RESUMO
BACKGROUND: Cultivated tea is one of the most important economic and ecological trees distributed worldwide. Cultivated tea suffer from long-term targeted selection of traits and overexploitation of habitats by human beings, which may have changed its genetic structure. The chloroplast is an organelle with a conserved cyclic genomic structure, and it can help us better understand the evolutionary relationship of Camellia plants. RESULTS: We conducted comparative and evolutionary analyses on cultivated tea and wild tea, and we detected the evolutionary characteristics of cultivated tea. The chloroplast genome sizes of cultivated tea were slightly different, ranging from 157,025 to 157,100 bp. In addition, the cultivated species were more conserved than the wild species, in terms of the genome length, gene number, gene arrangement and GC content. However, comparing Camellia sinensis var. sinensis and Camellia sinensis var. assamica with their cultivars, the IR length variation was approximately 20 bp and 30 bp, respectively. The nucleotide diversity of 14 sequences in cultivated tea was higher than that in wild tea. Detailed analysis on the genomic variation and evolution of Camellia sinensis var. sinensis cultivars revealed 67 single nucleotide polymorphisms (SNPs), 46 insertions/deletions (indels), and 16 protein coding genes with nucleotide substitutions, while Camellia sinensis var. assamica cultivars revealed 4 indels. In cultivated tea, the most variable gene was ycf1. The largest number of nucleotide substitutions, five amino acids exhibited site-specific selection, and a 9 bp sequence insertion were found in the Camellia sinensis var. sinensis cultivars. In addition, phylogenetic relationship in the ycf1 tree suggested that the ycf1 gene has diverged in cultivated tea. Because C. sinensis var. sinensis and its cultivated species were not tightly clustered. CONCLUSIONS: The cultivated species were more conserved than the wild species in terms of architecture and linear sequence order. The variation of the chloroplast genome in cultivated tea was mainly manifested in the nucleotide polymorphisms and sequence insertions. These results provided evidence regarding the influence of human activities on tea.
Assuntos
Camellia sinensis , Camellia , Genoma de Cloroplastos , Camellia/genética , Camellia sinensis/genética , Genoma de Cloroplastos/genética , Humanos , Filogenia , CháRESUMO
BACKGROUND Ginkgo biloba extract (EGb761), a standard extract of the Chinese traditional medicine Ginkgo biloba, plays an anti-tumor role in various cancers. However, whether EGb761 is involved in the invasion and metastasis of gastric cancer remains unclear. MATERIAL AND METHODS In the current study, cell viability assay, Western blotting, wound-healing assay, Transwell invasion assay, and orthotopic transplantation model were performed to explore the effects of EGb761 on gastric cancer. RESULTS In vitro, the results showed that EGb761 suppressed the proliferation of gastric cancer cells in a dose-dependent manner. Furthermore, the migration and invasiveness were weakened and the protein levels of p-ERK1/2, NF-kappaB P65, NF-kappaB p-P65, and MMP2 were decreased by EGb761 or U0126 (an inhibitor of ERK signaling pathway) exposure in gastric cancer cells. Moreover, the combined treatment with EGb761 and U0126 significantly inhibited ERK, NF-kappaB signaling pathway, and the expression of MMP2 than those of single drug. In vivo, EGb761 inhibited the tumor growth and hepatic metastasis of gastric cancer in the mouse model. Results of immunohistochemistry indicated that the expression of ERK1/2, NF-kappaB P65 and MMP2 were decreased by EGb761 in the tumor tissues. CONCLUSIONS EGb761 plays a vital role in the suppression of metastasis and ERK/NF-kappaB signaling pathway in gastric cancer.
