Molecular Aspects of Cardiometabolic Diseases: From Etiopathogenesis to Potential Therapeutic Targets.

Cardiometabolic diseases (CMDs) encompass a range of prevalent, often preventable, non-communicable illnesses, including myocardial infarction, stroke, cardiac insufficiency, arterial hypertension, obesity, type 2 diabetes mellitus, insulin resistance, chronic renal dysfunction, non-alcoholic fatty liver disease, and rare metabolic disorders [...].

The effect of zofenopril on the cardiovascular system of spontaneously hypertensive rats treated with the ACE2 inhibitor MLN-4760.

BACKGROUND: Angiotensin converting enzyme 2 (ACE2) plays a crucial role in the infection cycle of SARS-CoV-2 responsible for formation of COVID-19 pandemic. In the cardiovascular system, the virus enters the cells by binding to the transmembrane form of ACE2 causing detrimental effects especially in individuals with developed hypertension or heart disease. Zofenopril, a H2S-releasing angiotensin-converting enzyme inhibitor (ACEI), has been shown to be effective in the treatment of patients with essential hypertension; however, in conditions of ACE2 inhibition its potential beneficial effect has not been investigated yet. Therefore, the aim of the study was to determine the effect of zofenopril on the cardiovascular system of spontaneously hypertensive rats, an animal model of human essential hypertension and heart failure, under conditions of ACE2 inhibition induced by the administration of the specific inhibitor MLN-4760 (MLN). RESULTS: Zofenopril reduced MLN-increased visceral fat to body weight ratio although no changes in systolic blood pressure were recorded. Zofenopril administration resulted in a favorable increase in left ventricle ejection fraction and improvement of diastolic function regardless of ACE2 inhibition, which was associated with increased H2S levels in plasma and heart tissue. Similarly, the acute hypotensive responses induced by acetylcholine, L-NAME (NOsynthase inhibitor) and captopril (ACEI) were comparable after zofenopril administration independently from ACE2 inhibition. Although simultaneous treatment with zofenopril and MLN led to increased thoracic aorta vasorelaxation, zofenopril increased the NO component equally regardless of MLN treatment, which was associated with increased NO-synthase activity in aorta and left ventricle. Moreover, unlike in control rats, the endogenous H2S participated in maintaining of aortic endothelial function in MLN-treated rats and the treatment with zofenopril had no impact on this effect. CONCLUSIONS: Zofenopril treatment reduced MLN-induced adiposity and improved cardiac function regardless of ACE2 inhibition. Although the concomitant MLN and zofenopril treatment increased thoracic aorta vasorelaxation capacity, zofenopril increased the participation of H2S and NO in the maintenance of endothelial function independently from ACE2 inhibition. Our results confirmed that the beneficial effects of zofenopril were not affected by ACE2 inhibition, moreover, we assume that ACE2 inhibition itself can lead to the activation of cardiovascular compensatory mechanisms associated with Mas receptor, nitrous and sulfide signaling.

Taxifolin Reduces Blood Pressure via Improvement of Vascular Function and Mitigating the Vascular Inflammatory Response in Spontaneously Hypertensive Rats.

The effect of a 10-day-long treatment with taxifolin (TAX, 20 mg/kg/day p.o.) was investigated on spontaneously hypertensive rats (SHRs) with a focus on the vascular functions of isolated femoral arteries and thoracic aortas. TAX reduced blood pressure in SHRs. In femoral arteries, TAX increased acetylcholine-induced relaxation, reduced the maximal NA-induced contraction, and reduced acetylcholine-induced endothelium-dependent contraction (EDC); however, TAX had no effect on the vascular reactivity of isolated thoracic aortas. In addition, TAX elevated the total nitric oxide synthase (NOS) activity and iNOS protein expression but reduced cyclooxygenase-2 (COX2) protein expression in the tissue of the abdominal aorta without changes in Nos2 and Ptgs2 gene expressions. TAX also increased the gene expression of the anti-inflammatory interleukin-10 (Il10). In addition, in vitro studies showed that TAX has both electron donor and H atom donor properties. However, TAX failed to reduce superoxide production in the tissue of the abdominal aorta after oral administration. In conclusion, our results show that a decrease in the blood pressure in TAX-treated SHRs might be attributed to improved endothelium-dependent relaxation and reduced endothelium-dependent contraction. In addition, the results suggest that the effect of TAX on blood pressure regulation also involves the attenuation of COX2-mediated pro-inflammation and elevation of anti-inflammatory pathways.

