博来霉素诱导的硬皮病模型是建立SSc(系统性硬化症)小鼠模型的可靠方法。在皮肤结缔组织疾病领域,越来越多的临床和动物实验证据表明,TLRs(Toll样受体)在多种疾病中起重要作用.本研究旨在确定TLR7(Toll样受体7)和TLR9(Toll样受体9)在SSc免疫异常和纤维化机制中的作用。该研究使用TLR7-KO小鼠(具有balb/c背景的TLR7敲除小鼠)和TLR9-KO小鼠(具有balb/c背景的TLR9敲除小鼠)以及WT小鼠(野生型balb/c小鼠)。3种小鼠均采用BLM(博来霉素)诱导硬皮病模型作为实验组,用PBS(磷酸盐缓冲盐水)处理的WT小鼠用作对照组。我们使用流式细胞术分析了SSc疾病模型中TLR7缺陷和TLR9缺陷小鼠的纤维化表型和免疫异常表型,RT-PCR(逆转录-聚合酶链反应),组织学检查,和IHC(免疫组织化学染色)。在SSc疾病的小鼠模型中,TLR7的缺失减轻了皮肤和肺纤维化,而TLR9的缺失加剧了皮肤和肺纤维化。TLR7的缺失导致皮肤中各种促炎和纤维化细胞和细胞因子的浸润和表达相对减少。另一方面,TLR9的缺失导致皮肤中各种促炎和细胞因子抑制细胞和细胞因子的浸润和表达相对增加。在pDC(浆细胞样树突状细胞)的影响下,Beff/Breg(IL-6+CD19+B细胞/IL-10+CD19+B细胞)的平衡,Th17/Treg(IL-17A+CD4+T细胞/Foxp3+CD25+CD4+T细胞),M1/M2(CD86+巨噬细胞/CD206+巨噬细胞),和Th1/Th2(TNFα+CD3+CD4+T细胞/IL-4+CD3+CD4+T细胞)由于TLR7缺失而偏向于抑制炎症和纤维化。相对而言,由于TLR9缺失,平衡偏向于促进炎症和纤维化.在SSc模型中,TLR7促进炎症和纤维化进展,TLR9起保护作用。这些结果表明,TLR7和TLR9在引发SSc产生免疫系统异常和皮肤纤维化中起相反的作用。
The bleomycin-induced scleroderma model is a well-established and dependable method for creating a mouse model of SSc (systemic sclerosis). In the field of skin connective tissue diseases, increasing evidence from clinical and animal experiments suggests that TLRs (Toll-like receptors) play an important role in several diseases. This study aimed to determine the role of
TLR7 (Toll-like receptor 7) and TLR9 (Toll-like receptor 9) in the mechanisms of immune abnormalities and fibrosis in SSc. This study used
TLR7-KO mice (
TLR7-knockout mice with a balb/c background) and TLR9-KO mice (TLR9-knockout mice with a balb/c background) as well as WT mice (wild-type balb/c mice). All three kinds of mice were induced by BLM (bleomycin) in a scleroderma model as the experimental group; meanwhile, WT mice treated with PBS (phosphate-buffered saline) were used as the control group. We analyzed the fibrotic phenotype and the immunological abnormality phenotype of
TLR7-deficient and TLR9-deficient mice in the SSc disease model using flow cytometry, RT-PCR (reverse transcription-polymerase chain reaction), a histological examination, and IHC (immunohistochemical staining). In a mouse model of SSc disease, the deletion of TLR7 attenuated skin and lung fibrosis, while the deletion of TLR9 exacerbated skin and lung fibrosis. The deletion of TLR7 resulted in a relative decrease in the infiltration and expression of various pro-inflammatory and fibrotic cells and cytokines in the skin. On the other hand, the deletion of TLR9 resulted in a relative increase in the infiltration and expression of various pro-inflammatory and cytokine-inhibiting cells and cytokines in the skin. Under the influence of pDCs (plasmacytoid dendritic cells), the balances of Beff/Breg (IL-6 + CD19 + B cell/IL-10 + CD19 + B cell), Th17/Treg (IL-17A + CD4 + T cell/Foxp3 + CD25 + CD4 + T cell), M1/M2 (CD86 + macrophage/CD206 + macrophage), and Th1/Th2 (TNFα + CD3 + CD4 + T cell/IL-4 + CD3 + CD4 + T cell) were biased towards the suppression of inflammation and fibrosis as a result of the TLR7 deletion. Comparatively, the balance was biased towards promoting inflammation and fibrosis due to the TLR9 deletion. In the SSc model,
TLR7 promoted inflammation and fibrosis progression, while TLR9 played a protective role. These results suggest that
TLR7 and TLR9 play opposite roles in triggering SSc to produce immune system abnormalities and skin fibrosis.