肺腺癌脑膜转移患者脑脊液蛋白质组分析及临床诊断价值研究

Proteome analysis of cerebrospinal fluid in patients with lung adenocarcinoma with leptomeningeal metastasis and its clinical diagnostic value

目的:肺腺癌脑膜转移(leptomeningeal metastasis,LM)是一种肺腺癌细胞转移至脑膜的严重病变,本研究拟采用液质联用技术分析LM患者脑脊液中蛋白质组的变化,分析其潜在诊断标志物。方法:选取2019年1月至2021年12月天津市环湖医院肺腺癌脑膜转移患者脑脊液30例(试验组)与同期非炎症性神经系统疾病患者脑脊液30例(对照组),采用非标记定量液相色谱-串联质谱法(liquid chromatography-mass spectrometry,LC-MS/MS)检测受试者脑脊液中蛋白质组,并通过基因本体(gene ontolgy,GO)分析差异蛋白相关的生物学过程(biological Process,BP)和细胞组成(cellular component,CC)分布,通过京都基因与基因组百科全书(Kyoto encyclopedia of genes and genomes,KEGG)分析预测差异蛋白生物学功能,通过速率散射比浊试验、免疫透射比浊试验和酶联免疫吸附试验(ELISA)检测差异蛋白验证生物信息学结果,通过受试者工作特征曲线(receiver operating characteristic,ROC)分析差异蛋白检验效能。结果:与对照组患者脑脊液比较,试验组脑脊液蛋白组学结果显示13个蛋白显著上调,6个蛋白显著下调,生物信息学分析发现急性炎症应答和创伤应答是富集差异蛋白较多的生物学过程,细胞外膜结合细胞器、细胞外外泌体和细胞外囊泡等细胞组成分布发生了改变,速率散射比浊试验、免疫透射比浊试验和ELISA结果验证患者组脑脊液中α1微球蛋白(alpha-1-microglobulin,AMBP)、补体C1q/2/7/8B、IgG、肽聚糖识别蛋白2(peptidoglycan recognition protein 2,PGLYRP2)、血清淀粉样蛋白A4(serum amyloid A4,SAA4)显著上调(均P<0.05),ROC分析显示补体C2联合PGLYRP2的曲线下面积(area under curve,AUC)为0.97,灵敏度为83.33%,特异度为96.67%。结论:肺腺癌脑膜转移患者脑脊液免疫应答显著上调,脑脊液补体C2联合PGLYRP2分析可能是潜在肺腺癌脑膜转移辅助诊断手段。

Objective:Leptomeningeal metastasis(LM)from lung adenocarcinoma is a serious disease caused by lung adenocarcinoma cells metastasizing to the meninges.This study aimed to analyze the proteome changes in the cerebrospinal fluid of LM patients by label-free liquid chromatography-mass spectrometry(LC-MS/MS),and identifying the potential diagnostic markers.Methods:Thirty LM patients(patient group),and 30 non-inflammatory neurological-disease patients(control group)were selected from those who visited Tianjin Huanhu Hospital,from 2019 January to 2021 December.The LC-MS/MS was used to detect the proteome in cerebrospinal fluid;Gene Ontology(GO)was used to analyze the biological process(BP)and cellular composition(CC),while their biological functions were predicted by Kyoto encyclopedia of genes and genomes(KEGG)analysis.The above mentioned analysis was certified by rate scatter turbidimetry,immune transmission turbidimetry,and enzyme linked immunosorbent assay(ELISA)in cerebrospinal fluid,while the diagnostic efficiency was analyzed by receiver operating characteristic(ROC)curve.Results:Thirteen and six proteins in the cerebrospinal fluid of the patient group were significantly up-and down-regulated,respectively,when compared with the control group.Bioinformatics analysis revealed the acute inflammatory response and response to wounding,to be the major enriched biological process,while the cell components such as extracellular membrane-bounded organelles,extracellular exosome and extracellular vesicle,were changed.The significantly up-regulated alpha-1-microglobulin(AMBP),complement C1QC/2/7/8B,IgG,peptidoglycan recognition protein 2(PGLYRP2),and serum amyloid A4(SAA4)(all P<0.05)in cerebrospinal fluid,were identified by rate scatter turbidimetry,immune transmission turbidimetry,and ELISA assay.The area under curve(AUC),sensitivity,and specificity of C2+PGLYRP2 were 0.97,83.33%,and 96.67%,respectively.Conclusions:The cerebrospinal fluid immune response is significantly up-regulated in patients with meningeal metastases of lung adenocarcinoma,with C2+PGLYRP2 being a potential adjuvant diagnostic marker.