RESUMO
Kaiso is a Pox Virus and Zinc Finger (POZ-ZF) transcription factor with bi-modal DNA-binding specificity. Here, we demonstrated that Kaiso expression is inversely correlated with glucocorticoid receptor (GR) expression in breast carcinomas. Knockdown of Kaiso increased GR expression, while overexpression of Kaiso inhibited GR expression in breast cancer cells. Furthermore, Kaiso repressed GR proximal promoter-reporter activity in a dose-dependent manner. Remarkably, ChIP experiments demonstrated that endogenous Kaiso was associated with the GR promoter sequence in a methylation-dependent manner. Since glucocorticoids inhibit chemotherapyinduced apoptosis and have been widely used as a co-treatment of patients with breast cancer, we assessed the role of Kasio in GR-mediated anti-apoptotic effects. We found that overexpression of Kaiso attenuated the anti-apoptotic effects of glucocorticoids in breast cancer cells. Our findings suggest that GR is a putative target gene of Kaiso and suggest Kaiso to be a potential therapeutic target in GC-combination chemotherapy in breast cancer. [BMB Reports 2016; 49(3): 167-172].
Assuntos
Apoptose/efeitos dos fármacos , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Glucocorticoides/farmacologia , Receptores de Glucocorticoides/metabolismo , Fatores de Transcrição/metabolismo , Apoptose/genética , Sequência de Bases , Neoplasias da Mama/metabolismo , Linhagem Celular Tumoral , Ilhas de CpG/genética , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Metilação/efeitos dos fármacos , Regiões Promotoras Genéticas/genética , Ligação Proteica/efeitos dos fármacos , Receptores de Glucocorticoides/genética , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Fatores de Transcrição/genéticaRESUMO
AIMS: Advanced oxidation protein products (AOPPs) are new independent risk factor for coronary artery disease. This study was to determine the effects and potential mechanisms of AOPPs on cholesterol efflux from human macrophage foam cells. METHODS: Human THP-1 monocytes were preincubated with Phorbol-12-myristate- 13-acetate (PMA) and oxidized low density lipoprotein (ox-LDL) to form foam cells. The protein and mRNA expression were examined by western immunoblotting assays and real-time quantitative PCR, respectively. Cellular cholesterol content was measured by HPLC. The cholesterol efflux was assessed by liquid scintillation counting. RESULTS: AOPPs significantly decreased the expression of ATP-binding membrane cassette transporter A-1 (ABCA1) and liver X receptor α (LXRα) and reduced cholesterol efflux from THP-1 macrophage- derived foam cells. AOPPs substantially activated NADPH oxidase and activated Janus kinase/signal transducers and activators of transcription (JAK/STAT) signal pathway in THP-1-derived foam-like cells. Inhibiting NADPH oxidase by diphenyliodonium (DPI) effectively abolished the AOPPs-induced decrease in cholesterol efflux and the expression of ABCA1. Inhibiting JAK/STAT activation by its specific inhibitor AG-490 or by siRNA could also block AOPPs action on THP-1 cells. CONCLUSIONS: AOPPs may first down-regulate the expression of LXRα and ABCA1 through JAK/STAT signal pathway activation and then inhibit cholesterol efflux in THP-1-derived foam-like cells; therefore, our study may be useful for understanding the critical effects of AOPPs on the pathogenesis of atherosclerosis.
Assuntos
Transportadores de Cassetes de Ligação de ATP/metabolismo , Colesterol/metabolismo , Regulação para Baixo , Janus Quinases/metabolismo , Proteínas/metabolismo , Fatores de Transcrição STAT/metabolismo , Transdução de Sinais , Transportador 1 de Cassete de Ligação de ATP , Linhagem Celular , HumanosRESUMO
AIM: To determine the effects and potential mechanisms of ibrolipim on ATP-binding membrane cassette transporter A-1 (ABCA1) and ATP-binding membrane cassette transporter G-1 (ABCG1) expression from human macrophage foam cells, which may play a critical role in atherogenesis. METHODS: Human THP-1 cells pre-incubated with ox-LDL served as foam cell models. Specific mRNA was quantified using real-time RT-PCR and protein expression using Western blotting. Cellular cholesterol handling was studied using cholesterol efflux experiments and high performance liquid chromatography assays. RESULTS: Ibrolipim 5 and 50 µmol/L significantly increased cholesterol efflux from THP-1 macrophage-derived foam cells to apoA-I or HDL. Moreover, it upregulated the expression of ABCA1 and ABCG1. In addition, LXRα was also upregulated by the ibrolipim treatment. In addition, LXRα small interfering RNA completely abolished the promotion effect that was induced by ibrolipim. CONCLUSION: Ibrolipim increased ABCA1 and ABCG1 expression and promoted cholesterol efflux, which was mediated by the LXRα signaling pathway.
