大鼠膀胱肿瘤热应激后热休克蛋白70肽复合物的纯化及其抑瘤效应  

The antitumor immunological effect in vivo of HSP70-peptides complex derived from rat orthotopic bladder tumor heated by non-invasive pelvic cavity regional moderate hyperthermia

作　　者：张旭[1] 叶章群[1] 米振国[2] 郭素堂[3] 王全红[2] 崔援朝[2] 任连生[3] 韩存芝[3] 

机构地区：[1]华中科技大学同济医学院附属同济医院泌尿外科 [2]山西省肿瘤医院 [3]山西省肿瘤研究所

出　　处：《中华实验外科杂志》2007年第6期717-720,共4页Chinese Journal of Experimental Surgery

基　　金：卫生部医药卫生科技发展研究中心项目;山西省回国留学人员科研项目(200282);山西省自然科学基金项目(2006011130)

摘　　要：目的分离纯化经亚高温盆腔区域热疗处理后大鼠膀胱肿瘤组织中的热休克蛋白70肽复合物(HSP70-PC),研究其对原位的大鼠膀胱肿瘤的治疗效应。方法经尿道膀胱灌注MNU诱导出大鼠膀胱肿瘤,用内生场热疗系统加热大鼠盆腔后取出膀胱肿瘤,用改良的液相色谱法分离纯化HSP70-PC,用SDS-PAGE、Western blot、SELDI-TOF-MS鉴定纯化的HSP70及与其结合的多肽。将大鼠分为肿瘤对照组、HSP70治疗组,治疗1月后进行病理研究。结果纯化获得的蛋白质经SDS-PAGE、Western Mot、SELDI-TOF-MS鉴定,证实为HSP70-PC。HSP70-PC治疗组膀胱及肿瘤湿重低于对照组、肿瘤分期低于对照组(P<0.01)。肿瘤分级两组闻差异无统计学意义(P>0.05)。膀胱肿瘤组织中CD8阳性细胞率、脾脏S-100蛋白阳性细胞率及CD8阳性细胞率、肿瘤细胞凋亡指数均高于对照组(P<0.01)。结论应用改良的液相色谱法可以纯化出高纯度的HSP70- PC,该蛋白复合物具有上调肿瘤鼠的细胞免疫状态,促进肿瘤细胞凋亡的作用。Objective To study the antitumor effect in vivo of HSP70-peptides complexes （HSP70-PC） derived from rat orthotopic bladder tumor heated by non- invasive pelvic cavity regional moderate hyperthermia. Methods Twenty-five F344 rats of six-week-old female inbred line were infused with MNU once every other week for 6 times. The pelves of 15 rats were heated twice at an interval of 24 h to 41℃ by NRL-00II Double RF Tumor Hyperthermia System in a lucite bracket and persisted an h internal. Bladder tumor cell lysate supernatant was injected to Mono Q column, Con-A Sepharose column and 2＇5＇ ADP-Sepharose 4B column in order. Identification of HSP70-PC by SDS-PAGE, Western blot and SELDITOF-MS. Ten rats were separated randomly to groups of HSP70-PC therapy and control. The pathologic changes of tumor and spleen were detected by light microscope respectively after a month treatment. Resuits We obtained high purity HSP70-PC detected by SDS-PAGE, Western blotting and SELDI-TOF-MS respectively. The peptides binding with HSP70 by SELDI-TOF-MS indicated that molecular weight of these peptides ranged from 500 to 2500 Dalton. The therapeutic results indicated that both wet weight of bladder containing tumors and stage of tumor of group of HSP70-PC therapy were significant lower than that of contrel （P 〈 0.01 ）. There was non-significant difference for grade of tumor （ P 〉 0.05 ）. The ratio of CD8 positive cells in tumor, the ratio of S-100 protein positive cells in spleen,the ratio of CD8 positive ceils in spleen and apoptosis index of tumor ceils were significant higher than that of control （ P 〈 0.01 ）. Conclusion HSP70-PC can be purified from rat bladder tumor heated by non-invasive pelvic cavity regional moderate hyperthermia with our improved methods. HSP70-PC can promote both cellular immunity of rat containing orthotopic bladder tumor and tumor cell apoptosis in vivo.

关 键 词：膀胱肿瘤 热休克蛋白70 色谱 模型 动物 

分 类 号：R737.14[医药卫生—肿瘤]