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Obstructive sleep apnea (OSA), characterized by episodes of intermittent hypoxia (IH), is highly prevalent in patients with abdominal aortic aneurysm (AAA). However, whether IH serves as an independent risk factor for AAA development remains to be investigated. Here, we determined the effects of chronic (6 months) IH on angiotensin (Ang II)-induced AAA development in C57BL/6J male mice, and IH underlying mechanisms in cultured vascular smooth muscle cells (SMCs). IH increased abdominal aortic diameter and the incidence of AAA in mice infused with Ang II as assessed by transabdominal ultrasound imaging. Importantly, IH with Ang II augmented aortic elastin degradation and expression of matrix metalloproteinase (MMP)s, mainly MMP8, MMP12 and a disintegrin and metalloproteinase-17 (ADAM17) as measured by histology and immunohistochemistry. Mechanistically, IH increased the activities of MMP2, MMP8, MMP9, MMP12, and ADAM17, while reducing the expression of the MMP regulator, reversion inducing cysteine rich protein with kazal motifs (RECK) in cultured SMCs. Aortic samples from human AAA were associated with decreased RECK and increased expression of ADAM17 and MMPs. These data suggest that IH promotes the development of AAA in association with an increased expression of MMPs and ADAM17, while decreased expression of RECK may be responsible for the increased protease activity. These findings support a potential causal link between OSA and AAA and provide a better understanding of the molecular mechanisms underlying the pathogenesis of AAA.
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Endothelial insulin resistance represents a causal factor in the pathogenesis of type 2 diabetes (T2D) and vascular disease, thus the need to identify molecular mechanisms underlying defects in endothelial insulin signaling. We previously have shown that a disintegrin and metalloproteinase-17 (ADAM17) is increased while insulin receptor α-subunit (IRα) is decreased in the vasculature of patients with T2D, leading to impaired insulin-induced vasodilation. We have also demonstrated that ADAM17 sheddase activity targets IRα; however, the mechanisms driving endothelial ADAM17 activity in T2D are largely unknown. Herein, we report that externalization of phosphatidylserine (PS) to the outer leaflet of the plasma membrane causes ADAM17-mediated shedding of IRα and blunting of insulin signaling in endothelial cells. Furthermore, we demonstrate that endothelial PS externalization is mediated by the phospholipid scramblase anoctamin-6 (ANO6) and that this process can be stimulated by neuraminidase, a soluble enzyme that cleaves sialic acid residues. Of note, we demonstrate that men and women with T2D display increased levels of neuraminidase activity in plasma, relative to age-matched healthy individuals, and this occurs in conjunction with increased ADAM17 activity and impaired leg blood flow responses to endogenous insulin. Collectively, this work reveals the neuraminidase-ANO6-ADAM17 axis as a novel potential target for restoring endothelial insulin sensitivity in T2D.NEW & NOTEWORTHY This work provides the first evidence that neuraminidase, an enzyme increased in the circulation of men and women with type 2 diabetes (T2D), promotes anoctamin-6 (ANO6)-dependent externalization of phosphatidylserine in endothelial cells, which in turn leads to activation of a disintegrin and metalloproteinase-17 (ADAM17) and consequent shedding of the insulin receptor-α from the cell surface. Hence, this work supports that consideration should be given to the neuraminidase-ANO6-ADAM17 axis as a novel potential target for restoring endothelial insulin sensitivity in T2D.
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Diabetes Mellitus Tipo 2 , Resistência à Insulina , Masculino , Humanos , Feminino , Células Endoteliais/metabolismo , Receptor de Insulina/metabolismo , Fosfatidilserinas/metabolismo , Neuraminidase/metabolismo , Insulina/metabolismo , Desintegrinas , Proteína ADAM17/metabolismo , Anoctaminas/metabolismoRESUMO
Background: Substantial sex differences exist in atherosclerosis. Excessive reactive oxygen species (ROS) formation could lead to endothelial dysfunction which is critical to atherosclerosis development and progression. Helicobacter pylori (H. pylori) infection has been shown to attenuate endothelial function via exosomes-mediated ROS formation. We have demonstrated that H. pylori infection selectively increases atherosclerosis risk in males with unknown mechanism(s). The present study was to test the hypothesis that H. pylori infection impaired endothelial function selectively in male mice through exosome-mediated ROS formation. Methods and results: Age-matched male and female C57BL/6 mice were infected with CagA+ H. pylori to investigate sex differences in H. pylori infection-induced endothelial dysfunction. H. pylori infection attenuated acetylcholine (ACh)-induced endothelium-dependent aortic relaxation without changing nitroglycerine-induced endothelium-independent relaxation in male but not female mice, associated with increased ROS formation in aorta compared with controls, which could be reversed by N-acetylcysteine treatment. Treatment of cultured mouse brain microvascular endothelial cells with exosomes from H. pylori infected male, not female, mice significantly increased intracellular ROS production and impaired endothelial function with decreased migration, tube formation, and proliferation, which could be prevented with N-acetylcysteine treatment. Conclusions: H. pylori infection selectively impairs endothelial function in male mice due to exosome-mediated ROS formation.
