NO Synthase Inhibitor Research Articles

Significance of the activity of liver arginase and Kupffer cells in the detoxification processes and the thyroid status formation in rats under alcoholic intoxication of different severity

Modern medicine faces the problem of a steady growth of alcoholic pathology. As you know, morbidity and mortality with a regular consumption of alcoholic beverages is associated with the toxic effects of ethanol on the most important human organs and, first of all, on the liver. To date, a sufficient number of facts are accumulated, indicating the importance of liver arginase and Kupffer cells in vital processes in health and disease. The aim of the study was to elucidate the significance of the activity of liver arginase and Kupffer cells in the detoxification processes and the thyroid status formation in rats with chronic ethanol intoxication of different severity. In rat experiments using modern physiological, biochemical research methods and a pharmacological approach, it was found that liver arginase and Kupffer cells participate in changes in the liver detoxification function and the thyroid status formation induced by chronic ethanol intoxication. The activity of liver arginase and Kupffer cells determines the severity of detoxification processes and the thyroid status formation in chronic alcohol intoxication. The direction and severity of changes in the arginase activity and the liver detoxification function during chronic alcoholism depends on the severity of chronic alcohol intoxication. Under the influence of daily intragastric administration for 60 days, a 30 % aqueous solution of ethanol (3.5 g 92 % ethanol per kg of body weight) in animals inhibited the activity of liver arginase and the detoxification function, but the introduction of a 10 % aqueous solution of ethanol (1.0 g 92 % ethanol per kg of body weight) within 2 months leads to an increase in the activity of liver arginase and detoxification processes. Kupffer cells depression by GdCl3 as the action in the body of the NO-synthase inhibitor methyl ester NG-nitro-L-arginine weakens and the arginase inhibitor Nω-hydroxy-nor-L-arginine contributes to the development of characteristic changes in the processes of detoxification and triiodothyronine level in plasma during chronic alcohol intoxication caused by intragastric introduction of ethanol at a dose of 3.5 g/kg for 60 days.

In vitro inhibition of phosphodiesterase type 4 enhances rat corpus cavernosum nerve-mediated relaxation induced by gasotransmitters

AimsNitric oxide (NO) and hydrogen sulfide (H2S) are involved in nerve-mediated corpus cavernosum (CC) relaxation. Expression of phosphodiesterase type 5 (PDE5) and type 4 (PDE4), cyclic guanosine monophosphate (cGMP)- and cyclic adenosine monophosphate (cAMP)-specific, respectively, has been described and PDE5- and PDE4-inhibitors induce cavernous smooth muscle relaxation. Whereas the NO/cGMP signaling pathway is well established in penile erection, the cAMP-mediated mechanism is not fully elucidated. The aim of this study is to investigate the localization and the functional significance of PDE4 in rat CC tone regulation. Main methodsWe performed immunohistochemistry for the detection of the PDE4A isoenzyme. Isometric tension recordings for roflumilast and tadalafil, PDE4 and PDE5 inhibitors, respectively, electrical field stimulation (EFS) and β-adrenoceptor agonist isoproterenol and endogenous H2S production measurement. Key findingsA marked PDE4A expression was detected mainly localized in the nerve cells of the cavernous smooth muscle. Furthermore, roflumilast and tadalafil exhibited strong corpus cavernous relaxations. Endogenous H2S production was decreased by NO and H2S synthase inhibitors and increased by roflumilast. Isoproterenol- and EFS-induced relaxations were increased by roflumilast. SignificanceThese results indicate that PDE4A is mainly expressed within the nerves cells of the rat CC, where roflumilast induces a potent corpus cavernous relaxation per se and potentiates the response induced by β-adrenoceptor activation. The fact that roflumilast enhances H2S production, as well as EFS-elicited responses suggests that PDE4 inhibitors modulate, in a positive feedback fashion, nerve-mediated relaxation induced by gasotransmitters, thus indicating a key role for neuronal PDE4 in penile erection.

