A study was conducted on the identifications of the degraded samples of sika deer （Cervus nippon） and red deer （Cervus elaphus） by phylogenetic and nucleotide distance analysis of partial Cytb and 12s rRNA genes s...A study was conducted on the identifications of the degraded samples of sika deer （Cervus nippon） and red deer （Cervus elaphus） by phylogenetic and nucleotide distance analysis of partial Cytb and 12s rRNA genes sequences. 402 bp Cytb genes were achieved by PCR-sequencing using DNA extracted from 8 case samples, and contrasted with 27 sequences of Cytb gene downloaded from GenBank database. The values of three nucleotide distance between three suspected samples and sika deer were identical （0.026±0.006）, which was smaller than the smallest nucleotide distance between eastern red deer and sika deer （0.036）. Furthermore, phylogenetic analysis of sika deer and red deer indicated that the evidences located within the same cluster as sika deer. The evidences were sika deer materials. As the same way, other three suspected samples were derived from red deer. The results were further confirmed by phylogenetic and nucleotide distance analysis of 387 bp 12s rRNA gene. The method was powerful and less time-consuming and helpful to reduce the related cases with wildlife.展开更多
[Objective]The aim was to examine the anti heptocarcinoma effects of human interleukin-12（hIL-12） from transgenic potatoes.[Method]Human heptocarcinoma cell line HepG22.2.15 was cocultured with human peripherial blo...[Objective]The aim was to examine the anti heptocarcinoma effects of human interleukin-12（hIL-12） from transgenic potatoes.[Method]Human heptocarcinoma cell line HepG22.2.15 was cocultured with human peripherial blood monocyte（PBMC）.Human IL-12 extracted from its transgenic patatoes was introduced to this coculture system.MTT method and laser confocal microscope were then employed to evaluate its survival rates and morphological changes of HepG22.2.15 cell line.[Result]The HepG22.2.15 survival rate of the plant produced hIL-12 treated group was significantly lower than that of the wild type control,while similar to that of commercial purified recombinant hIL-12 group.As indicated by AnnexinV/PI double staining laser confocal microscope,cocultured heptocarcinoma cells showed the typical early and final phase apoptotic morphological characteristics 48 hours after 2 × 10-4 mg/L potato secreted hIL-12 treatment.[Conclusion] These results demonstrated that Heptocarcinoma HepG22.2.15 cell growth could be actively inhibited by the transgenic potato expressed hIL-12 in vitro.展开更多
AIM: To detect the variations of mitochondrial 12S rRNA in patients with gastric carcinoma, and to study their significance and the relationship between these variations and the genesis of gastric carcinoma.METHODS: P...AIM: To detect the variations of mitochondrial 12S rRNA in patients with gastric carcinoma, and to study their significance and the relationship between these variations and the genesis of gastric carcinoma.METHODS: PCR amplified mitochondrial 12S rRNA of 44 samples including 22 from gastric carcinoma tissues and 22 from adjacent normal tissues, was detected by direct DNA sequencing. Then laser capture microdissection technique (LCM) was used to separate the cancerous cells and dysplasia cells with specific mutations. Denaturing high performance liquid chromatography (DHPLC) plus allele-specific PCR (ASPCR), nest-PCR and polyacrylamide gel electrophoresis (PAGE)were used to further evaluate this mutant property and quantitative difference of mutant type between cancerous and dysplasia cells. Finally, RNAdraw biosoft was used to analyze the RNA secondary structure of mutant-type 12S rRNA.RESULTS: Compared with Mitomap database, some new variations were found, among which np652 G insertion and np716 T-G transversion were found only in cancerous tissues.There was a statistic difference in the frequency of 12S rRNA variation between intestinal type (12/17, 70.59%) and diffusive type (5/17, 29.41%) of gastric carcinoma (P＜0.05).DHPLC analysis showed that 12S rRNA np652 G insertion and np716 T-G transversion were heteroplasmic mutations.The frequency of 12S rRNA variation in cancerous cells was higher than that in dysplasia cells (P＜0.01). 12S rRNA np652 G insertion showed obviously negative effects on the stability of 12S rRNA secondary structure, while others such as T-G transversion did not.CONCLUSION: The mutations of mitochondrial 12S rRNA may be associated with the occurrence of intestinal-type gastric carcinoma. Most variations exist both in gastric carcinomas and in normal tissues, and they might not be the characteristics of tumors. However, np652 G insertion and np716 T-G transversion may possess some molecular significance in gastric carcinogenesis. During the process from normality to dysplasia, then to carcinoma, 12S rRNA tends to convert from homoplasmy (wild type) to heteroplasmy,then to homoplasmy (mutant type, np717 T-G).展开更多
