Respiratory tract gene transfer

Transplantation of genetically modified T-lymphocytes directly to the respiratory epithelial surface

M. Fukayama, T. Kanno, S. L. Brody, M. Kirby, Ronald Crystal

Research output: Contribution to journalArticle

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Abstract

To evaluate the strategy for potentially treating respiratory disorders with genetically modified T-lymphocytes, the interleukin-2 (IL-2)-dependent murine T-cell line, CTLL2, was genetically altered with the Escherichia coli β-galactosidase (β-gal) gene (lacZ) in vitro with a retroviral vector and the modified T-cells were transplanted directly to the respiratory epithelial surface of syngeneic C57Bl/6 mice. Southern and Northern analyses confirmed that the neomycin-selected modified T-cells contained and expressed the lacZ gene. The fate of the modified T-cells (CTLL2/lacZ) was followed by flow cytometry with T-cell surface marker Thy 1.2 and fluorescent β-gal analysis. One day after transplantation (7.5 x 105 CTLL2/lacZ T-cells/g of body weight), 95 ± 3% of the Thy 1.2+ T-cells recovered from respiratory epithelial lining fluid (ELF) were β-gal+. Importantly, the modified T-cells remained in the lung for some time; at 3 days, Thy 1.2+ β-gal+ T-cells represented 63 ± 12% of ELF Thy 1.2+ T-cells and 59 ± 6% of Thy 1.2+ T-cells recovered from the whole lung. At 7 days, 33 ± 8% of the Thy 1.2+ cells in ELF and 75 ± 6% of the Thy 1.2+ cells in whole lung were Thy 1.2+β-gal+. In contrast, the proportion of the Thy 1.2+ β-gal+ T-cells in the spleen, the major extrapulmonary lymphatic organ, never rose above 3 ± 1% of the total Thy1.2+ cells. The number of Thy 1.2+ β-gal+ T-cells in the lung could be modified by the systemic administration of IL-2, with whole lung Thy 1.2+ β-gal+ T-cells increasing 4.6-fold 3 days after transplantation, compared with non-IL-2-treated animals. These studies suggest that direct transplantation of genetically modified T-cells into the lung is feasible and represents a viable strategy for lung-specific gene transfer.

Original languageEnglish
Pages (from-to)18339-18344
Number of pages6
JournalJournal of Biological Chemistry
Volume266
Issue number27
Publication statusPublished - 8 Nov 1991
Externally publishedYes

Fingerprint

Gene transfer
T-cells
Respiratory System
Transplantation
T-Lymphocytes
Genes
Lung
Linings
Lac Operon
Interleukin-2
Fluids
Galactosidases
Neomycin
Flow cytometry

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Respiratory tract gene transfer : Transplantation of genetically modified T-lymphocytes directly to the respiratory epithelial surface. / Fukayama, M.; Kanno, T.; Brody, S. L.; Kirby, M.; Crystal, Ronald.

In: Journal of Biological Chemistry, Vol. 266, No. 27, 08.11.1991, p. 18339-18344.

Research output: Contribution to journalArticle

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abstract = "To evaluate the strategy for potentially treating respiratory disorders with genetically modified T-lymphocytes, the interleukin-2 (IL-2)-dependent murine T-cell line, CTLL2, was genetically altered with the Escherichia coli β-galactosidase (β-gal) gene (lacZ) in vitro with a retroviral vector and the modified T-cells were transplanted directly to the respiratory epithelial surface of syngeneic C57Bl/6 mice. Southern and Northern analyses confirmed that the neomycin-selected modified T-cells contained and expressed the lacZ gene. The fate of the modified T-cells (CTLL2/lacZ) was followed by flow cytometry with T-cell surface marker Thy 1.2 and fluorescent β-gal analysis. One day after transplantation (7.5 x 105 CTLL2/lacZ T-cells/g of body weight), 95 ± 3{\%} of the Thy 1.2+ T-cells recovered from respiratory epithelial lining fluid (ELF) were β-gal+. Importantly, the modified T-cells remained in the lung for some time; at 3 days, Thy 1.2+ β-gal+ T-cells represented 63 ± 12{\%} of ELF Thy 1.2+ T-cells and 59 ± 6{\%} of Thy 1.2+ T-cells recovered from the whole lung. At 7 days, 33 ± 8{\%} of the Thy 1.2+ cells in ELF and 75 ± 6{\%} of the Thy 1.2+ cells in whole lung were Thy 1.2+β-gal+. In contrast, the proportion of the Thy 1.2+ β-gal+ T-cells in the spleen, the major extrapulmonary lymphatic organ, never rose above 3 ± 1{\%} of the total Thy1.2+ cells. The number of Thy 1.2+ β-gal+ T-cells in the lung could be modified by the systemic administration of IL-2, with whole lung Thy 1.2+ β-gal+ T-cells increasing 4.6-fold 3 days after transplantation, compared with non-IL-2-treated animals. These studies suggest that direct transplantation of genetically modified T-cells into the lung is feasible and represents a viable strategy for lung-specific gene transfer.",
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