The establishment of the immortalized mouse brain microvascular pericytes model and its preliminary application in screening of cerebrovascular toxicants
10.3760/cma.j.issn.0253-9624.2018.05.014
- VernacularTitle: 小鼠脑微血管周细胞永生化模型的建立及其在脑血管毒物筛查中的初步应用
- Author:
Heping ZHAO
1
;
Yanfang GAO
;
Dong XIA
;
Zhiqiang ZHAO
;
Sai WU
;
Xiaohui WANG
;
Huaixiang LIU
;
Chen XIAO
;
Xiumei XING
;
Yun HE
Author Information
1. Sun Yat-sen University School of Public Health, Guangzhou 510080, China
- Publication Type:Journal Article
- Keywords:
Pericytes;
Lead acetate;
Cadmium chloride;
Sodium arsenite;
Cell toxicity
- From:
Chinese Journal of Preventive Medicine
2018;52(5):538-544
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To establish the immortalized mouse brain microvascular pericytes model and to apply to the cerebrovascular toxicants screening study.
Methods:Brain pericytes were isolated from 3 weeks of mice by tissue digestion. Immortalized pericyte cell line was constructed by infecting with LT retrovirus. Monoclone was selected to purify the immortalized pericyte cell line. The pericyte characteristics and purity were explored by immunocytochemistry. Cell proliferation was measured by using the Pomega MTS cell Proliferation Colorimetric Assay Kit. Pericytes were treated with 0, 160, 320, 640, 1 280, 2 560 μmol/L lead acetate, 0, 5, 10, 20, 40, 80 μmol/L cadmium chloride and 0, 5, 10, 20, 40, 80 μmol/L sodium arsenite in 24 hours. Cell toxicity of each group was determined by MTS assay, median lethal dose (LD50) was calculated in linear regression.
Results:Mouse brain pericytes were successfully isolated by tissue separation and enzyme digestion method. After immortalized by LT retroviruses, monoclone was selected and expanded to establish pericyte cell line. The brain pericytes exhibited typical long spindle morphology and positive staining for α-SMA and Vimentin. The proliferation of brain pericytes cell lines was very slowly, and the doubling time was about 48 hours. The proliferation of immortalized brain pericytes cell lines was very quickly, and the doubling time was about 24 hours. After lead acetate, cadmium chloride and sodium arsenite treatment for 24 hours respectively, gradual declines in cell viability were observed. The LD50 of lead acetate was 2 025.0 μmol/L, the LD50 of cadmium chloride was 36.6 μmol/L, and the LD50 of sodium arsenite was 33.2 μmol/L.
Conclusion:The immortalized mouse brain microvascular pericyte model is established successfully by infecting with LT retrovirus, and can be applied to screen cerebrovascular toxicants. The toxicity of these toxicants to immortalized mouse brain microvascular pericyte is in sequence: sodium arsenite,cadmium chloride, lead acetate.
