Schistosome infection stimulates host CD4(+) T helper cell and B-cell responses against a novel egg antigen, thioredoxin peroxidase - PubMed (original) (raw)

Schistosome infection stimulates host CD4(+) T helper cell and B-cell responses against a novel egg antigen, thioredoxin peroxidase

D L Williams et al. Infect Immun. 2001 Feb.

Abstract

Egg granuloma formation during schistosome infections is mediated by CD4(+) T helper (Th) cells sensitized to egg antigens; however, most of the relevant sensitizing egg antigens are still unknown. Here we show that schistosome thioredoxin peroxidase (TPx)-1 is a novel T- and B-cell egg antigen in schistosome-infected mice. CD4(+) Th cell responses to fractionated egg components identified a significant response against a 26-kDa antigen; a partial amino acid sequence of this antigen was found to be identical to that of Schistosoma mansoni TPx-1. The native TPx-1 elicited significant proliferative responses as well as gamma interferon (IFN-gamma), interleukin-2 (IL-2), IL-4, and IL-5 secretion in CD4(+) cells from 8.5-week-infected CBA and C57BL/6 mice. By comparison, recombinant TPx-1 elicited a smaller, more type 1-polarized response, with significant production of IFN-gamma and IL-2, less IL-5, and essentially no IL-4. In C57BL/6 mice the responses to TPx-1 were relatively more prominent than that directed against the major egg antigen, Sm-p40, whereas in CBA mice the reverse was true. B-cell responses were also monitored in infected C57BL/6, C3H, CBA, and BALB/c mice. All strains had significant antibody levels against the TPx-1 protein, but the most significant antibody production ensued following parasite oviposition. TPx-1 was localized in eggs and shown to be secreted by eggs. The identification of egg antigens is important to understand the specific basis of granuloma formation in schistosome infections and may prove to be useful in strategies to ameliorate pathological responses.

PubMed Disclaimer

Figures

FIG. 1

Identification of a 26-kDa antigen recognized by CD4+ Th cells from schistosome-infected BL/6 mice and the N-terminal amino acid sequence obtained from the 26-kDa fraction. The eluted SDS-PAGE fraction containing the 26-kDa component (Fr. E22) (3) from 20 μg of SEA was examined for purity on a silver-stained SDS–15% polyacrylamide gel and is shown next to total SEA and molecular mass marker standards.

FIG. 2

Proliferative response of CD4+ Th cells from BL/6 and CBA mice to the 26-kDa antigen. CD4+ Th cells were isolated from mesenteric lymph nodes of 8.5-week-infected mice. Culture conditions were as described in Materials and Methods. [3H]dThd incorporation was assessed by liquid scintillation spectroscopy. Data are expressed as the mean ± 1 standard deviation. Also shown for comparison are responses to Sm-p40 and SEA. Background radioactivity from cultures in the absence of antigen is subtracted. The same patterns of stimulation were observed when cells from 7.5-week-infected mice were used (not shown). The experiments were repeated at least three times with similar results.

FIG. 3

Cytokine production by CD4+ Th cells from BL/6 and CBA mice stimulated with the 26-kDa antigen. CD4+ Th cells were isolated from mesenteric lymph nodes of 8.5-week-infected mice and cultured with the indicated antigens in the presence of APC as described in Materials and Methods. The cytokines were measured in culture supernatants by ELISA. Data are expressed as means of triplicate determination ± 1 standard deviation. Also shown for comparison are responses to Sm-p40 and SEA. The experiments were repeated at least three times with similar results.

FIG. 4

Proliferative responses of CD4+ Th cells from infected BL/6 and CBA mice to rTPx-1. CD4+ Th cells were isolated from mesenteric lymph nodes of 8.5-week-infected mice and incubated with the indicated antigens in the presence of APC as described in Materials and Methods. [3H]dThd incorporation was assessed by liquid scintillation spectroscopy. Also shown for comparison are responses to Sm-p40 and SEA. Background radioactivity from cultures in the absence of antigen is subtracted. The same patterns of stimulation were observed when cells from 8 -week-infected mice were used (not shown). The experiments were repeated at least three times with similar results.

