人胃癌组织中分子伴侣蛋白的表达及检测

Expression of chaperone proteins in human gastric cancer

目的:检测人胃癌组织中GRP94/gp96,CRT的表达,分离并鉴定富含分子伴侣的细胞裂解物(CRCL),为研制和开发肿瘤来源的分子伴侣疫苗奠定基础.方法:采用免疫组织化学方法检测人胃癌组织中GRP94/gp96和CRT表达及其表达程度与胃癌病理因素的关系;等电聚焦技术从人胃癌组织匀浆中分离CRCL,经电泳及Westernblot进行蛋白分子质量及性质鉴定.结果:癌组织中GRP94/gp96和CRT表达的阳性率分别为86.3%和84.3%;而癌旁非癌组织中的阳性率分别为37.3%和43.1%,癌组织表达程度显著高于癌旁非癌组织(X2=25.946,18.704,P<0.01);癌组织中GRP94/gp96的表达程度与肿瘤分化程度、浸润深度及淋巴结转移有关,CRT的表达程度与肿瘤大小及淋巴结转移有关.分离的CRCL经鉴定为HSP70,HSP90,GRP94/gp96,CRT.并且这些分子伴侣蛋白沿pH梯度集中分布于5-11号收集管中,其余管中未见明显表达.pH跨度为4.8-5.6.不同组织类型中分子伴侣蛋白的分布及pH跨度具有一致性.结论:胃癌组织中稳定表达GRP94/gp96和CRT,其表达程度与胃癌的发生、发展及其生物学行为有关.

AIM： To detect the expression of GRP94/gp96 and calreticulin （CRT） in gastric cancer, and to extract and identify the chaperone-rich cell lysates （CRCL）. METHODS： Immunohistochemistry was used to detect GRP94/gp96 and CRT expression, and the relationship between the levels of expression and the pathological factors were analyzed. Isoelectric focusing technique was used to separate CRCL from the tumor tissue homogenate. The molecular weight and quality of the purified CRCL were identified by SDS-PAGE and Western blot. RESULTS： The positive rates of GRP94/gp96 and CRT expressive were 86.3% and 84.3% in gastric cancer, respectively, whereas they were 37.3% and 43.1% in cancer-adjacent tissues.The rates were markedly higher in gastric cancer than those in the cancer-adjacent tissues （χ^2 = 25.946, 18.704, both P〈0.01）. The levels of GRP94/ gp96 expression in gastric cancer were correlated with the differentiation, infiltration and lymph metastasis of the cancer and the levels of CRT expression were correlated with the size of tumor and lymph metastasis. Western blot showed that the isolated CRCL contained four kinds of chaperone proteins： GRP94/gp96, HSP90, HSP70 and CRT. The pH value ranged from 4.8 to 5.6 （lane 5-11）. These chaperone proteins clustering along the pH gradient were very typical and reproducible, and the distributions of the proteins and the pH range were accordant among the tissues of different types.