Effects of trans- and cis-resveratrol on Ca2+ handling in A7r5 vascular myocytes

Although the natural polyphenol resveratrol posses a direct vasorelaxant effect, its effects on cytoplasmic Ca(2+) concentration ([Ca(2+)](i)) in vascular cells remain still unclear. Here, we have investigated the effects of the isomers trans- and cis-resveratrol on agonist- and high-K(+)-induced [Ca(2+)](i) increases and on voltage-activated transmembrane Ca(2+) fluxes using imaging and patch-clamp techniques in vascular A7r5 myocytes. Arginine vasopressin (AVP) or angiotensin II caused a biphasic increase in [Ca(2+)](i) that was reduced by preincubation with trans-resveratrol and cis-resveratrol. Both isomers also reduced the agonist-induced increase in [Ca(2+)](i) in absence of extracellular Ca(2+). In high-K(+) Ca(2+)-free solution, reintroduction of Ca(2+) caused a sustained rise in [Ca(2+)](i) that was reduced by preincubation with trans-resveratrol or cis-resveratrol. When the isomers were applied during the plateau phase of the agonist- or the high-K(+)-induced response, a biphasic change in [Ca(2+)](i) was observed: a transient reduction of the plateau (10 min). Finally, trans-resveratrol and cis-resveratrol inhibited voltage-dependent L-type Ca(2+) currents (I(Ca(L))). In conclusion, resveratrol isomers exert a dual effect on [Ca(2+)](i) handling in A7r5 myocytes: 1) a blockade of I(Ca(L)) and 2) an increase in [Ca(2+)](i) by depletion of intracellular Ca(2+) stores (which interferes with the agonist-induced release of intracellular Ca(2+)) and influx of Ca(2+), mainly due to activation of capacitative Ca(2+) entry, although other Ca(2+)-permeable channels are also involved. Taken together, these effects may explain, in part, the endothelium-independent vasorelaxant effects of resveratrol in rat aorta

Gut microbiota, diet, and chronic diseases: the role played by oxidative stress

The authors gratefully acknowledge the Espirito Santo Research Foundation (FAPES) (Grant CNPq/FAPES No. 24/2018; Termo Outorga 569/2018) and the Brazilian National Council for Research and Development (CNPq) (CNPq Grant Bolsa Produtividade 2015-2019) for the financial support to our research projectS

Crambescin C1 Acts as A Possible Substrate of iNOS and eNOS Increasing Nitric Oxide Production and Inducing In Vivo Hypotensive Effect

Crambescins are guanidine alkaloids from the sponge Crambe crambe. Crambescin C1 (CC) induces metallothionein genes and nitric oxide (NO) is one of the triggers. We studied and compared the in vitro, in vivo, and in silico effects of some crambescine A and C analogs. HepG2 gene expression was analyzed using microarrays. Vasodilation was studied in rat aortic rings. In vivo hypotensive effect was directly measured in anesthetized rats. The targets of crambescines were studied in silico. CC and homo-crambescine C1 (HCC), but not crambescine A1 (CA), induced metallothioneins transcripts. CC increased NO production in HepG2 cells. In isolated rat aortic rings, CC and HCC induced an endothelium-dependent relaxation related to eNOS activation and an endothelium-independent relaxation related to iNOS activation, hence both compounds increase NO and reduce vascular tone. In silico analysis also points to eNOS and iNOS as targets of Crambescin C1 and source of NO increment. CC effect is mediated through crambescin binding to the active site of eNOS and iNOS. CC docking studies in iNOS and eNOS active site revealed hydrogen bonding of the hydroxylated chain with residues Glu377 and Glu361, involved in the substrate recognition, and explains its higher binding affinity than CA. The later interaction and the extra polar contacts with its pyrimidine moiety, absent in the endogenous substrate, explain its role as exogenous substrate of NOSs and NO production. Our results suggest that CC serve as a basis to develop new useful drugs when bioavailability of NO is perturbed.Fil: Rubiolo, Juan Andrés. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas; Argentina. Ministerio de Ciencia, Tecnologia E Innovacion Productiva (santa Fe). - Gobierno de la Provincia de Santa Fe. Ministerio de Ciencia, Tecnologia E Innovacion Productiva (santa Fe).; Argentina. Universidad de Santiago de Compostela; España. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario; ArgentinaFil: Lence, Emilio. Universidad de Santiago de Compostela; EspañaFil: González Bello, Concepción. Universidad de Santiago de Compostela; EspañaFil: Roel, María. Universidad de Santiago de Compostela; EspañaFil: Gil Longo, José. Universidad de Santiago de Compostela; EspañaFil: Campos Toimil, Manuel. Universidad de Santiago de Compostela; EspañaFil: Ternon, Eva. Université Nice Sophia Antipolis. Laboratoire Jean-alexandre Dieudonné.; FranciaFil: Thomas, Olivier P.. National University of Ireland Galway; IrlandaFil: González Cantalapiedra, Antonio. Universidad de Santiago de Compostela; EspañaFil: López Alonso, Henar. Universidad de Santiago de Compostela; EspañaFil: Vieytes, Mercedes R.. Universidad de Santiago de Compostela; EspañaFil: Botana, Luis M.. Universidad de Santiago de Compostela; Españ

