RESUMO
Zinc-based nanostructures are known for their numerous potential biomedical applications. In this context, the biosynthesis of nanostructures using plant extracts has become a more sustainable and promising alternative to effectively replace conventional chemical methods while avoiding their toxic impact. In this study, following a low-temperature calcination process, a green synthesis of Zn-hydroxide-based nanostructure has been performed using an aqueous extract derived from the leaves of Litchi chinensis, which is employed as a lignocellulose waste biomass known to possess a variety of phytocompounds. The biogenic preparation of Zn-hydroxide based nanostructures is enabled by bioactive compounds present in the leaf extract, which act as reducing and capping agents. In order to evaluate its physicochemical characteristics, the produced Zn-hydroxide-based nanostructure has been subjected to several characterization techniques. Further, the multifunctional properties of the prepared Zn-hydroxide-based nanostructure have been evaluated for antioxidant, antimicrobial, and anticancer activity. The prepared nanostructure showed antibacterial efficacy against Bacillus subtilis and demonstrated its anti-biofilm activity as evaluated through the Congo red method. In addition, the antioxidant activity of the prepared nanostructure has been found to be dose-dependent, wherein 91.52 % scavenging activity could be recorded at 200 µg/ml, with an IC50 value of 45.22 µg/ml, indicating the prepared nanostructure has a high radical scavenging activity. Besides, the in vitro cytotoxicity investigation against HepG2 cell lines explored that the as-prepared nanostructure exhibited a higher cytotoxic effect and 73.21 % cell inhibition could be noticed at 25.6 µg/ml with an IC50 of 2.58 µg/ml. On the contrary, it was found to be significantly lower in the case of HEK-293 cell lines, wherein ~47.64 % inhibition could be noticed at the same concentration. These findings might be further extended to develop unique biologically derived nanostructures that can be extensively evaluated for various biomedical purposes.
Assuntos
Litchi , Nanopartículas Metálicas , Nanoestruturas , Humanos , Antioxidantes/farmacologia , Antioxidantes/química , Litchi/química , Biomassa , Células HEK293 , Antibacterianos/farmacologia , Antibacterianos/química , Hidróxidos , Extratos Vegetais/farmacologia , Extratos Vegetais/química , Nanopartículas Metálicas/químicaRESUMO
Ultraviolet B (UVB) radiation induces regulatory T cells (Treg cells) and depletion of these Treg cells alleviates immunosuppression and inhibits photocarcinogenesis in mice. Here, we determined the effects of dietary grape seed proanthocyanidins (GSPs) on the development and activity of UVB-induced Treg cells. C3H/HeN mice fed a GSPs (0.5%, w/w)-supplemented or control diet were exposed to UVB (150 mJ/cm2) radiation, sensitized to 2,4-dinitrofluorobenzene (DNFB) and sacrificed 5 days later. FACS analysis indicated that dietary GSPs decrease the numbers of UVB-induced Treg cells. ELISA analysis of cultured sorted Treg cells indicated that secretion of immunosuppressive cytokines (interleukin-10, TGF-ß) was significantly lower in Treg cells from GSPs-fed mice. Dietary GSPs also enhanced the ability of Treg cells from wild-type mice to stimulate production of IFNγ by T cells. These effects of dietary GSPs on Treg cell function were not found in XPA-deficient mice, which are incapable of repairing UVB-induced DNA damage. Adoptive transfer experiments revealed that naïve recipients that received Treg cells from GSPs-fed UVB-irradiated wild-type donors that had been sensitized to DNFB exhibited a significantly higher contact hypersensitivity (CHS) response to DNFB than mice that received Treg cells from UVB-exposed mice fed the control diet. There was no significant difference in the CHS response between mice that received Treg cells from UVB-irradiated XPA-deficient donors fed GSPs or the control diet. Furthermore, dietary GSPs significantly inhibited UVB-induced skin tumor development in wild-type mice but not in XPA-deficient mice. These results suggest that GSPs inactivate Treg cells by promoting DNA repair in dendritic cells in UVB-exposed skin.
