Biomedical subjects

V B Schini-Kerth

Publications and source records attributed to V B Schini-Kerth.

At least 19 recordsLinked to original sources

PubMed2006

Endothelium-dependent vasodilator effect of Euterpe oleracea Mart. (Açaí) extracts in mesenteric vascular bed of the rat.

Açai (Euterpe oleracea Mart.) a fruit from the Amazon region, largely consumed in Brazil is rich in polyphenols. Experiments were undertaken to determine whether hydro-alcoholic extract obtained from stone of açaí induces a vasodilator effect in the rat mesenteric vascular bed precontracted with norepinephrine (NE) and, if so, to elucidate the underlying mechanism. Açai stone extract (ASE, 0.3-100 microg) induced a long-lasting endothelium-dependent vasodilation that was significantly reduced by N(G)-nitro-l-arginine methyl ester (l-NAME) and (1)H-[1,2,3] oxadiazolo [4,4-a] quinoxalin-l-one (ODQ) and abolished by KCl (45 mM) plus l-NAME. In vessels precontrated with NE and KCl (45 mM) or treated with K(Ca)(+2) channel blockers (charybdotoxin plus apamin), the effect of ASE was significantly reduced. However this effect is not affect by indomethacin, glybenclamide and 4-aminopiridine. Atropine, pyrilamine, yohimbine and HOE 140 significantly reduced the vasodilator effect of acetylcholine, histamine, clonidine and bradykinin, respectively, but did not change the vasodilator effect of ASE. In cultured endothelial cells ASE (100 microg/mL) induced the formation of NO that was reduced by N(G)-nitro-l-arginine (l-NA, 100 microM). The present study demonstrates that the vasodilator effect of ASE is dependent on activation of NO-cGMP pathway and may also involve endothelium-derived hyperpolarizing factor (EDHF) release. The vasodilator effect suggest a possibility to use ASE as a medicinal plant, in the treatment of cardiovascular diseases.

Animals↗

PubMed2006

Delphinidin and cyanidin inhibit PDGF(AB)-induced VEGF release in vascular smooth muscle cells by preventing activation of p38 MAPK and JNK.

BACKGROUND AND PURPOSE: Red wine polyphenols (RWPs) inhibit the expression of vascular endothelial growth factor (VEGF), a major pro-angiogenic and pro-atherosclerotic factor, in vascular smooth muscle cells (VSMCs). The aim of this study was to identify which red wine polyphenols were inhibitory and to determine the mechanism underlying the inhibitory effects. EXPERIMENTAL APPROACH: Release of VEGF stimulated by platelet derived growth factor(AB) (PDGF(AB)), from human aortic VSMCs was measured by immunoassay and phosphorylation of kinases by Western blot analysis. The direct antioxidant properties of polyphenols were determined by electron paramagnetic resonance and the cellular formation of reactive oxygen species (ROS) by dichlorofluorescein. KEY RESULTS: The inhibitory effect of RWPs on PDGF(AB)-induced release of VEGF was mimicked by delphinidin but not by quercetin, catechins, resveratrol, gallic acid or caffeic acid. In the anthocyanin class, not only delphinidin but also cyanidin prevented VEGF release whereas malvidin and peonidin were without effect. RWPs, delphinidin and cyanidin directly scavenged ROS and prevented the PDGF(AB)-induced formation of ROS in VSMCs. Malvidin and peonidin did not scavenge ROS but prevented the cellular formation of ROS. Although the p38 MAPK, ERK1/2 and JNK pathways have been involved in the PDGF(AB)-induced expression of VEGF, in our experiments, only phosphorylation of p38 MAPK and JNK was inhibited by RWPs, delphinidin and cyanidin. CONCLUSIONS AND IMPLICATIONS: Anthocyanins presenting a hydroxyl residue at position 3' are able to inhibit PDGF(AB)-induced VEGF expression by preventing activation of p38 MAPK and JNK in VSMCs.

