RESUMO
High molecular weight DNA isolated from 14 primary tumour tissues of human oral carcinoma patients was analysed for transforming activity by NIH3T3 co-transfection assay using pSV2neo gene as a selectable marker, followed by nude mouse tumorigenicity assay. Ten of the patient tumour tissues demonstrated molecular lesions in myc, ras or/and EGF-R genes, whereas 4 patients did not show tumour associated aberrations in these oncogenes. The G418-resistant transfected cells from 12 of 14 individual patients demonstrated transforming potential by colony formation in soft agar and tumour induction in nude mice within 25-80 days. DNAs from the transfected cells, consequent nude mice tumours and corresponding cell lines, contained human Alu sequences. Southern blot hybridisation with ras, myc, EGF-R oncogenes demonstrated the presence of human H-ras oncogene in one of the 12 sets of nude mice tumours. In contrast, DNA from the other 11 sets of nude mice tumours indicated absence of c-myc, N-myc, L-myc, H-ras, K-ras, N-ras and EGF-R genes on Southern analysis. Further, DNAs from five first cycle tumorigenic transformants were subjected to a second cycle of transfection, and induced tumours in nude mice with a shorter latency period of 21-50 days. The secondary transformants contained discrete human Alu sequences; however, the DNA did not hybridise with myc/ras/EGF-R probes. A genomic library was constructed from a second cycle nude mice tumour, using EMBL-3 as the vector. Four human Alu sequence positive clones were isolated on screening 2 x 10(5) plaques, and one of the recombinant clones subjected to fine restriction mapping using 16 restriction enzymes. The lack of association of the nude mice tumour DNA with myc/ras/EGF-R showing aberrations in the primary human tumour, implies activation of an alternative potent transforming gene(s) in the chewing tobacco-related oral carcinomas in India.
Assuntos
Carcinoma de Células Escamosas/genética , Transformação Celular Neoplásica/genética , Neoplasias Bucais/genética , Oncogenes/genética , Plantas Tóxicas , Tabaco sem Fumaça/efeitos adversos , Células 3T3 , Animais , Northern Blotting , Southern Blotting , DNA de Neoplasias/genética , Genes erbB-1/genética , Genes myc/genética , Genes ras/genética , Humanos , Camundongos , Camundongos Nus , TransfecçãoRESUMO
Female nu/+ or BALB/c mice were immunized with human peripheral blood lymphocytes (PBL) before and during pregnancy. Pups born to these mothers were inoculated with human PBL or human fetal bone marrow and thymus cells. Tolerization of the pups to human PBL was observed without graft-versus-host reaction. Presence of human immunoglobulins was observed in the pups for 3-4 weeks. Human T cells also could be detected for a period of 3-4 months in these mice.
