HSP70 and mucin 5B: novel protein targets of N,N'-dinitrosopiperazine-induced nasopharyngeal tumorigenesis.

N,N'-Dinitrosopiperazine (DNP) induces nasopharyngeal carcinoma (NPC) and shows organ specificity to the nasopharyngeal epithelium. To investigate its mechanism, the rat NPC model was induced using DNP. Rat NPC and normal nasopharyngeal cells were obtained from the NPC model using laser capture. The total proteins from these cell samples were separated with two-dimension polyacrylamide gel electrophoresis techniques, and highly expressed proteins (> five-fold) were analyzed using matrix-assisted laser desorption/ionization time of flight and bioinformatics. The results showed that HSP70 and mucin 5B expression increased not only in rat NPC but also in atypical hyperplasia nasopharyngeal tissues, a precancer stage of NPC. High-expression of heat shock protein 70 (HSP70) and mucin 5B was further supported by western blot analysis. The immunofluorescence and western-blotting studies further showed that DNP induced the expression of HSP70 and mucin 5B in a dosage-dependent manner in normal nasopharyngeal epithelia cells. Our data indicate that DNP triggers over-expression of HSP70 and mucin 5B, and is involved in nasopharyngeal tumorigenesis. HSP70 and mucin 5B may be important targets in nasopharyngeal tumorigenesis induced by DNP.

[Activation of anti-tumor cytotoxic T lymphocytes by fusion of human dendritic cells and melanoma cells].

OBJECTIVE: To determine the effect of activation of specific anti-tumor cytotoxic T lymphocytes (CTL) and the ability of cross-presentation in vitro by fusion of HLA-A2+ human dendritic cells (DCs) with HLA-A2- melanoma cells. METHODS: The HLA-A2+ human dendritic cells and HLA-A2- melanoma cells were fused by PEG and were cultivated in complete RPMI1640 media containing FCS (10%) and GM-CSF for 24-48 h, and then co-cultured fusion cells with Melan-A specific T cells. HLA-A2- melanoma cells were negative control,While T2 cells and DC+Pts were positive control. The activation of anti-tumor CTL elicited by the fusion cells was detected by intracellular cytokine staining. RESULTS: The immature DC could express CD80, CD83, CD86, HLA-DR, and HLA-ABC,but the mature DC induced by TNF-alpha, PGE-2, and CD40L further highly expressed above molecules. The rate of specific CTL cells primed by the fusion cells was 16.72%+/-4.26%, negative control was 0.21%+/-1.84%,and positive control was 28.60%+/-5.67%. The CTL from vaccine by fusing DC and LAR6 induced lysis of HLA-A2+ LAR1 cells. CONCLUSION: The HLA-A2 restricted specific anti-tumor CTL can be induced in vitro by fusion of HLA-A2+ human dendritic cells with HLA-A2- melanoma cells.

[Establishment and characterization of a melanoma cell line HME1].

OBJECTIVE: To establish a melanoma cell line and identify its characteristics in vitro. METHODS: The tissues from biopsies of melanoma were performed primary culture. After growing to 90% confluence, the cells were detached and transferred to another flask for subculture, and then we identified their characteristics including growth kinetic, morphology and tumorigenecity. RESULTS: This cell line was cultivated for more than 90 times. Its characteristics were as follows:The pathological morphology of the tumor transplanted in nude mice were similar to that of the original lesion;The growth curves of the 20th passage were determined, and the population doubling time calculated was 56.9 h; The cloning efficiency in soft agar was 19.1%; Karyo-type analysis showed aneuploidy with the modal chromosomal number 85-102; Electronmicroscopical observation showed that there were rich microvilli on the surface of the cells, abundant ribosomes and melanoid grain; SP immunohistochemical staining showed that the cells expressed HMB-45. CONCLUSION: The Melanoma cells were immortalized after being cultured in vitro and a new melanoma cell line was established.