AIM: To determine the role of p38 MAP kinase signal transduction pathways in diallyl disulfide (DADS)-induced G2/M arrest in human gastric cancer MGC803 cells.METHODS: MGC803 cell growth inhibition was measured by MTT...AIM: To determine the role of p38 MAP kinase signal transduction pathways in diallyl disulfide (DADS)-induced G2/M arrest in human gastric cancer MGC803 cells.METHODS: MGC803 cell growth inhibition was measured by MTT assay. Phase distribution of cell cyde was analyzed by flow cytometry. Expression of Cdc25C, p38, phosphorylation of p38 (pp38) were determined by Western blotting.RESULTS: MT1- assay showed that SB203580, a specific p38 MAPK inhibitor blocked DADS-induced growth inhibition.Flow cytometry analysis revealed that treatment of MGC803 cells with 30 mg/L DADS increased the percentage of cells in the G2/M phase from 9.3% to 39.4% (P<0.05), whereas inhibition of p38 activity by SB203580 abolished induction of G2/M arrest by DADS. Western blotting showed that phosphorylation of p38 was increased 3.52-fold following treatment of MGC803 cells with 30 mg/L DADS for 20 min(P<0.05), whereas Cdc25C was decreased 68% following treatment of MGCS03 cells with 30 mg/L DADS for 24 h(P<0.05). Decreased Cdc25C protein expression by DADS was attenuated by SB203580 (P<0.05).CONCLUSION: DADS-induced G2/M arrest of MGC803 cells involves activation of p38 MAP kinase pathways.Decreased Cdc25C protein expression by p38 MAPK played a crucial role in G2/M arrest after treatment with DADS.展开更多
AIM: To investigate the pathogenic mechanism of Hirschsprung's disease (HD) at the molecular level and to elucidate the relationship between RET oncogene and Chinese patients with HD.METHODS: Exon 13 of RET oncoge...AIM: To investigate the pathogenic mechanism of Hirschsprung's disease (HD) at the molecular level and to elucidate the relationship between RET oncogene and Chinese patients with HD.METHODS: Exon 13 of RET oncogene from 20 unrelated HD patients was analyzed with polymerase chain reactionsingle strand conformation polymorphism (PCR-SSCP). The positive amplifying products were then sequenced. According to the results of SSCP and DNA sequence, SSCP was done as well for the samples from the family other members of some cases with mutated RET gene.RESULTS: SSCP analysis indicated that mobility abnormality existed in 4 unrelated HD patients. Direct DNA sequence analysis identified a missense mutation, T to G at the nucleotide 18 888 and a frameshift mutation at the nucleotide 18 926 insG. In a HD family, the sicked child and his father were the same heterozygous missense mutation (T to G at nucleotide 18 888).CONCLUSION: Among Chinese HD patients, RET gene mutations may exist in considerable proportion with different patterns. These new discoveries indicate that RET mutations may play an important role in the pathogenesis of unrelated HD in the Chinese population. PCR-SSCP combined with DNA sequence can be used as a tool in the genetic diagnosis of HD.展开更多
Objective To construct hu-PBL/SCID chimeras and to investigate the development of lymphoma and oncogenicity of the Epstein-Barr virus (EBV). Methods Human peripheral blood lymphocytes (PBLs) were isolated from healthy...Objective To construct hu-PBL/SCID chimeras and to investigate the development of lymphoma and oncogenicity of the Epstein-Barr virus (EBV). Methods Human peripheral blood lymphocytes (PBLs) were isolated from healthy adult donors and transplanted intraperitoneally into severe combined immunodeficient(SCID) mice. Mice with hu-PBL engraftment from healthy EBV seronegative donors were injected intraperitoneally with EBV-containing supernatant from suspension culture of B95-8 cell line (active infection), whereas mice receiving lymphocytes from healthy EBV seropositive donors were not re-infected with B95-8 derived EBV (latent infection). Pathological examination and molecular analysis were performed on experime-ntal animals and induced neoplasms. Results In the early stage of this experiment, 12 mice died of acute graft-versus-host disease, mortality was 34.3% (12/35 mice) with an average life span of 17.5 days. In 19 survival hu-PBL/SCID chimeric recipients from 12 healthy donors, tumor incidence was 84.2% (16/19 mice). The average survival time of tumor-bearing mice was 65.5 days. EBV-related neoplasms in SCID mice were nodular tumors with aggressive and fatal features. Histological morphology of tumors exhibited diffuse large cell lymphomas. Immunohistochemistry revealed that LCA (CD45) and L26 (CD20) were positive, but both PS1 (CD3) and UCHL-1 (CD45RO) were negative, and EBV products ZEBRA, LMP1, and EBNA2 were expressed in a small number of tumor cells. EB virus particles were seen in the nuclei of some tumor cells by electron microscopy, and EBV DNA could be amplified in the tumor tissues by PCR. In situ hybridization indicated that the nuclei of tumor cells contained human-specific Alu sequence. Conclusions EBV-induced tumors were human B-cell malignant lymphomas. We obtained direct causative evidence dealing with EBV-associated tumor deriving from normal human cells.展开更多