Hypertension and Cardiovascular Diseases: From Etiopathogenesis to Potential Therapeutic Targets.

Cardiovascular diseases (CVDs) are the top cause of death worldwide, and arterial hypertension per se remains the major preventable cause of CVDs [...].

The Effect of a Single Dose of Dark Chocolate on Cardiovascular Parameters and Their Reactivity to Mental Stress.

Objective: This study investigated the effect of a single administration of dark or milk chocolate on blood pressure (BP), heart rate (HR), and double product (DP) in young healthy women at rest and during acute mental stress.Method: Measurements consisted of anthropometry, BP, and HR. Mean arterial BP (MAP) and DP were computed. The relative reactivity of individual variables was quantified as to their percentage change during the rest or test of mental arithmetic (MA) with respect to the respective baseline value. All subjects underwent two tests of MA-one before chocolate administration and the second one 2 hours after chocolate (1 mg/g of body weight) ingestion.Results: Two hours after ingestion at rest, dark chocolate administration resulted in a significant increase in relative values of systolic BP and DP by 5.1% ± 1.4% and 13.7% ± 3.2%, respectively, compared to the responses in the milk chocolate group (-2.4% ± 1.6% and 0.6% ± 3.4%, respectively, p < 0.04 for both comparisons) without changes in diastolic BP, HR, and MAP. During MA-induced acute stress, the relative magnitude of the reactivity of diastolic BP, HR, MAP, and DP decreased by about 10, 16, 8, and 23 percentage points, respectively, 2 hours after ingestion of dark chocolate compared to the relative reactivity determined before dark chocolate ingestion. Milk chocolate failed to affect any of the above-mentioned parameters at rest or during stress.Conclusions: The single oral intake of 85% dark chocolate increased relative values of systolic BP and DP at rest but buffered the reactivity of diastolic BP, HR, MAP, and DP during mental stress, which was not found after ingestion of milk chocolate. Thus, dark chocolate might have a beneficial effect during acute stress due to its ability to buffer cardiovascular reactivity in young healthy women.

The Molecular Mechanisms of Iron Metabolism and Its Role in Cardiac Dysfunction and Cardioprotection.

Iron is an essential mineral participating in different functions of the organism under physiological conditions. Numerous biological processes, such as oxygen and lipid metabolism, protein production, cellular respiration, and DNA synthesis, require the presence of iron, and mitochondria play an important role in the processes of iron metabolism. In addition to its physiological role, iron may be also involved in the adaptive processes of myocardial "conditioning". On the other hand, disorders of iron metabolism are involved in the pathological mechanisms of the most common human diseases and include a wide range of them, such as type 2 diabetes, obesity, and non-alcoholic fatty liver disease, and accelerate the development of atherosclerosis. Furthermore, iron also exerts potentially deleterious effects that may be manifested under conditions of ischemia/reperfusion (I/R) injury, myocardial infarction, heart failure, coronary artery angioplasty, or heart transplantation, due to its involvement in reactive oxygen species (ROS) production. Moreover, iron has been recently described to participate in the mechanisms of iron-dependent cell death defined as "ferroptosis". Ferroptosis is a form of regulated cell death that is distinct from apoptosis, necroptosis, and other types of cell death. Ferroptosis has been shown to be associated with I/R injury and several other cardiac diseases as a significant form of cell death in cardiomyocytes. In this review, we will discuss the role of iron in cardiovascular diseases, especially in myocardial I/R injury, and protective mechanisms stimulated by different forms of "conditioning" with a special emphasis on the novel targets for cardioprotection.

Consequences of lipopolysaccharide and n-3 polyunsaturated fatty acid administration on aortic function of spontaneously hypertensive rats.

