RESUMEN
Choriocarcinoma, a trophoblastic neoplasia, occurs in women as an incidence of abnormal pregnancy. BeWo choriocarcinoma cells derived from the abnormal placentation are a suitable model system to study the factors associated with differentiation, invasion and other cellular events as an alternative to clinical samples. Many protein kinases orchestrate the complex events of cell cycle and in case of malignancy such regulators are found to be mutated. In the present study, BeWo cells treated with forskolin (Fo) and phorbol 12-myristate 13-acetate (PMA) were used to study the role of PKA (protein kinase A) and PKC (protein kinase C), respectively, on the expression pattern of differentiation-related genes, membrane markers, PKC isoforms and cell cycle regulators. The effect of Fo and PMA on the cell proliferation was assessed. Progressive induction of alkaline phosphatase level and formation of multinucleated differentiated cells were observed in the cells treated with Fo. Exposure of cells to Fo and PMA induced the mRNA transcripts of α-hCG, ß-hCG and endoglin and down-regulates E-cadherin at mRNA and protein levels. Synergistic levels of both up- and down-regulated genes/proteins were observed when cells were treated with the combination of Fo and PMA. The mRNA levels of cyclin D1, cyclin E1, p21, Rb, p53, caspase-3 and caspase-8 decreased gradually during differentiation. Fo significantly inhibited the protein levels of PCNA, Rb, PKC-α and PMA stimulated mRNA expression of PKC-ε and PKC-δ. Further, failure in the activation of essential components of the cell cycle machinery caused G2/M phase arrest in differentiating BeWo cells.
Asunto(s)
Proteínas de Ciclo Celular/metabolismo , Ciclo Celular , Diferenciación Celular , Coriocarcinoma/patología , Proteínas Quinasas/metabolismo , Proteínas de Ciclo Celular/genética , Proliferación Celular , Coriocarcinoma/enzimología , Humanos , Proteínas Quinasas/genética , Células Tumorales CultivadasRESUMEN
Androgen receptor (AR) signaling axis plays a vital role in the development of prostate and critical in the progression of prostate cancer. Androgen withdrawal initially regresses tumors but eventually develops into aggressive castration-resistant prostate cancer (CRPC). Activator Protein-1 (AP-1) transcription factors are most likely to be associated with malignant transformation in prostate cancer. Hence, to determine the implication of AR and AP-1 in promoting the transition of prostate cancer to the androgen-independent state, we used AR-positive LNCaP and AR-negative PC-3 cells as an in vitro model system. The effect of dihydrotestosterone or anti-androgen bicalutamide on the cell proliferation and viability was assessed by MTT assay. Expression studies on AR, marker genes-PSA, TMPRSS2, and different AP-1 factors were analyzed by semi-quantitative RT-PCR and expressions of AR and Fra-1 proteins were analyzed by Western blotting. Dihydrotestosterone induced the cell proliferation in LNCaP with no effect on PC-3 cells. Bicalutamide decreased the viability of both LNCaP and PC-3 cells. Dihydrotestosterone induced the expression of AR, PSA, c-Jun, and Fra-1 in LNCaP cells, and it was c-Jun and c-Fos in case of PC-3 cells, while bicalutamide decreased their expression. In addition, constitutive activation and non-regulation of Fra-1 by bicalutamide in PC-3 cells suggested that Fra-1, probably a key component, involved in transition of aggressive androgen-independent PC-3 cells with poor prognosis.
