Baculovirus vectors repress phenobarbital-mediated gene induction and stimulate cytokine expression in primary cultures of rat hepatocytes

N. B. Beck, J. S. Sidhu, Curtis John Omiecinski

Research output: Contribution to journalArticle

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Abstract

Baculovirus transfection strategies have proven successful at transferring foreign DNA into hepatoma cells and primary hepatocytes. When testing the utility of these methodologies in cultured hepatocytes, we discovered that the presence of baculovirus disrupts the phenobarbital (PB) gene induction process, a potent transcriptional activation event characteristic of highly differentiated hepatocytes, and repressed expression of the albumin gene. In concert with previous reports from our laboratory demonstrating that increased cAMP levels can completely repress the induction of specific cytochrome P450 (CYP) genes, cAMP concentrations and PKA activities were measured in the primary hepatocytes subsequent to baculovirus exposure. However, neither parameter was affected by the presence of the virus. To evaluate whether immune response modulation was triggered by baculovirus exposure, RNase protection assays were performed and demonstrated that baculovirus infection activates TNF-α, IL-1α and IL-1β expression in the primary hepatocyte cultures. Immunocytochemical experiments indicated that the production of cytokines was likely due to the presence of small numbers of Kupffer cells present in the culture populations. Exogenously added TNF-α was also effective in repressing PB induction, consistent with other reports indicating that inflammatory cytokines are capable of suppressing expression of biotransformation enzyme systems. Comparative studies demonstrated the specificity of these effects since exposures of hepatocytes to adenoviral vectors did not result in down-regulation of hepatic gene responsiveness. These results indicate that baculovirus vectors enhance the expression of inflammatory cytokines in primary hepatocyte cultures, raising concerns as to whether these properties will compromise the use of baculovirus vectors for study of cytochrome P450 gene regulation, as well as for liver-directed gene therapy in humans.

Original languageEnglish (US)
Pages (from-to)1274-1283
Number of pages10
JournalGene Therapy
Volume7
Issue number15
DOIs
StatePublished - Jan 1 2000

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Baculoviridae
Phenobarbital
Hepatocytes
Cytokines
Genes
Interleukin-1
Cytochrome P-450 Enzyme System
Kupffer Cells
Liver
Ribonucleases
Biotransformation
Genetic Therapy
Transcriptional Activation
Transfection
Albumins
Hepatocellular Carcinoma
Down-Regulation
Viruses
Gene Expression
DNA

All Science Journal Classification (ASJC) codes

  • Molecular Medicine
  • Molecular Biology
  • Genetics

Cite this

Beck, N. B. ; Sidhu, J. S. ; Omiecinski, Curtis John. / Baculovirus vectors repress phenobarbital-mediated gene induction and stimulate cytokine expression in primary cultures of rat hepatocytes. In: Gene Therapy. 2000 ; Vol. 7, No. 15. pp. 1274-1283.
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Baculovirus vectors repress phenobarbital-mediated gene induction and stimulate cytokine expression in primary cultures of rat hepatocytes. / Beck, N. B.; Sidhu, J. S.; Omiecinski, Curtis John.

In: Gene Therapy, Vol. 7, No. 15, 01.01.2000, p. 1274-1283.

Research output: Contribution to journalArticle

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