The aim of the work was to study the delayed effect of lipopolysaccharide (LPS) administration on endothelial function of the aorta of rats with genetic hypertension. Further, the possibility to ameliorate LPS-induced changes by n-3 polyunsaturated fatty acids (n-3 PUFA) was tested. Rats received a bolus of 1 mg/kg LPS i.p.; n-3 PUFA were administered in the dose of 30 mg/kg daily for 10 days p.o.. Ten days after receiving of LPS, the body weight gain of rats was statistically lower compared to control rats (p < 0.05). n-3 PUFA administration to LPS rats had no effect on this parameter. The TBARS and NAGA concentrations in plasma were significantly increased in the LPS group (p < 0.05) and n-3 PUFA administration returned them to control values. In functional studies, phenylephrine (PE, 1 µmol/l) evoked contraction of aortas which was not statistically different among experimental groups. However, endothelium-dependent relaxation was depressed in the LPS group (p < 0.05) and n-3 PUFA slightly recovered it to control values. In conclusion, oxidative stress seems to be responsible for aortic endothelial dysfunction detected 10 days after administration of LPS to rats. n-3 PUFA slightly improved the function of the endothelium injured by LPS, probably thanks to their antioxidant properties. Prolonged administration of higher doses of n-3 PUFA should defend the vascular endothelium against detrimental effect of bacterial inflammation.

Effects of Red Palm Oil on Myocardial Antioxidant Enzymes, Nitric Oxide Synthase and Heart Function in Spontaneously Hypertensive Rats.

The purpose of this study was to investigate the effect of antioxidants rich red palm oil (RPO) supplementation on cardiac oxidative stress known as crucial factor deteriorating heart function in hypertension. 3-month-old, male spontaneously hypertensive rats (SHR) and normotensive Wistar Kyoto rats (WKY) were fed standard rat chow without or with RPO (0.2 mL/day/5 weeks). General characteristic of rats were registered. Left ventricular tissue (LV) was used to determine expression of superoxide dismutases (SOD1, SOD2) and glutathione peroxidases (Gpx) as well as activity of nitric oxide synthase (NOS). Functional parameters of the heart were examined during basal conditions and at the early-phase of post-ischemic reperfusion using Langendorff-perfused system. RPO intake significantly reduced elevated blood pressure and total NOS activity as well as increased lowered expression of mitochondrial SOD2 in SHR hearts during basal condition. Moreover, RPO supplementation resulted in suppression of elevated heart rate, increase of reduced coronary flow and enhancement of systolic and diastolic heart function at the early-phase of post-ischemic reperfusion. It is concluded that SHR benefit from RPO intake due to decrease of blood pressure, amelioration of oxidative stress and protection of heart function that was deteriorated by post-ischemic reperfusion.

Effect of crowding stress on tolerance to ischemia-reperfusion injury in young male and female hypertensive rats: molecular mechanisms.

Sex and social stress may represent risk factors in the etiology of hypertension and heart response to ischemia-reperfusion (I/R) injury. Phosphatidylinositol 3-kinase/protein kinase B (Akt) plays an important role in the processes associated with hypertension and myocardial tolerance to I/R, and may be involved in myocardial stress reaction. The impact of chronic stress on the response to I/R was investigated in the hearts of 7-week-old spontaneously hypertensive (SHR) and Wistar-Kyoto (WKY) rats of both sexes. Stress was induced by reducing living space to 70 cm(2)/100 g body mass of rat for 2 weeks, while the controls were kept at 200 cm(2)/100 g. Langendorff-perfused hearts, subjected to I/R, exhibited higher vulnerability to ventricular tachycardia in crowd-stressed SHR vs. the control rats, and this was more pronounced in the males. Myocardial infarction was not affected by crowding stress in any of the groups. Male and female SHR showed increased activation of cardiac Akt, whereas nitric oxide synthase activity (NOS) with pro-apoptotic signaling decreased in the males but was not altered in the females (vs. WKY rats). NOS was enhanced in the female SHR and WKY groups by comparison with the respective males. Stress only reduced NOS activity in the SHR groups, and without changes in apoptotic markers. In conclusion, we showed that stress in young SHR mainly affects the nonlethal markers for I/R, and has no impact on myocardial infarction and apoptosis, despite reduced NOS activity.

Pleiotropic preconditioning-like cardioprotective effects of hypolipidemic drugs in acute ischemia-reperfusion in normal and hypertensive rats.

