摘要
目的 使用蛋白质组学技术研究高脂饮食诱导下小鼠肾脏组织蛋白质的差异性表达。方法 22只雄性C57BL/6小鼠分为对照组(正常饮食,脂肪供能占比10%)、高脂组(高脂饮食,脂肪供能占比60%),每组11只,连续饲养24周。处死小鼠并眼眶取血,检测TC、TG、血糖(BG)、血肌酐(Scr)、血尿素氮(BUN)等指标。收集肾脏组织并采用HE染色、糖原染色(PAS)和电子显微镜观察病理改变。提取肾脏组织总蛋白,使用质谱仪检测并分析两组小鼠肾脏组织中的差异表达蛋白(DEPs),采用4D非标记定量蛋白质组学技术,以差异倍数>1.5(上调)或<1/1.5(下调)且P<0.05为标准筛选DEPs,对DEPs作功能注释和富集分析,通过蛋白质互作网络分析筛选核心蛋白。结果 高脂组体质量、TC、TG、BG、Scr、BUN水平均显著高于对照组(均P<0.05)。与对照组相比,高脂组肾小球体积稍大,系膜细胞和基质节段性略多,鲍曼囊壁层增厚,肾小管上皮细胞空泡变性明显。电镜检查显示高脂组小鼠足突部分融合,线粒体明显肿胀、畸形、大小不一,溶酶体样结构增多并出现自噬体。两组共鉴定出93种DEPs,其中上调蛋白49种,下调蛋白44种;DEPs主要参与脂质及碳水化合物的运输和代谢途径,显著富集的通路有过氧化物酶体和溶酶体相关通路以及初级胆汁酸生物合成、类固醇激素生物合成、肾素-血管紧张素系统等。与高脂诱导肾脏损害发生、发展密切相关的蛋白质主要有酰基辅酶A氧化酶2、载脂蛋白a4、植烷酰辅酶A羟化酶2、α-甲基酰基辅酶A消旋酶、载脂蛋白e、二羟基丙酮磷酸酰基转移酶、含胰蛋白酶域1。结论 高脂饮食诱导小鼠肾脏组织蛋白质发生改变,主要包括过氧化物酶体相关基质酶代谢通路。本研究为揭示脂质肾毒性提供了有价值的线索。
Objective To investigate the differentially expressed proteins(DEPs)in mouse kidney tissue induced by high-fat diet.Methods Twenty-two male C57BL/6 mice were divided into a control group(n=11)and a high-fat group(n=11);two groups were fed with normal diet(fat energy accounting for 10%)or high-fat diet(fat energy accounting for 60%)for 24 weeks,respectively.After the mice were euthanized,blood samples were collected from the eye socket,and the serum levels of total cholesterol(TC),triglycerides(TG),blood glucose(BG),serum creatinine(Scr),burea nitrogen(BUN)were measured.Kidney tissue samples were subjected to HE staining,periodic acid-Schiff(PAS)staining,and electron microscopy to observe pathological changes.Total proteins were extracted from the kidney tissue and detected by high performance liquid chromatography.The 4D label-free quantitative proteomic technique was employed,and the differential proteins were defined as those with a fold change>1.5(upregulated)or<1/1.5(downregulated).DEPs were screened,and functional annotation and enrichment analysis were performed.Core proteins were selected through protein interaction network analysis.Results The weight,and serum TC,TG,BG,Scr,and BUN levels in the high-fat group were significantly higher than those in the control group(all P<0.05).Compared to the control group,the high-fat group exhibited slightly larger glomerular volume,showing a slight increase of mesangial cells and matrix segments,thickening of Bowman's capsule wall,and significant vacuolar degeneration of renal tubular epithelial cells.Electron microscopy revealed fusion of foot processes in the high-fat group mice,noticeable swelling and deformation of mitochondria of varying sizes,increased lysosome-like structures,and the presence of autophagic bodies.A total of 93 DEPs were identified between the two groups,including 49 upregulated proteins and 44 downregulated proteins in high-fat group.Functional enrichment and pathway bioinformatics analysis indicated that DEPs were mainly involved in lipid and carbohydrate transport and metabolic pathways.Significantly enriched pathways included peroxisome and lysosome-related pathways,as well as primary bile acid biosynthesis,steroid hormone biosynthesis,and the renin-angiotensin system.Protein interaction network analysis revealed that proteins closely associated with the high-fat-induced renal damage included Acox2,ApoA4,Phyh,Amacr,ApoE,Gnpat,and Tysnd1.Conclusion High-fat diet can induce changes in the renal proteome,primarily involving peroxisome-related and matrix enzyme-related metabolic pathways,leading to the renal toxicity in mice.
作者
陈智鸿
王林
黄津华
CHEN Zhihong;WANG Lin;HUANG Jinhua(Department of Nephrology,Xiamen Branch,Zhongshan Hospital,Fudan University,Xiamen 361015,China;不详)
出处
《浙江医学》
CAS
2023年第24期2600-2604,I0005,I0006,共7页
Zhejiang Medical Journal
基金
复旦大学附属中山医院厦门医院孵化课题(2020ZSXMYS04)
厦门市医疗卫生指导性项目(3502Z20224ZD1080)。
