A simple and highly purified method for isolation of glomeruli from the mouse kidney

Author:

Wang Honglian12,Sheng Jingyi2,He Huijun2,Chen Xiaocui2,Li Jinhong3,Tan Ruizhi1,Wang Li1,Lan Hui-Yao2

Affiliation:

1. Research Center for Integrated Medicine, Affiliated Traditional Medicine Hospital of Southwest Medical University, Luzhou, Sichuan, China

2. Department of Medicine and Therapeutics and Li Ka Shing Institute of Health Sciences, the Chinese University of Hong Kong, Hong Kong, China

3. Department of Nephrology, the Seventh Affiliated Hospital, Sun Yat-sen University, Shenzhen, China

Abstract

Highly purified mouse glomeruli are of great value for studying glomerulus-associated kidney diseases. Here, we developed a simple and rapid procedure for mouse glomerular isolation with large quantity and high purity based on the combination of size-selective sieving and differential adhesion techniques, which we termed the “differential adhesion method.” In this method, mouse renal cortices were minced and digested with collagenase. Glomeruli were disassociated from tubules by successive sieving through 105-, 75-, and 40-μm cell strainers. The retained glomeruli-rich preparation on the 40-μm strainer was rinsed into a cell culture dish to allow tubules to adhere quickly to the dish while leaving most glomeruli floating (termed “differential adhesion”). The floating glomerular fraction was then subjected to another wash through the 40-μm strainer followed by an additional differential adhesion step to obtain highly purified glomeruli with yields of 8,357 ± 575 and purity of 96.1 ± 1.8% from one adult C57BL/6 mouse. The purity of the isolated glomeruli was further confirmed by high expression of the podocyte marker nephrin without detectable tubular marker cadherin-16. Importantly, we also found that although both the quantity and purity of the isolated glomeruli by this and the established Dynabeads method were comparable, glomeruli isolated by the current method showed much less inflammatory stress in terms of proinflammatory cytokine expression than the Dynabeads method. In conclusion, we established a newly mouse glomerular isolation method that is simple, rapid, cost effective, and productive. It provides an advanced methodology for research into glomerulus-related kidney diseases in the mouse.

Funder

Research Grants Council of Hong Kong

The Health and Medical Research Fund of Hong Kong

The Youth Scientist Innovation Workshop Foundation of Luzhou, Sichuan

Publisher

American Physiological Society

Subject

Physiology

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