Relaxation Of Rat Aorta Research Articles

Reduced Nitric Oxide–cGMP Signaling Contributes to Vascular Dysfunction Induced by the Adipokine Chemerin.

The new adipokine Chemerin has been implicated in the cardiovascular complications associated with obesity and metabolic syndrome. The present study was designed to test the hypothesis that the adipokine chemerin induces vascular dysfunction by decreasing nitric oxide (NO)–cGMP signaling. Isometric contraction was recorded in rat thoracic aortic rings incubated with chemerin (0.5ng/mL or 5ng/mL; 1 hour) or vehicle. Chemerin markedly reduced both sodium nitroprusside [pD2: 8,6±0,1 vs. chem 0,5ng/mL=7,4±0,1; chem 5ng/mL=6,9±0,1] and acetylcholine [Emax: vehicle=93,1±11,1; chem 0,5ng/mL=66,1±3,1; chem 5ng/mL= 63,8±12,6]‐induced relaxation in rat aorta. Chemerin increased eNOS phosphorylation [a.u.: 1 vs. 1.9±0.1], but also induced NOS uncoupling, as demonstrated by a decreased eNOS dimer/monomer ratio [a.u.: 1 vs. 0,5±0,1]. Chemerin decreased vascular expression of soluble guanylate cyclase [a.u.: 1 vs. 0.6±0.1] as well as cGMP levels [pmol/mg: 1±0,2]. Chemerin increased superoxide anion generation [rlu: 100 vs. 120,5±8,1], determined by lucigenin chemiluminescence assay in cultured VSMCs from rat aorta, an effect that was abrogated by incubation with L‐NAME [120,5 vs. 71,6±9,4] or tempol [120,5 vs. 73,3±7,5]. In conclusion, chemerin decreases NO‐cGMP signaling via ROS generation, NOS uncoupling and decreased GC activity. Our studies may contribute to a better understanding of the role of factors released by the visceral adipose tissue on vascular function and, consequently, on the vascular dysfunction in obesity and obesity‐associated diseases.

Induction of endothelium-dependent relaxation in rat aorta via nitric oxide production caused by the aqueous extract of Astragalus membranaceus Bunge leaves

Induction of endothelium-dependent relaxation in rat aorta via nitric oxide production caused by the aqueous extract of Astragalus membranaceus Bunge leaves

The novel ortho-chloro derivate propafenone induced relaxation in isolated rat aorta

The information on the inhibitory effect of propafenone in vascular smooth muscle is sparse. Propafenone acts through blockage of voltage-dependent cardiac Na+ channels, L-type Ca2+ channels, voltage-sensitive K+ (Kv) channels, as well as β-adrenergic receptors in the heart. The introduction of different chemical groups in the benzyl moiety of propafenone influences pharmacological properties of newly developed derivate of propafenone. Here we investigated the effect of new ortho-chloro derivate of propafenone (5OCl) on the vascular tone of precontracted rat aorta. 5OCl produced endothelium-independent relaxation of rat aorta. In order to test the involvement of different ion channels in 5OCl mechanism of action, antagonist of Na+, lidocaine, KV channels, 4-aminopyiridine (4-AP) and L-type Ca2+ channels, nifedipine were used. All tested antagonists of ion channels did not influence the relaxation of rat aorta induced by high a concentration of 5OCl (≥10 μM), but antagonized the relaxation induced by low concentrations of this propafenone derivate. Thus, 5OCl derivate has comparable potency and efficacy as propafenone. According to its interaction with lidocaine, 4-AP and nifedipine it seems that 5OCl partly shares the mechanism of action with propafenone. The mechanism of vasodilatation induced by high micromolar concentration of 5OCl is not defined and further investigations are necessary.

