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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 investigate the therapeutic effect and possible mechanism of fingolimod on mouse dermal collagen deposition. Methods The model of dermal collagen deposition in mice bearing B10.D2 → BALB / c allogeneic transplantation was established and randomly divided into two groups. The fingolimod treatment group received oral fingolimod 1 mg / (kg.d) for 50 days ; Model group oral equal volume of sterile PBS + ethanol (fingolimod solvent) mixed solution. At the same time BALB / c donor mouse cell suspension was transplanted to establish BALB / c mice with gene transfer control. The changes of body weight and status of the mice were recorded. The ear skin was taken for histopathological analysis. Ear RNA was extracted and the chemokines were detected by real-time fluorescence quantitative PCR, that is, the expression of RANTES, monocytes (MCP-1), and the expression of TGF-β1 and collagen Ⅰ in fibroblasts. Peripheral blood samples were collected from the peripheral blood of each group on the 4th and 8th week, and the peripheral blood was analyzed by flow cytometry Blood T, B lymphocyte level changes. Results At the end of the observation period (13 weeks), the body weight (18.67 ± 0.26) g in the fingolimod treatment group was significantly higher than that in the model group (15.68 ± 1.45) g (P <0.05). Histopathological results showed that, Compared with the model group, the mice in the fingolimod treatment group showed no significant thickening, collapsing and infiltration of inflammatory cells in the skin of the mice. Real-time fluorescent quantitative PCR confirmed that RANTES, collagen Ⅰ (P <0.05). The results of flow cytometry showed that the proportion of T lymphocytes in the peripheral blood of Fingolimod-treated mice at the 4th and 8th week was ( 22.8 ± 5.96)% decreased significantly compared with the model group (72.96 ± 14.97)% (P <0.05). Conclusion Fingolimod can improve the collagen deposition and inflammatory infiltration in the skin of the mice mainly by reducing the number of T lymphocytes in peripheral blood, inhibiting the activation of effector cells, decreasing the infiltration of inflammatory cells and the secretion of chemokines The quality of life in mice.