RESUMEN
A new cycloartane triterpene, yunnanterpene G (1), containing an oxaspiro[5.4]decane moiety, was purified from the roots of Cimicifuga foetida. The new structure was determined from spectroscopic data and the X-ray diffraction method. Biological evaluations revealed that compound 1 significantly inhibited the mRNA expression of the atherosclerosis-related adhesion molecule CD147 (extracellular matrix metalloproteinase inducer, EMMPRIN), and proteolytic enzymes matrix metalloproteinase 2 (MMP-2), MMP-9 and MMP-14, in a dose-dependent manner in phorbol-12-myristate-13-acetate-induced human monocytic THP-1 cells by quantitative real-time PCR method. At the same time, the migration ability of the induced THP-1 cells was potently inhibited. Furthermore, western blot experiments showed that compound 1 at 25 µM strongly suppressed phosphorylation of NF-κB p65 and p38 MAPK in the differentiated THP-1 cells.
RESUMEN
Patients with type 2 diabetes mellitus (T2DM) are characterized by insulin resistance and are subsequently at high risk for atherosclerosis. Hyperinsulinemia has been associated with proliferation, migration, and dedifferentiation of vascular smooth muscle cells (VSMCs) during the pathogenesis of atherosclerosis. Moreover, insulin-like growth factor-1 receptor (IGF-1R) and mammalian target of rapamycin (mTOR) have been demonstrated to be the underlying signaling pathways. Recently, microRNA-99a (miR-99a) has been suggested to regulate the phenotypic changes of VSMCs in cancer cells. However, whether it is involved in insulin-induced changes of VSCMs has not been determined. In this study, we found that insulin induced proliferation, migration, and dedifferentiation of mouse VSMCs in a dose-dependent manner. Furthermore, the stimulating effects of high-dose insulin on proliferation, migration, and dedifferentiation of mouse VSMCs were found to be associated with the attenuation of the inhibitory effects of miR-99a on IGF-1R and mTOR signaling activities. Finally, we found that the inducing effect of high-dose insulin on proliferation, migration, and dedifferentiation of VSMCs was partially inhibited by an active mimic of miR-99a. Taken together, these results suggest that miR-99a plays a key regulatory role in the pathogenesis of insulin-induced proliferation, migration, and phenotype conversion of VSMCs at least partly via inhibition of IGF-1R and mTOR signaling. Our results provide evidence that miR-99a may be a novel target for the treatment of hyperinsulinemia-induced atherosclerosis.
Asunto(s)
Células Endoteliales/efectos de los fármacos , Insulina/farmacología , MicroARNs/genética , Receptor IGF Tipo 1/genética , Serina-Treonina Quinasas TOR/genética , Animales , Desdiferenciación Celular/efectos de los fármacos , Movimiento Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Células Endoteliales/citología , Células Endoteliales/metabolismo , Regulación de la Expresión Génica , Insulina/metabolismo , Ratones , Ratones Endogámicos C57BL , MicroARNs/metabolismo , Imitación Molecular , Músculo Liso Vascular/citología , Músculo Liso Vascular/efectos de los fármacos , Músculo Liso Vascular/metabolismo , Oligorribonucleótidos/genética , Oligorribonucleótidos/metabolismo , Cultivo Primario de Células , Receptor IGF Tipo 1/metabolismo , Transducción de Señal , Serina-Treonina Quinasas TOR/metabolismoRESUMEN
OBJECTIVES: This study aimed to investigate the association between microRNA-155 (miR-155) and the severity and extent of coronary stenotic lesions. PATIENTS AND METHODS: We measured the miR-155 expression by real-time PCR in 110 consecutive patients undergoing coronary angiography for suspected coronary artery disease. The severity and extent of coronary stenotic lesions were evaluated on the basis of coronary angiography findings by the Gensini score. RESULTS: The miR-155 expression was significantly lower in 56 patients with coronary heart disease than those in 54 controls (P<0.01). The level of miR-155 in peripheral blood mononuclear cells or plasma was lower in patients with unstable angina pectoris and acute myocardial infarction than in patients with chest pain syndrome, whereas no statistically significant differences were observed between patients with stable angina pectoris and chest pain syndrome. Spearman's correlation analysis showed that the expression of miR-155 in plasma correlated positively with the expression in peripheral blood mononuclear cells. The levels of miR-155 in the patients with diseased vessels of two and three or more were significantly lower than in those with diseased vessel of zero and one. The levels of miR-155 were not significantly different among groups with diseased vessels of zero and one. miR-155 were associated negatively with Gensini scores (r = -0.663, P<0.001). The miR-155 expression was correlated significantly to age (r = -0.227), hypertension (r = -0.440), total cholesterol (r = 0.239), high-density lipoprotein cholesterol (r = 0.280), low-density lipoprotein cholesterol (r = -0.315), tobacco use (r = -0.363), angiotensin-converting enzyme inhibitor (r = -0.250), statins (r = -0.368), and high-sensitivity C-reactive protein (r = -0.515). CONCLUSION: miR-155 expression is associated inversely with complicated proatherogenic metabolic risk factors, and the severity of coronary stenotic lesions calculated by Gensini scores.
