Low-dose angiotensin II supplementation restores flow-induced dilation mechanisms in cerebral arteries of Sprague-Dawley rats on a high salt diet.

OBJECTIVE: Salt-induced suppression of angiotensin II contributes to impaired endothelium-dependent vascular reactivity. The present study investigated the effect of chronic low-dose angiotensin II (ANG II) supplementation on the mechanisms of flow-induced dilation (FID) and oxidative stress at the cellular and molecular level in middle cerebral arteries (MCA) of male Sprague-Dawley rats fed high salt diet. METHODS: Rats (10 weeks old) were randomly assigned to a low salt diet group (0.4% NaCl in rat chow); high salt diet group (7 days 4% NaCl in rat chow) or HS+ANG II group [7 days high salt diet with 3 days ANG II administration via osmotic minipumps (100 ng/kg per min on days 4-7)]. FID was determined in absence/presence of the NOS inhibitor L-NAME, the non-selective cyclooxygenase (COX-1,2) inhibitor indomethacin, a selective inhibitor of CYP450 epoxygenase activity (MS-PPOH) and the superoxide dismutase mimetic TEMPOL. Gene expression of antioxidative enzymes, and of genes and proteins involved in FID mechanisms were determined by RT-qPCR and western blot. Vascular nitric oxide and superoxide/reactive oxygen species levels were assessed by direct fluorescence. Serum systemic oxidative stress parameters were measured by spectrophotometry. RESULTS: Chronic low-dose ANG II supplementation in high salt fed rats restored FID of MCAs, which was nitric oxide, prostanoid and epoxyeicosatrienoic acid dependent. ANG II changed the protein/gene expression of COXs, HIF-1α and VEGF and significantly increased GPx4 and EC-SOD antioxidative enzyme expression, decreased systemic oxidative stress, decreased superoxide/ROS levels and increased nitric oxide bioavailability in the vascular wall. CONCLUSION: Physiological levels of circulating ANG II are crucial to maintain the HIF-1α dependent mechanisms of FID and vascular oxidative balance without affecting mean arterial pressure.

Arachidonic Acid Metabolites of CYP450 Enzymes and HIF-1α Modulate Endothelium-Dependent Vasorelaxation in Sprague-Dawley Rats under Acute and Intermittent Hyperbaric Oxygenation.

Acetylcholine-induced vasorelaxation (AChIR) and responses to reduced pO2 (hypoxia-induced relaxation (HIR), 0% O2) were assessed in vitro in aortic rings of healthy male Sprague-Dawley rats (N = 252) under hyperbaric (HBO2) protocols. The studied groups consisted of the CTRL group (untreated); the A-HBO2 group (single HBO2; 120 min of 100% O2 at 2.0 bars); the 24H-HBO2 group (examined 24 h after single exposure) and the 4D-HBO2 group (four consecutive days of single HBO2). AChIR, sensitivity to ACh and iNOS expression were decreased in the A-HBO2 group. HIR was prostanoid- and epoxyeicosatrienoic acid (EET)-mediated. HIF-1α expression was increased in the 24H-HBO2 and 4D-HBO2 groups. LW6 (HIF-1α inhibitor) decreased HIR in the 24H-HBO2 group. HBO2 affected the expression of COX-1 and COX-2. CYP2c11 expression was elevated in the 24H-HBO2 and 4D-HBO2 groups. Concentrations of arachidonic acid (AA) metabolites 14(15)-DiHET, 11(12)-DiHET and 8(9)-DiHET were increased in A-HBO2 and 24H-HBO2. An increased concentration of 8(9)-EET was observed in the A-HBO2 and 24h-HBO2 groups vs. the CTRL and 4D-HBO2 groups, and an increased concentration of 5(6)-DiHET was observed in the 24H-HBO2 group vs. the 4D-HBO2 group. The 20-HETE concentration was increased in the A-HBO2 group. All were determined by LC-MS/MS of the aorta. The results show that AChIR in all groups is mostly NO-dependent. HIR is undoubtedly mediated by the CYP450 enzymes' metabolites of AA, whereas HIF-1α contributes to restored HIR. Vasoconstrictor metabolites of CYP450 enzymes contribute to attenuated AChIR and HIR in A-HBO2.