Protective effects of L-arginine on cognitive deficits and biochemical parameters in an experimental model of type-2 diabetes mellitus induced Alzheimer's disease in rats.

Alzheimer's disease (AD) has been closely linked to type 2 diabetes mellitus (T2DM) and insulin resistance plays a key role in the onset and development of AD. It has also been reported that NO donors play an important role in diabetes and have neuroprotective activity. The present study evaluated the effects NOS substrate, L-arginine and NOS-blocker, NG-nitro-L-arginine methyl ester (L-NAME) on cognitive functions, brain amyloid β levels and other associated biochemical markers in diabetes (T2DM)-induced AD in rats. T2DM was induced by a combination of high fat diet (HFD) and streptozotocin (STZ) (35 mg/kg, i.p.). Pretreatment with L-arginine (100 mg/kg/day) improved diabetes related biochemical parameters viz. plasma glucose, triglycerides and cholesterol. In addition, L-arginine also improved insulin levels and glucose tolerance. On the other hand, NO synthase inhibitor, L-NAME (10 mg/kg), did not have much influence on these parameters. Further, L-arginine treatment showed ameliorative effects on cognitive deficits seen in the Morris water maze (MWM) test when compared to the vehicle (saline) treated group and similar results were observed in the passive avoidance test. These neurobehavioural changes were associated with predictable modulations in Aβ levels in hippocampus and cortex in HFD + STZ + saline group, which were attenuated in the L-arginine, but not in the L-NAME-treated group. Our results indicate that L-arginine could be a considered a potential therapeutic strategy to attenuate the diabetes induced cognitive deficits in AD.

Effects of Nitric Oxide on C2C12 Cell Inflammatory Responses and Notexin-Induced Muscle Injury

Skeletal muscle injury is an acute inflammatory condition caused by an inflammatory response. To reduce inflammatory cell infiltration and relieve skeletal muscle injury, efficient treatment is urgently needed. Nitric oxide is a free radical molecule reported to have anti-inflammatory effects. In this study, we showed that NO could inhibit the inflammatory response of C2C12 cells in vitro and protect rat skeletal muscle injury from notexin in vivo. NO synthase inhibitor (L-NG-Nitroarginine Methyl Este?L-NAME) and NO donor (sodium nitroprusside dehydrate ?SNP) were used to explore the vital role of lipopolysaccharides (LPSs) in LPS-stimulated C2C12 myoblasts.The expression of IL-18 and IL-1b was upregulated by L-NAME and downregulated by SNP, as indicated by the ELISA results. NO can reduce ASC, Caspase-1, and NLRP3 mRNA and protein levels. Furthermore, NO was detected in the rat model. The results of immunohistochemical staining showed that the production of DMD decreased. We conducted qRT-PCR and western blotting to detect the expression of Jo-1, Mi-2, TLR2, and TLR4 on day 6 post injury following treatment with L-NAME and SNP. The expression of Jo-1, Mi-2, TLR2, and TLR4 was upregulated by L-NAME and significantly reversed by SNP. NO can alleviate C2C12 cell inflammatory responses and protect rat skeletal muscle injury from notexin.

The effect of the long-term inhibition of hydrogen sulfide production on the reactivity of the cardiovascular system in Wistar rats.

In this study, we investigated the blood pressure responses of the peripheral bed invivo after chronic hydrogen sulfide (H2S) inhibition combined with acute nitric oxide (NO) deficiency. We also evaluated the role of endogenously produced H2S in the vasoactive responses of large- and medium-sized arteries invitro. Changes in integrated blood pressure responses were measured after chronic inhibition of cystathionine-γ-lyase, an enzyme involved in H2S synthesis, with DL-propargylglycine (PPG), and acute inhibition of NO-synthase with nonspecific L-NG-nitro arginine methyl ester (L-NAME), and vasoactive responses of the thoracic aorta (TA) and mesenteric artery (MA) were investigated after acute incubation with PPG. We confirmed that chronic H2S deficiency had no effect on blood pressure, heart trophycity, noradrenaline, and H2S donor vasoactive responses but induced renal hypertrophy and a decrease in acetylcholine-induced hypotensive and L-NAME-induced hypertensive responses. Acute H2S deficiency led to an increase in basal tone (MA) or active tone (TA), whereas endothelium-dependent vasorelaxation remained unaffected. Long-term administration of PPG revealed a role of endogenous H2S in the bioavailability of endothelial NO in peripheral arteries. When both H2S and NO were lacking, the activation of H2S-independent compensatory mechanisms plays an important role in maintaining the vasodilator responses of the cardiovascular system.