FIG. 5

Cytokine production by CD4+ Th cells from BL/6 and CBA mice stimulated with rTPX-1. CD4+ Th cells were isolated from mesenteric lymph nodes of 8.5-week-infected mice and cultured with the indicated antigens in the presence of APC as described in Materials and Methods. The cytokines were measured in culture supernatants by ELISA. Data are expressed as means of triplicate determinations ± 1 standard deviation. Also shown for comparison are responses to Sm-p40 and SEA. The experiments were repeated at least three times with similar results.

FIG. 6

B-cell response to schistosome TPx-1. Sera were obtained from mice at the times indicated after infection. Anti-TPx-1 antibodies in the sera were measured by ELISA using microtiter plates coated with recombinant S. mansoni TPx-1. Data are expressed as means of triplicate determinations ± standard error. Sera were from BL/6 mice (closed bars) and C3H mice (open bars). OD, optical density.

FIG. 7

Identification of TPx-1 protein in egg homogenates and secretory products. SDS–16% polyacrylamide gels were Coomassie blue stained (lanes 1 to 4) or transferred to membranes and probed with affinity-purified anti-S. mansoni TPx-1 antibodies (lanes 5 to 8). Lanes 1 and 5, 10 μg of adult male homogenate; lanes 2 and 6, 10 μg of adult female homogenate; lanes 3 and 7. 10 μg of egg homogenate; lanes 4 and 8, 3.75 μg of cultured egg secretory products. Protein size standards are shown on the left of each gel in kilodaltons. The figure was produced from scanned gels or membranes using Adobe Illustrator.

FIG. 8

Immunohistochemical localization of TPx-1 in schistosome eggs. A fluorescence micrograph of a 5-μg egg cryosection is shown. Green fluorescence surrounding the egg is due to autofluorescence of the eggshell. Orange-red Cy3-labeled secondary antibody fluorescence is found in the von Lichtenberg envelope surrounding the miracidium and underneath the eggshell. DAPI-stained nuclei of the miracidium appear blue. The image is an overlay of three separate color images created in Adobe Photoshop. No specific Cy3 fluorescence was seen when preimmune serum was used for staining (not shown).

References

1. Abdallahi O M S, Hanna S, De Reggi M, Gharib B. Visualization of oxygen radical production in mouse liver in response to infection with Schistosoma mansoni. Liver. 1999;19:495–500. - PubMed
1. Araki M, Nanri H, Ejima K, Murasato Y, Fujiwara T, Nakashima Y, Ikeda M. Antioxidant function of the mitochondrial protein SP-22 in the cardiovascular system. J Biol Chem. 1999;274:2271–2278. - PubMed
1. Asahi H, Hernandez H J, Stadecker M J. A novel 62-kilodalton egg antigen from Schistosoma mansoni induces a potent CD4+ T helper cell response in the C57BL/6 mouse. Infect Immun. 1999;67:1729–1735. - PMC - PubMed
1. Asahi H, Osman A, Cook R M, LoVerde P T, Stadecker M J. Schistosoma mansoni phosphoenolpyruvate carboxykinase (Sm-PEPCK), a novel egg antigen: immunological properties of the recombinant protein and identification of T-cell epitope. Infect Immun. 2000;68:3385–3393. - PMC - PubMed
1. Boros D L, Warren K S. Delayed hypersensitivity-type granuloma formation and dermal reaction induced and elicited by a soluble factor isolated from Schistosoma mansoni eggs. J Exp Med. 1970;132:488–507. - PMC - PubMed

Schistosome infection stimulates host CD4(+) T helper cell and B-cell responses against a novel egg antigen, thioredoxin peroxidase - PubMed (original) (raw)