The Physiological Effect of n-3 Polyunsaturated Fatty Acids (n-3 PUFAs) Intake and Exercise on Hemorheology, Microvascular Function, and Physical Performance in Health and Cardiovascular Diseases; Is There an Interaction of Exercise and Dietary n-3 PUFA Intake?

Physical activity has a beneficial effect on systemic hemodynamics, physical strength, and cardiac function in cardiovascular (CV) patients. Potential beneficial effects of dietary intake of n-3 polyunsaturated fatty acids (n-3 PUFAs), such as α-linolenic acid, eicosapentaenoic acid, and docosahexaenoic acid on hemorheology, vascular function, inflammation and potential to improve physical performance as well as other CV parameters are currently investigated. Recent meta-analysis suggests no effect of n-3 PUFA supplementation on CV function and outcomes of CV diseases. On the other hand, some studies support beneficial effects of n-3 PUFAs dietary intake on CV and muscular system, as well as on immune responses in healthy and in CV patients. Furthermore, the interaction of exercise and dietary n-3 PUFA intake is understudied. Supplementation of n-3 PUFAs has been shown to have antithrombotic effects (by decreasing blood viscosity, decreasing coagulation factor and PAI-1 levels and platelet aggregation/reactivity, enhancing fibrinolysis, but without effects on erythrocyte deformability). They decrease inflammation by decreasing IL-6, MCP-1, TNFα and hsCRP levels, expression of endothelial cell adhesion molecules and significantly affect blood composition of fatty acids. Treatment with n-3 PUFAs enhances brachial artery blood flow and conductance during exercise and enhances microvascular post-occlusive hyperemic response in healthy humans, however, the effects are unknown in cardiovascular patients. Supplementation of n-3 PUFAs may improve anaerobic endurance and may modulate oxygen consumption during intense exercise, may increase metabolic capacity, enhance endurance capacity delaying the onset of fatigue, and improving muscle hypertrophy and neuromuscular function in humans and animal models. In addition, n-3 PUFAs have anti-inflammatory and anti-nociceptive effects and may attenuate delayed-onset muscle soreness and muscle stiffness, and preserve joint mobility. On the other hand, effects of n-3 PUFAs were variably observed in men and women and they vary depending on dietary protocol, type of supplementation and type of sports activity undertaken, both in healthy and cardiovascular patients. In this review we will discuss the physiological effects of n-3 PUFA intake and exercise on hemorheology, microvascular function, immunomodulation and inflammation and physical performance in healthy persons and in cardiovascular diseases; elucidating if there is an interaction of exercise and diet.

Hyperbaric oxygenation affects the mechanisms of acetylcholine-induced relaxation in diabetic rats.

The effects of hyperbaric oxygenation (HBO2) on acetylcholine-induced vasorelaxation (AChIR) were evaluated in male Sprague-Dawley (SD) rats randomized into four groups: healthy controls (Ctrl), diabetic rats (DM), and control and diabetic rats that underwent hyperbaric oxygenation (Ctrl+HBO2 and DM+HBO2). AChIR was measured in aortic rings, with L-NAME, indomethacin, or MS-PPOH and a combination of inhibitors. mRNA expression of eNOS, iNOS, COX-1 and COX-2 was assessed by qPCR, and protein expression of CYP4A(1-3) by Western blot. Plasma antioxidative capacity and systemic oxidative stress were determined with the ferric reducing ability of plasma (FRAP) and thiobarbituric acid-reactive substances (TBARS) assays, respectively. AChIR was preserved in all groups of rats, but mediated with different mechanisms. In all experimental groups of rats, AChIR was mediated mainly by NO, with the contribution of CYP450 vasodilator metabolites. This effect was the most prominent in the DM+HBO2 group of rats. The TBARS was significantly higher in both DM and DM+HBO2 groups compared to respective controls. eNOS expression was upregulated in the DM+HBO2 group compared to other groups, COX-1 expression was upregulated in the DM+HBO2 group compared to the control. CYP450-4A1 / A2/A3protein expression was significantly higher expressed in both hyperbaric groups compared to their respective controls. In conclusion, HBO2 affected all three vasodilator pathways and shifted AChIR to CYP450 enzymes pathway.

Hyperbaric oxygenation modulates vascular reactivity to angiotensin-(1-7) in diabetic rats: potential role of epoxyeicosatrienoic acids.

