RESUMEN

The intestinal epithelium constitutes a first line of defense of the innate immune system. Epithelial dysfunction is a hallmark of intestinal disorders such as inflammatory bowel diseases (IBDs). The actin cytoskeleton controls epithelial barrier integrity but the function of actin regulators such as cortactin is poorly understood. Given that cortactin controls endothelial permeability, we hypothesized that cortactin is also important for epithelial barrier regulation. We found increased permeability in the colon of cortactin-KO mice that was accompanied by reduced levels of ZO-1, claudin-1, and E-cadherin. By contrast, claudin-2 was upregulated. Cortactin deficiency increased RhoA/ROCK1-dependent actomyosin contractility, and inhibition of ROCK1 rescued the barrier defect. Interestingly, cortactin deficiency caused increased epithelial proliferation without affecting apoptosis. KO mice did not develop spontaneous colitis, but were more susceptible to dextran sulfate sodium colitis and showed severe colon tissue damage and edema formation. KO mice with colitis displayed strong mucus deposition and goblet cell depletion. In healthy human colon tissues, cortactin co-localized with ZO-1 at epithelial cell contacts. In IBDs patients, we observed decreased cortactin levels and loss of co-localization with ZO-1. Thus, cortactin is a master regulator of intestinal epithelial barrier integrity in vivo and could serve as a suitable target for pharmacological intervention in IBDs.

Asunto(s)

Actomiosina/metabolismo , Colitis/inmunología , Cortactina/metabolismo , Enfermedades Inflamatorias del Intestino/inmunología , Mucosa Intestinal/patología , Quinasas Asociadas a rho/metabolismo , Proteína de Unión al GTP rhoA/metabolismo , Animales , Apoptosis , Proliferación Celular , Colitis/inducido químicamente , Cortactina/genética , Citoesqueleto/metabolismo , Sulfato de Dextran , Modelos Animales de Enfermedad , Humanos , Inmunidad Innata , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Proteína de la Zonula Occludens-1/metabolismo , Proteína de Unión al GTP rhoA/genética

RESUMEN

Beta-adrenergic receptor (ßAR)-dependent blood vessel relaxation is impaired in older animals and G protein activation has been suggested as the causative mechanism. Here, we investigated the role of ßAR subtypes (ß1AR, ß2AR, and ß3AR) and cAMP in maturation-dependent vasorelaxation impairment. Aortic rings from 15 Sprague-Dawley male rats (3 or 9 weeks old) were harvested and left intact or denuded of the endothelium. Vascular relaxation in aortic rings from younger and older groups was compared in the presence of ßAR subtype agonists and antagonists along with cAMP and cGMP antagonists. Isolated aortic rings were used to evaluate relaxation responses, protein expression was evaluated by western blot or real time PCR, and metabolites were measured by ELISA. Expression of ßAR subtypes and adenylyl cyclase was assessed, and cAMP activity was measured in vascular tissue from both groups. Isoproterenol- and BRL744-dependent relaxation in aortic rings with and without endothelium from 9-week-old rats was impaired compared with younger rats. The ß1AR antagonist CGP20712A (10-7 M) did not affect isoproterenol or BRL744-dependent relaxation in arteries from either group. The ß2AR antagonist ICI-118,551 (10-7 M) inhibited isoproterenol-dependent aortic relaxation in both groups. The ß3AR antagonist SR59230A (10-7 M) inhibited isoproterenol- and BRL744-dependent aortic ring relaxation in younger but not in older rats. All ßAR subtypes were expressed in both groups, although ß3AR expression was lower in the older group. Adenylyl cyclase (SQ 22536) or protein kinase A (H89) inhibitors prevented isoproterenol-induced relaxation in younger but not in older rats. Production of cAMP was reduced in the older group. Adenylyl cyclase III and RyR3 protein expression was higher in the younger group. In conclusion, altered expression of ß3AR and adenylyl cyclase III may be responsible for reduced cAMP production in the older group.

