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| Isolation and short term cultivation of swine hepatocytes for bioartificial liver support system |
| Jun Li, Lan-Juan Li, Hong-Cui Chao, Qian Yang, Xiao-Li Liu, Ji-Fang Sheng, Hai-Ying Yu and Jian-Rong Huang |
Hangzhou, China
Author Affiliations: Key Laboratory of Infectious Disease, Ministry of Public Health of China; Institute of Infectious Disease, Department of Infectious Diseases, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou 310003, China (Li J, Li LJ, Chao HC, Yang Q, Liu XL, Sheng JF, Yu HY and Huang JR)
Corresponding Author: Lan-Juan Li, MD, Department of Infectious Diseases, The First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou 310003, China (Tel: 86-571-87236759; Fax: 86-571-87236755; Email: lilj@zjwst.gov.cn) |
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Abstract BACKGROUND: The demand for the clinical use of hepatocytes is increasing. The aim of this study was to develop a method for procurement of high qualitative pig hepatocytes and to evaluate the state of freshly isolated and cultured hepatocytes.
METHODS: The domestic extracorporeal circulating perfusion apparatus was used to isolate and harvest swine hepatocytes by the two-step perfusion method with EDTA and collagenase. The viability, function and morphology of the freshly isolated and cultured cells were evaluated and observed by the trypan blue exclusion test, biochemical measurements, phase contrast microscopy and transmission electron micrography (TEM).
RESULTS: The total yield of isolated hepatocytes reached to 1.5(±0.4)×1010 per liver with a viability of 92(±5)%, and the purity of hepatocytes reached to 98%. Immediately after isolation, phase-contrast microscope and TEM showed that undamaged hepatocytes appeared bright, translucent and spherical in shape, with a characteristic well-contrasted border. After 24 hours, the concentrations of alanine aminotransferase (ALT), aspartate aminotransferase (AST), lactate dehydrogenase (LDH), albumin (ALB), creatinine (Cr) and blood urea nitrogen (BUN) in the fluid of culture were declined significantly.
CONCLUSION: This method of procuring swine hepatocytes could get high quality cells with active metabolic function.
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2005, Vol. 4 
