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Detection of Porcine Endogenous Retrovirus in Porcine Hepatocytes and the Supernatant of Cultured Porcine Hepatocytes
Peng-Peng Tian 1, Zhong Chen 1,2*, Hua Huang 2, Zhen-Yu Wu 2, Hong-Xun Shen 2, Gen-Xi Li 1, Yi-Tao Ding 3
1State Key Laboratory of Pharmaceutical Biotechnology, School of Life Sciences, Nanjing University, Nanjing 210093, China; 2Department of General Surgery, Affiliated Hospital, Nantong Medical College, Nantong 226001, China; 3
Abstract: In this study， we constructed the determination methods of porcine endogenous retrovirus (PERV) in porcine hepatocytes and the supernatant and studied the significance in the application of bioartificial liver based on porcine hepatocytes. Proviral PERV gag sequences in Chinese experimental miniature pig hepatocytes were detected by PCR using specific primers. PERV RNA in the sera from pigs， canines， rats and HBV patients and the supernatant of cultured porcine hepatocytes at 6 h， 24 h was detected by RT-PCR using specific primers. In the same time， porcine mitochondrial DNA (mtDNA) was detected in porcine hepatocytes. The results showed that proviral PERV gag sequences and porcine mtDNA were detectable in porcine hepatocytes from 5 Chinese experimental miniature pigs. PERV RNA was detectable in the porcine serum and the supernatant of cultured porcine hepatocytes at 24 h. Negative results were showed in the supernatant at 6 h and sera from 5 pigs， 5 canines， 5 rats and 5 HBV patients. Therefore， Chinese experimental miniature pig hepatocytes contain PERV and PERV particles can be released into the serum. PERV particles have been released into the supernatant of cultured porcine hepatocytes at 24 h. PCR and RT-PCR methods we used are of high specificity and convenience.
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