Previously, a facilitating effect of hyperbaric oxygenation (HBO2) on aortic ring responses to angiotensin-(1-7) in healthy rats was reported, with epoxyeicosatrienoic acids (EETs) possibly playing an important role. The aim of this study was to assess whether HBO2 exerts similar effects in diabetic rats and to further explore the role of specific cytochrome P450 (CYP) enzymes in changes induced by HBO2. Aortic relaxation to angiotensin-(1-7) was significantly higher in HBO2 diabetic rats compared to control diabetic rats, while HBO2 had no effect on angiotensin II contraction. N-methylsulphonyl-6-(2-propargyloxyphenyl/hexanamide inhibited the facilitation of angiotensin-(1-7) responses in HBO2 rats, suggesting an important role of EETs in this modulation. mRNA expression of CYP2J3 and protein expression of CYP2C11 were significantly upregulated in HBO2 diabetic rats, whereas CYP4A1, CYP4A2 and CYP4A3 mRNA and CYP2J3 protein expression was similar between groups. Mean arterial pressure, ferric reducing ability of plasma and Thiobarbituric Acid Reactive Substances levels and serum angiotensin-(1-7) concentrations were not significantly changed.

Restoring vascular function with hyperbaric oxygen treatment: recovery mechanisms.

Treatment with hyperbaric oxygen can be a beneficial adjuvant therapy in various disorders characterized by compromised tissue oxygenation and perfusion. However, the effects of hyperbaric oxygenation cannot be simply explained as a compensation of the oxygen deficit. Hyperbaric oxigenation has a much broader influence and has the ability to alter protein expression, modulate signaling pathways and affect vascular structure and function. We discuss some of the most important uses of hyperbaric oxigenation for clinical conditions that involve abnormal vascular function. We present recent studies and insights into the mechanisms and effects of hyperbaric oxygen in the vasculature.

Blood pressure, acid-base and blood gas status and indicators of oxidative stress in healthy male rats exposed to acute hyperbaric oxygenation.

Different protocols of hyperbaric oxygenation (HBO2) are used for research purposes; however, data on the changes in blood pressure, oxidative stress and acid-base and gas status induced by various oxygenation protocols are scarce and conflicting. The aim of this study was to examine the effects of an acute session of HBO2 [2 bar (200 kPa) for two hours] on arterial systolic and diastolic blood pressure, arterial blood gases and acid-base status, and oxidative stress in rats. Sprague-Dawley rats (12-15 weeks) were examined prior to, immediately and 24 hours after a two-hour HBO2 exposure at 2 bars. The femoral artery was cannulated to determine blood pressure, and blood samples were collected to measure blood gases and acid-base status, Ferric reducing antioxidant power ability of plasma (FRAP) and thiobarbituric acid reactive substances (TBARS). Immediately after HBO2 systolic and diastolic blood pressure significantly decreased (from 138 +/- 14/103 +/- 13 to 113 +/- 12/72 +/- 16 mmHg). However, these values were still inside the normal physiological range. pH decreased (from 7.34 +/- 0.05 to 7.28 +/- 0.05), pCO2 decreased (from 7.07 +/- 0.89 to 5.76 +/- 0.50 kPa), pO2 increased (from 12.48 +/- 0.88 to 13.68 +/- 2.4 kPa), plasma bicarbonate decreased (from 27.04 +/- 3.25 to 20.52 +/- 3.02 mmol/L). Exposure to HBO2 immediately increased TBARS levels (from 0.17 +/- 0.09 to 21.79 +/- 1.05 microM/MDA), while FRAP levels were not significantly changed. Measurements on separate animals 24 hours after a single HBO2 exposure showed no differences in comparison to control animals, except for pO2, which was significantly lower (11.10 +/- 0.31 kPa). The results define values of important parameters, serving as a necessary basis for complex analysis of HBO2 effects in research on rat animal models.

Effects of hyperbaric oxygenation on vascular reactivity to angiotensin II and angiotensin-(1-7) in rats.

