RESUMO
OBJECTIVE: We examined the effects of treatment with soluble epoxide hydrolase inhibitor (sEHi) and epoxyeicosatrienoic acids (EETs) analogue (EET-A), given alone or combined, on blood pressure (BP) and ischemia/reperfusion myocardial injury in rats with angiotensin II (ANG II)-dependent hypertension. METHODS: Ren-2 transgenic rats (TGR) were used as a model of ANG II-dependent hypertension and Hannover Sprague-Dawley rats served as controls. Rats were treated for 14 days with sEHi or EET-A and BP was measured by radiotelemetry. Albuminuria, cardiac hypertrophy and concentrations of ANG II and EETs were determined. Separate groups were subjected to acute myocardial ischemia/reperfusion injury and the infarct size and ventricular arrhythmias were determined. RESULTS: Treatment of TGR with sEHi and EET-A, given alone or combined, decreased BP to a similar degree, reduced albuminuria and cardiac hypertrophy to similar extent; only treatment regimens including sEHi increased myocardial and renal tissue concentrations of EETs. sEHi and EET-A, given alone or combined, suppressed kidney ANG II levels in TGR. Remarkably, infarct size did not significantly differ between TGR and Hannover Sprague-Dawley rats, but the incidence of ischemia-induced ventricular fibrillations was higher in TGR. Application of sEHi and EET-A given alone and combined sEHi and EET-A treatment were all equally effective in reducing life-threatening ventricular fibrillation in TGR. CONCLUSION: The findings indicate that chronic treatment with either sEHi or EET-A exerts distinct antihypertensive and antiarrhythmic actions in our ANG II-dependent model of hypertension whereas combined administration of sEHi and EET-A does not provide additive antihypertensive or cardioprotective effects.
Assuntos
Anti-Hipertensivos/farmacologia , Ácidos Araquidônicos/farmacologia , Arritmias Cardíacas/metabolismo , Pressão Sanguínea/efeitos dos fármacos , Hipertensão/metabolismo , Albuminúria/metabolismo , Angiotensina II/metabolismo , Animais , Ratos , Ratos Sprague-Dawley , Ratos TransgênicosRESUMO
AIM: We designed a study to evaluate the cardioprotective effect of two soluble epoxide hydrolase (sEH) inhibitors, 1-(1-propanoylpiperidin-4-yl)-3-(4-trifluoromethoxy)phenyl)urea (TPPU) and trans-4-{4-[3-(4-trifluoromethoxyphenyl)-ureido]cyclohexyloxy}benzoic acid (t-TUCB), in ischemia-reperfusion (IR) model. METHODS: Cardioprotective effects of the sEH inhibitors were evaluated against IR-induced myocardial damage in hearts from normal, hypertensive, and diabetic rats using Langendorff's apparatus. In addition, the effect of sEH inhibitors on endothelial function was evaluated in vitro and ex vivo using isolated rat thoracic aorta. RESULTS: Ischemia-reperfusion (IR) increased the myocardial damage in hearts from normal rats. IR-induced myocardial damage was augmented in hearts isolated from hypertensive and diabetic rats. Myocardial damage as evident from increase in the activities of lactate dehydrogenase (LDH) and creatine kinase-MB (CK-MB) in heart perfusate was associated with significant decrease in the heart rate and developed tension, and increase in the resting tension in isolated heart. Both sEH inhibitors protected the heart in normal, hypertensive, and diabetic rats subjected to IR injury. The sEH inhibitor t-TUCB relaxed phenylephrine precontracted aorta from normal rats. Relaxant effect of acetylcholine (ACh) was reduced in aortas from diabetic and hypertensive rats compared to normal rats. Pretreatment of sEH inhibitors to diabetic and hypertensive rats increased relaxant effect of ACh on aortas isolated from these rats. CONCLUSIONS: Prophylactic treatment with sEH inhibitors decreased myocardial damage due to IR, hypertension and diabetes, and decreased endothelial dysfunction created by diabetes and hypertension. Therefore, inhibitors of sEH are useful probes to study cardiovascular pathology, and inhibition of the sEH is a potential approach in the management of IR-induced cardiac damage and endothelial dysfunction-related cardiovascular disorders.