The microRNA-7-mediated reduction in EPAC-1 contributes to vascular endothelial permeability and eNOS uncoupling in murine experimental retinopathy

To investigate the consequences of oxidative stress and hypoxia on EPAC-1 expression during retinopathy. Oxygen-induced retinopathy was induced in mice and EPAC-1 expression investigated by immunofluorescence. In silico analyses were used to identify a link between EPAC-1 expression and microRNA-7-5p in endothelial cells and confirmed by western blot analyses on cells expressing microRNA-7-5p. In vitro, endothelial cells were either incubated at 2% oxygen or transfected with microRNA-7-5p, and the effects of these treatments on EPAC-1 expression, endothelial hyperpermeability and NO production were assessed. In the Ins2Akita mouse model, levels of EPAC-1 expression as well as microRNA-7-5p were assessed by qPCR. Endothelial nitric oxide synthase was assessed by immunoblotting in the Ins2Akita model. Hypoxia induces the expression of microRNA-7-5p that translationally inhibits the expression of EPAC-1 in endothelial cells, resulting in hyperpermeability and the loss of eNOS activity. Activation of EPAC-1 by the cAMP analogue 8-pCPT-2'-O-Me-cAMP reduced the sensitivity of EPAC-1 to oxidative stress and restored the endothelial permeability to baseline levels. Additionally, 8-pCPT-2'-O-Me-cAMP rescued eNOS activity and NO production. In mouse models of retinopathy, i.e., oxygen-induced retinopathy and the spontaneous diabetic heterozygous Ins2(Akita) mice, EPAC-1 levels are decreased which is associated with an increase in microRNA-7-5p expression and reduced eNOS activity. In retinopathy, EPAC-1 expression is decreased in a microRNA-7-mediated manner, contributing to endothelial dysfunction. Pharmacological activation of remnant EPAC-1 rescues endothelial function. Collectively, these data indicate that EPAC-1 resembles an efficacious and druggable target molecule for the amelioration of (diabetic) retinopathy