Asunto(s)
Angiografía Coronaria , Estenosis Coronaria/diagnóstico , Vasos Coronarios/diagnóstico por imagen , Pruebas Genéticas , MicroARNs/sangre , Anciano , Angina Inestable/diagnóstico por imagen , Angina Inestable/genética , Estudios de Casos y Controles , Estenosis Coronaria/sangre , Estenosis Coronaria/diagnóstico por imagen , Estenosis Coronaria/genética , Progresión de la Enfermedad , Femenino , Marcadores Genéticos , Pruebas Genéticas/métodos , Humanos , Masculino , Persona de Mediana Edad , Infarto del Miocardio/diagnóstico por imagen , Infarto del Miocardio/genética , Valor Predictivo de las Pruebas , Reacción en Cadena en Tiempo Real de la Polimerasa , Índice de Severidad de la EnfermedadRESUMEN
Macrophage apoptosis is a prominent feature of advanced atherosclerotic plaques. Here, we examined the hypothesis that the apoptotic machinery is regulated by microRNA-155 (miR-155). Constitutive expression of miR-155 was detected in RAW264.7 cells, which was increased following stimulation with oxidized low-density lipoprotein (OxLDL) in a dose- and time-dependent manner. OxLDL-treated RAW264.7 cells showed a marked time- and dose-dependent increase in apoptosis, which was suppressed in the presence of mimics and increased with antagonists of miR-155. Bioinformatics analysis revealed Fas-associated death domain-containing protein (FADD) as a putative target of miR-155. Luciferase reporter assay and Western blot further disclosed that miR-155 inhibits FADD expression by directly targeting the 3'-UTR region. We propose that miR-155 attenuates the macrophage apoptosis, at least in part, through FADD regulation, since forced expression of FADD blocked the ability of miR-155 to inhibit apoptosis. Our results collectively suggest that miR-155 attenuates apoptosis of OxLDL-mediated RAW264.7 cells by targeting FADD, supporting a possible therapeutic role in atherosclerosis.
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Apoptosis/efectos de los fármacos , Lipoproteínas LDL/farmacología , Macrófagos/citología , Macrófagos/metabolismo , MicroARNs/metabolismo , Regiones no Traducidas 3'/genética , Animales , Apoptosis/genética , Línea Celular , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/genética , Citoprotección/efectos de los fármacos , Citoprotección/genética , Proteína de Dominio de Muerte Asociada a Fas/metabolismo , Regulación de la Expresión Génica/efectos de los fármacos , Macrófagos/efectos de los fármacos , Ratones , MicroARNs/genética , ARN Mensajero/genética , ARN Mensajero/metabolismoRESUMEN
BACKGROUND: Extracellular matrix metalloproteinase inducer (EMMPRIN), a 58-kDa cell surface glycoprotein, has been identified as a key receptor for transmitting cellular signals mediating metalloproteinase activities, as well as inflammation and oxidative stress. Clinical evidence has revealed that EMMPRIN is expressed in human atherosclerotic plaque; however, the relationship between EMMPRIN and atherosclerosis is unclear. To evaluate the functional role of EMMPRIN in atherosclerosis, we treated apolipoprotein E-deficient (ApoE(-/-)) mice with an EMMPRIN function-blocking antibody. METHODS AND RESULTS: EMMPRIN was found to be up-regulated in ApoE(-/-) mice fed a 12-week high-fat diet in contrast to 12 weeks of normal diet. Administration of a function-blocking EMMPRIN antibody (100 µg, twice per week for 4 weeks) to ApoE(-/-) mice, starting after 12 weeks of high-fat diet feeding caused attenuated and more stable atherosclerotic lesions, less reactive oxygen stress generation on plaque, as well as down-regulation of circulating interleukin-6 and monocyte chemotactic protein-1 in ApoE(-/-) mice. The benefit of EMMPRIN functional blockage was associated with reduced metalloproteinases proteolytic activity, which delayed the circulating monocyte transmigrating into atherosclerotic lesions. CONCLUSION: EMMPRIN antibody intervention ameliorated atherosclerosis in ApoE(-/-) mice by the down-regulation of metalloproteinase activity, suggesting that EMMPRIN may be a viable therapeutic target in atherosclerosis.