Assuntos
Benzoatos/farmacologia , Diabetes Mellitus Experimental/tratamento farmacológico , Diabetes Mellitus Tipo 1/tratamento farmacológico , Inibidores Enzimáticos/farmacologia , Epóxido Hidrolases/antagonistas & inibidores , Hipertensão/tratamento farmacológico , Traumatismo por Reperfusão Miocárdica/prevenção & controle , Miocárdio/enzimologia , Compostos de Fenilureia/farmacologia , Piperidinas/farmacologia , Animais , Aorta Torácica/efeitos dos fármacos , Aorta Torácica/enzimologia , Aorta Torácica/patologia , Aorta Torácica/fisiopatologia , Diabetes Mellitus Experimental/enzimologia , Diabetes Mellitus Experimental/patologia , Diabetes Mellitus Experimental/fisiopatologia , Diabetes Mellitus Tipo 1/enzimologia , Diabetes Mellitus Tipo 1/patologia , Diabetes Mellitus Tipo 1/fisiopatologia , Relação Dose-Resposta a Droga , Epóxido Hidrolases/metabolismo , Frequência Cardíaca/efeitos dos fármacos , Hipertensão/enzimologia , Hipertensão/patologia , Hipertensão/fisiopatologia , Preparação de Coração Isolado , Masculino , Contração Miocárdica/efeitos dos fármacos , Infarto do Miocárdio/enzimologia , Infarto do Miocárdio/etiologia , Infarto do Miocárdio/patologia , Infarto do Miocárdio/fisiopatologia , Traumatismo por Reperfusão Miocárdica/enzimologia , Traumatismo por Reperfusão Miocárdica/patologia , Traumatismo por Reperfusão Miocárdica/fisiopatologia , Miocárdio/patologia , Ratos Wistar , Vasodilatação/efeitos dos fármacosRESUMO
Soluble epoxide hydrolase (sEH) is an emerging therapeutic target in a number of diseases that have inflammation as a common underlying cause. sEH limits tissue levels of cytochrome P450 (CYP) epoxides derived from omega-6 and omega-3 polyunsaturated fatty acids (PUFA) by converting these antiinflammatory mediators into their less active diols. Here, we explored the metabolic effects of a sEH inhibitor (t-TUCB) in fat-1 mice with transgenic expression of an omega-3 desaturase capable of enriching tissues with endogenous omega-3 PUFA. These mice exhibited increased CYP1A1, CYP2E1, and CYP2U1 expression and abundant levels of the omega-3-derived epoxides 17,18-epoxyeicosatetraenoic acid (17,18-EEQ) and 19,20-epoxydocosapentaenoic (19,20-EDP) in insulin-sensitive tissues, especially liver, as determined by LC-ESI-MS/MS. In obese fat-1 mice, t-TUCB raised hepatic 17,18-EEQ and 19,20-EDP levels and reinforced the omega-3-dependent reduction observed in tissue inflammation and lipid peroxidation. t-TUCB also produced a more intense antisteatotic action in obese fat-1 mice, as revealed by magnetic resonance spectroscopy. Notably, t-TUCB skewed macrophage polarization toward an antiinflammatory M2 phenotype and expanded the interscapular brown adipose tissue volume. Moreover, t-TUCB restored hepatic levels of Atg12-Atg5 and LC3-II conjugates and reduced p62 expression, indicating up-regulation of hepatic autophagy. t-TUCB consistently reduced endoplasmic reticulum stress demonstrated by the attenuation of IRE-1α and eIF2α phosphorylation. These actions were recapitulated in vitro in palmitate-primed hepatocytes and adipocytes incubated with 19,20-EDP or 17,18-EEQ. Relatively similar but less pronounced actions were observed with the omega-6 epoxide, 14,15-EET, and nonoxidized DHA. Together, these findings identify omega-3 epoxides as important regulators of inflammation and autophagy in insulin-sensitive tissues and postulate sEH as a druggable target in metabolic diseases.