Intense exercise training is not effective to restore the endothelial NO-dependent relaxation in STZ-diabetic rat aorta

BackgroundThe aim of this study was to examine the effects of intense physical training on vascular function in streptozotocin-diabetic rats. We focused on the endothelium-dependent relaxation (EDR) induced by acetylcholine (ACh) and stable ADP adenosine-5′- O – (2-thiodiphosphate) (ADPβS).MethodsControl or diabetic male Wistar rats (n=44) were randomly assigned to sedentary or trained groups. The training program consisted in a regular period of running on a treadmill during 8 weeks (10° incline and up to 25 m/min, 60 min/day). The reactivity of isolated thoracic aorta rings of healthy, diabetic and/or trained has been tested.ResultsACh and ADPβS-induced EDR were observed in phenylephrine (PE) pre-contracted vessels. As compared to sedentary control group, diabetic rats showed an increase in PE-induced contraction and a decrease in ACh and ADPβS-induced EDR (p<0.05). Moreover, there were no increase in ACh and ADPβS-induced EDR in diabetic rats. N-Nitro-L-Arginine Methyl Ester inhibited the nitric oxide synthase in diabetic and control rats, thereby resulting in a strong inhibition of the EDR induced by ACh and ADPβS (10-6 M).ConclusionDiabetes induced an endothelium dysfunction. Nevertheless, our intense physical training was not effective to restore the aorta endothelial function.

Na +/H + exchanger inhibitor induces vasorelaxation through nitric oxide production in endothelial cells via intracellular acidification-associated Ca2 + mobilization

The objective of this study was to determine the mechanism by which Na+/H+ exchanger (NHE) inhibitors induce vasodilatation. The NHE inhibitors, 5-(N,N-dimethyl)-amiloride (DMA), cariporide, and amiloride, evoked endothelium-dependent relaxation in rat aortas with ED50 values of 16, 89, and 148μM, respectively, and these effects were abolished by treatment with NG-nitro-l-arginine methyl ester (L-NAME). The relaxation effects induced by DMA and cariporide were strongly attenuated in aortas of the endothelial NO synthase (eNOS)-deficient mice, as compared to the effects in wild-type mice. The DMA-induced relaxation in rat aorta was attenuated by a calmodulin (CaM) inhibitor, calmidazolium, and a soluble guanylyl cyclase inhibitor, [1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one, but was not affected by a phosphoinositide 3-kinase inhibitor, wortmannin. Immunoblots for endothelial eNOS on immunoprecipitated CaM complexes showed that DMA enhanced the association of eNOS with CaM in rat aortas. Both DMA and cariporide induced the reduction of intracellular pH (pHi) in bovine aortic endothelial cells (BAECs), which was accompanied by a sustained elevation of cytosolic Ca2+ ([Ca2+]i). This DMA-induced rise of [Ca2+]i was not affected by removing external Ca2+ from the buffer, but was abolished in thapsigargin-pretreated BAECs. These results suggest that lowering of pHi by NHE inhibitors in endothelial cells induces the mobilization of Ca2+ from the thapsigargin-sensitive stores of endoplasmic reticulum, which in turn stimulates NO production via the CaM-dependent activation of eNOS.

Modulation of thiopental-induced vascular relaxation and contraction by perivascular adipose tissue and endothelium

Thiopental induces relaxation of vascular smooth muscle cells through its direct and/or indirect vasodilator effects. The perivascular adipose tissue (PVAT) and the endothelium are known to attenuate vascular contraction, and we have recently reported that PVAT potentiates the relaxation effect of propofol through endothelium-dependent and -independent mechanisms. Here, we studied the mechanisms of thiopental-induced vascular responses in relation to the involvement of PVAT and endothelium. Thoracic aortic rings from male Wistar rats were prepared with or without PVAT (PVAT+ and PVAT-) and with an intact endothelium (E+) or with the endothelium removed (E-) for functional studies. The contraction and relaxation responses of these vessels to thiopental in the presence of agonists and various receptor antagonists and channel blockers were studied. In vessels pre-contracted with phenylephrine or KCl, thiopental-induced relaxation was highest in vessels denuded of both PVAT and the endothelium. PVAT attenuated the relaxation response to thiopental, and this attenuation effect was reduced by both angiotensin II (Ang II) type 1 receptor antagonists CV-11974 (2-n-butyl-4-choloro-5-hydroxymethyl-1-[2'-(1H-tetrazol-5-yl)biphenyl-methyl]-imidazole) or losartan and the angiotensin-converting enzyme inhibitor enalaprilat. Thiopental at high concentration (3 × 10(-3) M) caused a contraction through an endothelin-dependent mechanism. Thiopental induced relaxation in rat aorta through an endothelium-independent pathway and the presence of PVAT, endothelium, or both attenuated this relaxation response through Ang II-dependent and endothelin-dependent mechanisms, respectively.