Assuntos
Reparo do DNA por Junção de Extremidades/efeitos dos fármacos , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/fisiologia , Extrato de Sementes de Uva/farmacologia , Proantocianidinas/farmacologia , Fenômenos Fisiológicos da Pele/efeitos da radiação , Pele/efeitos da radiação , Linfócitos T Reguladores/efeitos dos fármacos , Linfócitos T Reguladores/fisiologia , Transferência Adotiva , Animais , Biomarcadores , Citocinas/metabolismo , Imunomodulação , Imunofenotipagem , Interferon gama/metabolismo , Camundongos , Camundongos Knockout , Neoplasias Cutâneas/etiologia , Neoplasias Cutâneas/metabolismo , Neoplasias Cutâneas/patologia , Microambiente Tumoral , Raios UltravioletaRESUMO
Lung cancer and its metastasis is the leading cause of cancer-related mortality world-wide. Non-small cell lung cancer (NSCLC) accounts for about 90% of total lung cancer cases. Despite advancements in therapeutic approaches, only limited improvement has been achieved. Therefore, alternative strategies are required for the management of lung cancer. Here we report the chemotherapeutic effect of silymarin, a phytochemical from milk thistle plant (Silybum marianum L. Gaertn.), on NSCLC cell migration using metastatic human NSCLC cell lines (A549, H1299 and H460) together with the molecular targets underlying these effects. Using an in vitro cell migration assay, we found that treatment of human NSCLC cells (A549, H1299 and H460) with silymarin (0, 5, 10 and 20 µg/mL) for 24 h resulted in concentration-dependent inhibition of cell migration, which was associated with the inhibition of histone deacetylase (HDAC) activity and reduced levels of class 1 HDAC proteins (HDAC1, HDAC2, HDAC3 and HDAC8) and concomitant increases in the levels of histone acetyltransferase activity (HAT). Known HDAC inhibitors (sodium butyrate and trichostatin A) exhibited similar patterns of therapeutic effects on the lung cancer cells. Treatment of A549 and H460 cells with silymarin reduced the expression of the transcription factor ZEB1 and restored expression of E-cadherin. The siRNA knockdown of ZEB1 also reduced the expression of HDAC proteins and enhanced re-expression of the levels of E-cadherin in NSCLC cells. MicroRNA-203 (miR-203) acts as a tumor suppressor, regulates tumor cell invasion and is repressed by ZEB1 in cancer cells. Silymarin treatment restored the levels of miR-203 in NSCLC cells. These findings indicate that silymarin can effectively inhibit lung cancer cell migration and provide a coherent model of its mechanism of action suggesting that silymarin may be an important therapeutic option for the prevention or treatment of lung cancer metastasis when administered either alone or with standard cancer therapeutic drugs.
RESUMO
Melanoma is a highly aggressive form of skin cancer with poor survival rate. Aberrant activation of Wnt/ß-catenin has been observed in nearly one-third of human melanoma cases thereby indicating that targeting Wnt/ß-catenin signaling could be a promising strategy against melanoma development. In the present study, we determined chemotherapeutic effect of grape seed proanthocyanidins (GSPs) on the growth of melanoma cells and validated their protective effects in vivo using a xenograft mouse model, and assessed if ß-catenin is the target of GSP chemotherapeutic effect. Our in vitro data show that treatment of A375 and Hs294t human melanoma cells with GSPs inhibit the growth of melanoma cells, which was associated with the reduction in the levels of ß-catenin. Administration of dietary GSPs (0.2 and 0.5%, w/w) in supplementation with AIN76A control diet significantly inhibited the growth of melanoma tumor xenografts in nude mice. Furthermore, dietary GSPs inhibited the xenograft growth of Mel928 (ß-catenin-activated), while did not inhibit the xenograft growth of Mel1011 (ß-catenin-inactivated) cells. These observations were further verified by siRNA knockdown of ß-catenin and forced overexpression of ß-catenin in melanoma cells using a cell culture model.
Assuntos
Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Extrato de Sementes de Uva/farmacologia , Melanoma/tratamento farmacológico , Melanoma/metabolismo , Proantocianidinas/farmacologia , beta Catenina/metabolismo , Animais , Linhagem Celular Tumoral , Feminino , Humanos , Camundongos , Camundongos Nus , Ensaios Antitumorais Modelo de Xenoenxerto/métodosRESUMO
Here, we report the chemotherapeutic effect of honokiol, a phytochemical from Magnolia plant, on human head and neck squamous cell carcinoma (HNSCC). Treatment of HNSCC cell lines from different sub-sites, SCC-1 (oral cavity), SCC-5 (larynx), OSC-19 (tongue) and FaDu (pharynx) with honokiol inhibited their cell viability, which was associated with the: (i) induction of apoptosis, (ii) correction of dysregulatory cell cycle proteins of G0/G1 phase. Honokiol decreased the expression levels of epidermal growth factor receptor (EGFR), mTOR and their downstream signaling molecules. Treatment of FaDu and SCC-1 cell lines with rapamycin, an inhibitor of mTOR pathway, also reduced cell viability of HNSCC cells. Administration of honokiol by oral gavage (100 mg/kg body weight) significantly (P < 0.01-0.001) inhibited the growth of SCC-1 and FaDu xenografts in athymic nude mice, which was associated with: (i) inhibition of tumor cell proliferation, (ii) induction of apoptosis, (iii) reduced expressions of cyclins and Cdks, and (iv) inhibition of EGFR signaling pathway. Molecular docking analysis of honokiol in EGFR binding site indicated that the chemotherapeutic effect of honokiol against HNSCC is mediated through its firm binding with EGFR, which is better than that of gefitinib, a commonly used drug for HNSCC treatment.