Assuntos
Tecido Adiposo/metabolismo , Epóxido Hidrolases/antagonistas & inibidores , Ácidos Graxos Ômega-3/metabolismo , Obesidade/metabolismo , Células 3T3-L1 , Tecido Adiposo/efeitos dos fármacos , Tecido Adiposo/patologia , Animais , Autofagia/fisiologia , Benzoatos/farmacologia , Caderinas/genética , Caderinas/metabolismo , Citocromo P-450 CYP1A1/metabolismo , Citocromo P-450 CYP2E1/metabolismo , Sistema Enzimático do Citocromo P-450/metabolismo , Inibidores Enzimáticos/farmacologia , Compostos de Epóxi/metabolismo , Ácidos Graxos Dessaturases/genética , Ácidos Graxos Dessaturases/metabolismo , Feminino , Inflamação/metabolismo , Inflamação/patologia , Fígado/efeitos dos fármacos , Fígado/metabolismo , Fígado/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Mutantes , Camundongos Transgênicos , Obesidade/patologia , Compostos de Fenilureia/farmacologiaRESUMO
Recent studies have shown that the long-term antihypertensive action of soluble epoxide hydrolase inhibition (sEH) in angiotensin-II (AngII)-dependent hypertension might be mediated by the suppression of intrarenal AngII levels. To test this hypothesis, we examined the effects of acute (2 days) and chronic (14 days) sEH inhibition on blood pressure (BP) in transgenic rats with inducible AngII-dependent hypertension. AngII-dependent malignant hypertension was induced by 10 days' dietary administration of indole-3-carbinol (I3C), a natural xenobiotic that activates the mouse renin gene in Cyp1a1-Ren-2 transgenic rats. BP was monitored by radiotelemetry. Acute and chronic sEH inhibition was achieved using cis-4-(4-(3-adamantan-1-yl-ureido)cyclohexyloxy) benzoic acid, given at doses of 0.3, 3, 13, 26, 60 and 130 mg/L in drinking water. At the end of experiments, renal concentrations of epoxyeicosatrienoic acids, their inactive metabolites dihydroxyeicosatrienoic acids and AngII were measured. Acute BP-lowering effects of sEH inhibition in I3C-induced rats was associated with a marked increase in renal epoxyeicosatrienoic acids to dihydroxyeicosatrienoic acids ratio and acute natriuresis. Chronic treatment with cis-4-(4-(3-adamantan-1-yl-ureido)cyclohexyloxy) benzoic acid in I3C-induced rats elicited dose-dependent persistent BP lowering associated with a significant reduction of plasma and kidney AngII levels. Our findings show that the acute BP-lowering effect of sEH inhibition in I3C-induced Cyp1a1-Ren-2 transgenic rats is mediated by a substantial increase in intrarenal epoxyeicosatrienoic acids and their natriuretic action without altering intrarenal renin-angiotensin system activity. Long-term antihypertensive action of cis-4-(4-(3-adamantan-1-yl-ureido)cyclohexyloxy) benzoic acid in I3C-induced Cyp1a1-Ren-2 transgenic rats is mediated mostly by suppression of intrarenal AngII concentration.
Assuntos
Anti-Hipertensivos/farmacologia , Citocromo P-450 CYP1A1/metabolismo , Epóxido Hidrolases/antagonistas & inibidores , Epóxido Hidrolases/metabolismo , Renina/metabolismo , Angiotensina II/metabolismo , Animais , Pressão Sanguínea/efeitos dos fármacos , Hipertensão/tratamento farmacológico , Hipertensão/metabolismo , Indóis/metabolismo , Rim/efeitos dos fármacos , Rim/metabolismo , Masculino , Camundongos , Natriurese/efeitos dos fármacos , Ratos , Ratos Endogâmicos F344 , Ratos Transgênicos , Sistema Renina-Angiotensina/efeitos dos fármacosRESUMO
Tumor suppressor p53 plays an important role in mediating growth inhibition upon telomere dysfunction. Here, we show that loss of the p53 target gene cyclin-dependent kinase inhibitor 1A (CDKN1A, also known as p21(WAF1/CIP1)) increases apoptosis induction following telomerase inhibition in a variety of cancer cell lines and mouse xenografts. This effect is highly specific to p21, as loss of other checkpoint proteins and CDK inhibitors did not affect apoptosis. In telomerase, inhibited cell loss of p21 leads to E2F1- and p53-mediated transcriptional activation of p53-upregulated modulator of apoptosis, resulting in increased apoptosis. Combined genetic or pharmacological inhibition of telomerase and p21 synergistically suppresses tumor growth. Furthermore, we demonstrate that simultaneous inhibition of telomerase and p21 also suppresses growth of tumors containing mutant p53 following pharmacological restoration of p53 activity. Collectively, our results establish that inactivation of p21 leads to increased apoptosis upon telomerase inhibition and thus identify a genetic vulnerability that can be exploited to treat many human cancers containing either wild-type or mutant p53.