Assuntos
Transportadores de Cassetes de Ligação de ATP/metabolismo , Benzamidas/farmacologia , Células Espumosas/efeitos dos fármacos , Ativadores de Lipase de Lipoproteínas/farmacologia , Compostos Organofosforados/farmacologia , Receptores Nucleares Órfãos/metabolismo , Transportador 1 de Cassete de Ligação de ATP , Membro 1 da Subfamília G de Transportadores de Cassetes de Ligação de ATP , Transporte Biológico , Linhagem Celular Tumoral , Proliferação de Células , Colesterol/metabolismo , Células Espumosas/metabolismo , Regulação da Expressão Gênica , Humanos , Receptores X do Fígado , Receptores Nucleares Órfãos/genética , RNA Mensageiro/metabolismo , Transdução de Sinais , Regulação para CimaRESUMO
Adenosine triphosphate-binding cassette transporter A1 (ABCA1) plays a crucial role in apolipoprotein A-I (apoA-I) binding activity and promotes cellular cholesterol efflux. ApoA-I mimetic peptide D4-F has reported to have the similar ability as apoA-I. However, the detailed mechanisms of ABCA1 regulation by D4-F are not understood. In the present study, we investigated the effects of D4-F on ABCA1 expression and ABCA1-dependent cholesterol efflux and examined the role of Cdc42/cyclic adenosine monophosphate (cAMP)/protein kinase A (PKA) pathway on the regulation of ABCA1 by D4-F in THP-1 macrophage-derived foam cells. Results showed that D4-F stabilized ABCA1 protein and enhanced ABCA1-dependent cholesterol efflux but had no effect on ABCA1 messenger RNA expression. We also revealed that D4-F enhanced cAMP level and PKA activity and ABCA1 serine phosphorylation. Short interfering RNA of PKA led to reduction of ABCA1 serine phosphorylation and ABCA1-mediated cholesterol efflux compensated by D4-F. PKA-specific activation by PKA agonist enhanced the upregulation of ABCA1 serine phosphorylation and ABCA1-mediated cholesterol efflux by D4-F. However, ABCA1 expression did not change by treatment with PKA agonist or PKA-short interfering RNA. We found that secramine B of Cdc42 inhibitor reduced the cAMP level compensated by D4-F. These results provide evidence that D4-F enhances ABCA1 serine phosphorylation and ABCA1-dependent cholesterol efflux through Cdc42/cAMP/PKA pathway in THP-1 macrophage-derived foam cells.
Assuntos
Transportadores de Cassetes de Ligação de ATP/biossíntese , Apolipoproteína A-I/farmacologia , Colesterol/metabolismo , Células Espumosas/efeitos dos fármacos , Transportador 1 de Cassete de Ligação de ATP , Transportadores de Cassetes de Ligação de ATP/genética , Transporte Biológico , Linhagem Celular , AMP Cíclico/metabolismo , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Ativação Enzimática , Células Espumosas/metabolismo , Expressão Gênica , Humanos , Fosforilação , RNA Mensageiro/biossíntese , Serina/metabolismo , Transdução de Sinais , Fatores de Tempo , Regulação para Cima , Proteína cdc42 de Ligação ao GTP/metabolismoRESUMO
AIM: High density lipoprotein (HDL) and its apolipoproteins can promote cholesterol efflux from macrophage foam cells via the ATP-binding cassette transporter A1 (ABCA1), ABCG1, and scavenger receptor class B type I (SR-BI). Liver X receptors (LXRs) operate as cholesterol sensors which may protect from cholesterol overload by stimulating cholesterol efflux from cells to HDL through ABCA1, ABCG1 and SR-BI. The regulation of ABCA1, ABCG1 and SR-BI expression by cytokines present within the microenvironment of the atheroma may play an important role in determining the impact of reverse cholesterol transport on the atherosclerotic lesion. In the current study, we examined the effect of transforming growth factor-beta1 (TGF-beta1) on expressions of ABCA1, ABCG1 and SR-BI and explored the role of LXR alpha in the regulation of ABCA1, ABCG1 and SR-BI in THP-1 macrophage-derived foam cells. METHODS AND RESULTS: TGF-beta1 significantly increased expressions of ABCA1, ABCG1 and SR-BI at both transcriptional and translational levels in a dose-dependent and time-dependent manner. Cellular cholesterol content was decreased while cholesterol efflux was increased by TGF-beta1 treatment. Moreover, LXR alpha was up-regulated by TGF-beta1 treatment. In addition, LXR alpha small interfering RNA completely abolished the promotion effect induced by TGF-beta1. CONCLUSION: These results provide evidence that TGF-beta1 up-regulates expressions of ABCA1, ABCG1 and SR-BI through the LXR alpha pathway in THP-1 macrophage-derived foam cells.
