Fas-mediated apoptosis is involved in the elimination of gene-transduced hepatocytes with E1/E3-deleted adenoviral vectors

T. Okuyama, X. K. Li, N. Funeshima, M. Fujino, K. Sasaki, Y. Kita, M. Kosuga, M. Takahashi, Hidetsugu Saito, S. Suzuki, M. Yamada

Research output: Contribution to journalArticle

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Abstract

Gene-transduced hepatocytes with E1/E3-deleted adenoviral vectors are eliminated immediately and the expression of transduced genes disappears rapidly following the vector administration. In this report, we analysed the involvement of apoptotic cell death in the elimination of hepatocytes infected with adenoviral vectors. An E1/E3-deleted adenoviral vector expressing Escherichia coli β-galactosidase (LacZ) was injected via the portal vein into congenitally Fas-deficient mice (Ipr), Fas ligand-deficient mice (gld) and their control mice, MRL and C3H. 5-Bromo-4-chloro-3-indolyl- β-Dgalactoside (X-gal) staining of the liver specimens showed that 80-100% of hepatocytes were LacZ positive at 7 days after virus administration, suggesting that most of the hepatocytes received the injected adenoviral vectors. In normal mice, the number of LacZ-positive cells decreased dramatically at 14 and 21 days after transduction and few positive cells were observed at day 28. β-Galactosidase activity, quantified by the O- nitrophenyl-beta-D-galactopyranoside assay, gave comparable results to X-gal staining. At days 14 or 21, many apoptotic hepatocytes and apoptotic infiltrating cells were detected with the terminal deoxyribonucleotidyl transferase-mediated dUTP-digoxigenin nick end-labelling (TUNEL) in situ apoptosis detection method. This observation suggested that the apoptotic process was associated with the elimination of adenovirus-infected hepatocytes. To test the involvement of the Fas-Fas ligand interaction in this apoptotic process, the period of transgene expression was measured in lpr and gld mice, which had received the same amount of AxCALacZ. X-Gat histochemical analysis detected many LacZ-positive cells in lpr or gld mice liver even at 21 or 28 days after AxCALacZ injection. There were significant differences in the reduction rates of β-galactosidase activity of liver homogenates between lpr and MRL, or gld and C3H mice. Based on these observations, we conclude that the Fas-mediated apoptotic process is involved in the elimination of hepatocytes infected with E1/E3-deleted adenoviral vectors.

Original languageEnglish
JournalJournal of Gastroenterology and Hepatology (Australia)
Volume13
Issue numberSUPPL.
Publication statusPublished - 1998
Externally publishedYes

Fingerprint

Hepatocytes
Apoptosis
Galactosidases
Genes
Fas Ligand Protein
Inbred C3H Mouse
Liver
Nitrophenylgalactosides
Staining and Labeling
Digoxigenin
DNA Nucleotidylexotransferase
In Situ Nick-End Labeling
Portal Vein
Transgenes
Adenoviridae
Cell Death
Escherichia coli
Viruses
Gene Expression
Injections

Keywords

  • Fas-mediated apoptosis
  • Liver-directed gene therapy
  • Recombinant adenovirus

ASJC Scopus subject areas

  • Gastroenterology
  • Hepatology

Cite this

Fas-mediated apoptosis is involved in the elimination of gene-transduced hepatocytes with E1/E3-deleted adenoviral vectors. / Okuyama, T.; Li, X. K.; Funeshima, N.; Fujino, M.; Sasaki, K.; Kita, Y.; Kosuga, M.; Takahashi, M.; Saito, Hidetsugu; Suzuki, S.; Yamada, M.

In: Journal of Gastroenterology and Hepatology (Australia), Vol. 13, No. SUPPL., 1998.