OBJECTIVE: To assess and elucidate the mechanisms of hyperbaric oxygenation (HBO2) effects on vascular reactivity to angiotensin-(1-7) [ANG-(1-7)] and angiotensin II (ANG II). METHODS: Rat aortic rings (HBO2 vs. control group) were used to test responses to ANG II, ANG II+ ANG-(1-7) or ANG-(1-7) after noradrenaline precontraction in the presence/absence of MS-PPOH, a specific CYP 450-epoxygenase inhibitor, and glibenclamide, a KATP channels inhibitor. mRNA expression studies of specific CYP isozymes have been conducted as well. RESULTS: The mean contraction (expressed as percent of maximal contraction) for ANG II was similar between groups. Contraction for ANG II + ANG-(1-7) was 15% +/- 10 (HBO2) and 20% +/- 9 (control). There was a significant decrease between the contraction response to ANG II (HBO2) and the response to ANG II + ANG-(1-7) in the HBO2 group, without such a difference within the control group. Mean percentage of noradrenaline precontraction decrease after ANG-(1-7) addition was significantly different [10% +/- 9 (control) and 19% +/- 11 (HBO2)]. The epoxygenase inhibitor MS-PPOH in HBO2 animals reversed these changes. Glibenclamide had no effect on relaxation in response to ANG-(1-7). Expression of CYP4A2, CYP4A3 and CYP2J3 mRNA was not significantly altered with HBO, whereas CYP4A1 was significantly upregulated. CONCLUSIONS: Our results suggest a role for epoxyeicosatrienoic acids in modulating relaxation response to ANG-(1-7) with HBO2 that is conducted via potassium channels other than KATP channels. HBO2 increased the responses to ANG-(1-7) after precontraction with noradrenaline. The difference between the response to ANG II in the HBO2 group and ANG II + ANG-(1-7) in the HBO2 group (the contraction force of the peptide combination being lower), without such difference in the control group, suggests an influence ofHBO2 on vascular reactivity.

Influence of hyperbaric oxygen on blood vessel reactivity: concept of changes in conducted vasomotor response.

The actions of oxygen in the body are extremely complex, and are also involved in various signalling pathways. Hyperbaric oxygen is known to contribute to the improvement of conditions where tissue circulation is suboptimal, and has considerable usage in different treatment protocols and experimental investigations. However, the precise mechanism by which hyperbaric oxygen changes the functioning of coordinated blood vessel systems and microcirculation is still unknown. Taking into account the known facts, we suggest that hyperbaric oxygen induces changes in conducted vasomotor responses, and in that way influences vascular sensitivity and reactivity to vasodilators and vasoconstrictors. Conducted vasomotor responses are constrictions and dilations that are propagated along the vessel, leading to changes in vessel diameter on a certain distance of the initial site of vasoactive substance activity. Because these vascular responses are of substantial significance in physiological processes, their modification would subsequently cause alterations of blood vessel function and tissue perfusion that could explain observed effects of hyperbaric oxygen. We also discuss potential molecular targets of hyperbaric oxygen, investigation of which could presumably help in the eventual clarification of hyperbaric oxygen action.

The effect of hyperbaric oxygen therapy on blood vessel function in diabetes mellitus.

Prolonged untreated diabetes mellitus leads to microangiopathy, tissue hypoxia and ischemic lesions; it increases the risk for stroke and exacerbates brain tissue damage following ischemia. Patients exhibit advanced atherosclerosis in coronary and cerebral arteries as well as enhanced vascular responsiveness to vasoconstrictors, an attenuated response to vasodilators and impaired autoregulation of cerebral blood flow. Altered endothelial function of arterioles and an impaired vasomotor function of resistance vessels could contribute to altered regulation of regional blood flow and insufficient tissue perfusion in diabetes mellitus. Hyperbaric oxygen therapy is shown to contribute to the healing of ischemic ulcerations in diabetic patients and to improvement of several other pathologic conditions. However, information about the mechanism of how this therapy works is still very limited. We postulate that hyperbaric oxygen therapy has an effect on vascular function by modulating mechanisms of vascular responses to various dilator and constrictor agonists in cerebral resistance vessels, leading to restored vascular reactivity. In accordance to this, the therapy affects production of vasodilators and vasoconstrictors, as well as the vessel-sensitivity to these factors. Furthermore, we hypothesize that hyperbaric oxygen therapy would restore cerebral blood flow regulation that is impaired in diabetics, whereas in contrast to that, chronic intermittent hypoxia would lead to impaired cerebral blood flow. These proposed mechanisms would, if confirmed, represent a valuable advancement in the understanding of this subject.