基于蛋白质组学对脊柱结核患者外周血浆差异蛋白筛选及功能分析  

Screening and functional analysis of differential proteins in peripheral plasma of spinal tuberculosis patients based on proteomics

作　　者：娄才立 马宏宝 任智博 王立楠 张旭[2] 董辉[2] 俞梦楚 牛宁奎[3] LOU Caili;MA Hongbao;REN Zhibo;WANG Linan;ZHANG Xu;DONG Hui;YU Mengchu;NIU Ningkui(Clinical Medical School,Ningxia Medical University,Yinchuan,750004,China;Institute of Medical Sciences,General Hospital of Ningxia Medical University,Yinchuan,750004,China;Department of Spinal Surgery,General Hospital of Ningxia Medical University,Yinchuan,750004,China)

机构地区：[1]宁夏医科大学临床学院,银川市750004 [2]宁夏医科大学总医院科研设备管理中心,银川市750004 [3]宁夏医科大学总医院脊柱骨科,银川市750004

出　　处：《中国脊柱脊髓杂志》2022年第9期814-822,共9页Chinese Journal of Spine and Spinal Cord

基　　金：国家自然科学基金项目(编号:81860395);宁夏科学自然基金项目(编号:2020AAC03391);2022年自治区重点研发计划项目(2022BEG03099)。

摘　　要：目的:利用蛋白质组学技术筛选脊柱结核(spinal tuberculosis,STB)患者外周血浆中表达差异的蛋白质,探讨STB致病机制及代谢途径。方法:收集30例STB患者和30例健康志愿者的外周血,取其中3例STB患者(观察组)和3例健康志愿者(对照组)的外周血浆通过蛋白质组学同位素标记相对及绝对定量技术和液相色谱-质谱分析技术鉴定差异蛋白,筛选出表达差异显著(差异倍数>1.2,P<0.05)的蛋白质,并进行基因本体(gene ontology,GO)和京都基因与基因组百科全书(Kyoto Encyclopedia of Genes and Genomes,KECG)富集分析;然后将30例STB患者和30例健康志愿者的血浆进行实时荧光定量逆转录聚合酶链反应(real-time quantitative polymerase chain reaction,RT-PCR)和酶联免疫吸附测定法(enzyme-linked immunosorbent assay,ELISA)验证其差异基因及蛋白。结果:STB患者外周血浆中共筛选出16个与对照组存在显著性差异的蛋白;其中11个蛋白[脂多糖蛋白(lipopolysaccharide-binding protein,LBP)、S100-A8蛋白(protein S100-A8)、S100-A9蛋白(protein S100-A9)、淀粉样蛋白A-2(serumamyloid A-2protein,SAA2)、富含亮氨酸α-2糖蛋白(leucine-rich alpha-2-glycoprotein,LRG2)、蛋白聚糖4(proteoglycan 4,PRG4)、补体因子I(complement factor I,CFI)、α-1-抗胰蛋白酶(alpha-1-antitrypsin,SERPINA1)、组织蛋白酶D(cathepsin D,CTSD)、血浆铜蓝蛋白(ceruloplasmin,CP)和入免疫球蛋白(lambda-immunoglobulin 4-60,IGLV4-60)]表达上调,5个蛋白[黏着斑蛋白(vinculin,VCL)、3-64D免疫球蛋白(immunoglobulin heavy variable 3-64D,IGHV3-64D)、2D-29免疫球蛋白(immunoglobulin kappa variable 2D-29,IGKV2D-29)、非功能性免疫球蛋白(non-functional immunoglobulin 6D-41,IGKV6D-41)和1-45免疫球蛋白(immunoglobulin heavy variable 1-45,IGHV1-45)]表达下调。LBP、S100-A8蛋白和S100-A9蛋白在STB患者外周血浆的基因和蛋白表达与对照组比较均显著性升高(P<0.05),主要参与白介素-17信号通路和自噬信号通路,Objectives:To screen the differentially expressed proteins in peripheral plasma of patients with spinal tuberculosis with proteomics technology to provide reference for the study of the pathogenic mechanism and metabolic pathway of spinal tuberculosis.Methods:Peripheral blood was collected from 30 patients with spinal tuberculosis and 30 healthy volunteers,and the peripheral plasma from three patients with spinal tuberculosis(observation group)and three healthy volunteers(control group)was identified by isobaric tags for relative and absolute quantification(iTRAQ)based proteomics technique and liquid chromatography-mass spectrometry analysis for differential proteins to screen the proteins with significant expression differences(FC>1.2,P<0.05),and enrichment analysis was performed through Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG).Then,plasma from spinal tuberculosis patients and healthy volunteers was tested by real-time fluorescent quantitative reverse transcription polymerase chain reaction(RT-PCR)and enzymelinked immunosorbent assay(ELISA)to verify the differential genes and proteins.Results:A total of 16 proteins were screened in the peripheral plasma of patients with spinal tuberculosis that were significantly different from those of the control group.Among them,11 were up-regulated[lipopolysaccharide-binding protein(LBP),S100-A8,S100-A9,serum amyloid A-2 protein(SAA2),leucine-rich alpha-2 glycoprotein(LRG2),proteoglycan 4(PRG4),complement factor I(CFI),alpha-1-antitrypsin(SERPINA1),cathepsin D(CTSD),ceruloplasmin(CP),and lambda-immunoglobulin variable 4-60(IGLV4-60)]and 5 proteins were down-regulated[vinculin(VCL),immunoglobulin heavy variable 3-64D(IGHV3-64D),immunoglobulin kappa variable 2D-29(IGKV2D-29),non-functional immunoglobulin kappa variable 6D-41(IGKV6D-41),and immunoglobulin heavy variable 1-45(IGHV1-45)].The gene and protein expressions of LBP,S100-A8,and S100-A9 proteins were significantly elevated in the peripheral plasma of patients with spinal tuberculosis compared w

关 键 词：脊柱结核 蛋白质组学 差异蛋白 信号通路 

分 类 号：R529.2[医药卫生—内科学] R446.7[医药卫生—临床医学]