Other Liver Disease
Synchronization isolation method for multiple types of cells from mouse liver
Gan Jian, Ji Cuifeng, Mao Xiaorong, Wang Jiangtao, Lyu Chunyan, Shi Yifan, Liao Yao, He Yali, Shu Lian, Li Long, Li Junfeng
Published 2023-05-20
Cite as Chin J Hepatol, 2023, 31(5): 532-537. DOI: 10.3760/cma.j.cn501113-20220827-00433
Abstract
ObjectiveTo explore a simple and feasible method for the isolation and purification of hepatocytes, hepatic stellate cells (HSC), and lymphocytes from mice.
MethodsThe cell suspension was obtained from male C57bl/6 mice by hepatic perfusion through the portal vein digestion method and then isolated and purified by discontinuous Percoll gradient centrifugation. Trypan blue exclusion was used to determine cell viability. Glycogen staining, cytokeratin 18, and transmission electron microscopy were used to identify hepatic cells. Immunofluorescence was used to detect α-smooth muscle actin combined with desmin in HSCs. Flow cytometry was used to analyze lymphocyte subsets in the liver.
ResultsAfter isolation and purification, about 2.7×107 hepatocytes, 5.7×105 HSCS, and 4.6×106 hepatic mononuclear cells were obtained from the liver of mice with a body weight of about 22g. The cell survival rate in each group was > 95%. Hepatocytes were apparent in glycogen deposited purple-red granules and cytokeratin 18. Electron microscopy showed that there were abundant organelles in hepatocytes and tight junctions between cells. HSC had expressed α-smooth muscle actin and desmin. Flow cytometry showed hepatic mononuclear cells, including lymphocyte subsets such as CD4, CD8, NKs, and NKTs.
ConclusionThe hepatic perfusion through the portal vein digestion method can isolate multiple primary cells from the liver of mice at once and has the features of simplicity and efficiency.
Key words:
Hepatocytes; Hepatic stellate cells; Lymphocytes; Isolation; Primary culture
Contributor Information
Gan Jian
The First Clinical Medical College of Lanzhou University, Lanzhou 730000, China
Ji Cuifeng
The First Clinical Medical College of Lanzhou University, Lanzhou 730000, China
Mao Xiaorong
The First Clinical Medical College of Lanzhou University, Lanzhou 730000, China
Wang Jiangtao
The First Clinical Medical College of Lanzhou University, Lanzhou 730000, China
Lyu Chunyan
The First Clinical Medical College of Lanzhou University, Lanzhou 730000, China
Shi Yifan
The First Clinical Medical College of Lanzhou University, Lanzhou 730000, China
Liao Yao
The First Clinical Medical College of Lanzhou University, Lanzhou 730000, China
He Yali
The First Clinical Medical College of Lanzhou University, Lanzhou 730000, China
Shu Lian
The First Clinical Medical College of Lanzhou University, Lanzhou 730000, China
Li Long
The First Clinical Medical College of Lanzhou University, Lanzhou 730000, China
Li Junfeng
The First Clinical Medical College of Lanzhou University, Lanzhou 730000, China