Assuntos
Antineoplásicos Fitogênicos/uso terapêutico , Compostos de Bifenilo/farmacologia , Carcinoma de Células Escamosas/tratamento farmacológico , Receptores ErbB/metabolismo , Neoplasias de Cabeça e Pescoço/tratamento farmacológico , Lignanas/farmacologia , Animais , Apoptose , Sítios de Ligação , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células , Separação Celular , Sobrevivência Celular , Relação Dose-Resposta a Droga , Receptores ErbB/química , Feminino , Citometria de Fluxo , Humanos , Magnolia/química , Camundongos , Camundongos Nus , Simulação de Acoplamento Molecular , Fitoterapia/métodos , Extratos Vegetais/química , Transdução de Sinais , Sirolimo/químicaRESUMO
Melanoma is a highly aggressive form of skin cancer and a leading cause of death from skin diseases mainly due to its propensity to metastasis. Due to metastatic tendency, melanoma is often associated with activation of Wnt/ß-catenin signaling mechanism. Blocking ß-catenin activation may be a good strategy to block melanoma-associated mortality. We have shown earlier that grape seed proanthocyanidins (GSPs) inhibit melanoma cell migration via targeting cyclooxygenase-2 (COX-2) overexpression. Here we explored further whether inhibition of inflammatory mediators-mediated activation of ß-catenin by GSPs is associated with the inhibition of melanoma cell migration. Our study revealed that PGE2 receptors (EP2 and EP4) agonists promote melanoma cell migration while PGE2 receptor antagonist suppressed the migration capacity of melanoma cells. GSPs treatment inhibit butaprost (EP2 agonist) or Cay10580 (EP4 agonist) induced migration of melanoma cells. Western blot analysis revealed that GSPs reduced cellular accumulation of ß-catenin, and decreased the expressions of matrix metalloproteinase (MMP)-2, MMP-9 and MITF, downstream targets of ß-catenin in melanoma cells. GSPs also reduced the protein expressions of PI3K and p-Akt in the same set of experiment. To verify that ß-catenin is a specific molecular target of GSPs, we compared the effect of GSPs on cell migration of ß-catenin-activated (Mel1241) and ß-catenin-inactivated (Mel1011) melanoma cells. GSPs inhibit cell migration of Mel1241 cells but not of Mel1011 cells. Additionally, in vivo bioluminescence imaging data indicate that dietary administration of GSPs (0.5%, w/w) in supplementation with AIN76A control diet inhibited the migration/extravasation of intravenously injected melanoma cells in lungs of immune-compromised nude mice, and that this effect of GSPs was associated with an inhibitory effect on the activation of ß-catenin and its downstream targets, such as MMPs, in lungs as a target organ.
RESUMO
The green tea polyphenol, (-)-epigallocatechin-3-gallate (EGCG), has been shown to have anti-carcinogenic effects in several skin tumor models, and efforts are continued to investigate the molecular targets responsible for its cytotoxic effects to cancer cells. Our recent observation that ß-catenin is upregulated in skin tumors suggested the possibility that the anti-skin carcinogenic effects of EGCG are mediated, at least in part, through its effects on ß-catenin signaling. We have found that treatment of the A431 and SCC13 human skin cancer cell lines with EGCG resulted in reduced cell viability and increased cell death and that these cytotoxic effects were associated with inactivation of ß-catenin signaling. Evidence of EGCG-induced inactivation of ß-catenin included: (i) reduced accumulation of nuclear ß-catenin; (ii) enhanced levels of casein kinase1α, reduced phosphorylation of glycogen synthase kinase-3ß, and increased phosphorylation of ß-catenin on critical serine(45,33/37) residues; and (iii) reduced levels of matrix metalloproteinase (MMP)-2 and MMP-9, which are down-stream targets of ß-catenin. Treatment of cells with prostaglandin E2 (PGE2) enhanced the accumulation of ß-catenin and enhanced ß-catenin signaling. Treatment with either EGCG or an EP2 antagonist (AH6809) reduced the PGE2-enhanced levels of cAMP, an upstream regulator of ß-catenin. Inactivation of ß-catenin by EGCG resulted in suppression of cell survival signaling proteins. siRNA knockdown of ß-catenin in A431 and SCC13 cells reduced cell viability. Collectively, these data suggest that induction of cytotoxicity in skin cancer cells by EGCG is mediated by targeting of ß-catenin signaling and that the ß-catenin signaling is upregulated by inflammatory mediators.
