Characterization of HSV-1 antigens expressed on human Langerhans cells

Abstract

Langerhans cells (LC) are immunocompetent antigen presenting cells (APC) which are involved with many infections including those caused by herpes simplex virus (HSV). Viral glycoproteins (g) on the surface of infected cells have been studied in established cell lines. The identification of which viral g participate with LC processing and antigen presentation on APC has not been studied. It is hypothesized that specific HSV-1 viral g species can be identified on the surface of LC very early after viral challenge of LC and are important antigens (Ag) involved in the human immune response to this virus. The purpose of this investigation has been to elucidate the expression of HSV-1 Ag, in vitro, using freshly isolated human LC. These experiments have established techniques for the isolation and enrichment of human LC that retain for up to 18 h post enrichment all of the ultrastructural characteristics and cell markers consistent with those of LC. These LC have been shown to adsorb and replicate HSV-1 virus at a low copy number. Immunofluorescence and ELISA have demonstrated the presence of viral antigens on the surface of membrane fractions of LC 8 h post infection using polyvalent antiserum. Specific viral epitopes related to gD and gE have been identified for the first time on HSV-1 challenged LC at 5 h post infection by two complementary techniques. SDS-PAGE of virally challenged LC membranes demonstrated the emergence of proteins consistent with the electrophoretic mobilities of gD, gE and possibly gH as early as 5 h post infection. Immunoblotting (IB) with monoclonal antibodies (Mab) confirmed the presence of these g in membrane preparations. Dual colloidal gold immunolabelling for transmission electronmicroscopy (TEM), utilizing Mab to LC markers and to epitopes found on viral g demonstrated visual confirmation of the presence of sites reactive with antibodies to gD and gE on the surface of LC 5 h after viral challenge. Although present later in infection larger proteins gB and gC do not appear to be involved in early membrane changes of LC. An epitope of gH was identified by IB, however, visual confirmation by TEM was not accomplished. The 5 h post infection LC, therefore, selectively express epitopes of gD, gE, and gH which may be involved in immunoactivation, particularly in light of the evidence supporting gD as a potent stimulator of neutralizing antibodies. Presentation of such viral antigens by LC would be an early step in this process.