1.浙江中医药大学动物实验研究中心;2.浙江中医药大学药学院;3.杭州利孚泰转化医学研究中心
浙江省基础公益研究计划(LTGD23C040003)。
1.Animal Experimental Research Center, Zhejiang Chinese Medical University;2.School of Pharmacy, Zhejiang Chinese Medical University;3.Hangzhou Lifutai Biotechnology Co., Ltd;4.Hangzhou Lifutai Biotechnology Co., Ltd., Hangzhou
目的 采用“氧嗪酸钾联合腺嘌呤加酵母饲料饲喂”的多因素诱导法建立高尿酸肾病(HN)大鼠模型,并观察芪苓颗粒(QLG)的干预作用。方法 取58只SPF级雄性SD大鼠,随机取10只大鼠作正常对照组,其余大鼠采用多因素诱导法建立HN大鼠模型,造模2周后颌下取血检测血清UA、CREA、BUN、TG和TC,筛选出血清UA和体重接近均值的HN大鼠40只,采用分层随机法分为模型对照组、QLG低、高剂量组和阳性对照组,每组10只。各组每日给予相应药物灌胃,连续给药4周后颌下采血检测血清UA、CREA、BUN、TG和TC,行安死术,取肝组织检测XOD、ADA活性,取肾组织进行HE、Gomori六胺银染色,并用免疫组化和WB法观察肾脏GLUT9、OAT1、VCAM-1和TGF-β的蛋白表达。结果 与正常对照组比,模型对照组血清UA、CREA、BUN、TC、TG水平和肝脏XOD、ADA活性均显著升高(P＜0.01),肾组织病变明显,肾小管尿酸盐含量和肾脏GLUT9、VCAM-1、TGF-β蛋白表达均显著升高(P＜0.01,P＜0.05)、OAT1表达显著降低(P＜0.01)。与模型对照组比,各给药组大鼠的血清UA水平和肝脏XOD、ADA活性以及肾脏VCAM-1蛋白表达均显著降低(P＜0.01,P＜0.05),QLG低剂量组大鼠的血清CREA、BUN水平和肾脏TGF-β蛋白表达亦显著降低(P＜0.05,P＜0.01),QLG高剂量组大鼠的血清CREA、BUN水平和肾脏GLUT9蛋白表达亦显著降低(P＜0.01,P＜0.05),各给药组大鼠的尿酸盐沉积及其引发的肾损伤均有不同程度的减轻但无显著差异(P＞0.05)。结论 给予大鼠酵母饲料饲喂的同时给予氧嗪酸钾与腺嘌呤联合灌胃可诱导建立稳定的HN大鼠模型；QLG可通过改善HN模型大鼠UA代谢紊乱、减轻肾脏炎症和尿酸盐沉积及其引发的肾损伤来有效治疗HN,其作用机制与降低血清UA、CREA、BUN、TG水平和肝脏XOD、ADA活性以及肾脏GLUT9、OAT1、VCAM-1、TGF-β蛋白的表达有关。
Objective To establish a rat model of hyperuricemic nephropathy (HN) using a multifactorial induction method of potassium oxazinate combined with adenine and yeast feed, and to observe the intervention effect of Qiling granules (QLG). Methods Fifty-eight SPF grade male SD rats were selected, and ten rats were randomly selected as the normal control group. The remaining rats were induced by multiple factors to establish HN rat models. After 2 weeks of modeling, submandibular blood samples were taken to detect serum UA, CREA, BUN, TG, and TC. Forty HN rats with bleeding clearance UA and body weight close to the mean were selected. They were randomly divided into model control group, QLG low and high dose groups, and positive control group using a stratified randomization method, with ten rats in each group. Each group was given corresponding drugs by gavage daily, and after continuous administration for 4 weeks, submandibular blood samples were taken to detect serum UA, CREA, BUN, TG, and TC. After euthanasia of rats, and liver tissue was taken to detect XOD and ADA activity. Renal tissue was taken for HE and Gomori hexamine silver staining, and the protein expression of GLUT9, OAT1, VCAM-1, and TGF-β in the kidneys was observed using immunohistochemistry and WB methods. Results Compared with the normal control group, the serum levels of UA, CREA, BUN, TC, and TG, as well as liver XOD and ADA activities, were significantly increased in model control Group (P<0.01). The renal tissue of rats shows significant pathological changes. The positive area of urate in renal tubules and the expression of GLUT9, VCAM-1, and TGF-β proteins in the kidneys were significantly increased (P<0.01, P<0.05), while the expression of OAT1 was significantly reduced (P<0.01). Compared with model control group, the serum UA levels, liver XOD, ADA activity, and renal VCAM-1 protein expression of rats in each treatment group were significantly reduced (P<0.01, P<0.05). The serum CREA, BUN levels, and renal TGF-β protein expression of rats in the low-dose QLG group were also significantly reduced (P<0.05, P<0.01). The serum CREA, BUN levels, and renal GLUT9 protein expression of rats in the high-dose QLG group were also significantly reduced (P<0.01, P<0.05), and the urate deposition and renal injury caused by them in each treatment group were reduced to varying degrees, but there was no significant difference (P>0.05). Conclusion A stable HN rat model can be induced by gavage of potassium oxyzinate and adenine in combination with yeast feed to rats; QLG can effectively treat HN by improving UA metabolic disorders, reducing renal inflammation and urate deposition, as well as causing renal damage in HN model rats. Its mechanism of action is related to reducing serum UA, CREA, BUN, TG levels, liver XOD and ADA activities, as well as the expression of GLUT9, OAT1, VCAM-1, and TGF-β proteins in the kidneys.