Although pleiotropy, which is defined as multiple effects derived from a single gene, was recognized many years ago, and considerable progress has since been achieved in this field, it is not very clear how much this feature of a drug is clinically relevant. During the last decade, beneficial pleiotropic effects from hypolipidemic drugs (as in, effects that are different from the primary ones) have been associated with reduction of cardiovascular risk. As with statins, the agonists of peroxisome proliferator-activated receptors (PPARs), niacin and fibrates, have been suggested to exhibit pleiotropic activity that could significantly modify the outcome of a cardiovascular ailment. This review examines findings demonstrating the impacts of treatment with hypolipidemic drugs on cardiac response to ischemia in a setting of acute ischemia-reperfusion, in relation to PPAR activation. Specifically, it addresses the issue of susceptibility to ischemia, with particular regard to the preconditioning-like cardioprotection conferred by hypolipidemic drugs, as well as the potential molecular mechanisms behind this cardioprotection. Finally, the involvement of PPAR activation in the mechanisms of non-metabolic cardioprotective effects from hypolipidemic drugs, and their effects on normal and pathologically altered myocardium (in the hearts of hypertensive rats) is also discussed.

Intake of Dark and Milk Chocolate Differently Affects Autonomic Nervous Control of the Heart During Rest and Mental Stress.

OBJECTIVE: Chocolate is a popular food that may affect the activity of the autonomic nervous system (ANS). The aim of this study was to determine the effect of a single dose of dark or milk chocolate on ANS cardiac control during rest and mental stress induced by the Stroop test (ST). METHODS: Healthy participants, divided into DARK or MILK chocolate groups, ingested corresponding type of chocolate (1 g/kg body weight). They underwent measurement of ANS during relaxation and ST before and 2 h after chocolate consumption. ANS control was assessed by determination of heart rate (HR) and heart rate variability using parameters related to complex autonomic modulation (TP, SDNN) or primary vagal modulation (HFnu, RMSSD). RESULTS: HR was always increased during ST in both groups. Relaxation HR values after chocolate ingestion were higher only in the DARK chocolate group. During ST, values of TP, SDNN and HFnu decreased before and after chocolate ingestion in the DARK group, but only before chocolate ingestion in the MILK group. RMSSD values decreased during ST before and after chocolate ingestion in both groups. Relaxation TP, RMSSD and HFnu values after chocolate ingestion were lower in the DARK but not in the MILK group. CONCLUSION: The results suggest that even a single dose of milk chocolate attenuates changes in ANS cardiac control induced by mental stress, whereas a single dose of dark chocolate has an activating effect on the heart via modification of ANS cardiac control at rest. Different levels of sugars and cocoa biologically active compounds in the two types of chocolate could explain the observed effects.

(-)-Epicatechin reduces blood pressure and improves vasorelaxation in spontaneously hypertensive rats by NO-mediated mechanism.

Studies in humans have found consumption of certain flavanoid-containing foods to be associated with improvement in endothelial function and with reduction of blood pressure (BP). (-)-Epicatechin is a compound representative of the flavanols (a subfamily of flavonoids), abundant in cocoa seeds, which is preserved during the industrialization process to chocolate. The antihypertensive effect of dietary (-)-epicatechin was investigated on spontaneously hypertensive rats (SHRs). Consumption of (-)-epicatechin-supplemented diet (3 g (-)-epicatechin/kg diet) decreased BP in SHR by 27 and 23 mm Hg on days 2 and 6, respectively. On day 6, a 173% increase of nitric oxide synthase (NOS) activity was observed in the aorta of EPI-SHR as compared to nonsupplemented SHR (P < 0.05). Responses to acetylcholine (ACh) were then examined in femoral arteries in the absence and the presence of L-NAME, a nonselective NOS inhibitor, to assess the ACh-mediated relaxation ascribed to NO-dependent and -independent mechanisms. Acetylcholine-induced endothelium-dependent relaxation in the femoral artery was significantly higher in EPI-SHR than in SHR, with a predominance of the NO-dependent component of this relaxation. The endothelium-independent relaxation, assayed by using the NO donor sodium nitroprusside, resulted in nonsignificant difference in the three experimental groups, demonstrating an unaffected function of vascular smooth muscle cells. These results give further support to the concept that (-)-epicatechin can modulate BP in hypertension by increasing NO levels in the vasculature.

Long-term social stress induces nitric oxide-independent endothelial dysfunction in normotensive rats.