Anthocyanins↗

PubMed2001

Sex steroids used in hormonal treatment increase vascular procoagulant activity by inducing thrombin receptor (PAR-1) expression: role of the glucocorticoid receptor.

BACKGROUND: The use of sex steroids in oral contraception or hormonal replacement therapy is associated with an increased risk of cardiovascular thromboembolic complications. Although both the estrogen and the progestin components have been involved, the underlying mechanisms responsible are unclear. METHODS AND RESULTS: This study examined whether sex steroids promote hemostasis indirectly by increasing the procoagulant activity of blood vessels. Treatment of vascular smooth muscle cells with several progestins (progesterone, 3-keto-desogestrel, gestodene, and medroxyprogesterone acetate) upregulated proteolytically activatable thrombin receptor (PAR-1) expression, resulting in a potentiated thrombin-induced tissue factor expression and surface procoagulant activity. In contrast, neither the progestins levonorgestrel, norethisterone, and norgestimate nor the synthetic estrogen 17alpha-ethinylestradiol had such effects. The effect of the stimulatory progestins, which induce glucocorticoid-like effects in several cell systems, was mimicked by dexamethasone and inhibited by the progesterone and glucocorticoid receptor antagonist RU-38486. In addition, long-term administration of progesterone, 3-keto-desogestrel, or medroxyprogesterone acetate to ovariectomized rats increased PAR-1 protein level in the arterial wall, resulting in an increased responsiveness of isolated aortic rings to thrombin. CONCLUSIONS: These data demonstrate that several progestins markedly potentiate the vascular procoagulant effects of thrombin by increasing the availability of membrane thrombin receptors in the smooth muscle, an effect that is most likely due to their glucocorticoid-like activity.

Animals↗

PubMed2001

Thrombin activates the hypoxia-inducible factor-1 signaling pathway in vascular smooth muscle cells: Role of the p22(phox)-containing NADPH oxidase.

The heterodimeric transcription factor hypoxia-inducible factor-1 (HIF-1) is activated under hypoxic conditions, resulting in the upregulation of its target genes plasminogen activator inhibitor-1 (PAI-1) and vascular endothelial growth factor (VEGF). PAI-1 and VEGF are also induced in response to vascular injury, which is characterized by the activation of platelets and the coagulation cascade as well as the generation of reactive oxygen species (ROS). However, it is not known whether HIF-1 is also stimulated by thrombotic factors. We investigated the role of thrombin, platelet-associated growth factors, and ROS derived from the p22(phox)-containing NADPH oxidase in the activation of HIF-1 and the induction of its target genes PAI-1 and VEGF in human vascular smooth muscle cells (VSMCs). Thrombin, platelet-derived growth factor-AB (PDGF-AB), and transforming growth factor-beta(1) (TGF-beta(1)) upregulated HIF-1alpha protein in cultured and native VSMCs. This response was accompanied by nuclear accumulation of HIF-1alpha as well as by increased HIF-1 DNA-binding and reporter gene activity. The thrombin-induced expression of HIF-1alpha, PAI-1, and VEGF was attenuated by antioxidant treatment as well as by transfection of p22(phox) antisense oligonucleotides. Inhibition of p38 mitogen-activated protein kinase and phosphatidylinositol-3-kinase significantly decreased thrombin-induced HIF-1alpha, PAI-1, and VEGF expression. These findings demonstrate that the HIF-1 signaling pathway can be stimulated by thrombin and platelet-associated growth factors and that a redox-sensitive cascade activated by ROS derived from the p22(phox)-containing NADPH oxidase is crucially involved in this response.

Antioxidants↗

PubMed2001

Thrombin causes vascular endothelial growth factor expression in vascular smooth muscle cells: role of reactive oxygen species.

