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目的摸索鼠肝癌Ｈ２２细胞膜热休克蛋白７０（ＨＳＰ７０）最适的诱导条件，并明确其体内外的抗瘤机理。方法用免疫荧光及ＦＣＭ法检测热休克Ｈ２２细胞膜ＨＳＰ７０蛋白的动态表达。并用ＭＭＣ灭活、热休克后的Ｈ２２细胞作抗原进行体内肿瘤免疫保护试验及体外肿瘤杀伤试验。结果小鼠肝癌Ｈ２２细胞经４２℃、４３℃热休克１２小时后，其胞膜ＨＳＰ７０蛋白表达率分别达到５９．５％，９６．８％。均较对照组显著增高（Ｐ＜０．００１）。用４３℃休克１２小时的Ｈ２２细胞免疫的Ｃ３Ｈ小鼠能抵抗同种野生型肿瘤细胞的攻击，其肿瘤发生率比野生型Ｈ２２免疫组及未免疫组均显著降低，分别为１／９、７／８和８／８。同时小鼠荷瘤生存期显著延长，中位生存期分别为＞９０天，７３．５天及３９．５天。体外杀伤试验表明，经热休克Ｈ２２诱导７天后的Ｃ３Ｈ脾淋巴细胞对热休克型Ｈ２２细胞杀伤率高达６３．３８％（４３℃，２小时）和６７．８４％（４３℃，１２小时），且这种杀伤活性可被抗鼠ＨＳＰ７０的单抗所阻断。分析效应细胞的表型发现，ＣＤ４＋、ＣＤ８＋淋巴细胞在整个体外诱导过程中并不增加，而ＴＣＲγδ＋淋巴细胞随诱导时间的延长与杀伤活性同步增加，诱导第７天为３０．４３％，第１４天?
Objective To explore the optimal conditions for the induction of heat shock protein 70 (HSP70) in hepatoma H22 cells and to determine its anti-tumor mechanism in vitro and in vivo. Methods The dynamic expression of HSP70 protein in heat shock H22 cell membrane was detected by immunofluorescence and FCM. The M22-inactivated and heat-shocked H22 cells were used as antigens to perform in vivo tumor immunoprotection assay and in vitro tumor killing assay. Results After H22 cells were treated with heat shock at 42°C and 43°C for 12 hours, the HSP70 protein expression rate of membrane H22 cells reached 59.5% and 96.8%, respectively. Both were significantly higher than the control group (P<0.001). C3H mice immunized with H22 cells shocked at 43°C for 12 hours were able to resist the challenge of the same type of wild-type tumor cells, and the tumor incidence was significantly lower than that of the wild-type H22 immunized group and the non-immune group, which were 1/9 and 7 respectively. /8 and 8/8. At the same time, the tumor survival time of mice was significantly prolonged. The median survival time was >90 days, 73.5 days and 39.5 days, respectively. In vitro killing tests showed that the killing rate of heat shock H22 cells in C3H spleen lymphocytes was as high as 63.38% (43°C, 2 hours) and 67.84% (43°C, 12 hours) after 7 days of heat shock H22 induction. And this killing activity can be blocked by the monoclonal antibody against mouse HSP70. Analysis of the phenotype of effector cells revealed that CD4+ and CD8+ lymphocytes did not increase during in vitro induction, whereas TCRγδ+ lymphocytes increased synchronously with the prolongation of induction time and killing activity. The induction day was 30.43%. day?