The Novel Phenylpropiophenone Derivates Induced Relaxation of Isolated Rat Aorta

Our aim was to define how different chemical properties of newly developed phenylpropiophenone derivates (PhPds) influenced their potency and efficacy to relax rat aorta. A contribution of ion channels in the PhPds and propafenone mechanism of vasodilatation was tested. PhPds were syntethysed by substitution in the benzyl moiety with -F, -CH3 or -CF3 groups on the ortho or para position. The vasodilatation by PhPds was examined on the rings of rat aorta precontracted with phenylephrine. In order to test involvement of voltage-gated Na+ and K+ channels and L-type Ca2+ channels in a mechanism of action of PhPds, we used their blockers: lidocaine, nifedipine and 4-aminopiridine, respectively. Aorta was more sensitive to 5-ortho-trifluoromethyl derivate than to propafenone and other PhPds. The 5-para-methyl derivate had lower potency and efficacy than propafenone and other PhPds. Lidocaine did not influenced relaxation induced by PhPds, but slightly inhibited the effect of propafenone. The 4-aminopiridine only inhibited relaxation induced by 5-para-methyl derivate. Nifedipine inhibited relaxation of the rat aorta induced by 5-ortho-trifluoromethyl derivate and by propafenone. Introduction of 5-ortho-trifluoromethyl and 5-para-methyl group in the benzyl moiety of propafenone molecule changed its potency, efficacy and mechanism of action in the rat aorta. The 4-aminopiridine- and nifedipine sensitive ion channels are involved in mechanism of action of 5-para-methyl and 5-ortho-trifluoromethyl derivate. The introduction of other tested groups in the benzyl moiety does not affect pharmacological properties of the PhPds in relation to propafenone.

Carnosine analogues containing NO-donor substructures: Synthesis, physico-chemical characterization and preliminary pharmacological profile

The synthesis, physico-chemical, and biological characterisation of a short series of carnosine amides bearing NO-donor nitrooxy functionalities are described. The NO-donor carnosine analogues and their des-NO derivatives display carnosine-like properties, differing from the lead for their high serum stability. The newly-synthesised compounds are able to complex Cu2+ ions at physiological pH, displaying significant copper ion sequestering ability, and protect LDL from oxidation catalysed by Cu2+ ions. All products show moderately-potent HNE quenching activity. The NO-donor compounds 7c–f relaxed rat aorta strips via an NO-dependent mechanism. In vivo evaluation of organ protection in a model of cerebral ischaemia/reperfusion injury, using the selected NO-donor 7e and its des-NO analogue 7a, showed that both derivatives protect from hypoxia-induced brain damage, at lower concentrations than carnosine; 7e also decreased serum TNF-α levels. This class of NO-donor carnosine amides is worthy of further study as potential tools for treating a wide range of chronic vascular and neurodegenerative diseases in which NO-bioavailability is reduced.