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Aterosclerose , Exossomos , Infecções por Helicobacter , Helicobacter pylori , Masculino , Feminino , Animais , Camundongos , Espécies Reativas de Oxigênio , Células Endoteliais , Acetilcisteína , Infecções por Helicobacter/complicações , Camundongos Endogâmicos C57BL , Aterosclerose/complicações , EndotélioRESUMO
Adropin is a peptide largely secreted by the liver and known to regulate energy homeostasis; however, it also exerts cardiovascular effects. Herein, we tested the hypothesis that low circulating levels of adropin in obesity and type 2 diabetes (T2D) contribute to arterial stiffening. In support of this hypothesis, we report that obesity and T2D are associated with reduced levels of adropin (in liver and plasma) and increased arterial stiffness in mice and humans. Establishing causation, we show that mesenteric arteries from adropin knockout mice are also stiffer, relative to arteries from wild-type counterparts, thus recapitulating the stiffening phenotype observed in T2D db/db mice. Given the above, we performed a set of follow-up experiments, in which we found that 1) exposure of endothelial cells or isolated mesenteric arteries from db/db mice to adropin reduces filamentous actin (F-actin) stress fibers and stiffness, 2) adropin-induced reduction of F-actin and stiffness in endothelial cells and db/db mesenteric arteries is abrogated by inhibition of nitric oxide (NO) synthase, and 3) stimulation of smooth muscle cells or db/db mesenteric arteries with a NO mimetic reduces stiffness. Lastly, we demonstrated that in vivo treatment of db/db mice with adropin for 4 wk reduces stiffness in mesenteric arteries. Collectively, these findings indicate that adropin can regulate arterial stiffness, likely via endothelium-derived NO, and thus support the notion that "hypoadropinemia" should be considered as a putative target for the prevention and treatment of arterial stiffening in obesity and T2D.NEW & NOTEWORTHY Arterial stiffening, a characteristic feature of obesity and type 2 diabetes (T2D), contributes to the development and progression of cardiovascular diseases. Herein we establish that adropin is decreased in obese and T2D models and furthermore provide evidence that reduced adropin may directly contribute to arterial stiffening. Collectively, findings from this work support the notion that "hypoadropinemia" should be considered as a putative target for the prevention and treatment of arterial stiffening in obesity and T2D.
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Diabetes Mellitus Tipo 2 , Rigidez Vascular , Actinas , Animais , Células Endoteliais , Humanos , Artérias Mesentéricas , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Óxido Nítrico , Óxido Nítrico Sintase , Obesidade/complicações , Peptídeos/farmacologia , Rigidez Vascular/fisiologiaRESUMO
Inflammation and vascular insulin resistance are hallmarks of type 2 diabetes (T2D). However, several potential mechanisms causing abnormal endothelial insulin signaling in T2D need further investigation. Evidence indicates that the activity of ADAM17 (a disintegrin and metalloproteinase-17) and the presence of insulin receptor (IR) in plasma are increased in subjects with T2D. Accordingly, we hypothesized that in T2D, increased ADAM17 activity sheds the IR ectodomain from endothelial cells and impairs insulin-induced vasodilation. We used small visceral arteries isolated from a cross-sectional study of subjects with and without T2D undergoing bariatric surgery, human cultured endothelial cells, and recombinant proteins to test our hypothesis. Here, we demonstrate that arteries from subjects with T2D had increased ADAM17 expression, reduced presence of tissue inhibitor of metalloproteinase-3 (TIMP3), decreased extracellular IRα, and impaired insulin-induced vasodilation versus those from subjects without T2D. In vitro, active ADAM17 cleaved the ectodomain of the IRß subunit. Endothelial cells with ADAM17 overexpression or exposed to the protein kinase-C activator, PMA, had increased ADAM17 activity, decreased IRα presence on the cell surface, and increased IR shedding. Moreover, pharmacological inhibition of ADAM17 with TAPI-0 rescued PMA-induced IR shedding and insulin-signaling impairments in endothelial cells and insulin-stimulated vasodilation in human arteries. In aggregate, our findings suggest that ADAM17-mediated shedding of IR from the endothelial surface impairs insulin-mediated vasodilation. Thus, we propose that inhibition of ADAM17 sheddase activity should be considered a strategy to restore vascular insulin sensitivity in T2D.NEW & NOTEWORTHY To our knowledge, this is the first study to investigate the involvement of ADAM17 in causing impaired insulin-induced vasodilation in T2D. We provide evidence that ADAM17 activity is increased in the vasculature of patients with T2D and support the notion that ADAM17-mediated shedding of endothelial IRα ectodomains is a novel mechanism causing vascular insulin resistance. Our results highlight that targeting ADAM17 activity may be a potential therapeutic strategy to correct vascular insulin resistance in T2D.