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Anticuerpos Bloqueadores/farmacología , Apolipoproteínas E/deficiencia , Aterosclerosis/metabolismo , Basigina/metabolismo , Animales , Anticuerpos Bloqueadores/uso terapéutico , Aterosclerosis/etiología , Aterosclerosis/prevención & control , Basigina/inmunología , Basigina/fisiología , Dieta Alta en Grasa/efectos adversos , Dieta Alta en Grasa/métodos , Regulación hacia Abajo/genética , Masculino , Metaloproteasas/antagonistas & inhibidores , Metaloproteasas/biosíntesis , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Proteolisis , Distribución AleatoriaRESUMEN
UNLABELLED: The pathogenesis of acute coronary syndrome is rupture of vulnerable plaque. Extracellular matrix metalloproteinase inducer (EMMPRIN) is reported to have a important role in the destabilization of atheroma. OBJECTIVES: this investigation examined the effect of angiotensin II (Ang II) on EMMPRIN expression in atherosclerotic plaques in ApoE knockout mice. METHODS: ApoE knockout mice were fed a high fat diet to establish an atherosclerosis model then intervention was made with Ang II and valsartan. EMMPRIN gene and its protein expression were measured by real-time PCR immunohistochemistry, and Western blot. RESULTS: EMMPRIN gene and protein expression intervened with Ang II were significantly increased; valsartan could inhibit the effect of Ang II. CONCLUSION: Ang II up-regulated EMMPRIN expression in atherosclerotic plaque via AT1R, and valsartan could inhibit the effect of Ang II.
Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/metabolismo , Angiotensina II/farmacología , Aorta/patología , Apolipoproteínas E/deficiencia , Basigina/metabolismo , Placa Aterosclerótica/patología , Transducción de Señal/efectos de los fármacos , Animales , Apolipoproteínas E/metabolismo , Basigina/genética , Regulación de la Expresión Génica/efectos de los fármacos , Lípidos/sangre , Ratones , Ratones Noqueados , Placa Aterosclerótica/sangre , Placa Aterosclerótica/metabolismo , Transporte de Proteínas/efectos de los fármacos , ARN Mensajero/genética , ARN Mensajero/metabolismo , Coloración y EtiquetadoRESUMEN
AIM: To explore the expression of extracellular matrix metalloproteinase inducer (EMMPRIN) in THP-1 macrophages induced by angiotensin II (Ang II) and the mechanism of EMMPRIN expression. METHODS: THP-1 cells were cultured and induced into macrophages, then stimulated with 10(-6) mol/L Ang II. Levels of EMMPRIN gene and its protein were measured by real-time polymerase chain reaction and western blotting. Prostaglandin E(2) (PGE(2)) expression was assayed by enzyme-linked immunosorbent assay. Antagonists of the angiotensin type-1 receptor (AT(1)R) and angiotensin type-2 receptor (AT(2)R) were used to inhibit the effect of Ang II, and PGE(2) added to detail the mechanism of Ang II-induced EMMPRIN expression. RESULTS: Ang II clearly induced the expression of EMMPRIN mRNA and protein in macrophages; this expression peaked at 12 h and declined after 24 h. The tendency of enhancement of the levels of cyclooxygenase-2 (COX-2) and PGE(2) was coincident with EMMPRIN expression. AT(1)-receptor antagonists and COX-2 inhibitors inhibited the effect of Ang II, but AT(2)-receptor antagonists did not. CONCLUSION: Ang II can up-regulate EMMPRIN expression in THP-1 macrophages via the AT(1)/COX-2/PGE(2) signal transduction pathway, and the effect can be inhibited by losartan and NS-398.