IN VITRO TOLERANCE AFTER INCUBATION WITH NITROGLYCERIN (NTG) IS DUE TO INCREASED REACTIVE OXIGEN SPECIES IN THE ENDOTHELIAL CELLS

NTG may cause tolerance during long‐term administration, which is its major therapeutic limitation. However, the mechanisms underlying NTG‐induced tolerance and the contribution of endothelial cells (EC) remain unclear. We have verified that the endothelium modulates the vasodilatation induced by NTG in rat aorta. This work aimed to study the effects of nitric oxide (NO) and reactive oxygen species (ROS) on NTG‐induced relaxation in rat aorta and in isolated EC in order to investigate the cause of the in vitro tolerance. Relaxation curves for NTG were constructed in intact endothelium (E+) aortic rings. It was determined the efficacy (ME) and potency (pD2) of NTG. L‐NAME reduced the pD2 (from 8.70±0.13 to pD2 7.83±0.15) and ME from 102.7±2.7%, n=10 to 69.8± 3.2%, n=5 (p&lt;0.001). To study the tolerance in vitro, the E+ arteries were incubated for 5 min with NTG in the concentration that produced the ME: 100 μM. The ME and pD2 were reduced (from 101.9±1.6%; n=5 to 54.7±3.6%, n=5 p&lt;0.001; and from 8.88±0.20 to 7.63±0.04; n=5, p&lt;0.05), respectively. Following 5 min incubation, NTG enhanced the fluorescence intensity units (U) of superoxide (O2−) concentration by flow citometry (from basal 2.19±0.58U to 54.15±1.49U, p&lt;0.001). NO concentration was not changed in the EC. In conclusion, the in vitro tolerance is induced by 5 min incubation with NTG with increased production of superoxide in aortic EC. Supported by FAPESP and CNPq.

Gold Nanoparticle associated to Berberine induces vasorelaxation and cytosolic Ca2+ decrease in rat aorta

This study aimed to investigate the effect of berberine (BS) and modified surface of gold nanoparticles thioglycolic acid associated to berberine (AuNPTGA‐BS) on rat aortic vascular smooth muscle cells calcium ions concentration and aorta relaxation. Vascular reactivity was studied in denuded rat aortic rings and efficacy (ME) and potency (pD2) of BS in solution and in AuNPTGA and AuNPTGA‐BS were determined. Concentration‐effect curves for BS were constructed in phenylephrine (PE) pre‐contracted aorta. BS in solution, induced PE‐contraction reversal response with ME: 103.3 ± 4.2%, pD2: 5.47 ± 0.15 (n=6) and in AuNPTGA‐BS (ME: 81.9 ± 12.9%, pD2: 5.49 ± 0.25, n=5). The compound AuNP alone failed to induce rat aortic ring relaxation. The effects of the compounds on the cytosolic calcium ions concentration was measured by the fluorescence intensity decrease percentage (%ΔFI) of the calcium ions sensitive dye FLUO‐4AM by laser scanning confocal microscope. We have observed that the addition of BS decreased the fluorescence of the dye with (ΔIF: 39.5 ±5.4% (n=3) and for AuNPTGA‐BS (ΔIF= 65.0±4.7% n=3). On the other hand, AuNPTGA alone had no effect on the fluorescence emission. In conclusion, the vasorelaxation induced by BS and AuNPTGA‐BS is due to the cytosolic calcium ions concentration decrease. Supported by FAPDF, FAPESP, CAPES, CNPq, FINATEC and DPP‐UnB.

Betulinic acid inhibits superoxide anion-mediated impairment of endothelium-dependent relaxation in rat aortas

