Primary culture for synovial fibroblasts in mice
10.3969/j.issn.1671-8348.2016.02.026
- VernacularTitle:小鼠滑膜成纤维细胞的原代培养
- Author:
Jinjun ZHAO
;
Ziyou HU
;
Qingqing OUYANG
;
Jing WU
;
Yujiao CHEN
;
Min YANG
- Publication Type:Journal Article
- Keywords:
arthritis,rheumatoid;
fibroblasts;
synovial membrane;
cell culture techniques;
mice,inbred C57BL
- From:
Chongqing Medicine
2016;(2):228-231
- CountryChina
- Language:Chinese
-
Abstract:
Objective The primary culture of synovial fibroblasts is a convenient tool to study the pathology and physiology of synovial tissues .An improved method was constructed in this study by C57BL /6 mice to study the mechanism of rheumatoid ar-thritis(RA) .Methods The synovium around the hip joints were collected .Attention should be paid to eliminate the egg-yolk like yellow oval substance in the middle of the synovium .The synovium was transferred into a 1 .5 mL Eppendorf tube containing 0 .5%type Ⅳ collagenase and cut into 1 mm3 blocks or so .The Eppendorf tube was placed in 37 ℃ Constant temperature orbital shaker incubator for 60 min .After digestion ,the tube was placed on the Vortex for a high-speed oscillation for 1 .5 minutes to guarantee the separation of cells .Results Within about 1 week ,the first passage was performed by the trypsin digestion method .On day 10 , the number of synovial macrophages reached the maximum and then decreased gradually .After the third generation (day 15 to 20) , the synovial macrophages generally disappeared .Vimentin was suitable for the immunofluorescence cytochemical staining for the synovial fibroblasts .The cell purity was indicated as > 95% .The cytometric analysis indicated that purity of Vimentin and CD90 .2-labelled cells was over 95% ;the purity of CD54-labelled cells was 80% approximately .Conclusion It is a simple and effective method for primary culture of synovial fibroblasts in mice .
