Th22细胞干扰角质形成细胞分泌HB-EGF与其在银屑病发病中的作用探讨

Psoriasis is a common skin disease with unclear etiology that involves dysregulated interplay between immune cells and keratinocytes. Th22 cell was recently identified as another new subset of T cells generated in presence of TNF-α and IL-6, which produced IL-22. Th22 cells may participate in the pathogenesis of psoriasis. Heparin-binding epidermal-growth-factor-like growth factor (HB-EGF) plays an important role in keratinocyte hyperproliferation in psoriasis as identified in our previous studies. In this study, we will collect the lesions and peripheral blood of psoriatic patients, culture keratinocytes, culture skin model and mouse model as our targeted experimental materials. Cultured keratinocytes, cultured skin model and mouse model will be treated by supernatant obtained from culturing Th22 cells, IL-22 and anti-IL-22 Ab. We will use flow cytometry to detect Th22 cells in the peripheral blood of psoriatic patients, use in situ hybridization and real time quantitative RT-PCR to detect IL-22 and HB-EGF mRNA, use immunohistochemistry and Western-blot to detect IL-22 and HB-EGF protein. We will also observe the epidermal changes of cultured skin model and mouse model after treatment. We will analyze the relation of the number of Th22 cells with psoriatic severity, of expression level of IL-22 and HB-EGF with psoriatic severity. The objective of our study is to illustrate the function of Th22 cells in keratinocytes hyperproliferation, and the relation between Th22 cells and HB-EGF expression in psoriasis and prepare the ground for elucidating psoriatic pathogenesis and developing in its therapy.

银屑病是最常见的皮肤病之一，病因不清，难以治愈。目前认为银屑病的发病机制主要为由T细胞介导的以角质形成细胞（KC）为靶点的免疫应答反应。Th22细胞是新认识的一种T细胞亚群，参与银屑病的发生。课题组前期研究证明肝素结合表皮生长因子样生长因子（HB-EGF）在银屑病表皮KC异常增殖中发挥重要作用。我们推测Th22细胞可能通过干扰KC中HB-EGF的表达，进而影响银屑病的发生。本研究运用分子生物学，免疫学和细胞生物学等先进技术，以临床标本，Th22细胞及其细胞因子干预后培养的KC、皮肤类似物模型和银屑病动物模型为研究对象，检测银屑病患者皮损及外周血Th22细胞及其效应细胞因子的表达情况，检测干预前后KC增殖和凋亡的改变、观察皮肤类似物模型和银屑病动物模型表皮形态的改变，检测HB-EGF表达的改变，意欲阐明Th22细胞在银屑病表皮KC异常增殖中的作用，为揭示银屑病的发病机制奠定基础。

背景：银屑病是最常见的皮肤病之一，病因不清，难以治愈。在银屑病患者外周血及皮损中IL-22水平升高，且与银屑病的严重程度呈正相关。课题组前期研究表明，HB-EGF和TIG3在银屑病中发挥重要作用。因此，本课题研究IL-22对HaCaT细胞HB-EGF和TIG3表达的影响，并探讨其相关信号通路。. 研究内容：用不同浓度的IL-22和IL-22 siRNA作用于HaCaT细胞，用MTT法检测其对细胞增殖的影响；用流式细胞术检测其对细胞凋亡的影响；用Western-blot法、ELISA法和实时荧光定量RT-PCR法检测HB-EGF和TIG3表达水平的改变。另外，用咪喹莫特诱导小鼠背部皮肤产生银屑病样皮损，IL-22干预处理后观察HB-EGF的表达。. 结果：1.IL-22可剂量依赖诱导HaCaT细胞的增殖和抑制HaCaT细胞的凋亡。2.IL-22可剂量依赖诱导HaCaT细胞HB-EGF的表达。IL-22可诱导HaCaT细胞MAPK信号通路中P-ERK1/2蛋白的增加和JAK2/STAT3信号通路中P-JAK2和P-STAT3蛋白的升高。加入信号通路特异性抑制剂PD98059及AG490后HB-EGF蛋白和mRNA的表达较IL-22干预组明显降低。咪喹莫特持续作用于Bal B/c小鼠背部，可以成功诱导小鼠银屑病样皮损，IL-22皮下注射可引起正常Bal B/c小鼠背部及银屑病小鼠皮肤表皮中HB-EGF表达的升高。3.IL-22可剂量依赖抑制TIG3的表达。应用信号通路特异性抑制剂PD98059及AG490后IL-22 对TIG3抑制作用较IL-22组减少。差异有统计学意义。. 结论及意义：IL-22可促进HaCaT细胞增殖并抑制其凋亡，IL-22 siRNA可抑制HaCaT细胞增殖并促进其凋亡。IL-22可剂量依赖诱导HaCaT细胞HB-EGF的表达，其机制可能与MAPK-ERK1/2和JAK2/STAT3通路有关。IL-22可引起Bal B/c小鼠背部银屑病样皮损表皮中HB-EGF表达的升高。 IL-22可剂量依赖抑制HaCaT细胞TIG3的表达，其机制可能与MAPK-ERK1/2和JAK2/STAT3通路有关。进一步研究可能为治疗银屑病提供一个新的治疗途径。

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