In vitro studies on immunoglobulin transcytosis Fc receptor characterisation and specific antigen expression in rabbit yolk sac endoderm

Meads, Timothy James (1992) In vitro studies on immunoglobulin transcytosis Fc receptor characterisation and specific antigen expression in rabbit yolk sac endoderm. University of Southampton, Doctoral Thesis.

Abstract

Previous in vivo studies have shown that rabbit and human IgG are selectively transcytosed across the endodermal cell layer of the rabbit yolk sac splanchnopleur in an Fe receptor-mediated manner.

This transcytosis has been characterised in the present study in vitro, by culturing the intact yolk sac membrane in such a way that only the apical endodermal cell surface was exposed to IgG. The incubation of the intact membrane was necessitated by the failure of dissociated endodermal cells to reconstitute a differentiated polarised epithelium in vitro.

Human IgG, detected by immunofluorescence, was selectively transcytosed across the endodermal cell to the basement membrane and vascularised mesenchyme. Bovine IgG, added simultaneously with human IgG, remained within the endodermal cell, and was not transported. Human IgG transcytosis was also Fc mediated for whilst the Fc piece was transcytosed, Fab fragments remained confined to the endoderm. Transcytosis of human IgG occurred at 20^oC, albeit at a slow rate, and continued when the endodermal cell was exposed simultaneously to 10μM monensin, indicating that selective transcytosis is unlikely to be dependent upon acidified endocytic compartments. The extent of transcytosis was reduced if the endodermal cells were pre-exposed for 5 minutes to monensin before the addition of human IgG, and totally inhibited by 30 minutes pre-exposure to monensin. Human IgG also bound specifically to the apical endodermal cell surface chilled at 4^oC, indicating that the Fc receptor for IgG is probably expressed at the apical endodermal cell surface. Specific transcytosis of human IgM across the rabbit yolk sac endodermal cell has also been detected, but it occurs more slowly than transcytosis of human IgG. The ultrastructural route of IgG transcytosis has been studied both by exposing the endodermal cell to rabbit IgG adsorbed to colloidal gold, and by localising endogenous IgG in ultrathin sections of the yolk sac membrane. In both studies, IgG was detected within coated vesicles at the lateral endodermal cell plasma membrane, consistent with previous in vivo studies, but the nature of the selective event during transcytosis could not be addressed.

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