Cultivation of HepG2.2.15 on Cytodex-3: Higher yield of hepatitis B virus and less subviral particles compared to conventional culture methods

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• 作者：Joachim Lupberger ; Andreas Mund ; Josef Kock and Eberhard Hildt
• 刊名：Journal of Hepatology
• 出版年：2006
• 出版时间：October 2006
• 年：2006
• 卷：45
• 期：4
• 页码：547-552
• 全文大小：259 K

文摘

Background/Aims

Several novel systems are available to study human hepatitis B virus (HBV) replication in cell culture demanding for efficient cell culture based systems for HBV production. The aim was to enhance HBV production of the HBV stably producing cell line HepG2.2.15 by cultivation on spherical micro substrate.

Methods

HepG2.2.15 was cultivated on microcarrier substrate Cytodex-3. HBV specific transcripts, viral protein and genome secretion, cell proliferation and MAP kinase signaling were analyzed. Infectivity of HBV particles was analyzed using primary tupaia hepatocytes.

Results

Compared to stationary flask cultures, HepG2.2.15 on Cytodex-3 secreted 18-fold more HBV genomes, more HBeAg per culture volume and less HBV surface antigen per extracellular viral genome equivalent. This was reflected by a significantly higher infectivity of supernatant derived from carrier grown HepG.2.2.15 cells tested by infection of primary tupaia hepatocytes. The amount of phosphorylated ERK-2 was significantly elevated in cells cultivated on microcarrier.

Conclusions

The cultivation of HepG2.2.15 on Cytodex-3 increased production of infectious HBV particles and decreased secretion of subviral particles compared to the stationary cell cultivation. Microcarrier cultivation activates MAP kinase signaling that is crucial for HBV replication.