摘要
目的:基于Toll样受体9/髓样分化因子88/核转录因子-κB(TLR9/MyD88/NF-κB)信号通路研究白芍总苷(TGP对MRL/lpr狼疮模型小鼠肾脏损伤的干预作用,探讨TGP防治系统性红斑狼疮(SLE)的部分免疫学机制。方法:将SPF级MRL/lpr雌性小鼠随机分为4组,模型组、地塞米松组(0.15 g·kg^(-1))、TGP高、低剂量组(0.078、0.039 g·kg^(-1)),雌性C57BL/6J小鼠设为空白组,每组7只;各组小鼠每天同一时间段给予相应药物或生理盐水灌胃。4周后采集标本,称取肾脏、脾脏质量,计算脏器指数;生化分析法检测各组血清肌酐(SCr)、血尿素氮(BUN)水平;苏木素-伊红(HE)染色观察小鼠肾脏组织病理学变化;马松(Masson)染色评估小鼠肾脏组织纤维化程度;酶联免疫吸附测定法(ELISA)检测血清白细胞介素(IL)-2、干扰素-α(IFN-α)、IL-4和抗核抗体(ANA)的水平;实时荧光定量聚合酶链式反应(Real-time PCR)检测肾组织中TLR9、MyD88、NF-κB p65的mRNA表达;免疫荧光法检测肾组织中TLR9、NF-κB p65蛋白表达;蛋白免疫印迹法(Western blot)检测肾、脾组织中TLR9、MyD88、NF-κB p65蛋白的表达水平。结果:与空白组比较,模型组小鼠SCr、BUN及脾脏指数和肾脏指数明显升高(P<0.05),肾组织病理损伤和纤维化程度明显增强;血清IFN-α、IL-4及ANA水平显著升高,IL-2水平明显降低(P<0.05);肾脏和脾脏中TLR9、MyD88及NF-κB p65的mRNA和蛋白水平明显升高(P<0.05)。与模型组比较,TGP高、低剂量组小鼠SCr、BUN及脾脏指数和肾脏指数明显降低(P<0.05),肾组织病理损伤和纤维化程度明显减轻;血清IFN-α、IL-4及ANA水平明显下降(P<0.05),IL-2水平明显升高;肾脏和脾脏组织中TLR9、MyD88和NF-κB p65等分子mRNA和蛋白表达水平均明显降低(P<0.05)。结论:TGP可能通过调控TLR9/MyD88/NF-κB信号通路,进而抑制其下游相关炎症因子表达,发挥免疫调节及抗SLE肾脏损伤作用。
Objective:To investigate the intervention effect of total glucosides of paeony(TGP)on the renal injury of MRL/lpr mice based on the Toll-like receptor 9(TLR9)/myeloid differentiation factor 88(MyD88)/nuclear transcription factor-κB(NF-κB)signaling pathway and explore the immunological mechanism of TGP in preventing and treating systemic lupus erythematosus(SLE).Method:MRL/lpr female mice of SPF grade were randomly divided into a model group,a dexamethasone group(0.15 g·kg^(-1)),and high-(0.078 g·kg^(-1))and low-dose(0.039 g·kg^(-1))TGP groups,and female C57BL/6J mice were assigned to a blank group,with 7 mice in each group.Mice in each group were treated with corresponding drugs or normal saline by gavage at the same time every day.After 4 weeks,samples were collected.The kidney and spleen were weighed,and the organ index was calculated.Serum creatinine(SCr)and blood urea nitrogen(BUN)levels in each group were detected by biochemical assay.Hematoxylin-eosin(HE)staining was used to observe the histopathological changes in the kidney.The degree of renal fibrosis was evaluated by Masson staining.The serum levels of interleukin(IL)-2,interferon(IFN)-α,IL-4,and anti-nuclear antibody(ANA)were detected by enzymelinked immunosorbent assay(ELISA).The mRNA expression of TLR9,MyD88,and NF-κB p65 in renal tissues was detected by real-time quantitative polymerase chain reaction(Real-time PCR).The protein expression of TLR9 and NF-κB p65 in renal tissues was detected by immunofluorescence.The protein expression of TLR9,MyD88,and NF-κB p65 in renal and spleen tissues was tested by Western blot.Result:Compared with the blank group,the model group showed increased SCr,BUN,spleen index,and kidney index(P<0.05),deteriorated pathological injury and fibrosis in renal tissues,elevated serum levels of IFN-α,IL-4,and ANA,decreased level of IL-2(P<0.05),and up-regulated TLR9,MyD88,and NF-κB p65 mRNA and protein levels in the kidney and spleen(P<0.05).Compared with the model group,the TGP groups displayed reduced SCr,BUN,spleen index,and kidney index(P<0.05),relieved pathological damage and fibrosis in renal tissues,decreased serum levels of IFN-α,IL-4,and ANA(P<0.05),increased level of IL-2,and declining mRNA and protein expression levels of TLR9,MyD88,and NF-κB p65 in the kidney and spleen(P<0.05).Conclusion:TGP may inhibit the expression of downstream inflammatory factors to regulate immunity and resist SLE-induced renal injury by regulating the TLR9/MyD88/NF-κB signaling pathway.
作者
吴琼
于红红
罗瑞熙
何金涛
罗舒文
俞琦
WU Qiong;YU Hong-hong;LUO Rui-xi;HE Jin-tao;LUO Shu-wen;YU Qi(Guizhou University of Traditional Chinese Medicine,Guiyang 550025,China)
出处
《中国实验方剂学杂志》
CAS
CSCD
北大核心
2022年第12期103-110,共8页
Chinese Journal of Experimental Traditional Medical Formulae
基金
贵州省中医药、民族医药科学技术研究课题项目(QZYY-2016-002)。
