摘要
目的:探究黄连汤加味对过敏性紫癜性肾炎(HSPN)小鼠辅助性T细胞17(Th17)和细胞毒性T细胞17(Tc17)的影响。方法:选取40只昆明小鼠建立HSPN模型,并随机选取30只成功造模的小鼠,分为模型组[灌胃牛血清白蛋白(BSA)+尾静脉注射脂多糖(LPS)+皮下注射四氯化碳(CCl4)]、黄连汤加味低剂量组(7.5 g/kg)、黄连汤加味中剂量组(15.0 g/kg)、黄连汤加味高剂量组(30.0 g/kg)、霉酚酸酯组(0.3 g/kg)。另选6只小鼠作为对照组,灌胃等量双蒸水,尾静脉和皮下注射等量生理盐水。全自动生化分析仪检测小鼠尿液24 h尿蛋白水平、尿红细胞个数、血清肌酐(Scr)、白蛋白(ALB)和总蛋白(TP)水平;苏木精-伊红(HE)染色和碘酸雪夫(PAS)染色检测小鼠肾脏组织病理学变化;免疫组织化学法检测小鼠肾组织免疫球蛋白A(IgA)和补体成分3抗体(C3)水平;酶联免疫吸附测定(ELISA)检测小鼠血清肿瘤坏死因子-α(TNF-α)、白细胞介素-1β(IL-1β)、白细胞介素IL-6(IL-6)、白细胞介素-17(IL-17)和IgA水平;流式细胞术检测小鼠血液Th17和Tc17细胞比例。结果:对照组小鼠肾组织结构清晰,肾小球、肾小管形态正常,肾间质无充血及炎症细胞浸润;模型组小鼠肾小球体积增大,系膜区及毛细血管袢内细胞数量显著增多,肾小球肿大,系膜区增宽,系膜基质及细胞增生明显,PAS染色阳性表达面积百分率显著升高(P<0.01),肾间质可见明显充血,伴大量炎症细胞浸润;黄连汤加味各剂量组及霉酚酸酯组小鼠肾间质充血及炎症细胞浸润程度显著减轻,肾小球内系膜细胞和内皮细胞增生程度较模型组明显改善,系膜区增生程度均显著减轻,PAS染色阳性表达面积百分率显著降低(P<0.05)。模型组小鼠尿液24 h尿蛋白水平、尿红细胞个数,血清Scr水平,肾组织IgA和C3积分光密度(IOD)值,血清TNF-α、IL-1β、IL-6、IL-17、IgA水平,血液Th17细胞和Tc17细胞比例均高于对照组(P<0.05),血清ALB和TP水平均低于对照组(P<0.01)。黄连汤加味中、高剂量组和霉酚酸酯组小鼠24 h尿蛋白水平、尿红细胞个数,血清Scr水平,肾组织IgA IOD,血清TNF-α、IL-1β、IL-6、IL-17、IgA水平,血液Th17和Tc17细胞比例均低于模型组(P<0.05),血清ALB和TP水平高于模型组(P<0.01);黄连汤加味低剂量组小鼠血清TNF-α、IL-1β、IL-6、IL-17、IgA水平,血液Th17和Tc17细胞比例均低于模型组(P<0.05),血清ALB水平高于模型组(P<0.05)。黄连汤加味低、中剂量组小鼠尿液24 h尿蛋白水平、尿红细胞个数,血清Scr、TNF-α、IL-6、IL-17、IgA水平,血液Th17和Tc17细胞比例高于霉酚酸酯组(P<0.05);黄连汤加味低剂量组小鼠血清ALB和TP水平低于霉酚酸酯组(P<0.05),肾组织IgA IOD高于霉酚酸酯组(P<0.01);黄连汤加味高剂量组小鼠肾组织IgA和C3 IOD和血液Th17细胞比例低于霉酚酸酯组(P<0.05),血液Tc17细胞比例高于霉酚酸酯组(P<0.05)。结论:黄连汤加味可减轻HSPN小鼠肾脏损伤,减轻体内炎症反应,抑制免疫系统异常激活,其作用机制可能与抑制Th17细胞和Tc17细胞活化有关。
Objective:To explore the effect of modified Huanglian decoction on T helper cell 17(Th17)and T cytotoxic cell 17(Tc 17)in mice with henoch schonlein purpura nephritis(HSPN).Methods:Totally 40 Kunming mice were selected to establish a HSPN model.The 30 mice with successful modeling were randomly divided into model group[intragastric administration of bovine serum albumin(BSA)+tail vein injection of lipopolysaccharide(LPS)+subcutaneous injection of carbon tetrachloride(CCl4)],modified Huanglian decoction low-dose group(7.5 g/kg),modified Huanglian decoction medium-dose groups(15.0 g/kg),modified Huanglian decoction high-dose group(30.0 g/kg),and mycophenolate mofetil group(0.3 g/kg),with 6 mice in each group.In addition,6 mice were selected as the control group.The mice were given the same amount of double distilled water by gavage,tail vein and subcutaneous injection of the same amount of normal saline in control group.The 24-hour urine protein level,urine red blood cell count,serum creatinine(Scr),albumin(ALB)and total protein(TP)levels were detected by automatic biochemical analyzer.Hematoxylin-eosin(HE)and periodic acid-schiff stain(PAS)staining were used to detect the pathological changes of kidney tissue in mice.The levels of immunoglobulin A(IgA)and complement component 3(C3)in renal tissue of mice were detected by immunohistochemistry.The serum levels of tumor necrosis factor-alpha(TNF-α),interleukin-1β(IL-1β),interleukin-6(IL-6),interleukin-17(IL-17)and IgA were detected by enzyme linked immunosorbent assay(ELISA).The proportion of Th17 and Tc17 cells in the blood of mice was detected by flow cytometry.Results:The renal tissue structure was clear in control group,with normal morphology of glomeruli and renal tubules,and there was no congestion or inflammatory cell infiltration in the renal interstitium.In the model group,the volume of glomeruli