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Diabetes Mellitus Tipo 2 , Resistência à Insulina , Proteína ADAM17/genética , Proteína ADAM17/metabolismo , Estudos Transversais , Diabetes Mellitus Tipo 2/metabolismo , Desintegrinas , Células Endoteliais/metabolismo , Humanos , Insulina/metabolismo , Receptor de Insulina/metabolismo , Proteínas Recombinantes/metabolismo , Inibidor Tecidual de Metaloproteinase-3/metabolismoRESUMO
The glycocalyx is a polysaccharide structure that protrudes from the body of a cell. It is primarily conformed of glycoproteins and proteoglycans, which provide communication, electrostatic charge, ionic buffering, permeability, and mechanosensation-mechanotransduction capabilities to cells. In blood vessels, the endothelial glycocalyx that projects into the vascular lumen separates the vascular wall from the circulating blood. Such a physical location allows a number of its components, including sialic acid, glypican-1, heparan sulfate, and hyaluronan, to participate in the mechanosensation-mechanotransduction of blood flow-dependent shear stress, which results in the synthesis of nitric oxide and flow-mediated vasodilation. The endothelial glycocalyx also participates in the regulation of vascular permeability and the modulation of inflammatory responses, including the processes of leukocyte rolling and extravasation. Its structural architecture and negative charge work to prevent macromolecules greater than approximately 70 kDa and cationic molecules from binding and flowing out of the vasculature. This also prevents the extravasation of pathogens such as bacteria and virus, as well as that of tumor cells. Due to its constant exposure to shear and circulating enzymes such as neuraminidase, heparanase, hyaluronidase, and matrix metalloproteinases, the endothelial glycocalyx is in a continuous process of degradation and renovation. A balance favoring degradation is associated with a variety of pathologies including atherosclerosis, hypertension, vascular aging, metastatic cancer, and diabetic vasculopathies. Consequently, ongoing research efforts are focused on deciphering the mechanisms that promote glycocalyx degradation or limit its syntheses, as well as on therapeutic approaches to improve glycocalyx integrity with the goal of reducing vascular disease. © 2022 American Physiological Society. Compr Physiol 12: 1-31, 2022.
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Glicocálix , Mecanotransdução Celular , Endotélio Vascular/fisiologia , Glicocálix/metabolismo , Glicocálix/patologia , Heparitina Sulfato/metabolismo , Humanos , Mecanotransdução Celular/fisiologia , Estresse MecânicoRESUMO
Consumption of diets high in fat, sugar, and salt (Western diet, WD) is associated with accelerated arterial stiffening, a major independent risk factor for cardiovascular disease (CVD). Women with obesity are more prone to develop arterial stiffening leading to more frequent and severe CVD compared with men. As tissue transglutaminase (TG2) has been implicated in vascular stiffening, our goal herein was to determine the efficacy of cystamine, a nonspecific TG2 inhibitor, at reducing vascular stiffness in female mice chronically fed a WD. Three experimental groups of female mice were created. One was fed regular chow diet (CD) for 43 wk starting at 4 wk of age. The second was fed a WD for the same 43 wk, whereas a third cohort was fed WD, but also received cystamine (216 mg/kg/day) in the drinking water during the last 8 wk on the diet (WD + C). All vascular stiffness parameters assessed, including aortic pulse wave velocity and the incremental modulus of elasticity of isolated femoral and mesenteric arteries, were significantly increased in WD- versus CD-fed mice, and reduced in WD + C versus WD-fed mice. These changes coincided with respectively augmented and diminished vascular wall collagen and F-actin content, with no associated effect in blood pressure. In cultured human vascular smooth muscle cells, cystamine reduced TG2 activity, F-actin:G-actin ratio, collagen compaction capacity, and cellular stiffness. We conclude that cystamine treatment represents an effective approach to reduce vascular stiffness in female mice in the setting of WD consumption, likely because of its TG2 inhibitory capacity.NEW & NOTEWORTHY This study evaluates the novel role of transglutaminase 2 (TG2) inhibition to directly treat vascular stiffness. Our data demonstrate that cystamine, a nonspecific TG2 inhibitor, improves vascular stiffness induced by a diet rich in fat, fructose, and salt. This research suggests that TG2 inhibition might bear therapeutic potential to reduce the disproportionate burden of cardiovascular disease in females in conditions of chronic overnutrition.