Synthesis and Vasorelaxant Activity of Nitrate−Coumarin Derivatives

Due to the need for new chemical entities for cardiovascular diseases, we have synthesized a new series of nitrate−coumarins and evaluated their vasorelaxant activity in contraction-relaxation studies using rat aorta rings precontracted with phenylephrine or by depolarization with a high concentration of potassium chloride. Four of the new compounds were able to relax smooth vascular muscle with a similar profile and potency to glyceryl trinitrate (IC50=12.73 nM) and sodium nitroprusside (IC50=4.32 nM). Coumarin-7-yl-methyl nitrate (4), the best compound within the series, was able to relax smooth vascular muscle in the low nanomolar range (IC50=1.92 nM). The mechanisms of action have been explored, being the activation of sGC and the opening of K+ channels involved. Our studies indicate that the new nitrate derivatives are reversible and not deleterious for aortic rings, suggesting that these compounds have a potential interest for the development of new and highly efficient vasodilator drugsXunta de Galicia. Grant Number: ED431B 2020/26. Ministerio de Ciencia e Innovación Grant Numbers: PID2020-116076RJ-I00/AEI/10.13039/501100011033, PID2020-119178GB-I00. Fundação para a Ciência e Tecnologia. Grant Numbers: PTDC/ASP-PES/28397/2017, CEECIND/02423/2018, UIDB/00081/2020, LA/P/0056/2020, EXPL/BIA-BQM/0492/2021S

List of Medicines to Avoid in Primary Care Health and Their Application In Polymedicated Patients

[Resumen] Introduction: La alta prevalencia del uso de medicación inadecuada y sus importantes consecuencias para la salud requieren herramientas específicas y ágiles que ayuden a detectarla y evitarla. Objetivo de este trabajo fue elaborar un listado de medicamentos a evitar en AP y aplicarlo en pacientes polimedicados de un servicio asistencial de Atención Primaria (AP). Métodos: En la Base de datos del Consejo General de Colegios Oficiales de Farmacéuticos español (BOT-Plus) se comprobó el estado y disponibilidad de cada uno de los 93 medicamentos del Listado Prescrire 2019. Se elaboró el Listado de medicamentos a evitar en AP con aquellos que estaban comercializados en España y se excluyeron los que no estaban financiados y los de uso exclusivo hospitalario. 2 2 Con el listado de medicamentos a evitar en AP se hizo un análisis retrospectivo de todos (N=262) los informes de prescripción de polimedicados >10 medicamentos del año 2017 en un servicio asistencial de AP (5 centros de salud). Se realizó análisis de frecuencias, medidas de tendencia central y dispersión; se estimaron (IC 95%) y se utilizó X o exacta de Fisher para determinar la asociación entre variables y análisis de regresión logística. Resultados: Se observó una prevalencia de polimedicados de 1,2%, con una media de edad de 71,7 ± 12,4 años y una media de prescripciones de 12 ±1,7 medicamentos. El listado de medicamentos a evitar en AP incluyó 45 principios activos. Los fármacos a evitar más usados han sido: duloxetina, sitagliptina y olmesartán. El 50,4% de los polimedicados tenían al menos un medicamento a evitar y una edad media de 68,5±11,8 años. El sexo fue un factor de riesgo de prescripción inadecuada, el hecho de ser mujer incrementa con un OR=1,8 (IC 95%=1,3-3,0) la probabilidad de medicamentos a evitar. Conclusiones: Un alto porcentaje de pacientes polimedicados tienen prescripto al menos un medicamento a evitar. El listado de medicamentos a evitar en AP es una herramienta útil para identificar la medicación inadecuada y para uso de los profesionales de AP.[Abstract] Introduction: The high prevalence of inappropriate medication use and its important health consequences for health require specific and agile tools to detect and avoid it. The objective of this work was to elaborate a list of medications to avoid in Primary Care and to apply it on the polymedicated patients of a Primary Care assistance service. Methods: In the Database of the Spanish General Council of Official Associations of Pharmacists (BOT-Plus) the status and availability of each of the 93 MAE of the Prescrire 2019 List was checked. The list of medications to be avoided in Primary Care was drawn up with those that were marketed in Spain and excluded those that were not financed and those for exclusive hospital use. With the list of medicines to avoid in Primary Care, a retrospective analysis was made of all the prescription reports of polimedicated >10 medications for 2017 in a Primary Care services (N=262) in 5 health centers. Frequency analysis, central tendency measures and dispersion were carried out; they were estimated [CI: 95%] and X or Fisher’s exact was used to determine the association between variables and logistic regression analysis. Results: A prevalence of polymedicated drugs of 1.2% was observed, with a mean age of 71.7 years (DT± 12.4) and a mean prescription of 12 drugs (DT±1.7). The list of medications to be avoided in PC included 45 active ingredients. The 50.4% of the polymedicated had at least one drug to avoid and an average age of 68.5 years (DT±11.8). Sex was a risk factor for inappropriate prescription, the fact of being a woman increases with an OR=1.8 (IC95%=1, 3-3.0) the probability of having some medicines to avoid. The most commonly used drugs to avoid were: duloxetine, sitagliptin and olmesartan. Conclusions: A high percentage of polymedicated patients are prescribed at least one drug to avoid. The Primary Care medication avoidance list is a useful tool for identifying inappropriate medication and for the use by Primary Care professionals

