芬戈莫德对小鼠皮肤胶原沉着病的治疗作用及可能机制

Therapeutic effect of fingolimod on murine skin collage deposition disease and possible mechanisms

目的探讨芬戈莫德对小鼠皮肤胶原沉着病的治疗作用及可能机制。方法建立B10.D2→BALB/c异基因移植小鼠皮肤胶原沉着病模型,随机分为两组,芬戈莫德治疗组口服芬戈莫德1 mg/(kg.d),连续治疗50 d;模型组口服等体积的无菌PBS+无水乙醇(芬戈莫德溶剂)混合溶液。同时移植BALB/c供体小鼠细胞混悬液,建立BALB/c小鼠同基因移植对照。记录小鼠体质量和状态变化情况;取耳部皮肤进行组织病理学分析;提取耳部RNA,实时荧光定量PCR检测趋化因子,即调节正常T细胞表达和分泌因子(RANTES)、单核细胞趋化因子(MCP)-1,以及纤维生成相关的转化生长因子(TGF)-β1和胶原Ⅰ表达水平的变化;取第4周和第8周各组外周血,通过流式细胞仪分析外周血中T、B淋巴细胞水平的变化。结果观察期结束(13周)时,芬戈莫德治疗组小鼠体质量(18.67±0.26)g,明显高于模型组(15.68±1.45)g(P<0.05);组织病理学结果显示,与模型组相比,芬戈莫德治疗组小鼠皮肤无明显增厚变硬、胶原沉着及炎性细胞浸润;实时荧光定量PCR检测证实,芬戈莫德治疗组小鼠皮肤RANTES、胶原Ⅰ、TGF-β1以及MCP-1 mRNA转录水平较模型组显著下调(P<0.05);流式检测结果显示,第4、8周时芬戈莫德治疗组小鼠外周血T淋巴细胞所占比例为(22.8±5.96)%与模型组(72.96±14.97)%相比显著下降(P<0.05)。结论芬戈莫德主要通过降低外周血中T淋巴细胞的数量,抑制效应细胞的活化,下调炎症细胞的浸润以及趋化因子的分泌表达,从而改善小鼠皮肤胶原沉着及炎性浸润,显著提高小鼠的生存质量。

Objective To determine the therapeutic effect of fingolimod（FTY720） on murine skin collagen deposition and possible mechonisms.Methods Female BALB / c mice transplanted with male B10.D2 bone marrow and spleen cell mixture were used to construct a murine skin collagen deposition disease model.The allogenic transplantation mice were randomly divided into two groups.Mice in the experimental group were treated with fingolimod by oral administration while mice in the control group were administrated with the mixture of equal amount of PBS and ethanol.Meanwhile,in the syngenic transplantation group,mice were transplanted with bone marrow and spleen cell mixture from BALB / c mice.The body weight was recorded every day.Ear skin was depilated and collected for the pathological analysis and RNA isolation.Changes of gene expression level were analyzed by real-time PCR.The proportion of lymphocyte in peripheral blood was determined by flow cytometric analyses.Results The body weight of recipient mice treated with fingolimod gradually recovered,whereas mice in the control group went on weight loss until death.Histopathological evaluation of skin specimen showed skin thickening,collagen deposition,and mononuclear cell infiltration in control group.In contrast,no obvious skin lesions were observed in fingolimod-treated mice.Furthermore,the transcriptional expression of cutaneous regulate upon activation normal T cell expressed and secreted factor,collagenⅠ,transforming growth factor-β1,and monocyte chemotactic protein-1 was down-regulated in mice with fingolimod treatment as compared with those in control group.Finally,the proportion of the peripheral T cells significantly decreased in mice exposed to fingolimod as compared with those in control group.Conclusion Fingolimod inhibits the activation of effective cells and the infiltration of mononuclear cells in the skin through down-regulation of peripheral T cells,thus ameliorating the disease symptoms and improving the life quality of mice.