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Cistamina/farmacologia , Dieta Ocidental/efeitos adversos , Inibidores Enzimáticos/farmacologia , Proteína 2 Glutamina gama-Glutamiltransferase/antagonistas & inibidores , Rigidez Vascular/efeitos dos fármacos , Actinas/metabolismo , Animais , Aorta/metabolismo , Aorta/fisiologia , Células Cultivadas , Colágeno/metabolismo , Elasticidade , Feminino , Humanos , Artérias Mesentéricas/metabolismo , Artérias Mesentéricas/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/fisiologia , Miócitos de Músculo Liso/metabolismo , Miócitos de Músculo Liso/fisiologia , Análise de Onda de PulsoRESUMO
Obesity and insulin resistance stiffen the vasculature, with females appearing to be more adversely affected. As augmented arterial stiffness is an independent predictor of cardiovascular disease (CVD), the increased predisposition of women with obesity and insulin resistance to arterial stiffening may explain their heightened risk for CVD. However, the cellular mechanisms by which females are more vulnerable to arterial stiffening associated with obesity and insulin resistance remain largely unknown. In this study, we provide evidence that female mice are more susceptible to Western diet-induced endothelial cell stiffening compared with age-matched males. Mechanistically, we show that the increased stiffening of the vascular intima in Western diet-fed female mice is accompanied by enhanced epithelial sodium channel (ENaC) activity in endothelial cells (EnNaC). Our data further indicate that: (i) estrogen signaling through estrogen receptor α (ERα) increases EnNaC activity to a larger extent in females compared with males, (ii) estrogen-induced activation of EnNaC is mediated by the serum/glucocorticoid inducible kinase 1 (SGK-1), and (iii) estrogen signaling stiffens endothelial cells when nitric oxide is lacking and this stiffening effect can be reduced with amiloride, an ENaC inhibitor. In aggregate, we demonstrate a sexual dimorphism in obesity-associated endothelial stiffening, whereby females are more vulnerable than males. In females, endothelial stiffening with obesity may be attributed to estrogen signaling through the ERα-SGK-1-EnNaC axis, thus establishing a putative therapeutic target for female obesity-related vascular stiffening.
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Endotélio Vascular/fisiopatologia , Canais Epiteliais de Sódio/metabolismo , Obesidade/fisiopatologia , Caracteres Sexuais , Rigidez Vascular , Animais , Células Cultivadas , Endotélio Vascular/metabolismo , Feminino , Masculino , Camundongos Endogâmicos C57BL , Obesidade/metabolismoRESUMO
Recent clinical trials revealed that sodium-glucose co-transporter 2 (SGLT2) inhibitors significantly reduce cardiovascular events in type 2 diabetic patients, however, canagliflozin increased limb amputations, an effect not seen with other SGLT2 inhibitors. Since endothelial cell (EC) dysfunction promotes diabetes-associated vascular disease and limb ischemia, we hypothesized that canagliflozin, but not other SGLT2 inhibitors, impairs EC proliferation, migration, and angiogenesis. Treatment of human umbilical vein ECs (HUVECs) with clinically relevant concentrations of canagliflozin, but not empagliflozin or dapagliflozin, inhibited cell proliferation. In particular, 10 µM canagliflozin reduced EC proliferation by approximately 45%. The inhibition of EC growth by canagliflozin occurred in the absence of cell death and was associated with diminished DNA synthesis, cell cycle arrest, and a striking decrease in cyclin A expression. Restoration of cyclin A expression via adenoviral-mediated gene transfer partially rescued the proliferative response of HUVECs treated with canagliflozin. A high concentration of canagliflozin (50 µM) modestly inhibited HUVEC migration by 20%, but markedly attenuated their tube formation by 65% and EC sprouting from mouse aortas by 80%. A moderate 20% reduction in HUVEC migration was also observed with a high concentration of empagliflozin (50 µM), while neither empagliflozin nor dapagliflozin affected tube formation by HUVECs. The present study identified canagliflozin as a robust inhibitor of human EC proliferation and tube formation. The anti-proliferative action of canagliflozin occurs in the absence of cell death and is due, in part, to the blockade of cyclin A expression. Notably, these actions are not seen with empagliflozin or dapagliflozin. The ability of canagliflozin to exert these pleiotropic effects on ECs may contribute to the clinical actions of this drug.
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BACKGROUND: Arterial stiffness is emerging as an independent risk factor for the development of chronic kidney disease. The sodium glucose co-transporter 2 (SGLT2) inhibitors, which lower serum glucose by inhibiting SGLT2-mediated glucose reabsorption in renal proximal tubules, have shown promise in reducing arterial stiffness and the risk of cardiovascular and kidney disease in individuals with type 2 diabetes mellitus. Since hyperglycemia contributes to arterial stiffness, we hypothesized that the SGLT2 inhibitor empagliflozin (EMPA) would improve endothelial function, reduce aortic stiffness, and attenuate kidney disease by lowering hyperglycemia in type 2 diabetic female mice (db/db). MATERIALS/METHODS: Ten-week-old female wild-type control (C57BLKS/J) and db/db (BKS.Cg-Dock7m+/+Leprdb/J) mice were divided into three groups: lean untreated controls (CkC, n = 17), untreated db/db (DbC, n = 19) and EMPA-treated db/db mice (DbE, n = 19). EMPA was mixed with normal mouse chow at a concentration to deliver 10 mg kg-1 day-1, and fed for 5 weeks, initiated at 11 weeks of age. RESULTS: Compared to CkC, DbC showed increased glucose levels, blood pressure, aortic and endothelial cell stiffness, and impaired endothelium-dependent vasorelaxation. Furthermore, DbC exhibited impaired activation of endothelial nitric oxide synthase, increased renal resistivity and pulsatility indexes, enhanced renal expression of advanced glycation end products, and periarterial and tubulointerstitial fibrosis. EMPA promoted glycosuria and blunted these vascular and renal impairments, without affecting increases in blood pressure. In addition, expression of "reversion inducing cysteine rich protein with Kazal motifs" (RECK), an anti-fibrotic mediator, was significantly suppressed in DbC kidneys and partially restored by EMPA. Confirming the in vivo data, EMPA reversed high glucose-induced RECK suppression in human proximal tubule cells. CONCLUSIONS: Empagliflozin ameliorates kidney injury in type 2 diabetic female mice by promoting glycosuria, and possibly by reducing systemic and renal artery stiffness, and reversing RECK suppression.