Assuntos
MAP Quinases Reguladas por Sinal Extracelular/metabolismo , NF-kappa B/metabolismo , Extratos Vegetais/uso terapêutico , Transdução de Sinais , Neoplasias Gástricas/tratamento farmacológico , Neoplasias Gástricas/patologia , Animais , Butadienos/farmacologia , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Ginkgo biloba , Humanos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Metaloproteinase 2 da Matriz/metabolismo , Camundongos Nus , Invasividade Neoplásica , Metástase Neoplásica , Nitrilas/farmacologia , Fosforilação/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacosRESUMO
Systematic chemotherapy is indispensable for gastric cancer patients with advanced stage disease, but the occurrence of chemoresistance drastically limits treatment effectiveness. There is a tremendous need for identifying the underlying mechanism of chemoresistance. NIK and IKKßbinding protein (NIBP) (also known as TRAPPC9, trafficking protein particle complex 9) is a regulator of the cytokineinduced NFκB signaling pathway which has been proven to play pivotal roles in the progression of various malignancies. Nevertheless, it is still ambiguous whether NIBP is involved in the chemoresistance of gastric cancer. The aim of the present study was to investigate the effect of NIBP on chemotherapy resistance of gastric cancer (GC) and to research the mechanisms of Ginkgo biloba extract 761 (EGb 761®) on reversing chemoresistence which has been confirmed in our previous study. In the present study, the results of immumohistochemisty revealed that the positive staining rates of NIBP, NFκB p65 and NFκB pp65 in gastric cancer tissues were obviously higher than those in normal tissues. Furthermore, a close correlation was found to exist between the expression of NIBP and NFκB p65 (pp65) in gastric cancer tissues. Moreover, the overexpression of NIBP was closely related to tumor differentiation, depth of invasion, clinical stage and lymphatic metastasis in gastric cancer. Western blot analysis, realtime PCR, MTT assay and flow cytometric analysis were performed and the results demonstrated that compared with the gastric cancer SGC7901 cells, the expression of NIBP, NFκB p65, NFκB pp65 and mesenchymal marker vimentin were significantly increased in gastric cancer multidrugresistant SGC7901/CDDP cells, and the epithelial cell marker ZO1 was significantly decreased. Meanwhile, it was found that SGC7901/CDDP cells were accompanied by spindlelike mesenchymal appearance and upregulation of stem cell marker CD133 which has been verified to be an upstream regulatory gene of epithelialmesenchymal transition (EMT). Further research confirmed that downregulation of NIBP by Ginkgo biloba extract (EGb) 761 EGb 761 suppressed the cisdiamminedichloroplatinum(II) (CDDP)induced NFκB signaling pathway, EMT and the expression of CD133 in SGC7901 and SGC7901/CDDP cells. Altogether, these data indicate that the NIBPregulated NFκB signaling pathway plays a pivotal role in the chemoresistance of gastric cancer by promoting CD133induced EMT.
Assuntos
Proteínas de Transporte/metabolismo , Resistencia a Medicamentos Antineoplásicos , Transição Epitelial-Mesenquimal , Transdução de Sinais , Neoplasias Gástricas/metabolismo , Regulação para Cima , Adulto , Idoso , Proteínas de Transporte/genética , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Feminino , Ginkgo biloba , Humanos , Peptídeos e Proteínas de Sinalização Intercelular , Masculino , Pessoa de Meia-Idade , NF-kappa B/metabolismo , Extratos Vegetais/farmacologia , Transdução de Sinais/efeitos dos fármacos , Neoplasias Gástricas/genética , Regulação para Cima/efeitos dos fármacosRESUMO
Kinase suppressor of Ras 1 (KSR1) is a scaffold protein that modulates the activation of the oncogenic mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK) signaling pathway. Ginkgo biloba extract (EGb) 761 has been demonstrated to possess antitumor activity that may be related to the KSR1-mediated ERK signaling pathway. However, the roles and its underlying mechanism in gastric cancer are unclear. In the present study, 62 gastric cancer and matched normal tissues were exploited for immunohistochemistry and real-time fluorescent quantitative PCR detection. Results of the immunohistochemistry showed that the expression of ERK1/2 and p-ERK1/2 was correlated to the expression of KSR1 and p-KSR1 in the gastric cancer tissues, and the overexpression of KSR1, p-KSR1, ERK1/2 and p-ERK1/2 was significantly associated with histological grade, TNM stage, lymph node and distant metastasis. Compared with the normal tissues, the relative mRNA copy values of KSR1, ERK1 and ERK2 in the cancer tissues were 2.43 ± 0.49, 2.10 ± 0.44 and 3.65 ± 0.94. In addition, the expression of KSR1, p-KSR1, ERK1/2 and p-ERK1/2 in human gastric cancer multidrug resistant SGC-7901/CDDP cells was higher than that in the SGC-7901 cells as detected by the methods of immunocytochemistry and western blot analysis. EGb 761 not only suppressed expression of these proteins induced by cisplatin (CDDP) and etoposide in SGC-7901 cells, but also inhibited expression of these proteins in the SGC-7901/CDDP cells. Meanwhile, the proliferation-suppressing and apoptosis-inducing capacities of CDDP and etoposide were enhanced following combined treatment with EGb 761. Moreover, EGb 761 reduced the malondialdehyde (MDA) content and elevated the activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) in the tumor cells. These results confirmed that activation of the KSR1-mediated ERK1/2 signaling pathway may contribute to tumorigenesis, metastasis and chemoresistance of human gastric cancer. EGb 761 enhanced the chemotherapy sensitivity and reversed the chemoresistance through suppression of the KSR1-mediated ERK1/2 pathway in gastric cancer cells, and the underlying mechanism may be related to its antioxidative activity.