Vascular endothelial growth factor (VEGF) has been implicated in the reendothelialization of the vascular wall after balloon injury. This study investigated whether thrombin, which is formed during activation of the coagulation cascade at sites of vascular injury, upregulates VEGF expression in vascular smooth muscle cells (VSMCs). VEGF expression was assessed in native and cultured VSMCs by Northern blot analysis and reverse transcription-polymerase chain reaction and the release of VEGF protein by immunoassay. alpha-Thrombin time- and concentration-dependently increased VEGF mRNA levels, mainly that mRNA coding for the soluble splice variant VEGF(164/165), and stimulated the release of VEGF protein. These effects required the proteolytic activity of thrombin and were mimicked by a thrombin receptor activating-peptide. Upregulation of VEGF expression was also induced by conditioned medium from alpha-thrombin-stimulated VSMCs. Both the early and the delayed alpha-thrombin-induced VEGF expressions were attenuated by antioxidants and by diphenyleneiodonium. alpha-Thrombin-induced VEGF release was significantly reduced by a platelet-derived growth factor (PDGF)-, a transforming growth factor (TGF)-beta-, and a basic fibroblast growth factor (bFGF)-neutralizing antibody. Thrombin caused a redox-sensitive upregulation of expression of VEGF in VSMCs through a direct and an indirect effect, which was dependent on the endogenous formation of PDGF, TGF-beta, and bFGF. Upregulation of VEGF expression may represent an important mechanism by which the coagulation cascade contributes to the regeneration of the endothelial lining at sites of balloon injury.

Acetylcysteine↗

PubMed2000

The progestin levonorgestrel induces endothelium-independent relaxation of rabbit jugular vein via inhibition of calcium entry and protein kinase C: role of cyclic AMP.

The progestin and oestrogen component of oral contraceptives have been involved in the development of venous thromboembolic events in women. In the present study we determined the vasoactive effects of sex steroids used in oral contraceptives in isolated preconstricted rabbit jugular veins in the presence of diclofenac and examined the underlying mechanisms. The natural hormone progesterone, the synthetic progestins levonorgestrel, 3-keto-desogestrel, gestodene and chlormadinone acetate, and the synthetic estrogen 17 alpha-ethinyloestradiol induced concentration-dependent relaxations of endothelium-intact veins constricted with U46619. Levonorgestrel also inhibited constrictions evoked by either a high potassium (K(+)) solution or phorbol myristate acetate (PMA) in the absence and presence of extracellular calcium (Ca(2+)). In addition, levonorgestrel depressed contractions evoked by Ca(2+) and reduced (45)Ca(2+) influx in depolarized veins. Relaxations to levonorgestrel in U46619-constricted veins were neither affected by the presence of the endothelium nor by the inhibitor of soluble guanylyl cyclase, NS2028, but were significantly improved either by the selective cyclic AMP phosphodiesterase inhibitor rolipram or in the absence of diclofenac, and decreased by the protein kinase A inhibitor, Rp-8-CPT-cAMPS. Rolipram also potentiated relaxations to levonorgestrel in PMA-constricted veins in the presence, but not in the absence of extracellular Ca(2+). Levonorgestrel increased levels of cyclic AMP and inhibited PMA-induced activation of protein kinase C in veins. These findings indicate that levonorgestrel caused endothelium-independent relaxations of jugular veins via inhibition of Ca(2+) entry and of protein kinase C activation. In addition, the cyclic AMP effector pathway contributes to the levonorgestrel-induced relaxation possibly by depressing Ca(2+) entry.

15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5↗

PubMed2000

Oxidative stress and expression of p22phox are involved in the up-regulation of tissue factor in vascular smooth muscle cells in response to activated platelets.

Vascular injury after balloon angioplasty results in the rapid activation of platelets leading to the release of growth factors and vasoactive substances. In addition, up-regulation of tissue factor (TF) and an increased production of reactive oxygen species (ROS) have been detected at sites of vascular injury. We investigated whether platelet-derived products (PDP) released from activated human platelets increase ROS production, resulting in the induction of TF expression in vascular smooth muscle cells (SMC). PDP induced a time- and concentration-dependent increase in ROS generation in cultured SMC that was mediated mainly by PDGF-AB and TGF-beta1 and impaired by the flavin inhibitor diphenylene iodonium. Increased ROS formation was associated with enhanced mRNA levels of the small NAD(P)H oxidase subunit p22phox or its smooth muscle isoform. Transient transfection with a p22phox antisense vector decreased PDP-induced ROS generation. PDP up-regulated TF mRNA expression, which was redox sensitive and reduced by transfection of the p22phox antisense vector. In addition, PDP-stimulated reporter gene activity of two TF promoter constructs was decreased by coexpression of the p22phox antisense vector. These results indicate that activated platelets up-regulate TF expression and that this response involves ROS generation and a p22phox-containing NAD(P)H oxidase in SMC.