As chronic stress is a significant risk factor for several cardiovascular disorders, this study investigated the hypothesis that long-term stress produced by crowding may lead to alterations in nitric oxide (NO) production and NO-dependent relaxation in the course of stress, resulting in endothelial dysfunction and hypertension in Wistar-Kyoto (WKY) rats. For this purpose, male WKY rats were divided into control (480 cm2/rat, four rats/cage, n = 8) and crowded (200 cm2/rat, five rats/cage, n = 10) groups for 8 or 12 weeks. Vasorelaxation was evaluated in vitro as a response to acetylcholine (ACh) of femoral arteries pre-contracted by serotonin, before and after NO synthase inhibition (N (G)-nitro-l-arginine methyl ester, 300 µmol/l). Crowding increased plasma corticosterone concentration but failed to affect blood pressure (determined by tail-cuff plethysmography) of rats. NO production was unchanged in the hypothalamus and left ventricle of both stressed groups; however it was significantly elevated in the aorta. Maximal ACh-induced relaxation was elevated significantly after 8-week stress, but reduced after 12 weeks. Stress elevated the NO-dependent component and reduced the NO-independent component of ACh-induced relaxation in both crowded groups. However, a reduction in the NO-independent component was more pronounced after 12-week versus 8-week stress. In conclusion, elevated endothelium-dependent relaxation was observed after 8-week stress, while the extension of stress exposure resulted in a reduction in arterial relaxation associated with a more pronounced decrease of its NO-independent component. Thus, elevation of the NO-dependent component of relaxation can be considered as an adaptation mechanism, and impairment of NO-independent relaxation might be the initial step in chronic stress-induced cardiovascular disorders.

Impact of age and sex on response to ischemic preconditioning in the rat heart: differential role of the PI3K-AKT pathway.

Sex and aging represent important factors that determine morbidity and mortality due to cardiovascular diseases in the human population. This study aimed to investigate the impact of aging on the response to ischemia-reperfusion in male and female rat hearts, and to explore a potential role of the PI3K-Akt pathway in the cardioprotective effects of ischemic preconditioning (IPC) in the myocardium of younger and older adult males and females. Langendorff-perfused nonpreconditioned and preconditioned hearts of 12- and 18-week-old male and female Wistar rats were subjected to regional ischemia and reperfusion with or without prior perfusion with the PI3K inhibitor wortmannin for the evaluation of ischemia-induced arrhythmias and the size of myocardial infarction (infarct size; IS). Aging did not modify IS in both sexes; however, it markedly increased susceptibility to arrhythmias. Although IPC effectively reduced IS in males and females of both ages, only the hearts of males and 18-week-old females benefited from its antiarrhythmic effect. In the preconditioned 12-week-old females, but not the 18-week-old females, and in males of both ages, wortmannin blunted the anti-infarct effect of IPC. In conclusion, activation of the PI3K-Akt pathway plays an important role in protection against lethal injury conferred by IPC in males irrespective of age. The IS-limiting effect of IPC appears to be PI3K-Akt-dependent only in the 12-week-old females.

A Single Infusion of Polyethylene Glycol-Coated Superparamagnetic Magnetite Nanoparticles Alters Differently the Expressions of Genes Involved in Iron Metabolism in the Liver and Heart of Rats.

This study investigated genotype- and tissue-related differences in the biodistribution of superparamagnetic magnetite (Fe3O4) nanoparticles (IONs) into the heart and liver of normotensive Wistar Kyoto (WKY) and spontaneously hypertensive (SHR) rats after a single i.v. infusion of polyethylene glycol-coated IONs (~30 nm, 1mg Fe/kg) 100 min post-infusion. The effects of IONs on the expression of selected genes involved in the regulation of iron metabolism, including Nos, Sod and Gpx4, and their possible regulation by nuclear factor (erythroid-derived 2)-like 2 (NRF2, encoded by Nfe2l2) and iron-regulatory protein (encoded by Irp1) were investigated. In addition, superoxide and nitric oxide (NO) production were determined. Results showed reduced ION incorporations into tissues of SHR compared to WKY and in the hearts compared to the livers. IONs reduced plasma corticosterone levels and NO production in the livers of SHR. Elevated superoxide production was found only in ION-treated WKY. Results also showed differences in the regulation of iron metabolism on the gene level in the heart and liver. In the hearts, gene expressions of Nos2, Nos3, Sod1, Sod2, Fpn, Tf, Dmt1 and Fth1 correlated with Irp1 but not with Nfe2l2, suggesting that their expression is regulated by mainly iron content. In the livers, expressions of Nos2, Nos3, Sod2, Gpx4, and Dmt1 correlated with Nfe2l2 but not with Irp1, suggesting a predominant effect of oxidative stress and/or NO.