Assuntos
Proteína Quinase 1 Ativada por Mitógeno/biossíntese , Proteína Quinase 3 Ativada por Mitógeno/biossíntese , Extratos Vegetais/administração & dosagem , Proteínas Quinases/biossíntese , Neoplasias Gástricas/tratamento farmacológico , Idoso , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Cisplatino/uso terapêutico , Variações do Número de Cópias de DNA/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Etoposídeo , Regulação Neoplásica da Expressão Gênica , Ginkgo biloba/química , Humanos , Pessoa de Meia-Idade , Proteína Quinase 1 Ativada por Mitógeno/genética , Proteína Quinase 3 Ativada por Mitógeno/genética , Estresse Oxidativo/efeitos dos fármacos , Extratos Vegetais/química , Proteínas Quinases/genética , Transdução de Sinais/efeitos dos fármacos , Neoplasias Gástricas/genética , Neoplasias Gástricas/patologiaRESUMO
BACKGROUND: Norcantharidin, the demethylated analog of cantharidin derived from a traditional Chinese medicine, Mylabris, has been used in the treatment of anti-cancer effects. However, the detailed mechanisms underlying this process are generally unclear. The aim of this study was to investigate the mechanism of NCTD-induced apoptosis in HepG2 cells. METHODS: The cytotoxicity was measured by MTT assay for cellular viability and by flow cytometry. The mitochondrial membrane potential and reactive oxygen species production was evaluated by flow cytometry analysis. The role of caspase activities were assayed using caspase apoptosis detection kit . Western blot analysis was used to evaluate the level of Cyto-C, Bcl-2, Bax, Bid, caspase 3, -9, -8 and PARP expression RESULTS: After treatment with NCTD, a decrease in the viability of HepG2 cells and increase in apoptosis were observed. NCTD-induced apoptosis was accompanied by an increase in ROS production, loss of mitochondrial membrane potential and release of cytochrome c(cyto-c) from the mitochondria to the cytosol and down-regulation of anti-apoptotic protein Bcl-2 levels with concurrent up-regulation in pro-apoptotic protein Bax levels. However, another pro-apoptotic molecule, Bid, showed no change in such same treatment. NCTD-increased activity of caspase 9,caspase 3 and the subsequent cleavage caspase substrate PARP were also observed. The expression levels of pro-caspase-8 were not changed after NCTD treatment. CONCLUSION: These results indicate that NCTD induced cytotoxicity in HepG2 cells by apoptosis, which is mediated through ROS generation and mitochondrial pathway.
Assuntos
Antineoplásicos Fitogênicos/farmacologia , Apoptose/efeitos dos fármacos , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Medicamentos de Ervas Chinesas/farmacologia , Neoplasias Hepáticas/metabolismo , Mitocôndrias Hepáticas/efeitos dos fármacos , Proteína Agonista de Morte Celular de Domínio Interatuante com BH3/metabolismo , Western Blotting , Caspases/metabolismo , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Citocromos c/metabolismo , Relação Dose-Resposta a Droga , Citometria de Fluxo , Células Hep G2 , Humanos , Neoplasias Hepáticas/patologia , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Mitocôndrias Hepáticas/metabolismo , Mitocôndrias Hepáticas/patologia , Poli(ADP-Ribose) Polimerases/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Fatores de Tempo , Proteína X Associada a bcl-2/metabolismoRESUMO
OBJECTIVE: To research the hereditary stability of Dendrobium huoshanense which were subcultured 7-8 times in the same tissue culture system. METHOD: Using three primers of arbitrary decamer oligonucleotide sequences from 20 primers for DNA amplification, random amplified polymorphic DNA (RAPD) were performed. RESULT: The genetic similarity coefficient was varied from 85.4% to 98.4%. The variation of protocorm, germination and monoleaf were rather more notable than that of bileaf and inflorescence. CONCLUSION: The variation of the same pod is quite small. It is feasible to set up the rapid propagation system of D. huoshanense
