穿心莲内酯延缓急性胰腺炎相关肾损伤的代谢组学研究

Metabonomics analysis of andrographolide in delaying acute pancreatitis associated kidney injury

目的探讨穿心莲内酯(AG)对AP相关肾损伤的影响及其可能机制。方法将32只C57BL/6小鼠随机分为对照组、AP组、AG治疗组和AG对照组。AP组采用腹腔注射左旋精氨酸联合脂多糖的方法制备AP模型,AG治疗组在制模前8 h静脉注射AG注射液,对照组腹腔注射等体积生理盐水,AG对照组在腹腔注射生理盐水前8 h静脉注射AG注射液。采集小鼠眼内眦静脉血,检测血清淀粉酶、肌酐、尿素氮水平;取胰腺和双肾组织,应用比色法检测胰腺组织髓过氧化物酶(MPO)活性,光镜下观察小鼠胰腺及肾组织病理变化并进行病理评分;采用超高效液相色谱-四级杆-飞行时间串联质谱系统进行肾组织代谢组学分析。结果与对照组比较,AP组血清淀粉酶、肌酐、尿素氮水平和胰腺组织MPO活性明显升高[(13.78±6.01)U/L比(0.23±0.04)U/L、(79.81±24.03)μmol/L比(9.02±2.87)μmol/L、(34.76±14.53)mmol/L比(8.52±2.55)mmol/L、(16.55±4.23)U/g pro比(2.32±0.55)U/g pro],胰腺、肾组织病理评分亦显著升高[(2.70±0.26)分比(0.20±0.12)分、(3.00±0.35)分比(0.30±0.12)分],差异均有统计学意义(P值均<0.05);而AG治疗组血清淀粉酶、肌酐、尿素氮和胰腺组织MPO活性较AP组显著降低[(8.26±3.87)U/L比(13.78±6.01)U/L、(55.42±17.25)μmol/L比(79.81±24.03)μmol/L、(20.66±10.30)mmol/L比(34.76±14.53)mmol/L、(11.51±3.29)U/g pro比(16.55±4.23)U/g pro],胰腺、肾组织病理评分亦显著下降[(1.40±0.19)分比(2.70±0.26)分、(1.70±0.26)分比(3.00±0.35)分],差异均有统计学意义(P值均<0.05)。代谢组学分析共获得31个差异代谢物,这些代谢物主要参与氨基酸代谢和脂肪酸代谢,少部分还参与糖代谢、核苷酸代谢、维生素代谢以及胆汁酸代谢。结论AG可能通过改变AP相关肾损伤过程中差异代谢物的水平对肾损伤起到保护作用。

Objective To elucidate the effects of andrographolide(AG)against acute pancreatitis(AP)-associated kidney injury and explore the potential mechanism.Methods Thirty-two C57BL/6 mice were divided into control group,AP group,AG treatment group and AG control group.In AP group,AP model was established by intraperitoneal injection of L-arginine and LPS.AG treatment group was intravenously injected with AG 8 h before the model establishment.The control group was intraperitoneally injected with an equal volume of normal saline.The AG control group was intravenously injected with AG 8 h before the intraperitoneal administration of normal saline.Venous blood of inner canthus in mice was collected,and the levels of amylase,creatinine(Scr)and urea nitrogen(BUN)in serum were detected.Pancreatic and double kidney tissues were also collected.Pancreatic myeloperoxidase(MPO)activity was detected by colorimetry.Histopathological changes of pancreas and kidneys were observed under light microscope and pathological score was evaluated.Ultra-high-performance liquid chromatography coupled with quadrupole-time-of-flight mass spectrometry was used to analyze metabonomics in renal tissue.Results Compared to the control group,the levels of amylase,Scr and BUN in serum and pancreatic MPO activity in the AP group were obviously increased[(13.78±6.01)U/L to(0.23±0.04)U/L,(79.81±24.03)μmol/L to(9.02±2.87)μmol/L,(34.76±14.53)mmol/L to(8.52±2.55)mmol/L,(16.55±4.23)U/g pro to(2.32±0.55)U/g pro];histopathologic scores of pancreas and kidney in the AP group were also increased(2.70±0.26 to 0.20±0.12,3.00±0.35 to 0.30±0.12),and the differences were statistically significant(all P<0.05).The levels of amylase,Scr and BUN in serum and pancreatic MPO activity in the AG treatment group were decreased than the AP group[(8.26±3.87)U/L to(13.78±6.01)U/L,(55.42±17.25)μmol/L to(79.81±24.03)μmol/L,(20.66±10.30)mmol/L to(34.76±14.53)mmol/L,(11.51±3.29)U/g pro to(16.55±4.23)U/g pro];histopathologic scores of pancreas and kidney were also decreased in the AG treatment group than those in the AP group(1.40±0.19 to 2.70±0.26、1.70±0.26 to 3.00±0.35),and the differences were statistically significant(all P<0.05).Metabonomics analysis detected a total of 31 metabolites.Most of metabolites were involved in amino acid and fatty acid metabolism,and a few of them were also involved in glucose,nucleotide,vitamin and bile acid metabolism.Conclusions AG exerts protective effects on AP-associated kidney injury by altering the levels of multiple metabolites.