Objective. To investigate the anti- tumor effects of human single chain interleukin- 12 (hscIL- 12). Method. pcDNA/ hscIL- 12 recombinant was transfected into human hepatic carcinoma cells (7721 cells) by lipofectin m...Objective. To investigate the anti- tumor effects of human single chain interleukin- 12 (hscIL- 12). Method. pcDNA/ hscIL- 12 recombinant was transfected into human hepatic carcinoma cells (7721 cells) by lipofectin method. The 7721/hscIL- 12 cells which secrete hscIL- 12 stably, were obtained via G418 selection, and in vitro the influence of hscIL- 12 gene transduction on the growth of tumor cells was evaluated by cell cycle analysis. In vivo, genetically engineered 7721 cells (7721/hscIL- 12, 7721/pcDNA) and parental cells were implanted into BALB/c nude mice,respectively. 7721/pcDNA and 7721/hscIL- 12 groups were divided into two sub- groups on day 8: one was administered with hPBL twice, 6 days at interval; the other was given equal volume of PBS. Mice were sacrificed on day 26, and spleens and tumors were taken out for histologic assay. Results. hscIL- 12 produced stably by 7721/hscIL- 12 cells had bioactivity, and it was proved by Western blot, immunocytochemistry, and in situ hybridization. In vitro, compared with 7721 and 7721/pcDNA, the 7721/hscIL- 12 grew much more slowly. FACS assay showed apparent G1 arrest of 7721/hscIL- 12 cells. In animal experiment, on day 8 after inoculation, the tumors of 7721 and 7721/pcDNA group were up to 5～ 7mm,while those of 7721/hscIL- 12 group were 2～ 4mm.When treated with hPBL, the tumor of 7721/hscIL- 12 group disappeared completely. Histologically, the tumors from 7721/hscIL- 12 without hPBL treatment had numerous lymphocyte infiltration, the tumor cells displayed depression looking, atrophy, focal necrosis and apoptosis , whereas the tumors of 7721 and 7721/pcDNA groups grew thrivingly. Conclusion. hscIL- 12 transduced 7721 cells could induced significant antitumor immune response which resulted in tumor regression totally when the hPBL was inoculated, and also hscIL- 12 has certain effects on mice immune system. These findings suggest that hscIL- 12 and hscIL- 12 gene therapy might have promising prospects in clinical application.展开更多
文摘A study was conducted on the identifications of the degraded samples of sika deer （Cervus nippon） and red deer （Cervus elaphus） by phylogenetic and nucleotide distance analysis of partial Cytb and 12s rRNA genes sequences. 402 bp Cytb genes were achieved by PCR-sequencing using DNA extracted from 8 case samples, and contrasted with 27 sequences of Cytb gene downloaded from GenBank database. The values of three nucleotide distance between three suspected samples and sika deer were identical （0.026±0.006）, which was smaller than the smallest nucleotide distance between eastern red deer and sika deer （0.036）. Furthermore, phylogenetic analysis of sika deer and red deer indicated that the evidences located within the same cluster as sika deer. The evidences were sika deer materials. As the same way, other three suspected samples were derived from red deer. The results were further confirmed by phylogenetic and nucleotide distance analysis of 387 bp 12s rRNA gene. The method was powerful and less time-consuming and helpful to reduce the related cases with wildlife.
基金Supported by the Key Project of Education Department of Guizhou Province "Establishment of Human Interleukin-12 Plant Expression System" (2004118)~~
文摘[Objective]The aim was to examine the anti heptocarcinoma effects of human interleukin-12（hIL-12） from transgenic potatoes.[Method]Human heptocarcinoma cell line HepG22.2.15 was cocultured with human peripherial blood monocyte（PBMC）.Human IL-12 extracted from its transgenic patatoes was introduced to this coculture system.MTT method and laser confocal microscope were then employed to evaluate its survival rates and morphological changes of HepG22.2.15 cell line.[Result]The HepG22.2.15 survival rate of the plant produced hIL-12 treated group was significantly lower than that of the wild type control,while similar to that of commercial purified recombinant hIL-12 group.As indicated by AnnexinV/PI double staining laser confocal microscope,cocultured heptocarcinoma cells showed the typical early and final phase apoptotic morphological characteristics 48 hours after 2 × 10-4 mg/L potato secreted hIL-12 treatment.[Conclusion] These results demonstrated that Heptocarcinoma HepG22.2.15 cell growth could be actively inhibited by the transgenic potato expressed hIL-12 in vitro.