Interplay between nitric oxide and gonadotrophin-releasing hormone in the neuromodulation of the corpus luteum during late pregnancy in the rat

BackgroundNitric oxide and GnRH are biological factors that participate in the regulation of reproductive functions. To our knowledge, there are no studies that link NO and GnRH in the sympathetic ganglia. Thus, the aim of the present work was to investigate the influence of NO on GnRH release from the coeliac ganglion and its effect on luteal regression at the end of pregnancy in the rat.MethodsThe ex vivo system composed by the coeliac ganglion, the superior ovarian nerve, and the ovary of rats on day 21 of pregnancy was incubated for 180 min with the addition, into the ganglionic compartment, of L-NG-nitro arginine methyl ester (L-NAME), a non-selective NO synthase inhibitor. The control group consisted in untreated organ systems.ResultsThe addition of L-NAME in the coeliac ganglion compartment decreased NO as well as GnRH release from the coeliac ganglion. In the ovarian compartment, and with respect to the control group, we observed a reduced release of GnRH, NO, and noradrenaline, but an increased production of progesterone, estradiol, and expression of their limiting biosynthetic enzymes, 3β-HSD and P450 aromatase, respectively. The inhibition of NO production by L-NAME in the coeliac ganglion compartment also reduced luteal apoptosis, lipid peroxidation, and nitrotyrosine, whereas it increased the total antioxidant capacity within the corpora lutea.ConclusionCollectively, the results indicate that NO production by the coeliac ganglion modulates the physiology of the ovary and luteal regression during late pregnancy in rats.

Testosterone positively regulates vagina NO-induced relaxation: an experimental study in rats

PurposeFemale sexual response involves a complex interplay between neurophysiological mechanisms and the nitric oxide (NO)-mediated relaxation of clitoris and vagina. The aim of this study was to evaluate sex steroids regulation of the relaxant pathway in vagina, using a validated animal model.MethodsSubgroups of OVX Sprague–Dawley rats were treated with 17β-estradiol, testosterone, or testosterone and letrozole, and compared with a group of intact animals. Masson’s trichrome staining was performed for morphological evaluation of the distal vaginal wall, in vitro contractility studies investigated the effect of OVX and in vivo treatments on vaginal smooth muscle activity. RNA from vaginal tissue was analyzed by semi-quantitative RT-PCR.ResultsImmunohistochemical analysis showed that OVX induced epithelial and smooth muscle structural atrophy, testosterone and testo + letrozole increased the muscle bundles content and organization without affecting the epithelium while 17β-estradiol mediated the opposite effects. In vitro contractility studies were performed on noradrenaline pre-contracted vaginal strips from each experimental group. Acetylcholine (0.001–10 µM) stimulation induced a concentration-dependent relaxation, significantly reduced by NO-synthase inhibitor L-NAME and by guanylate cyclase inhibitor ODQ. OVX resulted in a decreased responsiveness to acetylcholine, restored by testosterone, with or without letrozole, but not by 17β-estradiol. OVX sensitivity to the NO-donor SNP was higher than in the control. Vardenafil, a PDE5 inhibitor, enhanced SNP effect in OVX + testosterone as well as in control, as supported by RNA expression analysis.ConclusionsOur study demonstrates that testosterone improves the NO-mediated smooth muscle vaginal cells relaxation confirming its role in maintaining the integrity of muscular relaxant machinery.