Assuntos
Anticarcinógenos/administração & dosagem , Catequina/análogos & derivados , Polifenóis/administração & dosagem , Neoplasias Cutâneas , Chá , beta Catenina/antagonistas & inibidores , Catequina/administração & dosagem , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Relação Dose-Resposta a Droga , Sistemas de Liberação de Medicamentos/métodos , Humanos , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Neoplasias Cutâneas/tratamento farmacológico , Neoplasias Cutâneas/metabolismo , beta Catenina/metabolismoRESUMO
Lung cancer remains a leading cause of death due to its metastasis to distant organs. We have examined the effect of honokiol, a bioactive constituent from the Magnolia plant, on human non-small cell lung cancer (NSCLC) cell migration and the molecular mechanisms underlying this effect. Using an in vitro cell migration assay, we found that treatment of A549, H1299, H460 and H226 NSCLC cells with honokiol resulted in inhibition of migration of these cells in a dose-dependent manner, which was associated with a reduction in the levels of cyclooxygenase-2 (COX-2) and prostaglandin E2 (PGE2). Celecoxib, a COX-2 inhibitor, also inhibited cell migration. Honokiol inhibited PGE2-enhanced migration of NSCLC cells, inhibited the activation of NF-κB/p65, an upstream regulator of COX-2, in A549 and H1299 cells, and treatment of cells with caffeic acid phenethyl ester, an inhibitor of NF-κB, also inhibited migration of NSCLC cells. PGE2 has been shown to activate ß-catenin signaling, which contributes to cancer cell migration. Therefore, we checked the effect of honokiol on ß-catenin signaling. It was observed that treatment of NSCLC cells with honokiol degraded cytosolic ß-catenin, reduced nuclear accumulation of ß-catenin and down-regulated matrix metalloproteinase (MMP)-2 and MMP-9, which are the down-stream targets of ß-catenin and play a crucial role in cancer cell metastasis. Honokiol enhanced: (i) the levels of casein kinase-1α, glycogen synthase kinase-3ß, and (ii) phosphorylation of ß-catenin on critical residues Ser(45), Ser(33/37) and Thr(41). These events play important roles in degradation or inactivation of ß-catenin. Treatment of celecoxib also reduced nuclear accumulation of ß-catenin in NSCLC cells. FH535, an inhibitor of Wnt/ß-catenin pathway, inhibited PGE2-enhanced cell migration of A549 and H1299 cells. These results indicate that honokiol inhibits non-small cell lung cancer cells migration by targeting PGE2-mediated activation of ß-catenin signaling.
Assuntos
Compostos de Bifenilo/farmacologia , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Movimento Celular/efeitos dos fármacos , Dinoprostona/metabolismo , Lignanas/farmacologia , Neoplasias Pulmonares/metabolismo , Transdução de Sinais/efeitos dos fármacos , beta Catenina/metabolismo , Compostos de Bifenilo/química , Carcinoma Pulmonar de Células não Pequenas/patologia , Linhagem Celular Tumoral , Movimento Celular/genética , Núcleo Celular/metabolismo , Ciclo-Oxigenase 2/metabolismo , Inibidores de Ciclo-Oxigenase 2/farmacologia , Medicamentos de Ervas Chinesas/química , Medicamentos de Ervas Chinesas/farmacologia , Técnicas de Silenciamento de Genes , Humanos , Lignanas/química , Neoplasias Pulmonares/patologia , Invasividade Neoplásica , Interferência de RNA , Sulfonamidas/farmacologia , Fator de Transcrição RelA/metabolismoRESUMO
Ultraviolet (UV) radiation-induced immunosuppression has been implicated in skin carcinogenesis. Grape seed proanthocyanidins (GSPs) have anti-skin carcinogenic effects in mice and GSPs-fed mice exhibit a reduction in UV-induced suppression of allergic contact hypersensitivity (CHS), a prototypic T-cell-mediated response. Here, we report that dietary GSPs did not inhibit UVB-induced suppression of CHS in xeroderma pigmentosum complementation group A (XPA)-deficient mice, which lack nucleotide excision repair mechanisms. GSPs enhanced repair of UVB-induced DNA damage (cyclobutane pyrimidine dimers) in wild-type, but not XPA-deficient, dendritic cells (DC). Co-culture of CD4(+) T cells with DCs from UVB-irradiated wild-type mice resulted in suppression of T-cell proliferation and secretion of T-helper (TH) 1-type cytokines that was ameliorated when the DCs were obtained from GSP-fed mice, whereas DCs obtained from GSP-fed XPA-KO mice failed to restore T-cell proliferation. In adoptive transfer experiments, donor DCs were positively selected from the draining lymph nodes of UVB-exposed donor mice that were sensitized to 2,4,-dinitrofluorobenzene were transferred into naïve recipient mice and the CHS response assessed. Naïve recipients that received DCs from UVB-exposed wild-type donors that had been fed GSPs exhibited a full CHS response, whereas no significant CHS was observed in mice that received DCs from XPA-KO mice fed GSPs. These results suggest that GSPs prevent UVB-induced immunosuppression through DNA repair-dependent functional activation of dendritic cells in mice. Cancer Prev Res; 6(3); 242-52. ©2013 AACR.