Protective effects of kefir in the angiotensin II-dependent hypertension

Recently, we have reported cardiovascular protective effects of the probiotic kefir in a model of primary hypertension. Now, we evaluated the beneficial effects of kefir in a model of secondary hypertension under hyperactivation of the renin-angiotensin-system by partially clipping one kidney artery (2K1C) for 60 days and compared with Sham rats. Maximum levels of arterial pressure were reached 7–14 days post-clipping in both 2K1C and 2K1C-Kefir, but after that time the values were approximately 20% lower in 2K1C-Kefir rats. Also, kefir attenuated the angiotensin converting enzyme activity (intrarenal-40%/plasma-25%) preventing the increase of angiotensin II in both samples. Isolated aortic rings showed an impaired relaxation to acetylcholine in 2K1C (-38%) compared to the Sham group and this difference was attenuated in 2K1C-Kefir rats (~15%). Additional analysis revealed that kefir protected kidney and vascular endothelium against the synergistic oxidative stress/angiotensin II-axis. Thus, kefir is an effective nutraceutical therapy for prevention/treatment of hypertensionThis work was supported by the CNPq/FAPES -Brazil (PRONEX CNPq # 24/2018; Termo Outorga 569/2018); FAPES-Universal (# 21/2018, Termo Outorga 120/2019); FAPES (BPC 552/2018;120/2019) and CNPq (BVN 160990/2019-0; SSM 312056/2018-5, TMCP 309277/2019-1 and ECV 305740/2019-9)S

The Treatment With the SGLT2 Inhibitor Empagliflozin Modifies the Hepatic Metabolome of Male Zucker Diabetic Fatty Rats Towards a Protective Profile

[Abstract] The EMPA-REG OUTCOME (Empagliflozin, Cardiovascular Outcome Event Trial in patients with Type 2 Diabetes Mellitus (T2DM)) trial evidenced the potential of sodium-glucose cotransporter 2 (SGLT2) inhibitors for the treatment of patients with diabetes and cardiovascular disease. Recent evidences have shown the benefits of the SGLT2 inhibitor empagliflozin on improving liver steatosis and fibrosis in patients with T2DM. Metabolomic studies have been shown to be very useful to improve the understanding of liver pathophysiology during the development and progression of metabolic hepatic diseases, and because the effects of empagliflozin and of other SGLT2 inhibitors on the complete metabolic profile of the liver has never been analysed before, we decided to study the impact on the liver of male Zucker diabetic fatty (ZDF) rats of a treatment for 6 weeks with empagliflozin using an untargeted metabolomics approach, with the purpose to help to clarify the benefits of the use of empagliflozin at hepatic level. We found that empagliflozin is able to change the hepatic lipidome towards a protective profile, through an increase of monounsaturated and polyunsaturated glycerides, phosphatidylcholines, phosphatidylethanolamines, lysophosphatidylinositols and lysophosphatidylcholines. Empagliflozin also induces a decrease in the levels of the markers of inflammation IL-6, chemerin and chemerin receptor in the liver. Our results provide new evidences regarding the molecular pathways through which empagliflozin could exert hepatoprotector beneficial effects in T2DM.This work was supported by Boehringer Ingelheim Pharma GmbH and Co., by the National Institute of Health “Fondo de Investigaciones Sanitarias del Instituto de Salud Carlos III” Madrid, Spain (PI15/00681, PI17/00409, PI18/00821, PI20/00902, RETICS Programme RD16/0012/0014 and CIBER de Enfermedades Cardiovasculares (CIBERCV)); European Regional Development Fund (FEDER) and European Union framework MSCA-RISE-H2020 Programme (Project number 734899). AH-A was funded by predoctoral research grants from Xunta de Galicia and FPU Program of the Spanish Ministry of Science, Innovation and Universities (Spain); MF-S was funded by the predoctoral research grants “Programa Científico do Centro de Investigación en Medicina Molecular e Enfermidades Crónicas (CiMUS) (Spain) and Xunta de Galicia; and AV-L was funded by the predoctoral research grant from the PFIS Program of the Spanish Ministry of Science and Instituto de Salud Carlos III (Spain

