The Impact of H2-DM on Humoral Immune Responses

Abstract

H2-DM (DM, previously H2-M) facilitates the exchange of peptides bound to MHC class II molecules. In this study, we have used H2-DM-deficient (DM−/−) mice to analyze the influence of DM in the priming of B cell responses in vivo and for Ag presentation by B cells in vitro. After immunization, IgG Abs could be raised to a T-dependent Ag, 4-hydroxy-5-nitrophenylacetyl-OVA, in DM−/− mice, but closer analysis revealed the IgG response to be slower, diminished in titer, and composed of low-affinity Abs. The Ab response correlated with a vast reduction in the number of germinal centers in the spleen. The presentation of multiple epitopes by H2-Ab from distinct Ags was found to be almost exclusively DM-dependent whether B cells internalized Ags via fluid phase uptake or using membrane Ig receptors. The poor B cell response in vivo could be largely, but not completely restored by expression of a H2-Ead transgene, despite the fact that Ag presentation by H2-Ed/b molecules was found to be highly DM dependent. Hence, while substantial Ab responses can be raised in the absence of DM, this molecule is a crucial factor both for Ag processing and for the normal maturation of T-dependent humoral immune responses in vivo.

Increased antigen presenting cell‐mediated T cell activation in mice and patients without the autoimmune regulator

“…Aire -/-DC over-stimulate CD4 + T cell hybridomas Since VCAM-1 expression was increased on the surface of Aire -/-DC, we wanted to address whether the cause of increased T cell stimulation was in part due to VCAM-1 [49]. Naturally processed OVA by DC will create a low-affinity interaction with TCR hybridomas, since these peptide fragments are different from the trypsin-derived fragments.…”

“…We measured IL-2 secretion from three different T cell hybridoma lines (Ob2, Ob4 and Ob15) that recognize trypsin fragments of OVA protein presented by H2-IA b [49]. Naturally processed OVA by DC will create a low-affinity interaction with TCR hybridomas, since these peptide fragments are different from the trypsin-derived fragments.…”

“…Adherent cells were pulsed in triplicate with OVA for 1 h at 37 C followed by three washes with media. Incubation with T cell hybridomas recognizing different trypsin fragments of OVA (Ob2, Ob4 and Ob15) [49] (gift from Dr L. Karlsson) at 1 Â 10 5 cells/well was performed for 36 h. Blocking experiment was performed with the addition of 0.1 lg/well anti-VCAM-1 antibody or isotype control (BD Pharmingen). Supernatants were collected and analyzed for IL-2 secretion by ELISA (BD Pharmingen).…”

NADPH Oxidase Modifies Patterns of MHC Class II–Restricted Epitopic Repertoires through Redox Control of Antigen Processing

“…In vitro assessment of Ag processing and presentation (25,26) were stably transfected with a pcDNA3 construct that contained enhanced GFP (eGFP) under the control of a NFATenhanced IL-2 promoter (pNFATeGFP) as previously described (25)(26)(27)(28). Hence, upon productive Ag presentation with the hybridoma's cognate I-A b -restricted Ag, the hybridoma would express eGFP.…”

“…To further interrogate the ability of NOX2 to alter processing patterns of a single Ag, we measured the efficiency of WT and Cybb 2/2 BMMfs to process four distinct HEL epitopes (HEL [20][21][22][23][24][25][26][27][28][29][30][31][32][33][34][35] , HEL [31][32][33][34][35][36][37][38][39][40][41][42][43][44][45][46][47] , HEL [48][49][50][51][52][53][54][55][56][57][58][59][60][61][62] , and HEL 74-90 ) from whole HEL protein and present them to epitope-specific CD4 + T cell hybridomas (Hb1.9, H30.44, H46.13, and B04, respectively) (26). To achieve this, T cell hybridomas...…”