Assuntos
Transportadores de Cassetes de Ligação de ATP/metabolismo , Células Espumosas/metabolismo , Receptores Nucleares Órfãos/metabolismo , Receptores Depuradores Classe B/metabolismo , Fator de Crescimento Transformador beta1/fisiologia , Regulação para Cima/fisiologia , Transportador 1 de Cassete de Ligação de ATP , Membro 1 da Subfamília G de Transportadores de Cassetes de Ligação de ATP , Western Blotting , Linhagem Celular , Cromatografia Líquida de Alta Pressão , Inativação Gênica , Humanos , Receptores X do Fígado , Receptores Nucleares Órfãos/genética , Reação em Cadeia da PolimeraseRESUMO
ATP binding cassette transporter G1 (ABCG1) is a membrane half-transporter which is the member of ATP-binding cassette (ABC) transporter super-family, it has an important role of regulating the cholesterol and phospholipids effluence. ABCG1 and ABCA1 synergize to mediate cholesterol effluence to HDL (high density lipoprotein). The expression of ABCG1 is mainly regulated by the liver X receptor/the lactochrome receptor system (LXR/RXR). Although the ABCG1 plays an important role in balancing the lipids, its role in cardiovascular disease (CVD) in the animal studies is still controversial. The issue focuses the ABCG1 on the structure, the function, the regulation and the contribution to CVD.
Assuntos
Transportadores de Cassetes de Ligação de ATP/fisiologia , Membro 1 da Subfamília G de Transportadores de Cassetes de Ligação de ATP , Animais , Doenças Cardiovasculares/fisiopatologia , Humanos , Lipoproteínas/fisiologia , Receptores X do Fígado , Receptores Nucleares Órfãos/fisiologia , Receptores X de Retinoides/fisiologiaRESUMO
BACKGROUND: The Niemann-Pick C1 (NPC1) protein regulates the transport of cholesterol from late endosomes/lysosomes to other compartments responsible for maintaining intracellular cholesterol homeostasis. Liver X receptors (LXRs) operate as cholesterol sensors which may protect from cholesterol overload by increasing the amount of free cholesterol in the plasma membrane through inducing NPC1 expression. NO-1886 has been proven to be highly effective at increasing liver X receptor alpha expression and promoting cellular cholesterol efflux. In this study, the effects of NO-1886 on NPC1 expression were investigated in THP-1 macrophage-derived foam cells. METHODS AND RESULTS: Results showed that NO-1886 markedly increased expression of NPC1 at both mRNA level and protein level in a dose-dependent and time-dependent manner. Cellular cholesterol content was decreased while cholesterol efflux was increased by NO-1886 treatment. In addition, LXR alpha was also up-regulated by NO-1886 treatment. And LXR alpha small interfering RNA completely abolished the promotion effect which was induced by NO-1886. CONCLUSION: These results provide evidence that NO-1886 up-regulates expression of NPC1 through LXR alpha pathway in THP-1 macrophage- derived foam cells.