Objectives:To investigate the protective effect of betulinic acid (BA) on endothelium-dependent relaxation (EDR) in rat aortas exposed to pyrogallol-produced superoxide anion and its underlying mechanism.Materials and Methods:The thoracic aorta of male Sprague-Dawley rats was isolated to mount in the organ bath system and the effect of BA on acetylcholine (ACh)-induced EDR, nitric oxide (NO) level, reactive oxygen species (ROS) level, nitric oxide synthase (NOS) activity, and superoxide dismutase (SOD) activity of aortic rings exposed to pyrogallol (500 μM) for 15 min were measured.Results:BA evoked a concentration-dependent EDR in aortas, and pretreatment with EC50 (2.0 μM) concentration of BA markedly enhanced ACh-induced EDR of aortas exposed to pyrogallol-produced superoxide anion (Emax rose from 23.91 ± 5.41% to 42.45 ± 9.99%), which was markedly reversed by both Nw -nitro-L-arginine methyl ester hydrochloride (L-NAME) and methylene blue, but not by indomethacin. Moreover, BA significantly inhibited the increase of ROS level, as well as the decrease of NO level, the endothelial NOS (eNOS) activity, and the SOD activity in aortas induced by pyrogallol-derived superoxide anion.Conclusion:These results indicate that BA reduces the impairment of EDR in rat aortas exposed to exogenous superoxide anion, which may closely relate to the reduction of oxidative stress and activation of eNOS–NO pathway.

Triplatin, a platelet aggregation inhibitor from the salivary gland of the triatomine vector of Chagas disease, binds to TXA(2) but does not interact with glycoprotein PVI.

Salivary glands from haematophagous animals express a notable diversity of negative modulators of platelet function. Triplatin is an inhibitor of collagen-induced platelet aggregation which has been described as an antagonist of glycoprotein VI (GPVI). Because triplatin displays sequence homology to members of the lipocalin family of proteins, we investigated whether triplatin mechanism of action could be explained by interaction with pro-haemostatic prostaglandins. Our results demonstrate that triplatin inhibits platelet aggregation induced by low doses of collagen, thromboxane A2 (TXA(2)) mimetic (U46619), and arachidonic acid (AA). On the other hand, it does not inhibit platelet aggregation by convulxin, PMA, or low-dose ADP. Isothermal titration calorimetry (ITC) revealed that triplatin binds AA, cTXA(2), TXB(2), U46619 or prostaglandin (PG)H(2) mimetic (U51605). Consistent with its ligand specificity, triplatin induces relaxation of rat aorta contracted with U46619. Triplatin also interacts with PGF(2α) and PGJ(2), but not with leukotrienes, AA or biogenic amines. Surface plasmon resonance experiments failed to demonstrate interaction of triplatin with GPVI; it also did to inhibit platelet adhesion to fibrillar or soluble collagen. Because triplatin displays sequence similarity to apolipoprotein D (ApoD) - a lipocalin associated with high-density lipoprotein, ApoD was tested as a putative TXA(2)-binding molecule. ITC failed to demonstrate binding of ApoD to all prostanoids described above, or to AA. Furthermore, ApoD was devoid of inhibitory properties towards platelets activation by AA, collagen, or U46619. In conclusion, triplatin mechanism of action has been elucidated without ambiguity as a novel TXA(2)- and PGF(2α)- binding protein. It conceivably blocks platelet aggregation and vasoconstriction, thus contributing to successful blood feeding at the vector-host interface.

Iron Sucrose Promotes Endothelial Injury and Dysfunction and Monocyte Adhesion/Infiltration

Background/Aims: Intravenous (IV) iron preparations are widely used in the management of anemia in ESRD populations. Recent changes in reimbursement policy have dramatically increased the use of IV iron to lower the use of costly erythropoiesis-stimulating agents. These preparations are frequently administered with insufficient attention to the total body iron stores or presence of inflammation which is aggravated by excess iron. Endothelial injury and dysfunction are critical steps in atherosclerosis, thrombosis and cardiovascular disease. IV iron preparations raise plasma non-transferrin-bound iron which can promote oxidative stress, endothelial damage and dysfunction. We explored the effect of an IV iron preparation on endothelial cells, monocytes and isolated arteries. Methods: Primary cultures of human aortic endothelial cells (HAEC) were treated with pharmacologically relevant concentrations of iron sucrose (10–100 µg/ml) for 4–24 h. Endothelial cell morphology, viability, and monocyte adhesion were tested. Endothelial function was assessed by measuring the vasorelaxation response to acetylcholine in normal rat thoracic aorta rings preincubated with iron sucrose (200 µg/ml). Results: In contrast to the control HAEC which showed normal cobblestone appearance, cells treated with iron sucrose (50–100 µg/ml) for 4 h showed loss of normal morphological characteristics, cellular fragmentation, shrinkage, detachment, monolayer disruption and nuclear condensation/fragmentation features signifying apoptosis. HAEC exposure to iron sucrose (10–100 µg/ml) increased monocyte adhesion 5- to 25-fold. Incubation in media containing 200 µg/ml iron sucrose for 3 h caused marked reduction in the acetylcholine-mediated relaxation in phenylephrine-precontracted rat aorta. Conclusion: Pharmacologically relevant concentration of iron sucrose results in endothelial injury and dysfunction and marked increase in monocyte adhesion.

