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目的 构建慢性肾脏病(chronic kidney disease,CKD)C57BL/6小鼠模型,探索其小管损伤指标和间质纤维化程度随顺铂造模剂量的改变,为研究从AKI到CKD进展过程提供动物实验依据。方法 将24只8周龄雄性C57BL/6小鼠随机平均分为对照组和低、中、高剂量顺铂模型组。模型组小鼠按5、7、10 mg/kg顺铂腹腔注射给药,每周1次,连续注射4周构建模型。处死小鼠后,留取标本进行相关检测。检测血浆肌酐和24 h尿蛋白排泌量来评估小鼠肾功能;PAS染色观察肾脏病理学变化;免疫组化检测肾损伤分子1(KIM-1)和尿液检测N-乙酰-β-D氨基葡萄糖苷酶(NAG)水平以评估肾小管损伤情况;免疫组化法检测肾脏CD3阳性T细胞和免疫荧光法检测F4/80阳性巨噬细胞浸润情况;天狼星红染色、免疫组化法检测胶原I和α-平滑肌肌动蛋白(α-SMA)表达以评估肾脏纤维化情况。结果 与正常对照组相比,随着注射顺铂浓度的升高,小鼠肾脏损伤越明显,其中10 mg/kg顺铂高剂量组最为显著。与对照组相比,顺铂高剂量组小鼠肾功能下降,表现为血浆肌酐浓度和24 h尿蛋白排泌量显著升高(P<0.05和P<0.001);肾小管上皮细胞坏死、空泡变性等病理学改变显著,肾组织KIM-1表达显著上升(P<0.05),尿NAG水平升高;肾组织浸润的CD3阳性T细胞和F4/80阳性巨噬细胞增多;肾组织天狼星红染色阳性胶原纤维区域显著增多(P<0.001),胶原I和α-SMA表达也明显增多(P<0.01),肾小管-间质发生纤维化。结论反复注射4周10 mg/kg顺铂可诱导小鼠慢性肾功能不全模型,可为研究AKI向CKD的转化机制提供了新的实验模型。
Objective To construct C57BL / 6 mouse model of chronic kidney disease (CKD) and explore the changes of tubulointerstitial fibrosis and tubulointerstitial fibrosis with the dose of cisplatin, and provide animal experiments for studying the progression from AKI to CKD in accordance with. Methods Twenty-four male C57BL / 6 mice, 8 weeks old, were randomly divided into control group and low, medium and high dose cisplatin model groups. The model mice were intraperitoneally injected with 5, 7 and 10 mg / kg cisplatin once a week for 4 weeks. After the mice were sacrificed, the specimens were taken for the relevant tests. Plasma creatinine and 24 h urinary protein excretion were measured to assess renal function; PAS staining was used to observe renal pathological changes; immunohistochemistry was used to detect KIM-1 and urine samples for N-acetyl-β-D The level of NAG was measured to evaluate the damage of renal tubules. CD3 positive T cells were detected by immunohistochemistry and the infiltration of F4 / 80 positive macrophages was detected by immunofluorescence staining. The collagen was detected by red staining with Sirius and immunohistochemistry I and α-smooth muscle actin (α-SMA) expression to assess renal fibrosis. Results Compared with the normal control group, the more severe the renal damage was, the higher the dose of cisplatin 10 mg / kg was. Compared with the control group, the renal function of high-dose cisplatin-treated mice decreased, showing a significant increase in plasma creatinine and 24-hour urinary protein excretion (P <0.05 and P <0.001), renal tubular epithelial cell necrosis, (P <0.05), and the level of urinary NAG increased; the infiltration of CD3 positive T cells and F4 / 80 positive macrophages in renal tissue increased; the expression of Sirius red The area of staining positive collagen fibers was significantly increased (P <0.001), the expression of collagen I and α-SMA was also significantly increased (P <0.01), and tubulointerstitial fibrosis. Conclusion Repeated injection of 10 mg / kg cisplatin for 4 weeks induces the model of chronic renal insufficiency in mice, which may provide a new experimental model for studying the mechanism of AKI conversion to CKD.