Assuntos
Compostos Benzidrílicos/farmacologia , Glicemia/efeitos dos fármacos , Diabetes Mellitus Tipo 2/tratamento farmacológico , Angiopatias Diabéticas/prevenção & controle , Nefropatias Diabéticas/prevenção & controle , Glucosídeos/farmacologia , Rim/irrigação sanguínea , Rim/efeitos dos fármacos , Circulação Renal/efeitos dos fármacos , Inibidores do Transportador 2 de Sódio-Glicose/farmacologia , Transportador 2 de Glucose-Sódio/metabolismo , Rigidez Vascular/efeitos dos fármacos , Albuminúria/etiologia , Albuminúria/prevenção & controle , Animais , Glicemia/metabolismo , Linhagem Celular , Diabetes Mellitus Tipo 2/sangue , Diabetes Mellitus Tipo 2/complicações , Diabetes Mellitus Tipo 2/genética , Angiopatias Diabéticas/etiologia , Angiopatias Diabéticas/fisiopatologia , Nefropatias Diabéticas/etiologia , Nefropatias Diabéticas/patologia , Nefropatias Diabéticas/fisiopatologia , Modelos Animais de Doenças , Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/metabolismo , Endotélio Vascular/fisiopatologia , Feminino , Proteínas Ligadas por GPI/metabolismo , Glicosúria/etiologia , Glicosúria/prevenção & controle , Humanos , Rim/metabolismo , Rim/patologia , Camundongos Endogâmicos C57BL , Camundongos Mutantes , Fluxo Pulsátil/efeitos dos fármacos , Resistência Vascular/efeitos dos fármacos , Vasodilatação/efeitos dos fármacosRESUMO
Hyperglycemia-induced production of endothelin (ET)-1 is a hallmark of endothelial dysfunction in diabetes. Although the detrimental vascular effects of increased ET-1 are well known, the molecular mechanisms regulating endothelial synthesis of ET-1 in the setting of diabetes remain largely unidentified. Here, we show that adapter molecule TRAF3 interacting protein 2 (TRAF3IP2) mediates high glucose-induced ET-1 production in endothelial cells and ET-1-mediated endothelial cell inflammation. Specifically, we found that high glucose upregulated TRAF3IP2 in human aortic endothelial cells, which subsequently led to activation of JNK and IKKß. shRNA-mediated silencing of TRAF3IP2, JNK1, or IKKß abrogated high-glucose-induced ET-converting enzyme 1 expression and ET-1 production. Likewise, overexpression of TRAF3IP2, in the absence of high glucose, led to activation of JNK and IKKß as well as increased ET-1 production. Furthermore, ET-1 transcriptionally upregulated TRAF3IP2, and this upregulation was prevented by pharmacological inhibition of ET-1 receptor B using BQ-788, or inhibition of NADPH oxidase-derived reactive oxygen species using gp91ds-tat and GKT137831. Notably, we found that knockdown of TRAF3IP2 abolished ET-1-induced proinflammatory and adhesion molecule (IL-1ß, TNF-α, monocyte chemoattractant protein 1, ICAM-1, VCAM-1, and E-selectin) expression and monocyte adhesion to endothelial cells. Finally, we report that TRAF3IP2 is upregulated and colocalized with CD31, an endothelial marker, in the aorta of diabetic mice. Collectively, findings from the present study identify endothelial TRAF3IP2 as a potential new therapeutic target to suppress ET-1 production and associated vascular complications in diabetes. NEW & NOTEWORTHY This study provides the first evidence that the adapter molecule TRAF3 interacting protein 2 mediates high glucose-induced production of endothelin-1 by endothelial cells as well as endothelin-1-mediated endothelial cell inflammation. The findings presented herein suggest that TRAF3 interacting protein 2 may be an important therapeutic target in diabetic vasculopathy characterized by excess endothelin-1 production.