A new NO donor failed to release NO and to induce relaxation in the rat basilar artery

Nitric oxide (NO)-donors are pharmacologically active substances that in vivo or in vitro release NO. Their most common side effect is headache caused by cerebral vasodilatation. We previously demonstrated that the new NO-donor Ru(terpy)(bdq)NO] 3+ (Terpy), synthesized in our laboratory, induces relaxation of rat aorta. This study aimed to verify the effect of Terpy and sodium nitroprusside (SNP) in basilar artery. We conducted vascular reactivity experiments on endothelium-denuded basilar rings. The concentrations of iron (Fe) and ruthenium (Ru) complex were analyzed in basilar artery lysates after incubation with NO donors by mass spectrometry. We also evaluated the NO released from SNP and Terpy by using confocal microscopy. Interestingly, Terpy did not induce relaxation of the basilar artery. SNP induced relaxation in a concentration-dependent way. NO donors cross the membrane of vascular smooth muscle and entered the cell. In spite of its permeability, Terpy did not release NO in the basilar artery. Otherwise, SNP released NO in the basilar artery cells cytoplasm. Taken together, our results demonstrate that the new NO donor (Terpy) failed to release NO and to induce relaxation in the basilar artery. The NO donor SNP induces vascular relaxation due to NO release in the vascular smooth muscle cells.

Divergent mechanisms involved in CO and CORM-2 induced vasorelaxation

Carbon monoxide (CO) may play an important physiological role in regulation of the vascular tone. CO-releasing molecule (CORM-2) is frequently used as a CO-donor to evaluate (patho)physiological properties of CO and its potential therapeutic applications. The aim of this study was to examine the molecular mechanisms underlying the vasodilatory properties of CORM-2 as this has not yet been extensively explored. Isometric tension recordings were performed using mice and rat isolated aortic ring segments as well as mice femoral artery ring segments. Responses to CO (10μmol/l–300μmol/l) and CORM-2 (30μmol/l–600μmol/l) were evaluated in the presence/absence of activators/inhibitors of different molecular pathways. CO was unable to relax mice blood vessels, whereas it induced concentration-dependent relaxations in rat aorta. The response to CO was inhibited by both the soluble guanylyl cyclase (sGC) inhibitor ODQ (10μmol/l) and potassium (K+) channel blocker tetraethyl-ammonium chloride (3mmol/l). CORM-2 relaxed both mice and rat isolated blood vessels in a concentration-dependent manner, however this response was only partially blocked by ODQ and tetraethyl-ammonium chloride. Interestingly, 4-aminopyridine (3mmol/l) inhibited the CORM-2 induced vasodilatation whereas iberiotoxin (100nmol/l) had no influence. The molecular mechanisms underlying CORM-2 induced relaxation differ from those of CO-induced relaxation. While CO relaxes vessels through activation of sGC and/or calcium-activated K+-channels, CORM-2 exerts its vasodilatory properties only partially through sGC or K+-channels activation. CORM-2 induced vasodilatation seems to involve voltage-dependent rather than calcium-activated K+-channels.