Assuntos
Apoptose , Inibidor de Quinase Dependente de Ciclina p21/antagonistas & inibidores , Neoplasias Experimentais/metabolismo , Telomerase/antagonistas & inibidores , Proteína Supressora de Tumor p53/metabolismo , Animais , Linhagem Celular Tumoral , Inibidor de Quinase Dependente de Ciclina p21/genética , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Humanos , Camundongos , Camundongos Nus , Mutação , Neoplasias Experimentais/genética , Neoplasias Experimentais/patologia , Neoplasias Experimentais/terapia , Telomerase/genética , Telomerase/metabolismo , Proteína Supressora de Tumor p53/genéticaRESUMO
OBJECTIVES: To determine the protective role of a soluble epoxide hydrolase(sEH) inhibitor, trans-4-{4-[3-(4-trifluoromethoxyphenyl)-ureido] cyclohexyloxy} benzoic acid (t-TUCB), in isoproterenol (ISO)-induced myocardial ischaemic injury in vivo. METHODS: Cardioprotective activity of t-TUCB was studied against ISO-induced myocardial ischaemic injury in male Wistar rats. Cardioprotection was assessed by measuring elecrocardiographic (EKG), serum lactate dehydrogenase (LDH) and creatine kinase (CK-MB) levels, cardiac calcium and antioxidant levels, and also by measuring infarct size in the cardiac tissue. KEY FINDINGS: Pretreatment with t-TUCB at 3, 10 and 30 mg/kg orally for a period of 14 days significantly prevented the changes in EKG parameters (QTc interval prolongation, ST height depression, pathological Q waves formation and T-wave inversion), serum cardiac biomarkers (CK-MB and LDH), relative heart weight, myocardial calcium levels, infarct size and the oxidative status in the cardiac tissue (lipid peroxidation, catalase and superoxide dismutase levels) when compared with the untreated control animals (P < 0.05). CONCLUSION: The sEH inhibitor t-TUCB significantly prevents ISO-induced myocardial ischaemic injury in rats. This study provides a preliminary confirmation of the efficacy of t-TUCB by oral administration in rats.
Assuntos
Benzoatos/uso terapêutico , Cardiotônicos/uso terapêutico , Epóxido Hidrolases/antagonistas & inibidores , Infarto do Miocárdio/tratamento farmacológico , Miocárdio , Estresse Oxidativo/efeitos dos fármacos , Compostos de Fenilureia/uso terapêutico , Traumatismo por Reperfusão/prevenção & controle , Animais , Antioxidantes/metabolismo , Antioxidantes/farmacologia , Antioxidantes/uso terapêutico , Benzoatos/farmacologia , Biomarcadores/sangue , Biomarcadores/metabolismo , Cardiotônicos/farmacologia , Coração/efeitos dos fármacos , Coração/fisiopatologia , Sistema de Condução Cardíaco/efeitos dos fármacos , Sistema de Condução Cardíaco/fisiopatologia , Isoproterenol , Peroxidação de Lipídeos/efeitos dos fármacos , Masculino , Infarto do Miocárdio/metabolismo , Infarto do Miocárdio/patologia , Miocárdio/metabolismo , Miocárdio/patologia , Tamanho do Órgão/efeitos dos fármacos , Compostos de Fenilureia/farmacologia , Ratos Wistar , Traumatismo por Reperfusão/metabolismo , Traumatismo por Reperfusão/patologiaRESUMO
1. The aim of the present study was to test the hypothesis that increasing kidney tissue concentrations of epoxyeicosatrienoic acids (EETs) by preventing their degradation to the biologically inactive dihydroxyeicosatrienoic acids (DHETEs) using blockade of soluble epoxide hydrolase (sEH) would attenuate the progression of chronic kidney disease (CKD). 2. Ren-2 transgenic rats (TGR) after 5/6 renal mass reduction (5/6 NX) served as a model of CKD associated with angiotensin (Ang) II-dependent hypertension. Soluble epoxide hydrolase was inhibited using cis-4-[4-(3-adamantan-1-yl-ureido)cyclohexyloxy]benzoic acid (c-AUCB; 3 mg/L drinking water) for 20 weeks after 5/6 NX. Sham-operated normotensive transgene-negative Hannover Sprague-Dawley (HanSD) rats served as controls. 3. When applied in TGR subjected to 5/6 NX, c-AUCB treatment improved survival rate, prevented the increase in blood pressure, retarded the progression of cardiac hypertrophy, reduced proteinuria and the degree of glomerular and tubulointerstitial injury and reduced glomerular volume. All these organ-protective actions were associated with normalization of the intrarenal EETs:DHETEs ratio, an index of the availability of biologically active EETs, to levels observed in sham-operated HanSD rats. There were no significant concurrent changes of increased intrarenal AngII content. 4. Together, these results show that 5/6 NX TGR exhibit a profound deficiency of intrarenal availability of active epoxygenase metabolites (EETs), which probably contributes to the progression of CKD in this model of AngII-dependent hypertension, and that restoration of intrarenal availability of EETs using long-term c-AUCB treatment exhibits substantial renoprotective actions.
