Impaired protective immunity and T helper 2 responses in alymphoplasia (aly) mutant mice infected with Trichinella spiralis

Abstract

The alymphoplasia (aly) mutation of mice prevents the development of systemic lymph nodes and Peyer's patches. The mutant homozygotes (aly/aly) are partially deficient in both humoral and cell-mediated immune functions. In the present study, we show that adult worm expulsion was slightly delayed and that T helper 2 (Th2)-type responses were partially defective in aly/aly mice after infection with Trichinella spiralis. Male aly/aly and aly/+ mice (8-weeks old) were infected with 400 muscle larvae. There was no difference in worm recovery between the two groups on day 5. However, worm recovery in aly/aly mice was significantly higher than that in aly/+ mice on day 14. Mucosal mast cells increased in number and peaked 14 days after infection in aly/+ mice. aly/aly mice were deficient in their mucosal mast cell response through out the primary infection. To examine the existence of mast cell precursors, aly/aly mice were treated with recombinant interleukin-3 (rIL-3) before infection. The mast cell response was poorly induced in aly/aly mice treated with rIL-3. An immunoglobulin E (IgE) response was not detected in aly/aly mice during the course of infection. Serum IgG1 levels in aly/aly mice were significantly lower than that of aly/+. The serum IgG2a levels increased in both strains of mice. However, IgG2a production in aly/aly mice on day 14 was half as much as that in aly/+mice. Stimulation of splenic T cells in vitro with anti-CD3 monoclonal antibody (mAb) showed that spleen cells from aly/+ mice on day 5 produced more IL-4 than spleen cells from aly/aly mice. IL-4 production from aly/aly mice on day 14 was half that from aly/+ mice. Interferon-gamma (IFN-gamma) was produced in both aly/aly and aly/+ mice on day 14. Proliferation assay showed that T cells of aly/aly mice responded poorly when cultured with antigen-presenting cells. These results suggest that aly gene is needed for the induction of protective immunity and Th2 responses in mice infected with T. spiralis.

Figure 1

Kinetics of mucosal mast cell responses in aly/aly and aly/+mice infected with Trichinella spiralis. Mice were infected with 400 muscle larvae. Values represent mean±se from four mice. (○) aly/aly; (•) aly/+. ***P < 0·001.

Figure 2

Mucosal mast cell responses in aly/aly and aly/+mice treated with rIL-3 before infection with Trichinella spiralis. Mice were injected intraperitoneally with a total of 10 000 units rIL-3 or saline for 5 days before infection with 400 muscle larvae. Values represent means±se from three mice. NE; not examined. Significantly different from aly/+ values. *P < 0·05.

Figure 3

Immunoglobulin levels in aly/aly and aly/+mice infected with Trichinella spiralis. Total serum IgE, IgG1, and IgG2a levels were detected with ELISA. Values represent means±se from three to four mice. ND; not detected (IgE < 30 ng/ml, IgG1 < 2 µg/ml, IgG2a < 200 ng/ml). Significantly lower than aly/+ values. *P < 0·05.

Figure 4

IL-4 and IFN-γ production of splenic T cells from aly/aly and aly/+mice infected with Trichinella spiralis. Spleen cells were separated into subsets using mAb and magnetic beads. Pooled cells (6·5 × 10⁵/150 µl/well) were cultured in 96-well plate with anti-CD3 mAb for 48 hr. Cytokines in the culture supernatant were detected by ELISA. Values represent means from duplicates. NE; not examined. Significantly different from aly/+ values. *P < 0·05.

Figure 5

Proliferation of splenic T cells cultured with antigen-presenting cells. aly/aly and aly/+mice were infected with Trichinella spiralis. The spleens were removed on day 8 after infection. After treatment with mitomycin C, 0·5 × 10⁵/well plastic adherent cells were used as antigen-presenting cells. T-enriched cells were prepared with mAb and magnetic microbeads. Cells (2 × 10⁵/200 µl/well) were cultured with antigen-pulsed or nonpulsed adherent cells for 4 days. Cell growth was measured with TetraColor ONE®. Values represent (absorbence of antigen-pulsed cells) − (absorbence of non-pulsed cells) from duplicates. *P < 0·05; **P < 0·01.