Animals↗

PubMed1999

Aggregating human platelets stimulate expression of vascular endothelial growth factor in cultured vascular smooth muscle cells through a synergistic effect of transforming growth factor-beta(1) and platelet-derived growth factor(AB).

BACKGROUND: Vascular endothelial growth factor (VEGF), an endothelial mitogen and chemoattractant, has been implicated in the recovery of the endothelium after balloon injury. The increased expression of VEGF in vascular smooth muscle cells (SMC) at sites of injury suggests that this cell type may be a major cellular source of VEGF. This study examined whether aggregating platelets stimulate VEGF expression in cultured SMC. METHODS AND RESULTS: ++VEGF expression in SMC was assessed by Northern blot analysis and by reverse transcription followed by polymerase chain reaction and the release of VEGF by Western blot analysis and immunoassay. Platelet-derived products (PDP) released by aggregating human platelets time-dependently and concentration-dependently enhanced VEGF mRNA levels, mainly that coding for the soluble splice variant VEGF(165/164), and stimulated the release of VEGF protein. These effects were potentiated by transient acidification of PDP, which release bioactive transforming growth factor (TGF)-beta(1), and mimicked by platelet-derived growth factor (PDGF)(AB) and TGF-beta(1) in a synergistic manner. Both a TGF-beta-neutralizing antibody and a PDGF-neutralizing antibody significantly attenuated the effect of acidified PDP on VEGF production. CONCLUSIONS: Aggregating human platelets induce VEGF mRNA expression in cultured SMC and the subsequent release of VEGF protein. This effect can be attributed to a supra-additive action of PDGF(AB) and TGF-beta(1) and may represent a novel mechanism by which platelets contribute to the recovery of the endothelial lining at sites of balloon-injured arteries.

Angioplasty, Balloon↗

PubMed1999

Proinflammatory mediators chronically downregulate the formation of the endothelium-derived hyperpolarizing factor in arteries via a nitric oxide/cyclic GMP-dependent mechanism.

BACKGROUND: Endothelium-dependent dilator responses mediated by NO and endothelium-derived hyperpolarizing factor (EDHF) are altered in arteriosclerosis and sepsis. The possibility that proinflammatory mediators that stimulate the expression of inducible NO synthase (NOS II) affect the generation of EDHF was examined in isolated arteries. METHODS AND RESULTS: Under combined blockade of NOS and cyclooxygenase, EDHF-mediated relaxation elicited by several agonists was significantly attenuated in rabbit carotid and porcine coronary arteries exposed to cytokines and lipopolysaccharide. The blunted relaxation was coincident with NOS II expression and was prevented by inhibition of NOS II as well as of global protein synthesis. The NO donor CAS 1609 and 8-bromo-cGMP mimicked the proinflammatory mediator effect. In contrast, long-term blockade of endothelial NO generation increased the relaxation in carotid but not in coronary arteries. Proinflammatory mediators reduced the synthesis of EDHF assessed as hyperpolarization of vascular smooth muscle cells elicited by the effluent from bradykinin-stimulated coronary arteries. Proinflammatory mediators induced NOS II expression in cultured endothelial cells and decreased the expression of cytochrome P450 enzymes, which are the most probable candidates for the synthesis of EDHF. CONCLUSIONS: Proinflammatory mediators inhibit the formation of EDHF in isolated arteries. This impairment is coincident with NOS II expression in the arterial wall and seems to be mediated through the induced generation of NO, which downregulates the putative EDHF-forming enzyme. Thus, a decreased formation of EDHF may contribute to the endothelial dysfunction in arteriosclerosis and sepsis.