Effects of nitric oxide modulators and antioxidants on endocrine and cellular markers of acute stress in rats

The effects of nitric oxide modulators (NO-modulators) and antioxidants on acute (RSx1) restraint stress induced endocrine, cellular and oxidative/nitrosative stress markers was studied in Wistar rats. The results of our study revealed that exposure to RS(x1) enhanced malondialdehyde (MDA), heat shock protein (HSP-70), corticosterone, nuclear factor kappa B (NF-κB) levels and suppressed glutathione (GSH), superoxide dismutase (SOD) and total nitrites and nitrates (NOx) levels. NO precursor and NO synthase inhibitors were found to differentially modulate stress mechanisms, by altering NF-κB, HSP-70 and corticosterone levels. l-Ascorbic acid significantly suppressed acute stress induced elevation of NF-κB and HSP-70 levels depicting protective effects, as also evidenced by reversal of elevated plasma corticosterone levels. Therefore, modulation of oxidative and nitrosative pathways, offers an approach in modulating stress induced changes associated with various disorders.

Region-specific effects of antenatal/early postnatal hypothyroidism on endothelial NO-pathway activity in systemic circulation.

BackgroundAntenatal/early postnatal hypothyroidism weakens NO-mediated anticontractile influence of endothelium in coronary arteries of adult rats, but it remains unclear whether this occurs in other vascular regions. We hypothesized that developmental thyroid deficiency is followed by region-specific changes in the endothelial NO-pathway activity in systemic vasculature. To explore this, we estimated the effects of antenatal/early postnatal hypothyroidism on NO-pathway activity and its potential local control mechanisms in rat mesenteric and skeletal muscle (sural) arteries.MethodsDams were treated with 6-propyl-2-thiouracil (PTU) in drinking water (0.0007%) during pregnancy and 2 weeks postpartum; control (CON) females received PTU-free water. Adult offspring (10–12-weeks) arteries were studied by wire myography, qPCR, and Western blotting.ResultsEndothelium removal or inhibition of NO-synthase with L-NNA augmented contractile responses to α1-adrenoceptor agonist methoxamine. In PTU compared to CON group, these effects were stronger in sural arteries, but did not differ in mesenteric arteries. The responses of both arteries to NO-donor DEA/NO were similar in CON and PTU rats. mRNA contents of deiodinase 2 and thyroid hormone receptor α were similar in mesenteric arteries of two groups but were elevated in sural arteries of PTU group compared to CON. The abundance of eNOS protein was higher in sural arteries of PTU compared to CON rats.ConclusionAntenatal/early postnatal hypothyroidism is followed by an increase in NO-mediated anticontractile influence in sural, but not in mesenteric arteries of adult animals. The diversity of hypothyroidism effects may be due to different alterations of local T3 synthesis/reception in different vascular beds.

Role of Amino Acid Arginine and Nitric Oxide in Mechanisms of Cytoprotective Effect of Non-Opiate Leu-Enkephalin Analogue In Vitro.

Incubation of primary culture of pulmonary fibroblasts with non-opiate analogue of leuenkephalin (NALE; Phe-D-Ala-Gly-Phe-Leu-Arg, 0.1 μM) reduced generation of superoxide anion-radical (by 20.7%) and decreased the number of p53+ cells (by 40.2%) induced by exposure to H2O2 (60 μM). The cytoprotective effect of NALE was potentiated by NO synthase inhibitor L-NAME (1 mM): the number of p53+ cells decreased by 65.3% and morphometric parameters of the cell nuclei and nucleoli were improved. Incubation of pulmonary fibroblasts culture with peptide G (Phe-D-Ala-Gly-Phe-Leu-Gly, 0.1 μM) also significantly reduced the damaging effect of H2O2: the number of p53+ cells decreased by 73.5%, the area of cell nuclei returned to normal, and generation of superoxide anion-radical decreased by 18.4%. These results indicate that C-terminal amino acid Arg and activation of NO synthase are not involved in the direct cytoprotective effect of NALE.

