Biochemical properties of MHC class II molecules endogenously synthesized and expressed by mouse Langerhans cells.
The cell surface expression and biosynthesis of Langerhans cells (LC)-derived major histocompatibility complex (MHC) class II molecules from epidermal cells (EC) prepared freshly and cultured for up to 3 days was investigated. Based on the constitutive expression of MHC class II determinants by LC, a panning and magnetic bead selection procedure was employed, yielding 65% and 86% of I-A+ cells, respectively. Phenotypical and cytochemical examinations revealed that the two LC preparations were free of contaminating macrophages as well as B and T cells. Freshly prepared enriched LC were highly efficient in the stimulation of protein antigen-specific T cell clones, while LC purified from short-term cultured EC suspensions proved to be more efficient allogeneic stimulator cells than fresh LC. Comparative analysis of LC obtained from freshly prepared and from short-term-cultured EC preparations indicated an up-regulation of MHC class II determinants during short-term culture. Radioiodination analysis of LC selected by magnetic beads demonstrated prominent class II alpha and beta chain signals with only a minute fraction of invariant chains p35 and p45 being expressed at the cell surface. Unlike class II complexes derived from B cells, those from LC contained invariant chain fragment p20 in association with alpha/beta heterodimers at the plasma membrane. No qualitative differences between freshly isolated and 3-day cultured LC in cell surface expressed MHC class II components were detectable. Metabolic labeling with subsequent two-dimensional electrophoresis revealed distinct features of LC-derived MHC class II molecules with a high proportion of invariant chains in particular gamma and p40 and their extensive sialylation. While fresh and 1-day cultured LC exhibited appreciable levels of newly synthesized class II molecules, a dramatic down-regulation in class II and invariant chain synthesis was measured after 3 days of continuous in vitro culture.
Becker
D
D
Institut für Immunologie der Joh. Gutenberg Universität, Mainz, FRG.
Reske-Kunz
A B
AB
Knop
J
J
Reske
K
K
D000818:Animals; D002478:Cells, Cultured; D005260:Female; D005434:Flow Cytometry; D000949:Histocompatibility Antigens Class II / Q000032:analysis* / Q000096:biosynthesis; D007457:Iodine Radioisotopes; D007801:Langerhans Cells / Q000276:immunology*; D008222:Lymphokines / Q000494:pharmacology; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008809:Mice, Inbred C3H; D010641:Phenotype
D000949:Histocompatibility Antigens Class II; D007457:Iodine Radioisotopes; D008222:Lymphokines