Assuntos
DNA de Plantas/genética , Dendrobium/genética , Variação Genética , Plantas Medicinais/genética , Análise de Variância , Primers do DNA , DNA de Plantas/análise , Dendrobium/crescimento & desenvolvimento , Germinação/genética , Plantas Medicinais/crescimento & desenvolvimento , Técnica de Amplificação ao Acaso de DNA Polimórfico , Fatores de Tempo , Técnicas de Cultura de TecidosRESUMO
Oxidative stress can be implicated as a cause of liver fibrosis. In this sense, Ginkgo Biloba Extract (EGB), an antioxidant, may be beneficial in restraining liver fibrosis. The aim of this study was to evaluate the effects of EGB on experimental liver fibrosis. Rat liver fibrosis was induced by intraperitoneal injection of carbon tetrachloride (CCl4) twice a week for 8 weeks. Three groups of rats received EGB (0.25, 0.5 and 1.0 g/kg, respectively) by stomach everyday. CCl4 administration induced liver fibrosis, which was inhibited by EGB in a dose-dependent manner. The histopathologic score of fibrosis, liver function and the levels of plasma hyaluronic acid (HA) and laminin (LN) were significantly improved in rats treated with CCl4 + EGB, compared with those treated with CCl4 only (p < 0.01 or p < 0.05). The activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) were notably elevated, while malondialdehyde (MDA) content was significantly decreased in the rats treated with CCl4 + EGB (p < 0.01 or p < 0.05). Inhibition of hepatic stellate cell (HSC) activation and nuclear factor kappaBP65 (NF-kappaBP65) expression was demonstrated in the livers of EGB-treated rats. The activation of NF-kappaB was significantly suppressed in EGB-treated rats determined by electrophoretic mobility shift assay (EMSA). Furthermore, EGB reduced expressions of transforming growth factor-beta1 (TGF-beta1) and collagen I mRNA. In conclusion, EGB is able to ameliorate liver injury and prevent rats from CCl4-induced liver fibrosis by suppressing oxidative stress. This process may be related to inhibiting the induction of NF-kappaB on HSC activation and the expression of TGF-beta1.
Assuntos
Ginkgo biloba , Cirrose Hepática/tratamento farmacológico , Extratos Vegetais/farmacologia , Alanina Transaminase/sangue , Animais , Aspartato Aminotransferases/sangue , Colágeno Tipo I/genética , Colágeno Tipo I/metabolismo , Citoglobina , Glutationa Peroxidase/metabolismo , Ácido Hialurônico/sangue , Laminina/sangue , Fígado/metabolismo , Cirrose Hepática/induzido quimicamente , Masculino , Malondialdeído/metabolismo , NF-kappa B/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Peroxidases/metabolismo , RNA Mensageiro/metabolismo , Ratos , Ratos Wistar , Superóxido Dismutase/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Fator de Crescimento Transformador beta1RESUMO
Inflammatory mediators play a critical role in ulcerative colitis immune and inflammatory processes. The aim of the study was to investigate the effects of Ginkgo biloba extract on inflammatory mediators (SOD, MDA, TNF-alpha, NF-kappaBp65, IL-6) in TNBS-induced colitis in rats. Colitis in rats was induced by colonic administration with 2,4,6-trinitrobenzene sulfonic acid (TNBS, 150 mg/kg). EGB in doses of (50, 100, 200 mg/kg) was administered for 4 weeks to protect colitis. The results showed that EGB could significantly ameliorate macroscopic and histological damage, evidently elevate the activities of SOD and reduce the contents of MDA, inhibit the protein and mRNA expressions of TNF-alpha, NF-kappaBp65, and IL-6 in the colon tissues of experimental colitis in a dose-dependent manner compared with the model group. We concluded that the probable mechanisms of EGB ameliorated inflammatory injury in TNBS-induced colitis in rats by its modulation of inflammatory mediators and antioxidation.