increased,and the number of cells in the mesangial area and capillary loops significantly increased.The glomeruli were swollen,and the mesangial area widened in model group.The mesangial matrix and cell proliferation were obvious,and the positive expression area percentage of PAS staining significantly increased in model group(P<0.01).Obvious congestion and a large number of inflammatory cell infiltrations were observed in the renal interstitium in model group.In the modified Huanglian decoction groups at various doses and the mycophenolate mofetil group,the degree of congestion and inflammatory cell infiltration in the renal interstitium was significantly reduced,and the degree of mesangial cell and endothelial cell proliferation in glomeruli was significantly improved compared with the model group.The degree of mesangial area proliferation was significantly reduced,with a significant decrease in the positive expression area percentage of PAS staining in modified Huanglian decoction groups at various doses and the mycophenolate mofetil group(P<0.05).The model group showed higher 24-hour urinary protein levels,urine red blood cell count,serum Scr levels,renal tissue IgA and C3 integrated optical density(IOD)values,serum TNF-α,IL-1β,IL-6,IL-17,IgA levels,and proportions of Th17 and Tc17 cells than control group(P<0.05),while lower serum ALB and TP levels than control group(P<0.01).The modified Huanglian decoction medium-dose group,modified Huanglian decoction high-dose group and mycophenolate mofetil group showed lower 24-hour urinary protein levels,urine red blood cell count,serum Scr levels,renal tissue IgA IOD,serum TNF-α,IL-1β,IL-6,IL-17,IgA levels,and proportions of Th17 and Tc17 cells than model group(P<0.05),while higher serum ALB and TP levels than model group(P<0.01).The modified Huanglian decoction low-dose group showed lower serum TNF-α,IL-1β,IL-6,IL-17,IgA levels,and proportions of Th17 and Tc17 cells than model group(P<0.05),while higher serum ALB levels than model group(P<0.05).The modified Huanglian decoction low-dose group and modified Huanglian decoction medium-dose group showed higher 24-hour urinary protein levels,urine red blood cell count,serum Scr,TNF-α,IL-6,IL-17,IgA levels,and proportions of Th17 and Tc17 cells than mycophenolate mofetil group(P<0.05).The modified Huanglian decoction low-dose group showed lower serum ALB and TP levels than mycophenolate mofetil group(P<0.05),while higher renal tissue IgA IOD than mycophenolate mofetil group(P<0.01).The modified Huanglian decoction high-dose group showed lower renal tissue IgA and C3 IOD values and proportion of Th17 cells than mycophenolate mofetil group(P<0.05),while higher proportion of Tc17 cells than mycophenolate mofetil group(P<0.05).Conclusions:Modified Huanglian decoction can alleviate kidney injury in HSPN mice,reduce systemic inflammatory responses,and suppress abnormal immune system activation.Its mechanism of action may be related to the inhibition of Th17 and Tc17 cell activation.
作者
胡琏
车虹
赵冰洁
肖利
刘培佳
易文静
刘松山
HU Lian;CHE Hong;ZHAO Bingjie;XIAO Li;LIU Peijia;YI Wenjing;LIU Songshan(Hospital of Chengdu University of Traditional Chinese Medicine,Chengdu Sichuan 610032,China)
出处
《中医药导报》
2025年第8期24-30,共7页
Guiding Journal of Traditional Chinese Medicine and Pharmacy
基金
四川省中医药管理局课题(2020JC0043)。
