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Primary culture and identification of rat glomerular microvascular endothelial cells

MA Hua-Gen¹, LIU Hai-Qin², LIU Zhao-De³, TANG Yuan-Yu^4,*

¹Department of Shang Han, College of Traditional Chinese Medicine, Beijing University of Chinese Medicine, Beijing 102488, China；²College of Integrated Traditional Chinese and Western Medicine, Fujian University of Traditional Chinese Medicine, Fuzhou 350122, China；³Basic Medical College, Nanjing Medical University, Nanjing 211116, China；⁴College of Traditional Chinese Medicine, Fujian University of Traditional Chinese Medicine, Fuzhou 350122, China

Abstract

The aim of the present study was to establish a simple and efficient method for isolation and culture of primary rat glomerular microvascular endothelial cells in vitro. The bilateral kidneys were taken from 7–10-day old Sprague-Dawley rats, and the renal cortex was separated. Glomeruli were obtained by cutting and continuously passing 200-mesh and 300-mesh sieves. After type IV collagenase digestion for 15–20 min, renal microvascular globules were collected for inoculation and culture. The cultured cells were identified by cell morphology observation and immunocytochemical staining with factor VIII related antigen. The results showed that the renal microvascular globules were irregularly spherical, without cysts, and the capillary loop structure was clear; after 3 days of primary culture, short spindle-shaped cells crawled out around the renal microvascular globules and gradually formed cell colonies, showing an “island-like shape” distribution; 4–5 days later, the cell colonies fused with each other; 6 days later, the cells covered the bottom of the dish, showing a typical monolayer, paving stone-like, mosaic arrangement. The immunocytochemical staining of factor VIII related antigen showed that the cytoplasm was lightly stained brownish red, and factor VIII related antigen-positive rate of cells was nearly 100%. The above results suggested that this study successfully established a method combining continuous screening and collagenase digestion for culture of primary rat glomerular microvascular endothelial cells in vitro. It provides an important tool cell for studying the mechanism of the occurrence and development of diabetic nephropathy.

Key words: glomerulus; microvascular endothelial cells; continuous screening method; collagenase digestion method; factor VIII related antigen; rats

Received: 2021-02-24　　Accepted: 2021-05-31

Corresponding author: 唐元瑜　　E-mail: 2422198977@qq.com

Citing This Article：

MA Hua-Gen, LIU Hai-Qin, LIU Zhao-De, TANG Yuan-Yu. Primary culture and identification of rat glomerular microvascular endothelial cells. Acta Physiol Sin 2021; 73 (6): 926-930 (in Chinese with English abstract).