Mechanisms of Action of Kefir in Chronic Cardiovascular and Metabolic Diseases

The gut microbiota maintains a complex mutual interaction with different organs of the host. Whereas in normal conditions this natural community of trillions of microorganisms greatly contributes to the human health, gut dysbiosis is related with onset or worsening of diverse chronic systemic diseases. Thus, the reestablishment of gut microbiota homeostasis with consumption of prebiotics and probiotics may be a relevant strategy to prevent or attenuate several cardiovascular and metabolic complications. Among these functional foods, the synbiotic kefir, which is a fermented milk composed of a mixture of bacteria and yeasts, is currently the most used and has attracted the attention of health care professionals. The present review is focused on reports describing the feasibility of kefir consumption to provide benefits in cardiometabolic diseases, including hypertension, vascular endothelial dysfunction, dyslipidemia and insulin resistance. Interestingly, recent studies show that mechanisms of actions of kefir in cardiometabolic diseases include recruitment of endothelial progenitor cells, improvement of the balance vagal/sympathetic nervous system, diminution of excessive generation of reactive oxygen species, angiotensin converting enzyme inhibition, anti-inflammatory cytokines profile and alteration of the intestinal microbiota. These findings provide a better understanding about the mechanisms of the beneficial actions of kefir and motivate further investigations to determine whether the use of this synbiotic could also be translated into clinical improvements in cardiometabolic diseases

Oxidative Stress and Dementia in Alzheimer’s Patients: Effects of Synbiotic Supplementation

Alzheimer’s disease (AD) is the most common cause of dementia in elderly patients. Recently, several studies have shown that inflammation and oxidative stress precede the cardinal neuropathological manifestations of AD. In view of the proven antioxidant effects of probiotics, we proposed that continuous dietary supplementation with milk fermented with kefir grains might improve cognitive and metabolic and/or cellular disorders in the AD patients. Methods. This study was designed as an uncontrolled clinical investigation to test the effects of probiotic-fermented milk supplementation (2 mL/kg/daily) for 90 days in AD patients exhibiting cognitive deficit. Cognitive assessment, cytokine expression, systemic oxidative stress levels, and blood cell damage biomarkers were evaluated before (T0) and after (T90) kefir synbiotic supplementation. Results. When the patients were challenged to solve 8 classical tests, the majority exhibit a marked improvement in memory, visual-spatial/abstraction abilities, and executive/language functions. At the end of the treatment, the cytometric analysis showed an absolute/relative decrease in several cytokine markers of inflammation and oxidative stress markers (⋅ O2 – , H2O2, and ONOO− , ~30%) accompanied by an increase in NO bioavailability (100%). In agreement with the above findings by using the same technique, we observed in a similar magnitude an improvement of serum protein oxidation, mitochondrial dysfunction, DNA damage/repair, and apoptosis. Conclusion. In conclusion, we demonstrated that kefir improves cognitive deficits, which seems to be linked with three important factors of the AD—systemic inflammation, oxidative stress, and blood cell damage—and may be a promising adjuvant therapy against the AD progression.This study was supported by the National Council for Scientific and Technological Development (CNPq) and the State Agency for the Development of Science and Technology (FAPES) through the Edital 24/2018 -PRONEx #84321148, TO 569/2018S