Identification of Schistosoma mansoni Antigens Recognized by T Cells of C57BL/6 Mice Immunized with Gamma-Irradiated Cercariae

Abstract

Immunization of C57Bl/6 mice with gamma-irradiated Schistosoma mansoni cercariae induces highly significant protection against subsequent challenge with unattenuated living cercariae. The antigens that evoke T cell-mediated immunity in this model are vaccine antigen candidates since T lymphocytes mediate antibody-independent immune responses and control the production of most antibodies. To identify these antigens, spleen cells of mice immunized twice with gamma-irradiated cercariae were tested for proliferative responsiveness and production of interleukins (Il) and interferon-gamma (IFN-gamma) following incubation with separated soluble schistosomular (SSA) and adult worm (SAWA) antigens in T cell western assays. Data obtained at 3, 4, 5, and 6 wk post-secondary immunization in 2 experiments indicated that SSA and SAWA bands of 62-60, 50, and 45 kDa reproducibly elicited T cell proliferation and production of Il-2, Il-4, and IFN-gamma by spleen cells from immunized, but not unimmunized, mice. Bands of 72-68, 29.5, and 28 kDa elicited proliferation and production of Il-2 and Il-4, but not IFN-gamma. Bands of 35-33 and 24 kDa induced either proliferation or Il production at some intervals.

A Schistosoma mansoni 62-KDA Band Is Identified as an Irradiated Vaccine T-Cell Antigen and Characterized as Calreticulin

“…BALB/ c mice were immunized twice, at 4-wk intervals, by tail exposure to 500-700 cercariae beginning within 30 min of irradiation. Antigen-free NC pieces (BO) were similarly treated and used as a negative control in every test (El Ridi et al, 1993, 1998Osman et al, 1994aOsman et al, , 1994b. T-cell western assays SAWA were prepared in the presence of protease inhibitors, resolved by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) under reducing conditions in 10% gels, and stained with Coomassie blue or electrotransferred onto nitrocellulose (NC) membrane.…”

“…At 3, 4, 5, and 6 wk after secondary immunization, spleen cells (SC) pooled from 5 or 6 immunized and 5 or 6 naive mice were enriched for T cells using paramagnetic beads coated with goat antibodies to mouse immunoglobulins (Advanced Magnetics Inc., Cambridge, Massachusetts) as described previously (El Ridi et al, 1993;Osman et al, 1994aOsman et al, , 1994b. SC were examined before and after fractionation for viability by trypan blue exclusion and for the percentage of CD4 and CD8 lymphocytes and macrophages by indirect membrane immunofluorescence (El Demellawy and El Ridi, 1992).…”

“…T-cell-activating antigens are required for antibody production. We have shown that the 62/60-and 50-kDa bands were the only ones recognized by both Swiss and C57BL/5 mice immunized twice with gamma-or ultraviolet-irradiated cercariae (El Ridi et al, 1993;Osman et al, 1994aOsman et al, , 1994b. Wide variability in the recognition of schistosome antigens was observed among the different strains.…”

“…Such immune sera have been used to identify and characterize potential vaccine antigens (Dalton et al, 1986(Dalton et al, , 1987Xu et al, 1989;Soisson et al, 1992). Hence, studies were conducted in several laboratories in an effort to identify the S. mansoni soluble adult worm antigen (SAWA) bands recognized by T lymphocytes of outbred Swiss (El Ridi et al, 1993) and inbred C57BL/6 (Reynolds et al, 1990;Mountford and Wilson, 1993;Richter et al, 1993;Osman et al, 1994aOsman et al, , 1994b and CBA (Richter et al, 1993) mice immunized twice with radiation-attenuated cercariae of S. mansoni. Hence, studies were conducted in several laboratories in an effort to identify the S. mansoni soluble adult worm antigen (SAWA) bands recognized by T lymphocytes of outbred Swiss (El Ridi et al, 1993) and inbred C57BL/6 (Reynolds et al, 1990;Mountford and Wilson, 1993;Richter et al, 1993;Osman et al, 1994aOsman et al, , 1994b and CBA (Richter et al, 1993) mice immunized twice with radiation-attenuated cercariae of S. mansoni.…”

Praziquantel binds Schistosoma mansoni adult worm actin

“…Approximately 2.5 × 10 5 peripheral blood mononuclear cells (PBMCs) were incubated with 200 L of medium-10% foetal calf serum (FCS) containing an antigen-free nitrocellulose (NC) piece or a NC piece to which were bound 2-3 g of the PZQ-45 kDa molecules, for 3 or 5 days for interleukins (ILs) and interferon-gamma (IFN-␥), respectively. This test is the T-cell Western assay and has been widely used in our laboratory and described by Osman et al [28,29]. Levels of IL-2, IL-4 and IFN-␥ in supernatants were evaluated by capture enzyme-linked immunosorbent assay (ELISA), using cytokine-specific capture and detecting (biotin-labelled) antibodies (Pharmingen, San Diego, CA), as described previously [30].…”