Animals↗

PubMed1999

Ginsenoside Rg3 mediates endothelium-dependent relaxation in response to ginsenosides in rat aorta: role of K+ channels.

The aim of the present study was to characterize the endothelium-dependent relaxation elicited by ginsenosides, a mixture of saponin extracted from Panax ginseng, in isolated rat aorta. Relaxations elicited by ginsenosides were mimicked by ginsenoside Rg1 and ginsenoside Rg1, two major ginsenosides of the protopanaxatriol group. Ginsenoside Rg3 was about 100-fold more potent than ginsenoside Rg1. The endothelium-dependent relaxation in response to ginsenoside Rg3 was associated with the formation of cycle GMP. These effects were abolished by N(G)-nitro-L-arginine and methylene blue. Relaxations in response to ginsenoside Rg3 were unaffected by atropine, diphenhydramine, [D-Pro2, D-Trp7,9]substance P, propranolol, nifedipine, verapamil and glibenclamide but were markedly reduced by tetraethylammonium. Tetraethylammonium modestly reduced the relaxation induced by sodium nitroprusside. These findings indicate that ginsenoside Rg3 is a major mediator of the endothelium-dependent nitric oxide-mediated relaxation in response to ginsenosides in isolated rat aorta, possibly via activation of tetraethylammonium-sensitive K+ channels.

Adrenergic beta-Antagonists↗

PubMed1999

The ginsenoside Rg3 evokes endothelium-independent relaxation in rat aortic rings: role of K+ channels.

The purpose of the present study was to characterize the mechanism underlying the direct relaxing activity of ginsenosides on vascular smooth muscle. The total ginsenoside mixture, ginsenosides from either the protopanaxadiol group or the protopanaxatriol group, and the ginsenoside Rg3 from the protopanaxatriol group caused a concentration-dependent relaxation of rat aortic rings without endothelium contracted with 25 x 10(-3) M KCl but affected only minimally those contracted with 60 x 10(-3) M KCl. Ginsenoside Rg3 was the most potent relaxing agonist. Relaxations elicited by ginsenoside Rg3 were markedly reduced by tetraethylammonium, a blocker of non-selective K+ channels, but not by glibenclamide, a blocker of ATP-sensitive K+ channels. Ginsenoside Rg3 significantly inhibited Ca2+-induced concentration-contraction curves and the 45Ca2+ influx in aortic rings incubated with 25 x 10(-3) M KCl whereas these responses were not affected in rings incubated with 60 x 10(-3) M KCl. Ginsenoside Rg3 caused a time- and concentration-dependent efflux of 86Rb from aortic rings that was inhibited by tetraethylammonium but not by glibenclamide. These findings indicate that ginsenoside Rg3 is a potent inhibitor of vascular smooth muscle tone and that this effect seems to be due to an inhibition of Ca2+ influx and stimulation of K+ efflux, possibly via activation of tetraethylammonium-sensitive K+ channels.

Animals↗

PubMed1999

Interleukin-1beta-induced nitric oxide synthesis in aortic rings from normal and hyperinsulinaemic rats: effect of physical exercise.

