Identification And Cloning Of SIL-36R And The Function Of SIL-36R And IL-36Ra In Inhibiting IL-36-induced REG3A Expression

Interleukin-36 (IL-36), which consists of IL-36 α, β and γ, is an important regulator of immunity and inflammation. As a proinflammatory factor, IL-36 induces inflammatory responses by binding to its receptor IL-36R and then initiating the downstream proinflammatory signaling pathway. IL-36 plays an important role in the regulation of innate immunity and host defense. IL-36R belongs to IL-1R family, and two different ways negatively regulate this signaling pathway. One is a receptor antagonist that antagonizes the receptor, the other is a soluble receptor or decoy that binds to the ligands. In IL-36R signaling, it has been reported that IL-36 receptor antagonist (IL-36Ra) inhibits inflammatory responses by binding to IL-36R competitively. However, whether there is a soluble receptor or decoy for IL-36R remains unknown. Moreover, as a highly expressed cytokine in skin, IL-36 and its antagonist IL-36Ra are studied extensively in the regulation of skin inflammation, while their functions in skin wound healing are not clear.In this study, we detected the mRNA expression of IL-36 and IL-36Ra in mouse skin wounds firstly, and found that the mRNA expression of IL-36 and IL-36Ra were upregulated. In addition to IL-36, the expression of RegⅢγ was increased in skin wounds, especially in skin wounds of IL-36Ra knockout mice. These data suggest that there might be a relationship among IL-36, IL-36Ra and RegⅢγ expression in skin wounds.To confirm that IL-36 regulates the expression of REG3A I cloned and purified human IL-36γ and IL-36Ra recombinant proteins and then IL-36γ was used to stimulate epidermal keratinocytes. IL-36γ induced the expression of REG3A and promoted the wound healing in keratinocytes, while IL-36Ra inhibited the upregulated expression of REG3A and delayed wound healing. These results demonstrate that IL-36γ promotes wound healing by inducing REG3A expression, and IL-36Ra regulates this process negatively.Moreover, I for the first time identified and cloned a truncated IL-36R splice variant in human skin keratinocytes and named it soluble IL-36R (sIL-36R). Overexpressed sIL-36R in keratinocytes decreased IL-36y-induced REG3A expression and inhibited wound healing promoted by IL-36y.In conclusion, our results demonstrate the role of IL-36Ra in regulating REG3A expression and skin wound healing, and identify a new IL-36R splice variant and determine its function in wound healing preliminarily. Our studies offer a new potential therapeutic target for the clinical treatment with non-healing skin wounds.