culture ; Vascular endothelial growth factor
• 刊名：BMC Cell Biology
• 出版年：2014
• 出版时间：December 2014
• 年：2014
• 卷：15
• 期：1
• 全文大小：1,886 KB
• 参考文献：1. Hashimoto, N, Phan, SH, Imaizumi, K, Matsuo, M, Nakashima, H, Kawabe, T, Shimokata, K, Hasegawa, Y (2010) Endothelial-mesenchymal transition in bleomycin-induced pulmonary fibrosis. Am J Respir Cell Mol Biol 43: pp. 161-172 CrossRef
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  3. Li, J, Bertram, JF (2010) Review: endothelial-myofibroblast transition, a new player in diabetic renal fibrosis. Nephrology (Carlton) 15: pp. 507-512 CrossRef
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• 刊物主题：Cell Biology; Biological Microscopy; Life Sciences, general;
• 出版者：BioMed Central
• ISSN：1471-2121

文摘

Background Endothelial-mesenchymal transition (EndoMT) has been shown to be a major source of myofibroblasts, contributing to kidney fibrosis. However, in vitro study of endothelial cells often relies on culture of isolated primary endothelial cells due to the unavailability of endothelial cell lines. Our recent study suggested that peritubular endothelial cells could contribute to kidney fibrosis through EndoMT. Therefore, successful isolation and culture of mouse peritubular endothelial cells could provide a new platform for studying kidney fibrosis. This study describes an immunomagnetic separation method for the isolation of mouse renal peritubular endothelial cells using anti-CD146 MicroBeads, followed by co-culture with mouse renal proximal tubular epithelial cells to maintain endothelial phenotype. Results Flow cytometry showed that after isolation and two days of culture, about 95% of cells were positive for endothelial-specific marker CD146. The percentage of other cells, including dendritic cells (CD11c) and macrophages (F4/80), was less than 1%. Maintenance of endothelial cell phenotype required vascular endothelial growth factor (VEGF) and co-culture with mouse proximal tubular epithelial cells. Conclusion In this study, we established a method for the isolation of mouse renal peritubular endothelial cells by using immunomagnetic separation with anti-CD146 MicroBeads, followed by co-culture with mouse renal proximal tubular epithelial cells to maintain phenotype.