TLR4, rather than TLR2, regulates wound healing through TGF-β and CCL5 expression

Background: Toll-like receptors (TLRs) have a crucial role in early host defense against invading pathogens. Recent studies suggest that TLRs play important roles in non-infections inflammation and tissue repair and regeneration. Objective: To determine the roles of TLR2 and TLR4 in mouse wound healing using TLR2-deficient (TLR2^-/-), TLR4-deficient (TLR4^-/-), and TLR2/TLR4-deficient (TLR2/4^-/-) mice. Methods: Open wounds made in TLR2^-/-, TLR4^-/-, and TLR2/4^-/- mice were examined clinically and histologically. Cytokine expression in the wounded skin was also investigated. TGF-β production from macrophages stimulated by hyaluronan, a ligand for TLR2 and TLR4, was evaluated by real-time PCR. Results: Wound areas in TLR2^-/-, TLR4^-/-, and TLR2/4^-/- mice were larger than wild-type mice both at days 3 and 7 after wounding, accompanied by decreased numbers of infiltrating macrophages in the dermis and decreased TGF-β and CCL5 mRNA expression in the wounded skin. Immunohistochemistry showed decreased numbers of macrophages expressing TGF-β and reduced CCL5 expression by keratinocytes in the wounded skin from TLR2^-/-, TLR4^-/-, and TLR2/4^-/- mice compared to wild-type mice. Moreover, TGF-β production from macrophages induced by hyaluronan stimulation in vitro was significantly decreased in the absence of TLRs, especially TLR4. Interestingly, macrophages and wounded skin from TLR2^-/- mice showed decreased TLR4 mRNA expression compared to wild-type mice, suggesting that the effect of TLR2 deficiency was at least partially dependent on decrease in TLR4. Topical application of TGF-β and CCL5 significantly improved wound healing in TLR-deficient mice. Conclusion: TLR4, rather than TLR2, regulates wound healing through TGF-β and CCL5 expression.