Reactivity of the mesenteric bed arteries of normotensive rats exposed to chronic social stress.

The aim of this study was to investigate the effects of chronic social stress on endothelium-dependent relaxation in the superior mesenteric artery (SMA) and its first branches (1MA) as well as on neurogenic contractions of SMA in adult, male Wistar-Kyoto (WKY) rats. Mesenteric arteries were isolated from control (living space: 480 cm(2)/rat) or stressed rats exposed to 8-week-lasting crowding stress (living space: 200 cm(2)/rat). Blood pressure (BP) and heart rate, determined by tail-cuff plethysmography, were not affected by crowding. Stress increased neurogenic contractions of SMA elicited by electrical stimulation of perivascular nerves and significantly elevated vasoconstriction induced by exogenous noradrenaline in SMA, without modulation of its endothelial function. In 1MA, nitric oxide (NO)-dependent component of endothelium-dependent relaxation to acetylcholine was investigated. In 1MA, stress failed to affect noradrenaline- and phenylephrine-induced vasoconstriction, total acetylcholine-induced relaxation as well as its NO-dependent and NO-independent components. Moreover, endothelium-independent sodium nitroprusside-induced relaxations of 1MA from the stressed rats did not differ from those of controls. In conclusion, chronic stress produced by crowding failed to induce an increase of BP, presumably because endothelial function of SMA and vascular function of small mesenteric arteries, which are rather important in BP regulation, remained preserved.

Preliminary Findings on the Effect of Ultrasmall Superparamagnetic Iron Oxide Nanoparticles and Acute Stress on Selected Markers of Oxidative Stress in Normotensive and Hypertensive Rats.

Several studies have reported that the administration of various nanoparticles in vivo can cause oxidative stress. The combination of ultrasmall superparamagnetic iron oxide nanoparticles (USPIONs) and acute stress was selected because, during intravenous application of a contrast agent, patients are exposed to psycho-emotional stress. This study was designed to investigate the effect of acute stress and USPIONs on selected markers of oxidative stress (antioxidant capacity, superoxide dismutase, glutathione peroxidase and catalase activities, levels of advanced oxidation protein products, protein carbonyls, lipoperoxides and 8-isoprostanes) in plasma and erythrocytes in normotensive Wistar-Kyoto rats (WKY) and spontaneously hypertensive rats (SHR). In the WKY and SHR groups, there was a significant main effect of genotype between groups on studied markers except protein carbonyls and lipoperoxides. In SHR, the combination of acute stress and USPIONs increased the antioxidant capacity of plasma and the selected enzyme activities of erythrocytes. In WKY, the combination of acute stress and USPIONs decreased the antioxidant capacity of erythrocytes and reduced levels of advanced oxidation protein products in plasma. Our study points to the fact that, when hypertensive subjects are treated with iron oxide nanoparticles, caution should be taken, especially in stress conditions, since they seem to be more vulnerable to oxidative stress produced by USPIONs.

Age- and Hypertension-Related Changes in NOS/NO/sGC-Derived Vasoactive Control of Rat Thoracic Aortae.

This study was aimed at examining the role of the NOS/NO/sGC signaling pathway in the vasoactive control of the thoracic aorta (TA) from the early to late ontogenetic stages (7 weeks, 20 weeks, and 52 weeks old) of normotensive Wistar-Kyoto (WKY) rats and spontaneously hypertensive rats (SHRs). Systolic blood pressure (SBP) and heart rate (HR) were significantly increased in SHRs compared to age-matched WKYs, which was associated with left heart ventricle hypertrophy in all age groups of rats. The plasma urea level was increased in 20-week-old and 52-week-old SHRs compared with WKYs without increasing creatinine and uric acid. The total cholesterol levels were lower in 20-week-old and 52-week-old SHRs than in WKYs, but triglycerides were higher in 7-week-old SHRs. The fructosamine level was increased in 52-week-old SHRs compared with age-matched WKYs and unchanged in other age groups. Superoxide production was increased only in 7-week-old SHRs compared to age-matched WKYs. The endothelium-dependent relaxation (EDR) of the TA deteriorated in both rat strains during aging; however, endothelial dysfunction already occurred in 20-week-old SHRs and was even more enhanced in 52-week-old rats. Our results also demonstrated increased activity of NOS in 52-week-old WKYs. Moreover, 7-week-old and 52-week-old WKY rats displayed an enhanced residual EDR after L-NMMA (NOS inhibitor) incubation compared with 20-week-old rats. Our results showed that in 7-week-old SHRs, the residual EDR after L-NMMA incubation was increased compared to that in other age groups. The activity of NOS in the TA was comparable in 7-week-old and 20-week-old SHRs, but it was reduced in 52-week-old SHRs compared to younger SHRs and 52-week-old WKYs. Thus, it seems that, in contrast to SHRs, the NOS/NO system in WKYs is probably able to respond to age-related pathologies to maintain endothelial functions and thus optimal BP levels even in later periods of life.