Chronic ethanol consumption reduces adrenomedullin-induced relaxation in the isolated rat aorta

Adrenomedullin (AM) is a peptide that displays cardiovascular protective activity. We investigated the effects of chronic ethanol consumption on vascular reactivity to AM and the expression of AM system components in the rat aorta. Male Wistar rats were treated with ethanol (20% vol/vol) for 6 weeks. Vascular reactivity experiments were performed in the isolated rat aorta. Metalloproteinase-2 (MMP-2) levels were determined by gelatin zymography. Nitrite and nitrate generation was measured by chemiluminescence. Protein and mRNA levels of pre-pro-AM, calcitonin receptor-like receptor (CRLR) and RAMP1, 2, and 3 (receptor-activity-modifying proteins) were assessed by western blot and quantitative real-time polymerase chain reaction, respectively. Ethanol intake reduced AM-induced relaxation in endothelium-intact rat aortas, whereas calcitonin gene-related peptide-, acetylcholine-, and sodium nitroprusside-induced relaxation were not affected by ethanol intake. N G-nitro- l-arginine-methyl-ester ( l-NAME), 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one, and tetraethylammonium reduced AM-induced relaxation in aortic rings from both control and ethanol-treated rats. Ethanol consumption did not alter basal levels of nitrate and nitrite, nor did it affect the expression of MMP-2 in the rat aorta. Ethanol consumption increased mRNA levels of pre-pro-AM and RAMP1. Protein levels of AM, CRLR, and RAMP1, 2, and 3 were not affected by ethanol consumption. The major findings of the present study are that ethanol consumption reduces the vascular relaxation induced by AM and changes the mRNA expression of the components of the AM system in the vasculature. This response could be one of the mechanisms by which ethanol predisposes individuals to vascular dysfunction and hypertension.

Inhibition of vascular endothelial growth factor (VEGF)-induced endothelial proliferation, arterial relaxation, vascular permeability and angiogenesis by dobesilate

Vascular endothelial growth factor (VEGF) is a key factor in angiogenesis and vascular permeability which is associated with many pathological processes. 2,5-hydroxybenzene sulfonate (DHBS; dobesilate) is a small molecule with anti-angiogenic activity that has been described as an inhibitor of fibroblast growth factors (FGF). The aim of the present study was to evaluate the effects of DHBS on VEGF-induced actions. The effects of DHBS were evaluated on VEGF-induced proliferation in human umbilical vein endothelial cells (HUVEC) and rat aorta relaxation, as well as on in vivo VEGF-induced skin vascular permeability and neovascularization in rats. DHBS at 50 and 100 μM concentration significantly inhibited the proliferation of HUVEC induced by VEGF (10 ng/ml), without significantly affecting HUVEC proliferation in the absence of VEGF. Rapid VEGF-induced activation of Akt in HUVEC was also prevented by DHBS (100 μM). Additionally, DHBS (2 μM) specifically inhibited the relaxation of rat aorta induced by VEGF (0.1 to 30 ng/ml), but not endothelium-dependent relaxation to acetylcholine (1 nM to 10 μM). The in vivo enhancement of vascular permeability caused by VEGF injection (50 μl at 10 ng/ml) in rat skin was also inhibited by DHBS co-administration (200 μM) (74.8 ± 3.8% inhibition of dye extravasation). Administration of DHBS (200 mg/kg/day; i.p.) also reduced VEGF-induced angiogenesis in vivo. DHBS inhibits main responses elicited in vitro and in vivo by VEGF. As a dual antagonist of VEGF and FGF activities, DHBS could be of therapeutic interest in the treatment of diseases related to VEGF/FGF overproduction and excessive angiogenesis.