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Angiopatias Diabéticas/induzido quimicamente , Células Endoteliais/efeitos dos fármacos , Endotelina-1/toxicidade , Glucose/toxicidade , Inflamação/induzido quimicamente , Peptídeos e Proteínas Associados a Receptores de Fatores de Necrose Tumoral/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Adesão Celular , Moléculas de Adesão Celular/metabolismo , Células Cultivadas , Técnicas de Cocultura , Citocinas/metabolismo , Diabetes Mellitus Experimental/genética , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Tipo 1/genética , Diabetes Mellitus Tipo 1/metabolismo , Angiopatias Diabéticas/genética , Angiopatias Diabéticas/metabolismo , Células Endoteliais/metabolismo , Endotelina-1/genética , Endotelina-1/metabolismo , Feminino , Humanos , Quinase I-kappa B/metabolismo , Inflamação/genética , Inflamação/metabolismo , Mediadores da Inflamação/metabolismo , Masculino , Camundongos Endogâmicos NOD , Proteína Quinase 8 Ativada por Mitógeno/metabolismo , Monócitos/metabolismo , Transdução de Sinais/efeitos dos fármacos , Peptídeos e Proteínas Associados a Receptores de Fatores de Necrose Tumoral/genéticaRESUMO
OBJECTIVE: Aortic vascular stiffness has been implicated in the development of cardiovascular disease (CVD) and chronic kidney disease (CKD) in obese individuals. However, the mechanism promoting these adverse effects are unclear. In this context, promotion of obesity through consumption of a western diet (WD) high in fat and fructose leads to excess circulating uric acid. There is accumulating data implicating elevated uric acid in the promotion of CVD and CKD. Accordingly, we hypothesized that xanthine oxidase(XO) inhibition with allopurinol would prevent a rise in vascular stiffness and proteinuria in a translationally relevant model of WD-induced obesity. MATERIALS/METHODS: Four-week-old C57BL6/J male mice were fed a WD with excess fat (46%) and fructose (17.5%) with or without allopurinol (125mg/L in drinking water) for 16weeks. Aortic endothelial and extracellular matrix/vascular smooth muscle stiffness was evaluated by atomic force microscopy. Aortic XO activity, 3-nitrotyrosine (3-NT) and aortic endothelial sodium channel (EnNaC) expression were evaluated along with aortic expression of inflammatory markers. In the kidney, expression of toll like receptor 4 (TLR4) and fibronectin were assessed along with evaluation of proteinuria. RESULTS: XO inhibition significantly attenuated WD-induced increases in plasma uric acid, vascular XO activity and oxidative stress, in concert with reductions in proteinuria. Further, XO inhibition prevented WD-induced increases in aortic EnNaC expression and associated endothelial and subendothelial stiffness. XO inhibition also reduced vascular pro-inflammatory and maladaptive immune responses induced by consumption of a WD. XO inhibition also decreased WD-induced increases in renal TLR4 and fibronectin that associated proteinuria. CONCLUSIONS: Consumption of a WD leads to elevations in plasma uric acid, increased vascular XO activity, oxidative stress, vascular stiffness, and proteinuria all of which are attenuated with allopurinol administration.
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Dieta Ocidental , Inflamação/induzido quimicamente , Proteinúria/induzido quimicamente , Ácido Úrico/sangue , Rigidez Vascular/efeitos dos fármacos , Alopurinol/administração & dosagem , Alopurinol/farmacologia , Animais , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Ácido Úrico/farmacologia , Xantina Oxidase/antagonistas & inibidoresRESUMO
The role of estrogen receptor-α (ERα) signaling in the vasculature of females has been described under different experimental conditions and our group recently reported that lack of endothelial cell (EC) ERα in female mice fed a Western diet (WD) results in amelioration of vascular stiffness. Conversely, the role of ERα in the male vasculature in this setting has not been explored. In conditions of overnutrition and insulin resistance, augmented arterial stiffness, endothelial dysfunction, and arterial remodeling contribute to the development of cardiovascular disease. Here, we used a rodent model of decreased ERα expression in ECs [endothelial cell estrogen receptor-α knockout (EC-ERαKO)] to test the hypothesis that, similar to our findings in females, loss of ERα signaling in the endothelium of insulin-resistant males would result in decreased arterial stiffness. EC-ERαKO male mice and same-sex littermates were fed a WD (high in fructose and fat) for 20 weeks and then assessed for vascular function and stiffness. EC-ERαKO mice were heavier than littermates but exhibited decreased vascular stiffness without differences in endothelial-dependent vasodilatory responses. Mesenteric arteries from EC-ERαKO mice had significantly increased diameters, wall cross-sectional areas, and mean wall thicknesses, indicative of outward hypertrophic remodeling. This remodeling paralleled an increased vessel wall content of collagen and elastin, inhibition of matrix metalloproteinase activation and a decrease of the incremental modulus of elasticity. In addition, internal elastic lamina fenestrae were more abundant in the EC-ERαKO mice. In conclusion, loss of endothelial ERα reduces vascular stiffness in male mice fed a WD with an associated outward hypertrophic remodeling of resistance arteries.
