应用定量蛋白质组学法筛选与斜坡脊索瘤骨侵袭程度相关的差异表达蛋白  

Screening of differentially expressed proteins associated bone invasion in patients with clivus chordoma using quantitative proteomics method

作　　者：王亮[1] 王科[2] 张扬[1] 郭正光 田凯兵[2] 张力伟[1] 张俊廷[1] 于春江[4] 万虹[5] 吴震[1] 

机构地区：[1]首都医科大学附属北京天坛医院神经外科,100050 [2]首都医科大学研究生院 [3]中国医学科学院基础医学研究所 [4]首都医科大学三博脑科医院神经外科 [5]北京市神经外科研究所神经损伤修复研究室

出　　处：《中华神经外科杂志》2015年第5期508-513,共6页Chinese Journal of Neurosurgery

基　　金：北京市自然科学基金（7142052）;国家自然科学基金（81472370）

摘　　要：目的 筛选与斜坡脊索瘤骨侵袭程度相关的蛋白.方法 收集起源于蝶岩交界区斜坡脊索瘤患者的肿瘤标本,共35例.12例行同位素标记相对和绝对定量（iTRAQ）蛋白质组学试验（试验组）,23例行免疫组化染色验证（验证组）.依照骨侵袭程度分型,试验组中膨胀内生型（Ⅰ型）和外生型（Ⅱ型）各6例;验证组Ⅰ和Ⅱ型分别有10例和13例.试验组应用iTRAQ法行差异蛋白检测,应用Panther软件行Gene Ontology功能分析,应用IPA软件行蛋白通路与网络分析.结果 共2 251个蛋白获得定量鉴定结果,Ⅰ、Ⅱ型斜坡脊索瘤有250个蛋白存在差异表达,与Ⅱ型相比,Ⅰ型59个蛋白表达上调,191个蛋白下调.Ⅰ和Ⅱ型脊索瘤细胞在哺乳动物雷帕霉素靶蛋白（mTOR）通路表达水平差异有统计学意义（P＜0.01）,Ⅰ型的炎性活性相关蛋白表达增强,细胞外基质蛋白和细胞骨架蛋白表达水平低于Ⅱ型.分子网络分析提示,可以调控细胞基质蛋白和骨架蛋白表达的转化生长因子β1（TGFβ1）表达水平降低.免疫组化显示,验证组中TGFβ1、第10号染色体缺失的磷酸酶和张力蛋白同源等位基因（PTEN）的表达阳性率分别为95.6％ （22/23）、69.6％ （16/23）,与Ⅰ型比较,Ⅱ型脊索瘤TGFβ1、PTEN的表达强度高（P =0.033,P=0.004）;验证组中mTOR的阳性表达率为80.7％ （20/23）,Ⅰ和Ⅱ型间mTOR的阳性表达率差异无统计学意义（P=0.092）.结论 TGFβ1可能在斜坡脊索瘤的骨侵袭过程中发挥重要作用,机制可能与其介导炎性细胞反应增高及细胞骨架蛋白表达下降有关.PI3K-AKt-mTOR信号通道可能与脊索瘤的骨侵袭发生机制有关,但mTOR蛋白可能与脊索瘤的骨侵袭程度无关.PTEN的表达水平可能与脊索瘤的骨侵袭程度有关.Objective To screen the proteins associated with the degree of bone invasion in clivus chordomas.Methods A total of 35 tumor specimens of patients with clivus chordoma originated from the spheno petrosal junction were collected.Twelve specimens of isobaric tags for relative and absolute quantitation （iTRAQ） proteomics experiment （experimental group） were performed,23 were validated by immunohistochemical staining （validated group）.According to the classification criteria of bone invasive degree,they were classified into endophytic type （Type Ⅰ） or exophytic type （Type Ⅱ） （n =6 in each group） in the experimental group.Type Ⅰ and type Ⅱ in the validated group were 10 and 13 specimens respectively.The iTRAQ technique was used for the detection of the difference proteins.Panther software was used for functional analysis of gene ontology （GO）.IPA software was used for protein pathway and network analysis.Results A total of 2251 proteins obtained quantitative identification results,250 proteins had difference expression in type Ⅰ and type Ⅱ clivus chordomas.Compared with type Ⅱ,59 proteins were up-regulated and 191 were down-regulated in type Ⅰ.There was significant difference in the expression levels of chordomas cells of the mammalian target of rapamycin （mTOR） pathway between the type Ⅰ and type Ⅱ chordoma cells （P 〈0.01）.The inflammatory activity-associated protein expression of type Ⅰ enhanced.The expression levels of extracellular matrix proteins and cytoskeletal proteins were lower than those of type Ⅱ.Molecular network analysis suggested that the expression levels of transforming growth factor beta 1 （TGFβ31） of regulating the cell matrix protein and cytoskeleton protein expression were low.Immunohistochemistry showed that the expression positive rates of TGFβ1,phosphatase and tensin homolog deleted on chromosome ten （PTEN） in the validated group were 95.6% （22/23） and 69.6% （16/23）.Compare with type Ⅰ,the expression intensity of

关 键 词：脊索瘤 蛋白质组学 同位素标记 骨侵袭性 

分 类 号：R738.1[医药卫生—肿瘤]