MLN-4760 Induces Oxidative Stress without Blood Pressure and Behavioural Alterations in SHRs: Roles of Nfe2l2 Gene, Nitric Oxide and Hydrogen Sulfide.

Reduced angiotensin 1-7 bioavailability due to inhibition of angiotensin-converting enzyme 2 (ACE2) may contribute to increased mortality in hypertensive individuals during COVID-19. However, effects of ACE2 inhibitor MLN-4760 in brain functions remain unknown. We investigated the selected behavioural and hemodynamic parameters in spontaneously hypertensive rats (SHRs) after a 2-week s.c. infusion of MLN-4760 (dose 1 mg/kg/day). The biochemical and molecular effects of MLN-4760 were investigated in the brainstem and blood plasma. MLN-4760 had no effects on hemodynamic and behavioural parameters. However, MLN-4760 increased plasma hydrogen sulfide (H2S) level and total nitric oxide (NO) synthase activity and conjugated dienes in the brainstem. Increased NO synthase activity correlated positively with gene expression of Nos3 while plasma H2S levels correlated positively with gene expressions of H2S-producing enzymes Mpst, Cth and Cbs. MLN-4760 administration increased gene expression of Ace2, Sod1, Sod2, Gpx4 and Hmox1, which positively correlated with expression of Nfe2l2 gene encoding the redox-sensitive transcription factor NRF2. Collectively, MLN-4760 did not exacerbate pre-existing hypertension and behavioural hyperactivity/anxiety in SHRs. However, MLN-4760-induced oxidative damage in brainstem was associated with activation of NO- and H2S-mediated compensatory mechanisms and with increased gene expression of antioxidant, NO- and H2S-producing enzymes that all correlated positively with elevated Nfe2l2 expression.

Effects of Taxifolin in Spontaneously Hypertensive Rats with a Focus on Erythrocyte Quality.

Oxidative stress and multiple erythrocyte abnormalities have been observed in hypertension. We focused on the effects of angiotensin-converting enzyme 2 (ACE2) inhibition by MLN-4760 inhibitor on angiotensin peptides, oxidative stress parameters, and selected erythrocyte quality markers in spontaneously hypertensive rats (SHR). We also investigated the potential effects of polyphenolic antioxidant taxifolin when applied in vivo and in vitro following its incubation with erythrocytes. SHRs were divided into four groups: control, taxifolin-treated, MLN-4760-treated, and MLN-4760 with taxifolin. MLN-4760 administration increased the blood pressure rise independent of taxifolin treatment, whereas taxifolin decreased it in control SHRs. Body weight gain was also higher in ACE2-inhibited animals and normalized after taxifolin treatment. However, taxifolin did not induce any change in angiotensin peptide concentrations nor a clear antioxidant effect. We documented an increase in Na,K-ATPase enzyme activity in erythrocyte membranes of ACE2-inhibited SHRs after taxifolin treatment. In conclusion, ACE2 inhibition deteriorated some selected RBC properties in SHRs. Although taxifolin treatment did not improve oxidative stress markers, our data confirmed the blood pressure-lowering potential, anti-obesogenic effect, and some "erythroprotective" effects of this compound in both control and ACE2-inhibited SHRs. In vitro investigations documenting different effects of taxifolin on erythrocyte properties from control and ACE2-inhibited SHRs accentuated the irreplaceability of in vivo studies.