Assuntos
Benzamidas/farmacologia , Proteínas de Transporte/biossíntese , Proteínas de Ligação a DNA/fisiologia , Células Espumosas/efeitos dos fármacos , Hipolipemiantes/farmacologia , Glicoproteínas de Membrana/biossíntese , Compostos Organofosforados/farmacologia , Receptores Citoplasmáticos e Nucleares/fisiologia , Proteínas de Transporte/genética , Linhagem Celular , Colesterol/metabolismo , Proteínas de Ligação a DNA/genética , Relação Dose-Resposta a Droga , Células Espumosas/metabolismo , Humanos , Peptídeos e Proteínas de Sinalização Intracelular , Receptores X do Fígado , Glicoproteínas de Membrana/genética , Proteína C1 de Niemann-Pick , Receptores Nucleares Órfãos , RNA Mensageiro/biossíntese , Receptores Citoplasmáticos e Nucleares/genética , Transdução de Sinais , Fatores de Tempo , Regulação para CimaRESUMO
ABCA1 is a key mediator of cholesterol efflux to apoA-I in cholesterol loaded macrophages, a first step of RCT in vivo. Unsaturated fatty acids can inhibit cholesterol efflux from macrophages by increasing degradation of ABCA1. However, the detailed mechanisms of ABCA1 regulation by unsaturated fatty acids are not fully understood. In the present study, we investigated the effects of EPA on ABCA1 expression and ABCA1-dependent cholesterol efflux and examined the role of cAMP/PKA pathway on the regulation of ABCA1 by EPA in THP-1 macrophage-derived foam cells. Results showed that EPA significantly destabilized ABCA1 protein and reduced ABCA1-dependent cholesterol efflux but had no effect on ABCA1 mRNA expression. We also revealed that EPA markedly reduced cAMP level and PKA activity and ABCA1 serine phosphorylation. PKA-specific activation by PKA agonist markedly compensated the down-regulation of ABCA1 serine phosphorylation and ABCA1-mediated cholesterol efflux by EPA, while, siRNA of PKA leaded to reduce of ABCA1 serine phosphorylation and ABCA1-mediated cholesterol efflux more significantly than EPA. However, EPA-Induced enhancement of degradation rate of ABCA1 protein did not change by treatment with PKA agonist or PKA-siRNA. These results provide evidence that EPA may have dual negative effects on ABCA1 activity by decreasing ABCA1 protein level and by reducing PKA-mediated ABCA1 serine phosphorylation in THP-1 macrophage-derived foam cells.
Assuntos
Transportadores de Cassetes de Ligação de ATP/metabolismo , Colesterol/metabolismo , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , AMP Cíclico/metabolismo , Ácido Eicosapentaenoico/metabolismo , Células Espumosas/enzimologia , Transdução de Sinais , 3',5'-AMP Cíclico Fosfodiesterases/metabolismo , Transportador 1 de Cassete de Ligação de ATP , Transportadores de Cassetes de Ligação de ATP/genética , Acil Coenzima A/metabolismo , Adenilil Ciclases/metabolismo , Transporte Biológico , Linhagem Celular , Proteínas Quinases Dependentes de AMP Cíclico/antagonistas & inibidores , Proteínas Quinases Dependentes de AMP Cíclico/genética , Regulação para Baixo , Ativadores de Enzimas/farmacologia , Células Espumosas/efeitos dos fármacos , Humanos , Fosforilação , Estabilidade Proteica , Interferência de RNA , RNA Mensageiro/metabolismo , Serina , Transdução de Sinais/efeitos dos fármacos , Fatores de TempoRESUMO
Interferon gamma (IFN-gamma) is an immunomodulatory and anti-microbial cytokine, which has a variety of proatherogenic effects. It has been reported that IFN-gamma can down-regulate ABCA1 expression. However, its mechanism is elusive. In the present study, we have investigated the effect of IFN-gamma on ABCA1 expression and cholesterol efflux in THP-1 macrophage-derived foam cells. IFN-gamma decreased ABCA1 expression at both transcriptional and translational levels in a dose-dependent manner. Cellular cholesterol content was increased while cholesterol efflux was decreased by IFN-gamma treatment. Liver X receptor alpha (LXRalpha), which can regulate the expression of ABCA1, was also down-regulated by IFN-gamma treatment. LXRalpha-specific activation by LXRalpha agonist almost compensated the down-regulation of ABCA1 expression by IFN-gamma, while siRNA of LXRalpha led to down-regulation of ABCA1 expression more significantly than IFN-gamma. IFN-gamma induced phosphorylation of STAT1 and expression of STAT1alpha in the nucleus, which was inhibited by a JAK inhibitor AG-490. Treatment with STAT1 siRNA further enhanced down-regulation of LXRalpha mRNA by IFN-gamma. Furthermore, AG-490 and STAT1 siRNA almost compensated the effect of IFN-gamma on ABCA1 expression and cholesterol efflux. In conclusion, IFN-gamma may first down-regulate expression of LXRalpha through the JAK/STAT1 signaling pathway and then decrease expression of ABCA1 and cholesterol efflux in THP-1 macrophage-derived foam cells. Therefore, our study may be useful in understanding the critical effect of IFN-gamma in pathogenesis of atherosclerosis.