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Dieta Ocidental/efeitos adversos , Receptor alfa de Estrogênio/genética , Remodelação Vascular/genética , Rigidez Vascular/genética , Animais , Células Cultivadas , Feminino , Masculino , Artérias Mesentéricas/fisiopatologia , Camundongos , Camundongos Knockout , Vasodilatação/genéticaRESUMO
Consumption of a diet high in fat and refined carbohydrates (Western diet [WD]) is associated with obesity and insulin resistance, both major risk factors for cardiovascular disease (CVD). In women, obesity and insulin resistance abrogate the protection against CVD likely afforded by estrogen signaling through estrogen receptor (ER)α. Indeed, WD in females results in increased vascular stiffness, which is independently associated with CVD. We tested the hypothesis that loss of ERα signaling in the endothelium exacerbates WD-induced vascular stiffening in female mice. We used a novel model of endothelial cell (EC)-specific ERα knockout (EC-ERαKO), obtained after sequential crossing of the ERα double floxed mice and VE-Cadherin Cre-recombinase mice. Ten-week-old females, EC-ERαKO and aged-matched genopairs were fed either a regular chow diet (control diet) or WD for 8 weeks. Vascular stiffness was measured in vivo by pulse wave velocity and ex vivo in aortic explants by atomic force microscopy. In addition, vascular reactivity was assessed in isolated aortic rings. Initial characterization of the model fed a control diet did not reveal changes in whole-body insulin sensitivity, aortic vasoreactivity, or vascular stiffness in the EC-ERαKO mice. Interestingly, ablation of ERα in ECs reduced WD-induced vascular stiffness and improved endothelial-dependent dilation. In the setting of a WD, endothelial ERα signaling contributes to vascular stiffening in females. The precise mechanisms underlying the detrimental effects of endothelial ERα in the setting of a WD remain to be elucidated.
Assuntos
Dieta Ocidental , Células Endoteliais/metabolismo , Receptor alfa de Estrogênio/metabolismo , Rigidez Vascular/fisiologia , Animais , Antígenos CD/genética , Antígenos CD/metabolismo , Aorta Torácica/metabolismo , Aorta Torácica/fisiologia , Caderinas/genética , Caderinas/metabolismo , Receptor alfa de Estrogênio/genética , Feminino , Artéria Femoral/fisiologia , Immunoblotting , Camundongos Knockout , Camundongos Transgênicos , Microscopia de Força Atômica , Análise de Onda de Pulso , Fator de Crescimento Transformador beta/metabolismo , Rigidez Vascular/genética , VasodilataçãoRESUMO
Inward remodeling of the resistance vasculature is strongly associated with life-threatening cardiovascular events. Previous studies have demonstrated that both actin polymerization and the activation of transglutaminases mediate early stages of the transition from a structurally normal vessel to an inwardly remodeled one. Ex vivo studies further suggest that a few hours of exposure to vasoconstrictor agonists induces inward remodeling in the absence of changes in intraluminal pressure. Here we report that a short, 10-min, topical exposure to serotonin (5-HT) + N(ω)-nitro-l-arginine methyl ester hydrochloride (l-NAME) was sufficient to initiate inward remodeling processes in rat cremasteric feed arterioles (100-200 µm lumen diameter), in vivo. Addition of the transglutaminase inhibitor, cystamine, blocked the in vivo remodeling. We further demonstrate that, in isolated arterioles, 5-HT + l-NAME activates transglutaminases and modulates the phosphorylation state of cofilin, a regulator of actin depolymerization. The 5-HT + l-NAME-induced remodeling process in isolated arterioles was also inhibited by an inhibitor of Lim Kinase, the kinase that phosphorylates and inactivates cofilin. Therefore, our results indicate that a brief vasoconstriction induced by 5-HT + l-NAME is able to reduce the passive structural diameter of arterioles through processes that are dependent on the activation of transglutaminases and Lim kinase, and the subsequent phosphorylation of cofilin.
Assuntos
Citoesqueleto de Actina/efeitos dos fármacos , Citoesqueleto de Actina/metabolismo , Arteríolas/efeitos dos fármacos , Serotonina/farmacologia , Transglutaminases/metabolismo , Fatores de Despolimerização de Actina/metabolismo , Animais , Cistamina/farmacologia , Ativação Enzimática/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Técnicas In Vitro , Masculino , Músculo Esquelético/irrigação sanguínea , Músculo Esquelético/efeitos dos fármacos , NG-Nitroarginina Metil Éster/farmacologia , Fosforilação , Ratos , Ratos Sprague-Dawley , Transglutaminases/antagonistas & inibidores , Vasoconstritores/farmacologiaRESUMO
The prevalence of obesity and associated medical disorders has increased dramatically in the United States and throughout much of the world in the past decade. Obesity, induced by excess intake of carbohydrates and fats, is a major cause of Type 2 diabetes, hypertension, and the cardiorenal metabolic syndrome. There is emerging evidence that excessive nutrient intake promotes signaling through the mammalian target of rapamycin (mTOR), which, in turn, may lead to alterations of cellular metabolic signaling leading to insulin resistance and obesity-related diseases, such as diabetes, cardiovascular and kidney disease, as well as cancer. While the pivotal role of mTOR signaling in regulating metabolic stress, autophagy, and adaptive immune responses has received increasing attention, there remain many gaps in our knowledge regarding this important nutrient sensor. For example, the precise cellular signaling mechanisms linking excessive nutrient intake and enhanced mTOR signaling with increased cardiovascular and kidney disease, as well as cancer, are not well understood. In this review, we focus on the effects that the interaction between excess intake of nutrients and enhanced mTOR signaling have on the promotion of obesity-associated diseases and potential therapeutic strategies involving targeting mTOR signaling.
