Expressions of Toll-like receptors 2 and 4 in mouse skin during early immune responses against Sporothrix

Abstract

Objective To explore the expressions of Toll-like receptors (TLRs) 2 and 4 in mouse skin during early immune responses against Sporothrix. Methods A total of 60 BALB/c mice were randomly and equally divided into an experimental group and a control group to be intracutaneously injected with Sporothrix conidium suspensions at a concentration of 1 × 106 cfu/ml and sodium chloride physiological solution respectively. Five mice were sacrificed before the injection, and at 6, 12, 24, 48, and 96 hours after the injection in each group, blood samples were obtained from the mice before sacrifice, and skin tissue specimens were resected from the area around the injection sites after sacrifice. Real-time fluorescence-based quantitative PCR was performed to quantify the mRNA expressions of TLR2 and TLR4, and immunohistochemical staining to observe the protein expressions of TLR2 and TLR4 in mouse skin specimens. Enzyme-linked immunosorbent assay (ELISA) was conducted to determine the levels of interleukin 12 (IL-12) and tumor necrosis factor α (TNFα) in serum samples from the mice. Results After injection of Sporothrix conidium suspensions, the mRNA expression level of TLR2 gradually increased and peaked at 24 hours, which was 18.8 times that in the control group at 6 hours and 34 times at 24 hours. In addition, the mRNA expression level of TLR4 in the experiment group reached a peak, and was 56.7 times that in the control group at 6 hours after injection, then gradually decreased and reached the nadir at 96 hours. As immunohistochemical staining revealed, TLR2 and TLR4 were apparently expressed in both keratinocytes and macrophages in skin specimens from the experimental group, but not obviously in those from the control group. No significant differences were observed between the experimental group and control group in serum levels of IL-12 or TNF-α at any of the sampling time points. Conclusion TLR2 and TLR4 may play a favoring role in immunological defense by participating in the recognition of Sporothrix by keratinocytes and macrophages in mouse skin. Key words: Sporothrix; Toll-like receptor 2; Toll-like receptor 4