Internalization and acidification of surface HLA-DR molecules by epidermal Langerhans cells: a paradigm for antigen processing

Abstract

CD4+ T lymphocytes recognize multi-molecular complexes, formed by major histocompatibility complex class II molecules and exogenous antigens, on the surface of antigen-presenting cells (APC). For most protein antigens, processing is required to produce immunogenic peptide fragments that can then form stable associations with class II molecules. These two processes, the modification of antigen and its coupling to class II molecules, are thought to occur in acidic endosomal compartments. Furthermore, membrane class II molecules are endocytosed in APC and may provide ligands for the immunogenic peptides. To gain insight into these processes, we examined the internalization and acidification of membrane HLA-DR molecules by three APC populations: 1) freshly isolated Langerhans cells (LC), 2) LC after 48-72 h of bulk epidermal cell culture, and 3) peripheral blood monocytes (PBM). Using FITC-conjugated anti-HLA-DR monoclonal antibodies (MoAb), endocytosis was studied by fluorescence microscopy and by flow cytometry (pulse width analysis), while acidification was assessed by exploiting the pH sensitivity of fluorescein fluorescence. We observed both freshly isolated LC and PBM to internalize surface HLA-DR molecules into acidic compartments with great efficiency. Endocytosis was inhibited by the addition of azide and 2-deoxy-D-glucose, whereas acidification was partially blocked by treatment with ammonium chloride or chloroquine. The degree of internalization and acidification of HLA-DR molecules was greatly influenced by the degree of Ab cross-linking. On the other hand, cultured LC were capable of internalizing HLA-DR molecules, but were not able to acidify the environments to which these molecules were delivered; this loss of acidification capacity was partially restored by treatment with phorbol 12-myristate 13-acetate.(ABSTRACT TRUNCATED AT 250 WORDS)