基金Supported by the National Natural Science Foundation of China,No.30371607
文摘AIM: To detect the variations of mitochondrial 12S rRNA in patients with gastric carcinoma, and to study their significance and the relationship between these variations and the genesis of gastric carcinoma.METHODS: PCR amplified mitochondrial 12S rRNA of 44 samples including 22 from gastric carcinoma tissues and 22 from adjacent normal tissues, was detected by direct DNA sequencing. Then laser capture microdissection technique (LCM) was used to separate the cancerous cells and dysplasia cells with specific mutations. Denaturing high performance liquid chromatography (DHPLC) plus allele-specific PCR (ASPCR), nest-PCR and polyacrylamide gel electrophoresis (PAGE)were used to further evaluate this mutant property and quantitative difference of mutant type between cancerous and dysplasia cells. Finally, RNAdraw biosoft was used to analyze the RNA secondary structure of mutant-type 12S rRNA.RESULTS: Compared with Mitomap database, some new variations were found, among which np652 G insertion and np716 T-G transversion were found only in cancerous tissues.There was a statistic difference in the frequency of 12S rRNA variation between intestinal type (12/17, 70.59%) and diffusive type (5/17, 29.41%) of gastric carcinoma (P＜0.05).DHPLC analysis showed that 12S rRNA np652 G insertion and np716 T-G transversion were heteroplasmic mutations.The frequency of 12S rRNA variation in cancerous cells was higher than that in dysplasia cells (P＜0.01). 12S rRNA np652 G insertion showed obviously negative effects on the stability of 12S rRNA secondary structure, while others such as T-G transversion did not.CONCLUSION: The mutations of mitochondrial 12S rRNA may be associated with the occurrence of intestinal-type gastric carcinoma. Most variations exist both in gastric carcinomas and in normal tissues, and they might not be the characteristics of tumors. However, np652 G insertion and np716 T-G transversion may possess some molecular significance in gastric carcinogenesis. During the process from normality to dysplasia, then to carcinoma, 12S rRNA tends to convert from homoplasmy (wild type) to heteroplasmy,then to homoplasmy (mutant type, np717 T-G).
文摘Objective. To investigate the anti- tumor effects of human single chain interleukin- 12 (hscIL- 12). Method. pcDNA/ hscIL- 12 recombinant was transfected into human hepatic carcinoma cells (7721 cells) by lipofectin method. The 7721/hscIL- 12 cells which secrete hscIL- 12 stably, were obtained via G418 selection, and in vitro the influence of hscIL- 12 gene transduction on the growth of tumor cells was evaluated by cell cycle analysis. In vivo, genetically engineered 7721 cells (7721/hscIL- 12, 7721/pcDNA) and parental cells were implanted into BALB/c nude mice,respectively. 7721/pcDNA and 7721/hscIL- 12 groups were divided into two sub- groups on day 8: one was administered with hPBL twice, 6 days at interval; the other was given equal volume of PBS. Mice were sacrificed on day 26, and spleens and tumors were taken out for histologic assay. Results. hscIL- 12 produced stably by 7721/hscIL- 12 cells had bioactivity, and it was proved by Western blot, immunocytochemistry, and in situ hybridization. In vitro, compared with 7721 and 7721/pcDNA, the 7721/hscIL- 12 grew much more slowly. FACS assay showed apparent G1 arrest of 7721/hscIL- 12 cells. In animal experiment, on day 8 after inoculation, the tumors of 7721 and 7721/pcDNA group were up to 5～ 7mm,while those of 7721/hscIL- 12 group were 2～ 4mm.When treated with hPBL, the tumor of 7721/hscIL- 12 group disappeared completely. Histologically, the tumors from 7721/hscIL- 12 without hPBL treatment had numerous lymphocyte infiltration, the tumor cells displayed depression looking, atrophy, focal necrosis and apoptosis , whereas the tumors of 7721 and 7721/pcDNA groups grew thrivingly. Conclusion. hscIL- 12 transduced 7721 cells could induced significant antitumor immune response which resulted in tumor regression totally when the hPBL was inoculated, and also hscIL- 12 has certain effects on mice immune system. These findings suggest that hscIL- 12 and hscIL- 12 gene therapy might have promising prospects in clinical application.