基金Supported by the Natural Science Foundation of Hunan Province,No.02JJY2026,01JJY2146 and the Foundation of Hunan ProvinceEducation Department,No.01A016
文摘AIM: To determine the role of p38 MAP kinase signal transduction pathways in diallyl disulfide (DADS)-induced G2/M arrest in human gastric cancer MGC803 cells.METHODS: MGC803 cell growth inhibition was measured by MTT assay. Phase distribution of cell cyde was analyzed by flow cytometry. Expression of Cdc25C, p38, phosphorylation of p38 (pp38) were determined by Western blotting.RESULTS: MT1- assay showed that SB203580, a specific p38 MAPK inhibitor blocked DADS-induced growth inhibition.Flow cytometry analysis revealed that treatment of MGC803 cells with 30 mg/L DADS increased the percentage of cells in the G2/M phase from 9.3% to 39.4% (P<0.05), whereas inhibition of p38 activity by SB203580 abolished induction of G2/M arrest by DADS. Western blotting showed that phosphorylation of p38 was increased 3.52-fold following treatment of MGC803 cells with 30 mg/L DADS for 20 min(P<0.05), whereas Cdc25C was decreased 68% following treatment of MGCS03 cells with 30 mg/L DADS for 24 h(P<0.05). Decreased Cdc25C protein expression by DADS was attenuated by SB203580 (P<0.05).CONCLUSION: DADS-induced G2/M arrest of MGC803 cells involves activation of p38 MAP kinase pathways.Decreased Cdc25C protein expression by p38 MAPK played a crucial role in G2/M arrest after treatment with DADS.
基金Excellent Young Talented Person by Chinese Health Ministry and Analysis and Testing Foundation of Zhejiang Province
文摘AIM: To investigate the pathogenic mechanism of Hirschsprung's disease (HD) at the molecular level and to elucidate the relationship between RET oncogene and Chinese patients with HD.METHODS: Exon 13 of RET oncogene from 20 unrelated HD patients was analyzed with polymerase chain reactionsingle strand conformation polymorphism (PCR-SSCP). The positive amplifying products were then sequenced. According to the results of SSCP and DNA sequence, SSCP was done as well for the samples from the family other members of some cases with mutated RET gene.RESULTS: SSCP analysis indicated that mobility abnormality existed in 4 unrelated HD patients. Direct DNA sequence analysis identified a missense mutation, T to G at the nucleotide 18 888 and a frameshift mutation at the nucleotide 18 926 insG. In a HD family, the sicked child and his father were the same heterozygous missense mutation (T to G at nucleotide 18 888).CONCLUSION: Among Chinese HD patients, RET gene mutations may exist in considerable proportion with different patterns. These new discoveries indicate that RET mutations may play an important role in the pathogenesis of unrelated HD in the Chinese population. PCR-SSCP combined with DNA sequence can be used as a tool in the genetic diagnosis of HD.
基金Supportedby the National Natural Science Foundation of China(39730200) and by a specialgrantfrom Departmentof Education,Hunan Province(02B040 ).
文摘Objective To construct hu-PBL/SCID chimeras and to investigate the development of lymphoma and oncogenicity of the Epstein-Barr virus (EBV). Methods Human peripheral blood lymphocytes (PBLs) were isolated from healthy adult donors and transplanted intraperitoneally into severe combined immunodeficient(SCID) mice. Mice with hu-PBL engraftment from healthy EBV seronegative donors were injected intraperitoneally with EBV-containing supernatant from suspension culture of B95-8 cell line (active infection), whereas mice receiving lymphocytes from healthy EBV seropositive donors were not re-infected with B95-8 derived EBV (latent infection). Pathological examination and molecular analysis were performed on experime-ntal animals and induced neoplasms. Results In the early stage of this experiment, 12 mice died of acute graft-versus-host disease, mortality was 34.3% (12/35 mice) with an average life span of 17.5 days. In 19 survival hu-PBL/SCID chimeric recipients from 12 healthy donors, tumor incidence was 84.2% (16/19 mice). The average survival time of tumor-bearing mice was 65.5 days. EBV-related neoplasms in SCID mice were nodular tumors with aggressive and fatal features. Histological morphology of tumors exhibited diffuse large cell lymphomas. Immunohistochemistry revealed that LCA (CD45) and L26 (CD20) were positive, but both PS1 (CD3) and UCHL-1 (CD45RO) were negative, and EBV products ZEBRA, LMP1, and EBNA2 were expressed in a small number of tumor cells. EB virus particles were seen in the nuclei of some tumor cells by electron microscopy, and EBV DNA could be amplified in the tumor tissues by PCR. In situ hybridization indicated that the nuclei of tumor cells contained human-specific Alu sequence. Conclusions EBV-induced tumors were human B-cell malignant lymphomas. We obtained direct causative evidence dealing with EBV-associated tumor deriving from normal human cells.
