Successive tick infestations selectively promote a T-helper 2 cytokine profile in mice

Abstract

Several studies have revealed that T lymphocytes and cytokines play a crucial role in determining the outcome of parasitic infections in terms of protective immunity. In this study we found that Rhipicephalus sanguineus tick saliva stimulates transforming growth factor-beta (TGF-beta), and reduces interleukin-12 (IL-12) secretion by cells from normal C3H/HeJ mice. Moreover, murine lymph node cells harvested 6 days after the fourth infestation with ticks presented an 82.4% decrease in their proliferative response to concanavalin A (Con A) compared with the response of control cells. In addition, lymph node cells cultured in the presence of Con A showed a T-helper 2-type (Th2-type) cytokine profile, represented by augmented IL-4 and IL-10 and TGF-beta. On the other hand, the IL-2, interferon-gamma (IFN-gamma) and IL-12 synthesis was significantly inhibited. These results indicate that ticks may modulate the host's immune response through saliva injection. Considering that C3H/HeJ mice develop no protective immunity to R. sanguineus infestation, our results suggest that tick-induced Th2-type cytokines and a decreased proliferative response probably lead the host to a susceptible state to both tick and tick-transmitted pathogens.

Figure 1

Effect of R. sanguineus saliva on Trypanosoma cruzi-induced IL-12 (a) and TGF-β (b) synthesis by normal mouse cells. Mouse macrophages or splenocytes were cultured in 24-well plates (2×10⁶ cells per well) in the presence of T. cruzi (Tc) (one parasite per cell) and/or tick saliva (64 μg/ml). Supernatants were collected after 48 hr and assayed for IL-12 and TGF-β by ELISA. Results are shown as mean cytokine production ± SD. *P < 0·05 and **P < 0·001compared with cells cultured with T. cruzi parasites alone. The experiments were repeated three times.

Figure 2

Effect of R. sanguineus tick infestations on Con A-induced cell proliferation in mice. Lymph node cells from mice infested once (1×Inf), four times (4×Inf) or non-infested (Control) with ticks were cultured (5×10⁵ cells per well) with or without Con A (2 μg/ml) and assayed for cell proliferation by methyl-[³H]-thymidine incorporation. A stimulation index was calculated for each experimental group as described in the Material and Methods. Results are shown as mean of Stimulation Index ± SD from two independent experiments.

Figure 3

Effect of repeated tick infestations with R. sanguineus on the production of IL-2 (a) and IFN-γ (b) by mice. Lymph node cells from mice infested once (1×Inf) or four times (4×Inf) with ticks were cultured (2×10⁶ cells per well) with or without Con A (2 μg/ml). After 48 hr the supernatants were collected and assayed by specific ELISA for IL-2 and IFN-γ. *P < 0·05 and **P < 0·001 compared with cells from control mice plus Con A. The experiments were performed in duplicate and repeated three times.

Figure 4

Effect of repeated tick infestations with R. sanguineus ticks on the production of IL-4 (a) and IL-10 (b) by mice. Lymph node cells from mice infested once (1×Inf) or four times (4×Inf) with ticks were cultured (2×10⁶ cells per well) with or without Con A (2 μg/ml). After 48 hr the supernatants were collected and assayed by specific ELISA for IL-4and IL-10. *P < 0·05 and **P < 0·001 compared with cells from control mice plus Con A. The experiments were performed in duplicate and repeated three times.

Figure 5

Effect of repeated tick infestations with R. sanguineus on the production of IL-12 by mice. Lymph node cells from mice infested once (1×Inf) or four times (4×Inf) with ticks were cultured (2×10⁶ cells per well) with or without Con A (2 μg/ml). After 48 hr the supernatants were collected and were assayed by specific ELISA for IL-12. **P < 0·001 compared with cells from control mice plus Con A. The experiments were performed in duplicate and repeated three times.

Figure 6

Effect of repeated tick infestations with R. sanguineus on the production of TGF-β by mice. Lymph node cells from mice infested once (1×Inf) or four times (4×Inf) with ticks were cultured (2×10⁶ cells per well) with or without Con A (2 μg/ml). After 48 hr the supernatants were collected and assayed by specific ELISA for TGF-β. *P < 0·05 and **P < 0·001 compared with cells from control mice plus Con A. The experiments were performed in duplicate and repeated three times.