Asunto(s)

Inhibidores de Adenilato Ciclasa/farmacología , Antagonistas Adrenérgicos beta/farmacología , Aorta Torácica/efectos de los fármacos , Aorta Torácica/fisiopatología , Vasodilatación/efectos de los fármacos , Vasodilatación/fisiología , Adenilil Ciclasas/fisiología , Factores de Edad , Albuterol/farmacología , Animales , Aorta Torácica/fisiología , Western Blotting , AMP Cíclico/análisis , AMP Cíclico/metabolismo , Dobutamina/farmacología , Ensayo de Inmunoadsorción Enzimática , Expresión Génica , Isoproterenol/farmacología , Masculino , Ratas Sprague-Dawley , Reacción en Cadena en Tiempo Real de la Polimerasa , Receptores Adrenérgicos beta/fisiología , Valores de Referencia , Factores de Tiempo

RESUMEN

Beta-adrenergic receptor (βAR)-dependent blood vessel relaxation is impaired in older animals and G protein activation has been suggested as the causative mechanism. Here, we investigated the role of βAR subtypes (β1AR, β2AR, and β3AR) and cAMP in maturation-dependent vasorelaxation impairment. Aortic rings from 15 Sprague-Dawley male rats (3 or 9 weeks old) were harvested and left intact or denuded of the endothelium. Vascular relaxation in aortic rings from younger and older groups was compared in the presence of βAR subtype agonists and antagonists along with cAMP and cGMP antagonists. Isolated aortic rings were used to evaluate relaxation responses, protein expression was evaluated by western blot or real time PCR, and metabolites were measured by ELISA. Expression of βAR subtypes and adenylyl cyclase was assessed, and cAMP activity was measured in vascular tissue from both groups. Isoproterenol- and BRL744-dependent relaxation in aortic rings with and without endothelium from 9-week-old rats was impaired compared with younger rats. The β1AR antagonist CGP20712A (10-7 M) did not affect isoproterenol or BRL744-dependent relaxation in arteries from either group. The β2AR antagonist ICI-118,551 (10-7 M) inhibited isoproterenol-dependent aortic relaxation in both groups. The β3AR antagonist SR59230A (10-7 M) inhibited isoproterenol- and BRL744-dependent aortic ring relaxation in younger but not in older rats. All βAR subtypes were expressed in both groups, although β3AR expression was lower in the older group. Adenylyl cyclase (SQ 22536) or protein kinase A (H89) inhibitors prevented isoproterenol-induced relaxation in younger but not in older rats. Production of cAMP was reduced in the older group. Adenylyl cyclase III and RyR3 protein expression was higher in the younger group. In conclusion, altered expression of β3AR and adenylyl cyclase III may be responsible for reduced cAMP production in the older group.

Asunto(s)

Animales , Masculino , Aorta Torácica/efectos de los fármacos , Aorta Torácica/fisiopatología , Vasodilatación/efectos de los fármacos , Vasodilatación/fisiología , Antagonistas de Receptores Adrenérgicos beta 1/farmacología , Inhibidores de Adenilato Ciclasa/farmacología , Aorta Torácica/fisiología , Factores de Tiempo , Expresión Génica , Adenilil Ciclasas/fisiología , Western Blotting , Factores de Edad , AMP Cíclico/análisis , AMP Cíclico/metabolismo , Albuterol/farmacología , Dobutamina/farmacología

RESUMEN

OBJECTIVE: This study aimed to evaluate the levels of the adipokines, resistin and adiponectin in normotensive and high normal blood pressure patients. METHODS: Circulating levels of the adipokines, resistin and adiponectin were measured by enzyme-linked immunosorbent assay (ELISA; R'D Systems, Minneapolis) in 20 high normal blood pressure patients and in 20 age-matched normotensive non-diabetic subjects. Statistical analysis was performed with analysis of variance (ANOVA). RESULTS: The control group showed non-significantly decreased levels of resistin when compared with patients with high normal blood pressure [systolic 130-139 mmHg; diastolic 85-89 mmHg] (12.25 vs 14.38 pg/mL, p = 0.40). There were significantly higher levels of adiponectin in the control group when compared with high normal blood pressure patients (11.3 vs 7.51 µg/mL, p = 0.028). CONCLUSIONS: High normal blood pressure patients have increased levels of resistin and lower values of adiponectin when compared with age-matched non-diabetic normotensive subjects. This may explain why those patients showed more progression to hypertension, atherosclerosis and cardiovascular risk than normotensive subjects.