Assuntos
Células Dendríticas/efeitos dos fármacos , Extrato de Sementes de Uva/farmacologia , Proantocianidinas/farmacologia , Pele/efeitos da radiação , Raios Ultravioleta/efeitos adversos , Animais , Southern Blotting , Western Blotting , Dano ao DNA/efeitos dos fármacos , Células Dendríticas/imunologia , Dermatite de Contato/imunologia , Feminino , Sistema Imunitário/efeitos dos fármacos , Sistema Imunitário/efeitos da radiação , Tolerância Imunológica/efeitos dos fármacos , Tolerância Imunológica/imunologia , Camundongos , Camundongos Knockout , Pele/efeitos dos fármacos , Pele/imunologia , Vitis/químicaRESUMO
Grape seed proanthocyanidins (GSPs) have been shown to have anti-skin carcinogenic effects in in vitro and in vivo models. However, the precise epigenetic molecular mechanisms remain unexplored. This study was designed to investigate whether GSPs reactivate silenced tumor suppressor genes following epigenetic modifications in skin cancer cells. For this purpose, A431 and SCC13 human squamous cell carcinoma cell lines were used as in vitro models. The effects of GSPs on DNA methylation, histone modifications and tumor suppressor gene expressions were studied in these cell lines using enzyme activity assays, western blotting, dot-blot analysis and real-time polymerase chain reaction (RT-PCR). We found that treatment of A431 and SCC13 cells with GSPs decreased the levels of: (i) global DNA methylation, (ii) 5-methylcytosine, (iii) DNA methyltransferase (DNMT) activity and (iv) messenger RNA (mRNA) and protein levels of DNMT1, DNMT3a and DNMT3b in these cells. Similar effects were noted when these cancer cells were treated identically with 5-aza-2'-deoxycytidine, an inhibitor of DNA methylation. GSPs decreased histone deacetylase activity, increased levels of acetylated lysines 9 and 14 on histone H3 (H3-Lys 9 and 14) and acetylated lysines 5, 12 and 16 on histone H4, and reduced the levels of methylated H3-Lys 9. Further, GSP treatment resulted in re-expression of the mRNA and proteins of silenced tumor suppressor genes, RASSF1A, p16(INK4a) and Cip1/p21. Together, this study provides a new insight into the epigenetic mechanisms of GSPs and may have significant implications for epigenetic therapy in the treatment/prevention of skin cancers in humans.
Assuntos
Antineoplásicos/farmacologia , Carcinoma de Células Escamosas/genética , Inativação Gênica/efeitos dos fármacos , Genes Supressores de Tumor/efeitos dos fármacos , Extrato de Sementes de Uva/farmacologia , Proantocianidinas/farmacologia , Neoplasias Cutâneas/genética , Azacitidina/análogos & derivados , Azacitidina/farmacologia , Western Blotting , Carcinoma de Células Escamosas/tratamento farmacológico , Linhagem Celular Tumoral , Metilação de DNA/efeitos dos fármacos , Decitabina , Histona Acetiltransferases/metabolismo , Humanos , Immunoblotting , Reação em Cadeia da Polimerase em Tempo Real , Neoplasias Cutâneas/tratamento farmacológicoRESUMO
Lung cancer remains the leading cause of cancer-related deaths worldwide, and non-small cell lung cancer (NSCLC) represents approximately 80% of total lung cancer cases. The use of non-toxic dietary phytochemicals can be considered as a chemotherapeutic strategy for the management of the NSCLC. Here, we report that grape seed proanthocyanidins (GSPs) induce apoptosis of NSCLC cells, A549 and H1299, in vitro which is mediated through increased expression of pro-apoptotic protein Bax, decreased expression of anti-apoptotic proteins Bcl2 and Bcl-xl, disruption of mitochondrial membrane potential, and activation of caspases 9, 3 and poly (ADP-ribose) polymerase (PARP). Pre-treatment of A549 and H1299 cells with the caspase-3 inhibitor (z-DEVD-fmk) significantly blocked the GSPs-induced apoptosis of these cells confirmed that GSPs-induced apoptosis is mediated through activation of caspases-3. Treatments of A549 and H1299 cells with GSPs resulted in an increase in G1 arrest. G0/G1 phase of the cell cycle is known to be controlled by cyclin dependent kinases (Cdk), cyclin-dependent kinase inhibitors (Cdki) and cyclins. Our western blot analyses showed that GSPs-induced G1 cell cycle arrest was mediated through the increased expression of Cdki proteins (Cip1/p21 and Kip1/p27), and a simultaneous decrease in the levels of Cdk2, Cdk4, Cdk6 and cyclins. Further, administration of 50, 100 or 200 mg GSPs/kg body weight of mice by oral gavage (5 d/week) markedly inhibited the growth of s.c. A549 and H1299 lung tumor xenografts in athymic nude mice, which was associated with the induction of apoptotic cell death, increased expression of Bax, reduced expression of anti-apoptotic proteins and activation of caspase-3 in tumor xenograft cells. Based on the data obtained in animal study, human equivalent dose of GSPs was calculated, which seems affordable and attainable. Together, these results suggest that GSPs may represent a potential therapeutic agent for the non-small cell lung cancer.