Assuntos
Epóxido Hidrolases/antagonistas & inibidores , Epóxido Hidrolases/metabolismo , Hipertensão/metabolismo , Ratos Transgênicos/metabolismo , Angiotensina II/farmacologia , Animais , Pressão Sanguínea/efeitos dos fármacos , Feminino , Hipertensão/tratamento farmacológico , Nefrectomia/métodos , Ratos , Ratos Sprague-Dawley , Insuficiência Renal Crônica/tratamento farmacológico , Insuficiência Renal Crônica/metabolismo , Taxa de SobrevidaRESUMO
OBJECTIVE: In the present study, we compared the effects of treatment with the novel soluble epoxide hydrolase (sEH) inhibitor (c-AUCB) with those of the AT1 receptor antagonist losartan on blood pressure (BP), autoregulation of renal blood flow (RBF) and on glomerular filtration rate (GFR) and the pressure-natriuresis relationship in response to stepwise reduction in renal arterial pressure (RAP) in Cyp1a1-Ren-2 transgenic rats. METHODS: Hypertension was induced in Cyp1a1-Ren-2 rats through dietary administration for 11 days of the natural xenobiotic indole-3-carbinol (I3C) which activates the renin gene. Treatment with c-AUCB and losartan was started 48âh before initiating administration of the diet containing I3C. Rats were prepared for renal functional studies to evaluate in-vivo renal autoregulatory efficiency when RAP was gradually decreased by an aortic clamp. RESULTS: I3C administration resulted in the development of severe hypertension which was associated with markedly lower basal RBF and GFR and substantially impaired autoregulatory efficiency as well as a suppression of the pressure-natriuresis relationship when compared with noninduced rats. Treatment with c-AUCB significantly decreased BP, improved autoregulatory efficiency of RBF and GFR and the slope of pressure-natriuresis relationship. Treatment with losartan completely prevented the impaired autoregulation and pressure-natriuresis relationship as well as the development of hypertension in I3C-induced rats. CONCLUSION: Our present findings indicate that chronic treatment with the sEH inhibitor c-AUCB substantially attenuates the development of malignant hypertension in I3C-induced rats likely via improvement of the renal autoregulatory efficiency and the pressure-natriuresis relationship.
Assuntos
Pressão Sanguínea/efeitos dos fármacos , Citocromo P-450 CYP1A1/genética , Inibidores Enzimáticos/farmacologia , Epóxido Hidrolases/antagonistas & inibidores , Hipertensão Maligna/prevenção & controle , Nefropatias/prevenção & controle , Natriurese/efeitos dos fármacos , Renina/genética , Bloqueadores do Receptor Tipo 1 de Angiotensina II/farmacologia , Animais , Pressão Sanguínea/fisiologia , Modelos Animais de Doenças , Taxa de Filtração Glomerular/efeitos dos fármacos , Taxa de Filtração Glomerular/fisiologia , Hipertensão Maligna/induzido quimicamente , Hipertensão Maligna/fisiopatologia , Indóis/efeitos adversos , Rim/irrigação sanguínea , Rim/patologia , Rim/fisiopatologia , Nefropatias/fisiopatologia , Losartan/farmacologia , Natriurese/fisiologia , Ratos , Ratos Transgênicos , Fluxo Sanguíneo Regional/efeitos dos fármacos , Fluxo Sanguíneo Regional/fisiologiaRESUMO
Epoxyeicotrienoic acids (EETs) are cytochrome P450-dependent anti-hypertensive and anti-inflammatory derivatives of arachidonic acid, which are highly abundant in the kidney and considered reno-protective. EETs are degraded by the enzyme soluble epoxide hydrolase (sEH) and sEH inhibitors are considered treatment for chronic renal failure (CRF). We determined whether sEH inhibition attenuates the progression of CRF in the 5/6-nephrectomy model (5/6-Nx) in mice. 5/6-Nx mice were treated with a placebo, an ACE-inhibitor (Ramipril, 40 mg/kg), the sEH-inhibitor cAUCB or the CYP-inhibitor fenbendazole for 8 weeks. 5/6-Nx induced hypertension, albuminuria, glomerulosclerosis and tubulo-interstitial damage and these effects were attenuated by Ramipril. In contrast, cAUCB failed to lower the blood pressure and albuminuria was more severe as compared to placebo. Plasma EET-levels were doubled in 5/6 Nx-mice as compared to sham mice receiving placebo. Renal sEH expression was attenuated in 5/6-Nx mice but cAUCB in these animals still further increased the EET-level. cAUCB also increased 5-HETE and 15-HETE, which derive from peroxidation or lipoxygenases. Similar to cAUCB, CYP450 inhibition increased HETEs and promoted albuminuria. Thus, sEH-inhibition failed to elicit protective effects in the 5/6-Nx model and showed a tendency to aggravate the disease. These effects might be consequence of a shift of arachidonic acid metabolism into the lipoxygenase pathway.