Asunto(s)
Regulación Neoplásica de la Expresión Génica , Neoplasias de la Próstata Resistentes a la Castración/metabolismo , Proteínas Proto-Oncogénicas c-fos/metabolismo , Factor de Transcripción AP-1/metabolismo , Línea Celular Tumoral , Humanos , Masculino , Neoplasias de la Próstata Resistentes a la Castración/genética , Neoplasias de la Próstata Resistentes a la Castración/patología , Proteínas Proto-Oncogénicas c-fos/genética , Factor de Transcripción AP-1/genéticaRESUMEN
Cadmium (Cd) is one of the well-known highly toxic environmental and industrial pollutants. Cd first accumulates in the nucleus and later interacts with zinc finger proteins of antiapoptotic genes and inhibit the binding of transcriptional factors and transcription. However, the role of Cd in oxidative stress and apoptosis is less understood. Hence, the present study was undertaken to unveil the mechanism of action. A549 cells were treated with or without Cd and cell viability was measured by MTT assay. Treatment of cells with Cd shows reduced viability in a dose-dependent manner with IC50 of 45 µM concentration. Cd significantly induces the reactive oxygen species (ROS), lipid peroxidation followed by membrane damage with the leakage of lactate dehydrogenase (LDH). Cells with continuous exposure of Cd deplete the antioxidant super oxide dismutase (SOD) and glutathione peroxidase (GSH-Px) enzymes. Further, analysis of the expression of genes involved in apoptosis show that both the extrinsic and intrinsic apoptotic pathways were involved. Death receptor marker tumor necrosis factor-α (TNF-α), executor caspase-8 and pro-apoptotic gene (Bax) were induced, while antiapoptotic gene (Bcl-2) was decreased in Cd-treated cells. Fluorescence-activated cell sorting (FACS) analysis further confirms the induction of apoptosis in Cd-treated A549 cells.
Asunto(s)
Apoptosis/efectos de los fármacos , Cadmio/toxicidad , Contaminantes Ambientales/toxicidad , Células Epiteliales/efectos de los fármacos , Pulmón/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , Células A549 , Antioxidantes/metabolismo , Técnicas de Cultivo de Célula , Supervivencia Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Células Epiteliales/metabolismo , Células Epiteliales/patología , Citometría de Flujo , Humanos , Peroxidación de Lípido/efectos de los fármacos , Pulmón/metabolismo , Pulmón/patología , Especies Reactivas de Oxígeno/metabolismoRESUMEN
Steroid hormones and their nuclear receptors play a major role in the development and progression of breast cancer. MCF-7 cells are triple-positive breast cancer cells expressing estrogen receptor (ER), progesterone receptor (PR), and glucocorticoid receptor (GR). However, interaction and their role in expression pattern of activator protein (AP-1) transcription factors (TFs) are not completely understood. Hence, in our study, MCF-7 cells were used as an in vitro model system to study the interplay between the receptors and hormones. MCF-7 cells were treated with estradiol-17ß (E2), progesterone (P4), and dexamethasone (Dex), alone or in combination, to study the proliferation of cells and expression of AP-1 genes. MTT assay results show that E2 or P4 induced the cell proliferation by more than 35 %, and Dex decreased the proliferation by 26 %. E2 and P4 are found to increase ERα by more than twofold and c-Jun, c-Fos, and Fra-1 AP-1 TFs by more than 1.7-fold, while Dex shows opposite effect of E2- or P4-induced effect as well as effect on the expression of nuclear receptors and AP-1 factors. E2 antagonist Fulvestrant (ICI 182,780) found to reduce proliferation and E2-induced expression of AP1-TFs, while P4 or Dex antagonist Mifepristone (RU486) is found to block GR-mediated expression of NRs and AP-1 mRNAs. Results suggest that E2 and P4 act synergistically, and Dex acts as an antagonist of E2 and P4.