Alpha Lipoic Acid Treatment Improved Endothelium-dependent Relaxation in Diabetic Rat Aorta

The aim of this study was to ascertain the effects of α-lipoic acid (ALA) treatment on relaxant responses of acetylcholine (ACh) and isoprenaline (ISO) in aortic rings precontracted with serotonin (5-HT, 10(-6) M) obtained from streptozotocin (STZ)-induced diabetic rats. Diabetes was induced in the rats by 50 mg/kg streptozotocin (STZ) via an intraperitoneal injection. Rat body and aorta weights were measured. The isometric tension to ACh (10(-9)-3×10(-6) M) and ISO (10(-9)-10(-4) M) of 5-HT-precontracted diabetic and non-diabetic rat (control), diabetic-ALA-treated, and ALA-treated aortas, in organ baths were recorded. Six weeks after STZ treatment blood glucose was elevated compared to control rats. In aortic rings from diabetic rats ACh and ISO-induced relaxations were impaired whereas endothelium-independent relaxation to sodium nitroprusside (SNP) was unaffected. ALA (100 mg/kg/day) treatment for 5 weeks enhanced ACh and ISO-induced relaxation in diabetic aortas. This recovering effect was via NO because prevented by incubating the vessels with N(G)-nitro-L-arginine methyl ester (L-NAME, a NOS inhibitor). It may be assumed that ALA treatment in vivo, can protect against impaired vascular responsiveness in STZ-induced diabetic rats.

Contractions induced by phenylephrine and noradrenaline are differently affected by endothelium-dependent relaxation in rat aorta.

In rings of rat aorta precontracted with phenylephrine (10 microM) or noradrenaline (10 microM), addition of carbachol (10 microM) produced an endothelium-dependent relaxation. However, regardless of the concentration of agonist tested, both the intensity and duration of the relaxation were significantly less when noradrenaline, rather than phenylephrine, was used as the precontracting agent. The different responses observed do not appear to be related to destruction of endothelium-derived relaxing factor by autoxidation of noradrenaline since neither EDTA (30 microM) nor superoxide dismutase (30 units mL-1) improved the relaxation to carbachol. In addition, in endothelium-free rings, the noradrenaline (1 microM)-induced contraction was less sensitive than the phenylephrine (1 microM)-induced contraction to sodium nitroprusside (0.1 microM) or to 8-Br-cGMP (300 microM). With phenylephrine-, but not noradrenaline-, induced contraction, the relaxation triggered by carbachol was significantly reduced by pretreatment of the aortic rings with chloroethylclonidine (50 microM), which inactivates a subpopulation of alpha 1-adrenoceptors. Thus, the results confirm that both alkylation sensitive and resistant alpha 1-adrenoceptors exist in rat aorta and indicate that EDRF may discriminate between these two alpha 1-adrenoceptor subtypes which are differently affected by phenylephrine and noradrenaline.

Bakuchicin induces Vascular Relaxation via Endothelium‐dependent NO‐cGMP Signaling

Bakuchicin is a furanocoumarin derived from the seeds of Psoralea corylifolia. The aim of the present study was to investigate the effect of bakuchicin on vascular tone in rat aortic tissue. Bakuchicin induced a dose-dependent relaxation of phenylephrine-precontracted rat aorta which was abolished by removal of the endothelium. Pretreatment of the endothelium-intact aortic tissues with NG-nitro-L-arginine methylester (L-NAME) or 1H-[1,2,4]-oxadiazole-[4,3-α]-quinoxalin-1-one (ODQ) significantly inhibited the vascular relaxation induced by bakuchicin. Incubation with bakuchicin increased the production of cGMP in a concentration-dependent manner, and this effect was blocked by pretreatment with both L-NAME and ODQ. Vascular relaxation induced by bakuchicin was significantly inhibited by pretreatment with verapamil and diltiazem, but not by several other inhibitors including tetraethylammonium (TEA), glibenclamide, indomethacin, atropine or propranolol. These results suggested that bakuchicin-induced vasodilatation is closely associated with the endothelium-dependent nitric oxide (NO)/cGMP signaling pathway, with the possible involvement of L-type Ca(2+) channels.