当归四逆汤对硬皮病模型小鼠血清炎症因子表达及TLR4/NF-κB信号通路的影响

Effects of Danggui Sini Decoction on the Expression of Inflammatory Factors and TLR4/NF-κB Signaling Pathway in Mice with Scleroderma

目的:观察当归四逆汤对硬皮病模型小鼠血清炎症因子表达的影响,并初步探讨当归四逆汤通过Toll样受体(TLR)4/核因子κB(NF-κB)信号通路影响硬皮病的调控机制。方法:选取BALB/c小鼠共60只,按照随机数字法分为空白对照组、模型对照组、阳性对照组及当归四逆汤治疗组(包括高、中和低剂量组),每组10只。模型对照组、阳性对照组及当归四逆汤治疗组小鼠均采用皮下注射盐酸博来霉素0.1 ml建立硬皮病模型小鼠,空白对照组小鼠注射0.9%氯化钠注射液0.1 ml,连续注射3周。造模结束后5 d,当归四逆汤高、中和低剂量组小鼠分别给予当归四逆汤16、8和4 g/kg灌胃治疗;阳性对照组小鼠给予泼尼松0.2 g/kg灌胃治疗;空白对照组、模型对照组小鼠给予等容积的蒸馏水5 ml进行灌胃,各组均1日给药1次,连续治疗3周。末次给药24 h后处死小鼠,测定各组小鼠肿瘤坏死因子α(TNF-α)、白细胞介素10(IL-10)及白细胞介素16(IL-16)的表达水平;检测组织中TLR4、NF-κB蛋白的表达变化。结果:造模后,模型对照组小鼠出现皮肤厚度不同程度增加,毛囊萎缩,真皮组织出现组织排列错乱;与空白对照组相比,模型对照组小鼠血清TNF-α、IL-10及IL-16含量明显升高,皮肤组织中TLR4、NF-κB蛋白表达水平明显升高,差异均有统计学意义(P<0.05)。与模型对照组相比,阳性对照组、当归四逆汤治疗组小鼠皮肤组织病理损伤不同程度减轻;且血清TNF-α、IL-10及IL-16含量明显降低,皮肤组织中TLR4、NF-κB蛋白表达水平明显降低,差异均有统计学意义(P<0.05);当归四逆汤的治疗效果具有剂量依赖性。结论:当归四逆汤可有效调控硬皮病模型小鼠血清TNF-α、IL-10及IL-16等炎症因子含量,调节TLR4/NF-κB信号蛋白表达,对硬皮病模型小鼠皮肤组织起到有效保护作用。

OBJECTIVE:To observe the effects of Danggui Sini decoction on the expression of inflammatory factors in mice with scleroderma,and to probe into the regulatory mechanism of Danggui Sini decoction affecting scleroderma through Toll-like receptor 4(TLR4)/nuclear factor kappa B(NF-κB)signal pathway.METHODS:A total of 60 BALB/c mice were extracted to be divided into the blank control group,the model control group,the positive control group and the Danggui Sini decoction group(high-dose,medium-dose and low-dose group)via the random number table,with 10 cases in each group.Mice in the model control group,positive control group and Danggui Sini decoction group were given given subcutaneous injection of bleomycin hydrochloride 0.1 ml to establish the sclerderma model mice,and the blank control group was given 0.1 ml 0.9%sodium chloride injection for 3 weeks.On the 5th day after the establishment of the model,mice in the high-dose,medium-dose and low-dose group were given 16,8 and 4 g/kg Danggui Sini decoction by gavage.mice in the positive control group were given prednisone 0.2 g/kg by gavage,and the blank control group and the model control group were given the same volume of 5 ml distilled water for gavage,each group was given drugs once a day,for 3 weeks.The mice were sacrificed 24 h after the last administration,and the expression levels of tumor necrosis factorα(TNF-α),interleukin 10(IL-10)and interleukin 16(IL-16)in each group were determined,changes of TLR4,NF-κB protein expression were detected.RESULTS:After modeling,mice in the model control group showed various degrees of increase in skin thickness,atrophy of hair follicles,and disordered organization of dermal tissues.Compared with the blank control group,the serum levels of TNF-α,IL-10 and IL-16 in the model control group were significantly increased,and the expression levels of TLR4 and NF-κB protein in skin tissues were significantly increased,with statistically significant differences(P<0.05).Compared with the model control group,the skin pathological damage in the positive control group and the Danggui Sini decoction group was alleviated to different degrees;the serum levels of TNF-α,IL-10 and IL-16 were significantly reduced,and the expression levels of TLR4 and NF-κB in skin tissues were significantly reduced,with statistically significant differences(P<0.05).The therapeutic effects of Danggui Sini decoction were dose-dependent.CONCLUSIONS:Danggui Sini decoction can effectively regulate the content of inflammatory factors such as TNF-α,IL-10 and IL-16 in scleroderma model mice,regulate the expression of TLR4/NF-κB signaling protein,and effectively protect the skin tissues of scleroderma model mice.