Assuntos
Apoptose/efeitos dos fármacos , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Extrato de Sementes de Uva/farmacologia , Neoplasias Pulmonares/tratamento farmacológico , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Proantocianidinas/farmacologia , Animais , Carcinoma Pulmonar de Células não Pequenas/patologia , Caspase 3/metabolismo , Ciclo Celular/efeitos dos fármacos , Proteínas de Ciclo Celular , Linhagem Celular Tumoral , Relação Dose-Resposta a Droga , Extrato de Sementes de Uva/administração & dosagem , Extrato de Sementes de Uva/uso terapêutico , Humanos , Neoplasias Pulmonares/patologia , Camundongos , Proantocianidinas/administração & dosagem , Proantocianidinas/uso terapêutico , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Melanoma is the most serious type of skin disease and a leading cause of death from skin disease due to its highly metastatic ability. To develop more effective chemopreventive agents for the prevention of melanoma, we have determined the effect of green tea catechins on the invasive potential of human melanoma cells and the molecular mechanisms underlying these effects using A375 (BRAF-mutated) and Hs294t (Non-BRAF-mutated) melanoma cell lines as an in vitro model. Employing cell invasion assays, we found that the inhibitory effects of green tea catechins on the cell migration were in the order of (-)-epigallocatechin-3-gallate (EGCG)>(-)-epigallocatechin>(-)-epicatechin-3-gallate>(-)-gallocatechin>(-)-epicatechin. Treatment of A375 and Hs294t cells with EGCG resulted in a dose-dependent inhibition of cell migration or invasion of these cells, which was associated with a reduction in the levels of cyclooxygenase (COX)-2, prostaglandin (PG) E(2) and PGE(2) receptors (EP2 and EP4). Treatment of cells with celecoxib, a COX-2 inhibitor, also inhibited melanoma cell migration. EGCG inhibits 12-O-tetradecanoylphorbol-13-acetate-, an inducer of COX-2, and PGE(2)-induced cell migration of cells. EGCG decreased EP2 agonist (butaprost)- and EP4 agonist (Cay10580)-induced cell migration ability. Moreover, EGCG inhibited the activation of NF-κB/p65, an upstream regulator of COX-2, in A375 melanoma cells, and treatment of cells with caffeic acid phenethyl ester, an inhibitor of NF-κB, also inhibited cell migration. Inhibition of melanoma cell migration by EGCG was associated with transition of mesenchymal stage to epithelial stage, which resulted in an increase in the levels of epithelial biomarkers (E-cadherin, cytokeratin and desmoglein 2) and a reduction in the levels of mesenchymal biomarkers (vimentin, fibronectin and N-cadherin) in A375 melanoma cells. Together, these results indicate that EGCG, a major green tea catechin, has the ability to inhibit melanoma cell invasion/migration, an essential step of metastasis, by targeting the endogenous expression of COX-2, PGE(2) receptors and epithelial-to-mesenchymal transition.