Sympathetic Nerve Activity and Baroreflex are Strongly Altered in a Context of Severe Hypertension Using the Spontaneously Hypertensive Rat Model Associated with Chronic Reduction of Nitric Oxide.

The aim of our study is to investigate the sympathetic output and baroreflex via renal sympathetic nerve activity (RSNA) recording in a model of severe hypertension which exhibits arterial, cardiac, and renal damages, the spontaneously hypertensive rat (SHR) under lowered NO bioavailability. SHR are treated from 18 to 20 weeks of age with a low dose of L-NAME, a NO synthase inhibitor, in drinking water (SHRLN) and compared to SHR and normotensive Wistar Kyoto (WKY) rats. After the two-week treatment, rats are anesthetized for RSNA, mean blood pressure (MBP), and heart rate (HR) recording. MBP is higher in SHR than in WKY and higher in SHRLN than in SHR. Compared to WKY, SHR displays an alteration in the baroreflex with a displacement of the sympathoinhibition curve to highest pressures; this displacement is greater in SHRLN rats. The bradycardic response is reduced in SHRLN compared to both SHR and WKY. In hypertensive rats, SHR and SHRLN, basal RSNA is modified, the maximal amplitude of burst is reduced, but minimal values are increased, indicating an increased basal RSNA with reduced bursting activity. The temporal correlation between RSNA and HR is preserved in SHR but altered in 10 SHRLN out of 10. The RSNA inhibition triggered by the Bezold–Jarisch reflex activation is not modified in hypertensive rats, SHR or SHRLN, in contrast to that triggered by the baroreflex. Histological analysis of the carotid bifurcation does not reveal any abnormality in SHRLN at the level of the carotid sinus. In conclusion, data indicate that the sympathetic outflow is altered in SHRLN with a strong reduction of the baroreflex sympathoinhibition and suggest that its central pathway is not involved. These additional results on SHRLN also confirm the usefulness of this model of severe hypertension with multiple target organ damages.

Citrus flavonoids effects on human umbilical vein

• Hesperidin is more potent than diosmin as vasorelaxant of Human Umbilical Vein (HUV). • K ATP channel activation is involved in hesperidin-dependent vasorelaxation only. • Endothelium-dependent vasorelaxation to hesperidin is gender-sensitive. • Hesperidin may be beneficial in maintaining vascular and endothelial functions in HUV. Flavonoids intake is associated to beneficial effects on the cardiovascular system. Aim of this study was to investigate the biological activities of hesperidin and diosmin on the vascular tone of human umbilical vein (HUV). Umbilical cords obtained at delivery from healthy women were set up for recording of isometric tension; vasorelaxing activity of flavonoids was assessed in serotonin-precontracted HUV rings. Hesperidin evoked a more profound vasorelaxation than diosmin, and inhibition of NO-synthase significantly impaired relaxant responses to hesperidin but completely abolished those to diosmin. The residual vasorelaxation to hesperidin was prevented by glybenclamide (a K ATP channel blocker). Endothelium-dependent responses to hesperidin were larger in female- when compared to male-derived HUV. β-estradiol and a direct NO-donor compound caused a marked vasorelaxation irrespective of gender of the HUV. Diosmin and, in particular, hesperidin may exert a beneficial effect on the vascular and endothelial function of HUV, and help preventing fetal vascular complications.

Differential hyperpolarization to substance P and calcitonin gene-related peptide in smooth muscle versus endothelium of mouse mesenteric artery.

