Models of adenoviral transfection and hypoxia/reoxygenation-induced injury in AMCMs of adult mouse cardiomyocytes
10.3969/j.issn.1005-4847.2024.04.003
- VernacularTitle:成年小鼠心肌细胞腺病毒转染及缺氧/复氧诱导损伤模型的建立
- Author:
Xiaoru LI
1
;
Xinye YAO
;
Jia LIU
;
Xiaoyu ZHANG
;
Yiman ZHANG
;
Baochang LAI
;
Qiang MA
;
Yidong WANG
;
Hongyan TIAN
;
Qian YIN
Author Information
1. 西安交通大学心血管研究中心,西安 710061
- Keywords:
AMCMs extraction;
adenovirus transfection;
hypoxia-reoxygenation
- From:
Acta Laboratorium Animalis Scientia Sinica
2024;32(4):435-443
- CountryChina
- Language:Chinese
-
Abstract:
Objective To construct models of viral transfection and hypoxia/reoxygenation induced cellular injury in adult mouse cardiomyocytes(AMCMs)isolated using a non-Langendorff method.Methods AMCMs were isolated,extracted,sedimented,and plated using a non-Langendorff method.The morphology and survival rate of the isolated cells were evaluated 2,24,48 and 72 h after plating,and their integrity was observed by immunofluorescence staining for α-actinin.The isolated AMCMs were infected with adenoviruses carrying an RFP-expressing vector and fluorescence images were obtained at 36 and 48 h post-infection and used to calculate transfection efficiency.The cells were cultured under hypoxic conditions for 45 min,reoxygenated for 24 h,and then stained with propidium iodide(PI)to verify establishment of the hypoxia/reoxygenation injury model.Results The survival rates of AMCMs at 2,24 and 48 h after plating were comparable,but survival was significantly reduced at 72 h.The integrity of the AMCMs was good and＞80%of the cells were transfected with adenovirus at 48 h.After hypoxia/reoxygenation treatment,42%of cells were stained by PI,suggesting successful establishment of the AMCM injury model.Conclusions In this study,we developed a non-Langendorff method for the fast and easy isolation of AMCMs with high cell viability.The isolated cells can be efficiently infected with adenovirus and respond to hypoxia/reoxygenation injury.These findings provide a systematic method for isolating AMCMs and for applying gene modification and hypoxia/reoxygenation injury in these cells.