Assuntos
Catequina/farmacologia , Ciclo-Oxigenase 2/metabolismo , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Melanoma/enzimologia , Melanoma/patologia , Receptores de Prostaglandina E/metabolismo , Chá/química , Catequina/análogos & derivados , Catequina/química , Celecoxib , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Inibidores de Ciclo-Oxigenase 2/farmacologia , Dinoprostona/biossíntese , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , NF-kappa B/metabolismo , Invasividade Neoplásica , Pirazóis/farmacologia , Receptores de Prostaglandina E Subtipo EP2/agonistas , Receptores de Prostaglandina E Subtipo EP2/metabolismo , Receptores de Prostaglandina E Subtipo EP4/agonistas , Receptores de Prostaglandina E Subtipo EP4/metabolismo , Sulfonamidas/farmacologia , Acetato de Tetradecanoilforbol/farmacologia , Fator de Transcrição RelA/metabolismoRESUMO
Melanoma is the leading cause of death from skin disease due, in large part, to its propensity to metastasize. We have examined the effect of grape seed proanthocyanidins (GSPs) on melanoma cancer cell migration and the molecular mechanisms underlying these effects using highly metastasis-specific human melanoma cell lines, A375 and Hs294t. Using in vitro cell invasion assays, we observed that treatment of A375 and Hs294t cells with GSPs resulted in a concentration-dependent inhibition of invasion or cell migration of these cells, which was associated with a reduction in the levels of cyclooxygenase (COX)-2 expression and prostaglandin (PG) E(2) production. Treatment of cells with celecoxib, a COX-2 inhibitor, or transient transfection of melanoma cells with COX-2 small interfering RNA, also inhibited melanoma cell migration. Treatment of cells with 12-O-tetradecanoylphorbol-13-acetate, an inducer of COX-2, enhanced the phosphorylation of ERK1/2, a protein of mitogen-activated protein kinase family, and subsequently cell migration whereas both GSPs and celecoxib significantly inhibited 12-O-tetradecanoylphorbol-13-acetate-promoted cell migration as well as phosphorylation of ERK1/2. Treatment of cells with UO126, an inhibitor of MEK, also inhibited the migration of melanoma cells. Further, GSPs inhibited the activation of NF-κB/p65, an upstream regulator of COX-2, in melanoma cells, and treatment of cells with caffeic acid phenethyl ester, an inhibitor of NF-κB, also inhibited cell migration. Additionally, inhibition of melanoma cell migration by GSPs was associated with reversal of epithelial-mesenchymal transition process, which resulted in an increase in the levels of epithelial biomarkers (E-cadherin and cytokeratins) while loss of mesenchymal biomarkers (vimentin, fibronectin and N-cadherin) in melanoma cells. Together, these results indicate that GSPs have the ability to inhibit melanoma cell invasion/migration by targeting the endogenous expression of COX-2 and reversing the process of epithelial-to-mesenchymal transition.
Assuntos
Dinoprostona/biossíntese , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Extrato de Sementes de Uva/farmacologia , Proantocianidinas/farmacologia , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Ciclo-Oxigenase 2/genética , Ciclo-Oxigenase 2/metabolismo , Transição Epitelial-Mesenquimal/genética , Humanos , Imunoensaio , TransfecçãoRESUMO
In the present study, we report the effects of honokiol, a phytochemical from Magnolia spp., on cancer cell migration capacity and the molecular mechanisms underlying these effects using breast cancer cell lines as an in vitro model. Using cell migration assays, we found that the treatment of human breast cancer cells (MCF-7) and murine mammary cancer cells (4T1) with honokiol resulted in a dose-dependent inhibition of migration of these cells, which was associated with a reduction in nitric oxide (NO) levels. The cell migration capacity was decreased in the presence of NG-nitro-L-arginine methyl ester (L-NAME), an inhibitor of NO synthase. Honokiol reduced the elevated levels of cyclic guanosine monophosphate (cGMP) in the cells, while the treatment of 4T1 cells with guanylate cyclase (GC) inhibitor 1-H-[1,2,4]oxadiaxolo[4,3-a]quinolalin-1-one (ODQ) reduced the migration of cells and the levels of cGMP. The presence of 8-bromoguanosine 3'5'-cyclic monophosphate, an analogue of cGMP, enhanced the migration of these cells, suggesting a role for GC in the migration of 4T1 cells. Honokiol also inhibited the levels of cyclooxygenase-2 (COX-2) and prostaglandin (PG) E2 in 4T1 cells. The transfection of 4T1 cells with COX-2 siRNA resulted in a reduction in cell migration. ODQ and L-NAME also decreased the levels of PGE2 in 4T1 cells suggesting a role for COX-2/PGE2 in cell migration. Moreover, honokiol inhibited the activation of nuclear factor κB (NF-κB), an upstream regulator of COX-2 and iNOS, in 4T1 cells. These results indicate that NO and COX-2 are the key targets of honokiol in the inhibition of breast cancer cell migration, an essential step in invasion and metastasis.