Research output: Contribution to journalArticle

Okuyama, T, Li, XK, Funeshima, N, Fujino, M, Sasaki, K, Kita, Y, Kosuga, M, Takahashi, M, Saito, H, Suzuki, S & Yamada, M 1998, 'Fas-mediated apoptosis is involved in the elimination of gene-transduced hepatocytes with E1/E3-deleted adenoviral vectors', Journal of Gastroenterology and Hepatology (Australia), vol. 13, no. SUPPL..
Okuyama, T. ; Li, X. K. ; Funeshima, N. ; Fujino, M. ; Sasaki, K. ; Kita, Y. ; Kosuga, M. ; Takahashi, M. ; Saito, Hidetsugu ; Suzuki, S. ; Yamada, M. / Fas-mediated apoptosis is involved in the elimination of gene-transduced hepatocytes with E1/E3-deleted adenoviral vectors. In: Journal of Gastroenterology and Hepatology (Australia). 1998 ; Vol. 13, No. SUPPL.
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abstract = "Gene-transduced hepatocytes with E1/E3-deleted adenoviral vectors are eliminated immediately and the expression of transduced genes disappears rapidly following the vector administration. In this report, we analysed the involvement of apoptotic cell death in the elimination of hepatocytes infected with adenoviral vectors. An E1/E3-deleted adenoviral vector expressing Escherichia coli β-galactosidase (LacZ) was injected via the portal vein into congenitally Fas-deficient mice (Ipr), Fas ligand-deficient mice (gld) and their control mice, MRL and C3H. 5-Bromo-4-chloro-3-indolyl- β-Dgalactoside (X-gal) staining of the liver specimens showed that 80-100{\%} of hepatocytes were LacZ positive at 7 days after virus administration, suggesting that most of the hepatocytes received the injected adenoviral vectors. In normal mice, the number of LacZ-positive cells decreased dramatically at 14 and 21 days after transduction and few positive cells were observed at day 28. β-Galactosidase activity, quantified by the O- nitrophenyl-beta-D-galactopyranoside assay, gave comparable results to X-gal staining. At days 14 or 21, many apoptotic hepatocytes and apoptotic infiltrating cells were detected with the terminal deoxyribonucleotidyl transferase-mediated dUTP-digoxigenin nick end-labelling (TUNEL) in situ apoptosis detection method. This observation suggested that the apoptotic process was associated with the elimination of adenovirus-infected hepatocytes. To test the involvement of the Fas-Fas ligand interaction in this apoptotic process, the period of transgene expression was measured in lpr and gld mice, which had received the same amount of AxCALacZ. X-Gat histochemical analysis detected many LacZ-positive cells in lpr or gld mice liver even at 21 or 28 days after AxCALacZ injection. There were significant differences in the reduction rates of β-galactosidase activity of liver homogenates between lpr and MRL, or gld and C3H mice. Based on these observations, we conclude that the Fas-mediated apoptotic process is involved in the elimination of hepatocytes infected with E1/E3-deleted adenoviral vectors.",
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AU - Okuyama, T.

AU - Li, X. K.

AU - Funeshima, N.

AU - Fujino, M.

AU - Sasaki, K.

AU - Kita, Y.

AU - Kosuga, M.

AU - Takahashi, M.

AU - Saito, Hidetsugu

AU - Suzuki, S.

AU - Yamada, M.

PY - 1998

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N2 - Gene-transduced hepatocytes with E1/E3-deleted adenoviral vectors are eliminated immediately and the expression of transduced genes disappears rapidly following the vector administration. In this report, we analysed the involvement of apoptotic cell death in the elimination of hepatocytes infected with adenoviral vectors. An E1/E3-deleted adenoviral vector expressing Escherichia coli β-galactosidase (LacZ) was injected via the portal vein into congenitally Fas-deficient mice (Ipr), Fas ligand-deficient mice (gld) and their control mice, MRL and C3H. 5-Bromo-4-chloro-3-indolyl- β-Dgalactoside (X-gal) staining of the liver specimens showed that 80-100% of hepatocytes were LacZ positive at 7 days after virus administration, suggesting that most of the hepatocytes received the injected adenoviral vectors. In normal mice, the number of LacZ-positive cells decreased dramatically at 14 and 21 days after transduction and few positive cells were observed at day 28. β-Galactosidase activity, quantified by the O- nitrophenyl-beta-D-galactopyranoside assay, gave comparable results to X-gal staining. At days 14 or 21, many apoptotic hepatocytes and apoptotic infiltrating cells were detected with the terminal deoxyribonucleotidyl transferase-mediated dUTP-digoxigenin nick end-labelling (TUNEL) in situ apoptosis detection method. This observation suggested that the apoptotic process was associated with the elimination of adenovirus-infected hepatocytes. To test the involvement of the Fas-Fas ligand interaction in this apoptotic process, the period of transgene expression was measured in lpr and gld mice, which had received the same amount of AxCALacZ. X-Gat histochemical analysis detected many LacZ-positive cells in lpr or gld mice liver even at 21 or 28 days after AxCALacZ injection. There were significant differences in the reduction rates of β-galactosidase activity of liver homogenates between lpr and MRL, or gld and C3H mice. Based on these observations, we conclude that the Fas-mediated apoptotic process is involved in the elimination of hepatocytes infected with E1/E3-deleted adenoviral vectors.

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