We sought to define how sensory neurotransmitters substance P and calcitonin gene-related peptide (CGRP) affect membrane potential of vascular smooth muscle and endothelium. Microelectrodes recorded membrane potential of smooth muscle from pressurized mouse mesenteric arteries (diameter, ~150µm) and in endothelial tubes. Resting potential was similar (~ -45mV) for each cell layer. Substance P hyperpolarized smooth muscle and endothelium ~ -15mV; smooth muscle hyperpolarization was abolished by endothelial disruption or NO synthase inhibition. Blocking KCa channels (apamin+charybdotoxin) attenuated hyperpolarization in both cell types. CGRP hyperpolarized endothelium and smooth muscle ~ -30mV; smooth muscle hyperpolarization was independent of endothelium. Blocking KCa channels prevented hyperpolarization to CGRP in endothelium but not smooth muscle. Inhibiting KATP channels with glibenclamide or genetic deletion of KIR 6.1 attenuated hyperpolarization in smooth muscle but not endothelium. Pinacidil (KATP channel agonist) hyperpolarized smooth muscle more than endothelium (~ -35 vs. ~ -20mV). Calcitonin gene-related peptide elicits greater hyperpolarization than substance P. Substance P hyperpolarizes both cell layers through KCa channels and involves endothelium-derived NO in smooth muscle. Endothelial hyperpolarization to CGRP requires KCa channels, while KATP channels mediate hyperpolarization in smooth muscle. Differential K+ channel activation in smooth muscle and endothelium through sensory neurotransmission may selectively tune mesenteric blood flow.

NADPH oxidase 4 has a crucial impact on the microcirculation of hypercholesteremic LDL receptor deficient mice

Abstract Introduction NADPH oxidase (NOX) 4-generated H2O2 has anti-atherosclerotic properties in conduit arteries like the aorta and carotids. However, the role of NOX4 on vascular function of small resistance arteries and blood pressure in a mouse model of familial hypercholesterolemia is unknown. Purpose We evaluated whether NOX4-generated H2O2 might play a role in perivascular adipose tissue of the thoracic aorta (tPVAT) and small resistance arteries on vascular function in a mouse model of familial hypercholesterolemia. Methods Aortic segments and mesenteric arteries from 26-week-old Ldlr−/− and Nox4−/− / Ldlr−/− mice were analysed by Mulvany myograph. In addition, vascular contraction and relaxation was analysed in the presence of L-NAME and catalase. Analysis of mRNA expression was performed in murine and human tissue by quantitative real-time PCR. Blood pressure was detected by tail cuff method in conscious, trained mice. Results Loss of NOX4 led to severe endothelial dysfunction in mesenteric arteries of Ldlr−/− mice. Blocking of NO synthases with L-NAME led to decreased endothelial relaxation in Ldlr−/− mice at the level of Nox4−/− / Ldlr−/− mice. However, incubation with L-NAME did not worsen the established endothelial dysfunction of the mesenteric arteries from Nox4−/− / Ldlr−/− mice. These results are strikingly different from the aorta, where inhibition of NO synthases led to a similarly impaired endothelial relaxation in both mouse strains. We detected a similar eNOS expression in the aorta of Ldlr−/− and Nox4−/− / Ldlr−/−, but a reduced eNOS expression in the mesenteric arteries of Nox4−/− / Ldlr−/− mice. H2O2 can induce eNOS expression. Therefore, we analysed the vascular function after catalase incubation and again found a significant reduction of endothelial function in the mesenteric arteries of Ldlr−/− mice. Finally, we analysed blood pressure of these mice and did not observe differences in systolic blood pressure, despite significant differences in endothelial function of resistant arteries. Conclusion NOX4 protects against severe endothelial dysfunction in the mesenteric artery in a model of hypercholesterolemia. Funding Acknowledgement Type of funding sources: Other. Main funding source(s): Ghanaian-German postgraduate training scholarship program (DAAD)

Effects of NO modulators and antioxidants on endocrine and cellular markers in rats under repetitive restraint stress.