Assuntos
Colite/tratamento farmacológico , Colite/metabolismo , Ginkgo biloba , Mediadores da Inflamação/metabolismo , Fitoterapia , Animais , Anti-Inflamatórios não Esteroides/administração & dosagem , Anti-Inflamatórios não Esteroides/farmacologia , Antioxidantes/administração & dosagem , Antioxidantes/farmacologia , Sequência de Bases , Colite/genética , Colite/patologia , Interleucina-6/genética , Interleucina-6/metabolismo , Masculino , Malondialdeído/metabolismo , Mesalamina/administração & dosagem , Mesalamina/farmacologia , Extratos Vegetais/administração & dosagem , Extratos Vegetais/farmacologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos , Ratos Wistar , Superóxido Dismutase/metabolismo , Fator de Transcrição RelA/genética , Fator de Transcrição RelA/metabolismo , Ácido Trinitrobenzenossulfônico/toxicidade , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/metabolismoRESUMO
OBJECTIVE: To evaluate the effects of Ginkgo biloba extract (EGB) on CCl(4)-induced liver fibrosis and to investigate the underlying mechanisms. METHODS: Rats were divided into the following groups: normal control group, CCl(4) model group, low dose EGB group, moderate dose EGB group and high dose EGB group. The rat liver fibrosis model was induced by intraperitoneal injection of CCl(4) twice a week for 8 weeks. The model rats of the three EGB treated groups were given 0.25 g/kg, 0.5 g/kg, 1.0 g/kg of EGB by stomach tubes every day. At the end of the eighth week, the blood and liver specimens were obtained. The expressions of nuclear factor kappaB (NF-kappaB) P65, and alpha-smooth muscle actin (alpha-SMA) were detected by immunohistochemistry. Radioimmunoassay was exploited to evaluate serum hyaluronic acid (HA) and laminin (LN) levels. Electrophoretic mobility shift assay (EMSA) was used to confirm the nuclear translocation activity of NF-kappaB in liver tissues. The mRNA expression of transforming growth factor-beta1 (TGFbeta1) and collagen I was determined by RT-PCR. Malondialdehyde (MDA), superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) in liver tissues and alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in the sera were also examined. RESULTS: CCl(4) administration induced liver fibrosis, which was inhibited by EGB in a dose-dependent manner. The histopathologic scores of liver fibrosis, the levels of serum ALT, AST, HA and LN were significantly lower in the rats treated with EGB compared with those not treated (P <0.01 or P <0.05). SOD and GSH-Px activities were notably elevated and MDA content was significantly lower in the rats treated with EGB (P <0.01 or P <0.05), indicating reduced oxidative stress. Immunohistochemical staining demonstrated inhibition of hepatic stellate cell (HSC) activation (in terms of alpha-SMA expression) and NF-kappaB P65 expression in the livers of the EGB-treated rats. As determined by EMSA and RT-PCR, activation of NF-kappaB, the mRNA expression of TGFbeta1 and collagen I were significantly higher in model group rats, but obviously lower in EGB treated rats. CONCLUSION: EGB is able to ameliorate liver injury and prevent rats from CCl(4)-induced liver fibrosis by suppressing oxidative stress. This process may be related to inhibiting the expression of TGFbeta1 and the induction of NF-kappaB on HSC activation.
Assuntos
Medicamentos de Ervas Chinesas/uso terapêutico , Ginkgo biloba , Cirrose Hepática Experimental/tratamento farmacológico , NF-kappa B/biossíntese , Fitoterapia , Fator de Crescimento Transformador beta/biossíntese , Animais , Tetracloreto de Carbono , Intoxicação por Tetracloreto de Carbono , Cirrose Hepática Experimental/induzido quimicamente , Masculino , NF-kappa B/genética , Folhas de Planta , Ratos , Ratos Wistar , Fator de Crescimento Transformador beta/genéticaRESUMO
AIM: To investigate the role of NF-kappaB in the pathogenesis of TNBS-induced colitis in rats. METHODS: Thirty-two healthy adult Sprague-Dawley (SD) rats were randomly divided into four groups of eight each: normal, NS, model I, model II groups in our study. Rat colitis model was established through 2-,4-,6-trinitrobenzene sulfonic acid (TNBS) enema. At the end of four weeks, the macroscopical and histological changes of the colon were examined and mucosa myeloperoxidase (MPO) activities assayed. NF-kappaB p65 expression was determined by Western blot assessment in cytoplasmic and nuclear extracts of colon tissue, and the expressions of TNF-alpha and ICAM-1 protein in colon tissue were examined by immunohistochemistry. The relativities between expression of NF-kappaB p65 and other parameters were analyzed. RESULTS: TNBS enema resulted in pronounced pathological changes of colonic mucosa in model II group (macroscopic and histological injury indices 6.25+/-1.39 and 6.24+/-1.04, respectively), which were in accordance with the significantly elevated MPO activity (1.69+/-0.11). And the nuclear level of NF-kappaB and expression of TNF-alpha, ICAM-1 in rats of model II group were higher than that of normal control (9.7+/-1.96 vs 1.7+/-0.15, 84.09+/-14.52 vs 16.03+/-6.21, 77.69+/-8.09 vs 13.41+/-4.91 P<0.01), Linear correlation analysis revealed that there were strong correlations between the nuclear level of NF-kappaB and the tissue positive expression of TNF-alpha and ICAM-1, MPO activities, macroscopical and histological indices in TNBS-induced colitis, respectively (r = 0.8235, 0.8780, 0.8572, 0.9152, 0.8247; P<0.05). CONCLUSION: NF-kappaB plays a pivotal role in the pathogenesis of ulcerative colitis, which might account for the up-regulation the expression of TNF-alpha and ICAM-1.