Expression of Retroviral Transduced Human CD18 in Murine Cells: AnIn VitroModel of Gene Therapy for Leukocyte Adhesion Deficiency

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Leukocyte adhesion deficiency (LAD) is an autosomal recessive disease caused by a defective CD18 gene. The cell-surface glycoprotein encoded by this gene CD18 is normally expressed in cells of the hematopoietic system. An in vitro murine model of CD18 gene replacement therapy was developed to investigate the feasibility of an in vivo murine hematopoietic stem cell gene therapy model. Human CD18-transducing retroviruses were used to transfer a functional human CD18 gene into a variety of cells including (i) murine lymphoblasts (which express murine CD11a and murine CD18), (ii) murine fibroblasts (which have no endogenous murine CD11a/CD18 expression), and (iii) murine fibroblasts, which have been stably transfected with a human CD11a gene. In murine lymphoblasts, human CD18 was expressed on the cell surface as a heterodimer with murine CD11a. Cell-surface expression of human CD18 had no apparent effect on the level of endogenous murine CD11a/CD18 expression. Immunoprecipitation of cell-surface labeled proteins in murine lymphoblasts with a human CD18 specific antibody co-precipitated murine CD11a. Human CD18 can be detected by immunochemistry in the cytoplasm of fibroblasts infected with CD18 encoding retrovirus, but coexpression with CD11a is required for cell-surface expression of either subunit in fibroblasts. These studies suggest that human CD18 will form a heterodimer with murine CD11a and that human CD18 is not expressed on the cell surface of cells not expressing CD11. This provides the basis for the development of a murine hematopoietic stem cell gene replacement therapy model for the treatment of LAD.