The aim of the study was to evaluate the effects of NO modulators and antioxidant treatments on endocrine (plasma corticosterone), cellular (heat shock protein 70 [HSP-70] and nuclear factor κB [NF-κB]), and oxidative stress markers in repetitively stressed rats. Repetitive (restraint) stress (RS 1hr/day × 21 days) enhanced the levels of cellular and endocrine stress markers in the rat blood and altered pro-oxidant-antioxidant balance differentially in the control and test groups. Exposure to repetitive RS enhanced malondialdehyde (MDA) levels, lowered reduced glutathione (GSH), and superoxide dismutase (SOD) levels as well as nitric oxide (NOx) levels. NO precursor L-arginine and NO synthase inhibitors were found to differentially modulate stress-induced mechanism in altering NF-κB, HSP-70, and corticosterone levels. The antioxidant L-ascorbic acid (L-AA) significantly suppressed RS(×21)-induced elevation of NF-κB and HSP-70 levels, depicting protective effects, as also evidenced by reversal of elevated corticosterone levels. The results suggest that NO modulators and antioxidants differentially influence repetitive stress-induced changes in endocrine and cellular markers, and the complex interaction between NO and cellular markers like HSP70 and NF-κB plays a crucial modulatory role in this phenomenon.

Effect of Cholecystokinin-8 (CCK-8) on Blood Pressure and Blood Content of Calcitonin-Gene-Related Peptide (CGRP) in Rats with Hypertension Caused by Fructose or Inhibition of Nitric Oxide Synthesis

We studied the effect of CCK-8 on BP and blood content of CGRP in rats with hypertension caused by fructose or inhibition of NO synthase with L-NAME. The decrease in the CGRP content was found during the development of fructose-induced hypertension, but not L-NAME-caused hypertension. Administration of CCK-8 to fructose-fed animals reduced BP and increased the content of CGRP. In rats with hypertension caused by NO deficit, CCK-8 lowered BP, but did not affect the content of CGRP. These findings suggest that CGRP mediates the hypotensive effect of CCK-8 in fructose-induced hypertension, but not in NO-deficient hypertension.

Noninvasive assessment of coronary microvascular dysfunction using SPECT myocardial perfusion imaging and myocardial perfusion entropy quantification in a rodent model of type 2 diabetes

Coronary microvascular dysfunction (CMVD) plays a major role in the occurrence of cardiovascular events (CVE). We recently suggested the clinical potential of myocardial perfusion entropy (MPE) quantification from SPECT myocardial perfusion images (MPI) for the prognosis of CVE occurrence. We hypothesized that the quantification of MPE from SPECT MPI would allow the assessment of CMVD-related MPE variations in a preclinical model of type 2 diabetes (T2D) including treatment with the anti-diabetic incretin liraglutide (LIR). Optimal conditions for the preclinical quantification of MPE using 201Tl SPECT MPI were determined in rats with a T2D-like condition induced by a high-fat diet and streptozotocin injection (feasibility study, n = 43). Using such conditions, echocardiography and post-mortem LV capillary density evaluation were then used in order to assess the effect of LIR and the ability of MPE to assess CMVD (therapeutic study, n = 39). The feasibility study identified dobutamine stress and acute NO synthase and cyclooxygenase inhibition as optimal conditions for the quantification of MPE, with significant increases in MPE being observed in T2D animals (P < 0.01 vs controls). In the therapeutic study, T2D rats were hyperglycemic (5.5 ± 0.5 vs 1.1 ± 0.3g/L for controls, P < 0.001) and had a significantly lower left ventricular ejection fraction (LVEF) (65 ± 4% vs 74 ± 9%, P < 0.01) and LV capillary density (2400 ± 300 vs 2800 ± 600mm-3, P < 0.05). LIR partially restored glycemia (3.9 ± 0.6g/L, P < 0.05 vs controls and T2D), totally prevented LVEF impairment (72 ± 7%, P = NS vs CTL), with no significant effect on capillary density. MPE was significantly increased in T2D rats (7.6 ± 0.5 vs 7.1 ± 0.5, P < 0.05), with no significant improvement in T2D-LIR rats (7.4 ± 0.4, P = NS vs controls and T2D). MPE quantification allowed the preclinical noninvasive assessment of CMVD. Both MPE and capillary density quantification suggested that LIR did not improve T2D-induced CMVD. The relevance of MPE for CMVD assessment warrants further clinical investigation.

