RESUMO
BACKGROUND: Because of increased resistance to apoptosis in tumor cells, inhibition of specific anti-apoptotic factors may provide a rational approach for the development of novel therapeutic strategies. Livin, a novel inhibitor of apoptosis protein family, has been found to be expressed in various malignancies and is suggested to have poorly prognostic significance. However, no data are available concerning the significance of livin in gastric cancer. In this study, we detected the expression of livin in human gastric carcinoma and investigated the apoptotic susceptibility of SGC - 7901 cell by shRNA-mediated silencing of the livin gene. METHODS: The mRNA and protein expression of livin were analyzed by RT-PCR and western blot assay. The relationship between livin expression and clinical pathologic parameters was investigated. The small interfering RNA eukaryotic expression vector specific to livin was constructed by gene recombination, and the nucleic acid was sequenced. Then it was transfected into SGC-7901 cells by Lipofectamin 2000. RT-PCR and Western blot assay were used to validate gene-silencing efficiency of livin in SGC-7901 cells. Stable clones were obtained by G418 screening. The cell apoptosis was assessed by flow cytometry (FCM). Cell growth state and 50 % inhibition concentration (IC50) of 5-FU and cisplatin was determined by MTT method. RESULTS: The expression of livin mRNA and protein were detected in 19 of 40 gastric carcinoma cases (47.5%) and SGC-7901 cells. No expression of livin was detected in tumor adjacent tissues and benign gastric lesion. The positive correlation was found between livin expression and poor differentiation of tumors as well as lymph node metastases (P<0.05). Four small interfering RNA eukaryotic expression vector specific to livin were constructed by gene recombination. And one of them can efficiently decrease the expression of livin, the inhibition of the gene was not less than 70% (P<0.01). The recombinated plasmids were extracted and transfected gastric cancer cells. The stable clones were obtained by G418 screening, and were amplified and cultured. When livin gene was silenced, the reproductive activity of the gastric cancer cells was significantly lower than the control groups(P<0.05). The study also showed that IC50 of 5-Fu and cisplatin on gastric cancer cells treated by shRNA was decreased and the cells were more susceptible to proapoptotic stimuli (5-Fu and cisplatin) (P<0.01). CONCLUSIONS: Livin is overexpressed in gastric carcinoma with a relationship to tumor differentiation and lymph node metastases, which is suggested to be one of the molecular prognostic factors for some cases of gastric cancer. ShRNA can inhibit livin expression in SGC-7901 cells and induce cell apoptosis. Livin may serve as a new target for apoptosis-inducing therapy of gastric cancer.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Regulação Neoplásica da Expressão Gênica , Inativação Gênica , Proteínas Inibidoras de Apoptose/genética , Proteínas de Neoplasias/genética , Neoplasias Gástricas/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Adenocarcinoma/genética , Adulto , Idoso , Antineoplásicos/administração & dosagem , Antineoplásicos/farmacologia , Apoptose/genética , Western Blotting , Linhagem Celular Tumoral , Cisplatino/administração & dosagem , Cisplatino/farmacologia , Feminino , Citometria de Fluxo , Fluoruracila/administração & dosagem , Fluoruracila/farmacologia , Humanos , Proteínas Inibidoras de Apoptose/metabolismo , Concentração Inibidora 50 , Metástase Linfática , Masculino , Pessoa de Meia-Idade , Proteínas de Neoplasias/metabolismo , RNA Mensageiro/metabolismo , RNA Interferente Pequeno/genética , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Two new benzo[j]fluoranthene-based secondary metabolites named daldinone C (1) and daldinone D (2), along with two known metabolites, altechromone A and (4S)-5,8-dihydroxy-4-methoxy-alpha-tetralone, were isolated from the CHCl3/MeOH (1:1) extract of a solid culture of the endophyte Hypoxylon truncatum IFB-18 harbored inside the symptomless stem tissue of Artemisia annua. The structures of the new compounds were elucidated by MS and 1D and 2D NMR spectra and by X-ray diffraction analysis. Their absolute configurations were determined unambiguously by a combination of their CD data and the established exciton chirality rule. Compounds 1 and 2 were substantially cytotoxic against SW1116 cells, with IC50 values of 49.5 and 41.0 microM, respectively, comparable to that (37.0 microM) of 5-fluorouracil. The biosynthetic pathway for 1 and 2 was postulated with the natural occurrence of benzo[j]fluoranthene analogues discussed in brief.
Assuntos
Antineoplásicos , Ascomicetos/química , Fluorenos , Antineoplásicos/química , Antineoplásicos/isolamento & purificação , Antineoplásicos/farmacologia , Artemisia annua , China , Cristalografia por Raios X , Ensaios de Seleção de Medicamentos Antitumorais , Fluorenos/química , Fluorenos/isolamento & purificação , Fluorenos/farmacologia , Humanos , Concentração Inibidora 50 , Conformação Molecular , Estrutura Molecular , Plantas MedicinaisRESUMO
In addition to 7-methoxy-2-methyl-3,4,5-trihydroxyanthraquinone (1), physcion (2), macrosporin (3), deoxybostrycin (4), altersolanol B (5) and dactylariol (6), a new hexahydroanthraquinone named pleospdione (7) was isolated from the culture of Pleospora sp . IFB-E006, an endophytic fungus residing in the normal stem of Imperata cylindrical (Gramineae). Structure determination of pleospdione was accomplished using IR, HR-ESI-MS, 1D and 2D NMR spectral analysis. Compounds 4 - 6 exhibited significant cytotoxic activity against human colon cancer (SW1116) and leukemia (K562) cell lines while compounds 1, 2 and 7 were only weakly or moderately active.