Assuntos
Doenças Cardiovasculares/etiologia , Obesidade/complicações , Transdução de Sinais , Serina-Treonina Quinases TOR/metabolismo , Animais , Doenças Cardiovasculares/imunologia , Doenças Cardiovasculares/metabolismo , Doenças Cardiovasculares/fisiopatologia , Doenças Cardiovasculares/prevenção & controle , Ingestão de Energia , Metabolismo Energético , Humanos , Resistência à Insulina , Terapia de Alvo Molecular , Obesidade/tratamento farmacológico , Obesidade/imunologia , Obesidade/metabolismo , Obesidade/fisiopatologia , Proteínas Quinases S6 Ribossômicas/metabolismo , Fatores de Risco , Transdução de Sinais/efeitos dos fármacosRESUMO
In this study, we examined the ability of vasoactive agonists to induce dynamic changes in vascular smooth muscle cell (VSMC) elasticity and adhesion, and tested the hypothesis that these events are coordinated with rapid remodelling of the cortical cytoskeleton. Real-time measurement of cell elasticity was performed with atomic force microscopy (AFM) and adhesion was assessed with AFM probes coated with fibronectin (FN). Temporal data were analysed using an Eigen-decomposition method. Elasticity in VSMCs displayed temporal oscillations with three components at approximately 0.001, 0.004 and 0.07 Hz, respectively. Similarly, adhesion displayed a similar oscillatory pattern. Angiotensin II (ANG II, 10(-6) M) increased (+100%) the amplitude of the oscillations, whereas the vasodilator adenosine (ADO, 10(-4) M) reduced oscillation amplitude (-30%). To test whether the oscillatory changes were related to the architectural alterations in cortical cytoskeleton, the topography of the submembranous actin cytoskeleton (100-300 nm depth) was acquired with AFM. These data were analysed to compare cortical actin fibre distribution and orientation before and after treatment with vasoactive agonists. The results showed that ANG II increased the density of stress fibres by 23%, while ADO decreased the density of the stress fibres by 45%. AFM data were supported by Western blot and confocal microscopy. Collectively, these observations indicate that VSMC cytoskeletal structure and adhesion to the extracellular matrix are dynamically altered in response to agonist stimulation. Thus, vasoactive agonists probably invoke unique mechanisms that dynamically alter the behaviour and structure of both the VSMC cytoskeleton and focal adhesions to efficiently support the normal contractile behaviour of VSMCs.
Assuntos
Miócitos de Músculo Liso/efeitos dos fármacos , Miócitos de Músculo Liso/fisiologia , Vasoconstritores/farmacologia , Actinas/metabolismo , Adenosina/farmacologia , Adenosina/fisiologia , Angiotensina II/farmacologia , Angiotensina II/fisiologia , Animais , Fenômenos Biomecânicos , Adesão Celular/efeitos dos fármacos , Adesão Celular/fisiologia , Citoesqueleto/efeitos dos fármacos , Citoesqueleto/fisiologia , Módulo de Elasticidade/efeitos dos fármacos , Módulo de Elasticidade/fisiologia , Elasticidade/efeitos dos fármacos , Elasticidade/fisiologia , Microscopia de Força Atômica , Microscopia Confocal , Ratos , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologiaRESUMO
Inward remodeling is the most prevalent structural change found in the resistance arteries and arterioles of hypertensive individuals. Separate studies have shown that the inward remodeling process requires transglutaminase activation and the polymerization of actin. Therefore, we hypothesize that inward remodeling induced via endogenous transglutaminase activation requires and depends on actin cytoskeletal structures. To test this hypothesis, isolated and cannulated rat cremaster arterioles were exposed to dithiothreitol (DTT) to activate endogenous transglutaminases. DTT induced concentration-dependent vasoconstriction that was suppressed by coincubation with cystamine or cytochalasin-D to inhibit tranglutaminase activity or actin polymerization, respectively. Prolonged (4 h) exposure to DTT caused arteriolar inward remodeling that was also blocked by the presence of cystamine or cytochalasin-D. DTT inwardly remodeled arterioles had reduced passive diameters, augmented wall thickness-to-lumen ratios and altered elastic characteristics that were reverted upon disruption of the actin cytoskeleton with mycalolide-B. In freshly isolated arterioles, exposure to mycalolide-B caused no changes in their passive diameters or their elastic characteristics. These results suggest that, in arterioles, the early stages of the inward remodeling process induced by prolonged endogenous transglutaminase activation require actin dynamics and depend on changes in actin cytoskeletal structures.