The study on the expression of membrane HSP70 protein in H22 cell and its immunoprotective mechanism against carcinoma.
The mRNA and protein expression of HSP70 were studied using RT-PCR and FCM techniques. It's immune protective effects in vivo and the cytotoxicity in vitro were also observed. Results showed that cell viability didn't change under 42-43 degrees C, but declined in 44-45 degrees C; the level of HSP70 mRNA decreased initially (0.5-4.0 hour) but gradually resumed and increased from 8 to 12 hours at 42 degrees C. The positive cells expressed membrane HSP70 were significantly higher in heat shocked group than in a control group (P<0.001); the highest positive rate was 96.8% at 43 degrees C. The C3H mice immunized with heat shocked H22 cells could resist a secondary subcutaneous inoculation of parental H22 cells and their tumorigenic rate was significantly lower than that in mice immunized with parental H22 cells and RPMI 1640 control mice, which were 1/9.7/8 and 8/8, respectively. Their median survival time was also longer than that of parental cell group and RPMI 1640 control group, which were >90 days, 73.5 days, and 39.5 days, respectively. Through heat-treated tumor cell and lymphocyte mixed culture (TLMC), the induced lymphocytes had higher cytotoxic activities than that of splenic cells. The cytotoxicity against H22 cells reached 65.38% (2 hrs, 42 degrees C) and 67.84% (12 hrs, 43 degrees C) and could be blocked by anti-HSP70 McAb. Phenotype analysis revealed that the rate of TCR gammadelta+ cells rose with increasing cytotoxic activity, but no similar changes could be found in the CD4+ CD8+ TCR alphabeta+ subset. These results suggest that proper heat shock conditions can improve the immunogenicity of tumor cells and CD4- CD8- TCR gammadelta+ T carried on cytotoxic function via the HSP70 molecule.
Wu
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WZ
Liver Cancer Institute, Zhong Shan Hospital, Shanghai Medical University, P.R. China.
Liu
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KD
Xie
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Wu
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HS
D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D002470:Cell Survival; D003602:Cytotoxicity, Immunologic; D018840:HSP70 Heat-Shock Proteins / Q000096:biosynthesis / Q000235:genetics*; D006358:Hot Temperature; D008114:Liver Neoplasms, Experimental / Q000235:genetics* / Q000276:immunology / Q000473:pathology* / Q000517:prevention & control; D007959:Lymphocyte Culture Test, Mixed; D008214:Lymphocytes / Q000276:immunology; D051379:Mice; D008809:Mice, Inbred C3H; D008810:Mice, Inbred C57BL; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured
D018840:HSP70 Heat-Shock Proteins; D012333:RNA, Messenger