Assuntos
Compostos de Bifenilo/farmacologia , Neoplasias da Mama/patologia , Carcinoma/patologia , Movimento Celular/efeitos dos fármacos , Ciclo-Oxigenase 2/metabolismo , Lignanas/farmacologia , Magnolia/química , Óxido Nítrico/metabolismo , Animais , Antineoplásicos Fitogênicos/farmacologia , Neoplasias da Mama/metabolismo , Carcinoma/metabolismo , Linhagem Celular Tumoral , Avaliação Pré-Clínica de Medicamentos , Medicamentos de Ervas Chinesas/química , Medicamentos de Ervas Chinesas/farmacologia , Feminino , Humanos , Camundongos , Modelos Biológicos , Terapia de Alvo Molecular , Óxido Nítrico/antagonistas & inibidores , Fitoterapia/métodosRESUMO
The inhibition of UVB-induced immunosuppression by dietary grape seed proanthocyanidins (GSP) has been associated with the induction of interleukin (IL)-12 in mice, and we now confirm that GSPs do not inhibit UVB-induced immunosuppression in IL-12p40 knockout (IL-12 KO) mice and that treatment of these mice with recombinant IL-12 restores the inhibitory effect. To characterize the cell population responsible for the GSP-mediated inhibition of UVB-induced immunosuppression and the role of IL-12 in this process, we used an adoptive transfer approach. Splenocytes and draining lymph nodes were harvested from mice that had been administered dietary GSPs (0.5%-1.0%, w/w), exposed to UVB, and sensitized by the application of 2,4-dinitrofluorobenzene (DNFB) onto the UVB-exposed skin. CD8(+) and CD4(+) T cells were positively selected and transferred into naive mice that were subsequently challenged by application of DNFB on the ear skin. Naive recipients that received CD8(+) T cells from GSP-treated, UVB-irradiated donors exhibited full contact hypersensitivity (CHS) response. Naive mice that received CD4(+) suppressor T cells from GSP-treated, UVB-exposed mice could mount a CHS response after sensitization and subsequent challenge with DNFB. On culture, the CD8(+) T cells from GSP-treated, UVB-exposed mice secreted higher levels (5- to 8-fold) of Th1 cytokines than CD8(+) T cells from UVB-irradiated mice not treated with GSPs. CD4(+) T cells from GSP-treated, UVB-exposed mice secreted significantly lower levels (80%-100%) of Th2 cytokines than CD4(+) T cells from UVB-exposed mice not treated with GSPs. These data suggest that GSPs inhibit UVB-induced immunosuppression by stimulating CD8(+) effector T cells and diminishing regulatory CD4(+) T cells.
Assuntos
Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Dermatite Irritante/imunologia , Extrato de Sementes de Uva/farmacologia , Tolerância Imunológica/efeitos dos fármacos , Subunidade p40 da Interleucina-12/fisiologia , Proantocianidinas/farmacologia , Raios Ultravioleta , Transferência Adotiva , Animais , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/imunologia , Células Dendríticas/efeitos da radiação , Dermatite Irritante/etiologia , Suplementos Nutricionais , Dinitrofluorbenzeno/farmacologia , Feminino , Extrato de Sementes de Uva/administração & dosagem , Linfonodos/citologia , Ativação Linfocitária/efeitos da radiação , Camundongos , Camundongos Endogâmicos CBA , Camundongos Knockout , Proantocianidinas/administração & dosagem , Pele/efeitos dos fármacos , Pele/imunologia , Pele/efeitos da radiação , Baço/citologia , Vitis/químicaRESUMO
In recent years, a considerable emphasis has been focused on the importance of the naturally available botanicals that can be consumed in an individual's everyday diet and that can also be useful as a chemopreventive or chemotherapeutic agent for certain diseases, including cancers. A wide variety of botanicals, mostly dietary flavonoids or polyphenolic substances, have been reported to possess substantial anti-carcinogenic and antimutagenic activities because of their antioxidant and anti-inflammatory properties. Proanthocyanidins are considered as one of them, and are abundantly available in various parts of the plants, such as fruits, berries, bark and seeds. Their modes of action were evaluated through a number of in vitro and in vivo studies which showed their potential role as anti-carcinogenic agent. We summarize and highlight the latest developments on anti-carcinogenic activities of proanthocyanidins from different sources, specifically from grape seeds, and their molecular targets, such as NF-kappaB, mitogen-activated protein kinases, PI3K/Akt, caspases, cytokines, angiogenesis and cell cycle regulatory proteins and other check points, etc. Although the bioavailability and metabolism data on proanthocyanidins is still largely unavailable, certain reports indicate that at least monomers and smaller oligomeric procyanidins are absorbed in the gut. The modulation of various molecular targets by proanthocyanidins in vitro and in vivo tumor models suggests their importance, contribution and mechanism of action to the prevention of cancers of different organs.