Asunto(s)
Neoplasias de la Mama/metabolismo , Proliferación Celular/efectos de los fármacos , Receptor alfa de Estrógeno/metabolismo , Proteínas de Neoplasias/metabolismo , Receptores de Glucocorticoides/metabolismo , Receptores de Progesterona/metabolismo , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Dexametasona/farmacología , Estradiol/farmacología , Receptor alfa de Estrógeno/agonistas , Receptor alfa de Estrógeno/genética , Femenino , Humanos , Células MCF-7 , Proteínas de Neoplasias/agonistas , Proteínas de Neoplasias/genética , Progesterona/farmacología , Receptores de Glucocorticoides/agonistas , Receptores de Glucocorticoides/genética , Receptores de Progesterona/agonistas , Receptores de Progesterona/genéticaRESUMEN
Apigenin is one of the plant flavonoids present in fruits and vegetables, acting as an important nutraceutical component. It is recognized as a potential antioxidant, antimicrobial, and anti-inflammatory molecule. In the present study, the mechanism of anti-inflammatory action of apigenin on lipopolysaccharide (LPS)-induced pro-inflammatory cytokines and activator protein-1 (AP-1) factors in human lung A549 cells was investigated. The anti-inflammatory activity of apigenin on LPS-induced inflammation was determined by analyzing the expression of pro-inflammatory cytokines, nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), and different AP-1 factors. Apigenin significantly inhibited the LPS-induced expression of iNOS, COX-2, expression of pro-inflammatory cytokines (IL-1ß, IL-2, IL-6, IL-8, and TNF-α), and AP-1 proteins (c-Jun, c-Fos, and JunB) including nitric oxide production. Study confirms the anti-inflammatory effect of apigenin by inhibiting the expression of inflammatory mediators and AP-1 factors involved in the inflammation and its importance in the treatment of lung inflammatory diseases.
Asunto(s)
Antiinflamatorios/farmacología , Antioxidantes/farmacología , Apigenina/farmacología , Células Epiteliales/metabolismo , Mucosa Respiratoria/metabolismo , Factor de Transcripción AP-1/metabolismo , Línea Celular , Supervivencia Celular/efectos de los fármacos , Ciclooxigenasa 2/biosíntesis , Citocinas/biosíntesis , Citocinas/genética , Humanos , Inflamación/tratamiento farmacológico , Inflamación/inmunología , Lipopolisacáridos/farmacología , Óxido Nítrico/biosíntesis , Óxido Nítrico Sintasa de Tipo II/biosíntesis , ARN Mensajero/genética , Mucosa Respiratoria/citologíaRESUMEN
Acute and chronic alveolar or bronchial inflammation is thought to be central to the pathogenesis of many respiratory disorders. Cytokines and granulocyte macrophage colony-stimulating factors (GM-CSF) play an important role in chronic inflammation. Activator protein-1 (AP-1) the superfamily of transcription factors is involved in proliferation, differentiation, apoptosis, and transformation including inflammation. Understanding the function and regulation of proinflammatory factors involved in inflammation may provide the novel therapeutic strategies in the treatment of inflammatory diseases. Our aim of the present study is to investigate the pro-inflammatory cytokines and pattern of AP-1 factors expressed during activation of lung adenocarcinoma A549 cells by Phorbol-12-myristate-13-acetate (PMA) and to understand the anti-inflammatory effect of apigenin. A549 cells were treated with and without PMA or apigenin, and the cell viability was assessed by MTT assay. Expressions of inflammatory mediators and different AP-1 factors were analyzed by semi-quantitative RT-PCR. IL-6 protein secreted was analyzed by ELISA, and expressions of IL-1ß, c-Jun, and c-Fos proteins were analyzed by Western blotting. Activation of A549 cells by PMA, induced the expression of pro-inflammatory cytokine (IL-1ß, IL-2, IL-6, IL-8, and TNF-α) mRNAs and secretion of IL-6 and the expression of specific AP-1 factors (c-Jun, c-Fos, and Fra-1). Treatment of cells with apigenin, significantly inhibited PMA-stimulated mRNA expression of above pro-inflammatory cytokines, AP-1 factors, cyclooxygenase-2, and secretion of IL-6 protein. Results suggested that the AP-1 factors may be involved in inflammation and apigenin has anti-inflammatory effect, which may be useful for therapeutic management of lung inflammatory diseases.