Endothelial deficiency of insulin-like growth factor-1 receptor (IGF1R) impairs neurovascular coupling responses in mice, mimicking aspects of the brain aging phenotype.

Age-related impairment of neurovascular coupling (NVC; or "functional hyperemia") compromises moment-to-moment adjustment of regional cerebral blood flow to increased neuronal activity and thereby contributes to the pathogenesis of vascular cognitive impairment (VCI). Previous studies established a causal link among age-related decline in circulating levels of insulin-like growth factor-1 (IGF-1), neurovascular dysfunction and cognitive impairment. Endothelium-mediated microvascular dilation plays a central role in NVC responses. To determine the functional consequences of impaired IGF-1 input to cerebromicrovascular endothelial cells, endothelium-mediated NVC responses were studied in a novel mouse model of accelerated neurovascular aging: mice with endothelium-specific knockout of IGF1R (VE-Cadherin-CreERT2/Igf1rf/f). Increases in cerebral blood flow in the somatosensory whisker barrel cortex (assessed using laser speckle contrast imaging through a cranial window) in response to contralateral whisker stimulation were significantly attenuated in VE-Cadherin-CreERT2/Igf1rf/f mice as compared to control mice. In VE-Cadherin-CreERT2/Igf1rf/f mice, the effects of the NO synthase inhibitor L-NAME were significantly decreased, suggesting that endothelium-specific disruption of IGF1R signaling impairs the endothelial NO-dependent component of NVC responses. Collectively, these findings provide additional evidence that IGF-1 is critical for cerebromicrovascular endothelial health and maintenance of normal NVC responses.

Caffeic Acid Phenethyl Ester (CAPE), Active Phenolic Compound of Propolis Attenuates Endothelin, Prostaglandin F2α and U46619 Elicited Contractions of Isolated Human Umbilical Artery

Background: Propolis is a product of honeybees that contains a variety of different compounds, including caffeic acid phenethyl ester (CAPE). Propolis and its bioactive compounds are widely used in folk medicine and as a dietary supplement. Previously it has been shown that CAPE has antioxidant, anti-inflammatory, antimicrobial, antiviral, immunomodulatory, and anticancer activity. Objective: This in vitro study was designed to investigate the vasoactive effects of CAPE on quiescent and precontracted human umbilical arteries. Methods: Umbilical artery strips were suspended in aerated organ baths containing a buffer solution. The strips were randomly allocated to study groups (n=8). Via a transducer and computer, changes in isometric tension were recorded. The effects of cumulative CAPE (10-8-10-4M) on basal tone of the artery, and in different groups of strips, the vasodilatory effect of cumulative CAPE on the constriction elicited by endothelin (ET-1), prostaglandin F2α (PGF2α) and U46619, and the effect of incubation with NO synthase inhibitor L-NAME, were recorded. Conclusion: CAPE elicits concentration-dependent relaxation on precontracted human umbilical artery strips depending on the constrictor agent. NO plays significant role in CAPE’s vasorelaxant effect.

Role of H2S in Regulation of Vascular Tone in Metabolic Disorders.

We studied the effect of the H2S donor (NaHS, 1-500 μM) on the contractile responses of isolated aortic smooth muscle segments from rats with metabolic syndrome induced by high-fat, high-carbohydrate diet. It was found that the vasorelaxing effect of NaHS (5-100 μM) decreased in under conditions of MS. The endothelial NO synthase inhibitor L-NAME (100 μM) suppressed the effect of NaHS, while cystathionine-gamma-lyase inhibitor PAG (100 μM) decreased the vasodilating effects of acetylcholine (0.1-100 μM). Application of endogenous NO precursor L-arginine (1 mM) potentiated in the effects of H2S donor NaHS. Thus, the contractile activity of vascular smooth muscles in metabolic syndrome is determined by not only the effect of H2S, but also the influence of NO.