Insulin has been suggested to prevent the induction of nitric oxide synthase (NOS) in vitro in arterial smooth muscle, but whether such a mechanism is operative in vivo is not known. Therefore, we evaluated the sensitivity of smooth muscle NOS to induction by interleukin-1beta (IL-1beta) in aortic rings of lean and obese Zucker rats, a model of experimental hyperinsulinaemia. In order to modulate the insulin and glucose balance of the rats, a 22-week-long treadmill exercise was included in the study. The training attenuated weight gain and reduced blood glucose in the obese and lean rats, whereas the abnormally high plasma insulin of the obese rats remained unaffected. A 6-h incubation of aortic rings with IL-1beta (10 ng/ml) increased cyclic GMP in smooth muscle by approximately threefold in all groups, and this effect was prevented by methylene blue. The contractile sensitivity of endothelium-denuded aortic rings to phenylephrine was reduced by incubation with IL-1beta (1 ng/ml and 10 ng/ml) in the exercised obese and lean rats, whereas no significant change was observed in the sedentary groups. The aortic maximal contractile force induced by phenylephrine was reduced in sedentary and exercised obese rats by incubation with IL-1beta, while no change was detected in the lean rats. The aortic relaxation to exogenous L-arginine was augmented by IL-1beta in all groups, while the relaxation sensitivity to L-arginine after induction by IL-1beta was enhanced by exercise in the obese but not in the lean rats. Finally, the relaxation to nitroprusside was not significantly affected by IL-1beta in any of the study groups. In conclusion, since maximal contractile force generation to phenylephrine was reduced by IL-1beta in the obese but not in the lean rats, the sensitivity of NOS to induction by IL-1beta was higher in arterial smooth muscle of the obese than the lean Zucker rats. Thus, this model of hyperinsulinaemia was not associated with reduced sensitivity of smooth muscle NOS to induction by IL-1beta. Regular exercise did not change plasma insulin concentrations, but it enhanced the action of insulin in both strains as reflected by reduced blood glucose, and increased the sensitivity of smooth muscle NOS to induction by IL-1beta.

Animals↗

PubMed1999

Vascular endothelial growth factor up-regulates nitric oxide synthase expression in endothelial cells.

OBJECTIVE: Vascular endothelial growth factor (VEGF), secreted by vascular cells and a variety of tumour cells, is a potent angiogenic factor. Since nitric oxide (NO) seems to play a key role in the VEGF-induced proliferation of endothelial cells, the aim of the present study was to determine whether VEGF stimulates endothelial NO synthase (eNOS) expression and hence results in a maintained increase in NO formation. METHODS: Experiments were performed using cultured human umbilical vein endothelial cells (HUVEC) and isolated rat aortic rings. eNOS expression was assessed by Western blotting and RT-PCR analysis. RESULTS: Exposure of either confluent HUVEC or rat aortic rings to VEGF165 significantly increased eNOS mRNA and protein levels. This stimulatory effect of VEGF165 on eNOS expression was associated with an elevation in the basal production of cGMP in HUVEC, and with a leftward shift of the concentration-relaxation curve to acetylcholine in aortic rings. The VEGF-induced increase in eNOS mRNA levels was abolished by tyrosine kinase inhibitors suggesting involvement of a tyrosine kinase-dependent pathway. Since eNOS mRNA levels remained elevated in VEGF-treated cells in the presence of actinomycin D. it is likely that the VEGF-induced up-regulation of eNOS expression may be a consequence of a post-transcriptional effect on eNOS mRNA stability. CONCLUSION: The results demonstrate that VEGF enhances the expression of eNOS in native and cultured endothelial cells, an effect which may be important in the process of VEGF-induced angiogenesis.

Analysis of Variance↗

PubMed1999

Vascular biosynthesis of nitric oxide: effect on hemostasis and fibrinolysis.

Nitric oxide (NO) is a multifunctional effector molecule that plays a central role in the regulation of vascular homeostasis. NO is synthesized from L-arginine by a family of enzymes called NO synthases. The principal source of NO in the vascular system of healthy mammals is the constitutively expressed NO synthase in endothelial cells. The basal endothelial formation of NO can be increased by receptor-dependent agonists (i.e., bradykinin) in a calcium-calmodulin-dependent manner, and also by physical forces (i.e., shear stress), predominantly without changes in the intracellular concentration of free calcium. Nitric oxide can diffuse toward the blood vessel wall where the major target is the smooth muscle cell. NO regulates vascular tone, and the free radical is also a potent inhibitor of smooth muscle cell proliferation, migration and synthesis of extracellular matrix proteins. NO can also diffuse toward the lumen of the blood vessel where it helps maintain blood fluidity. NO inhibits platelets' and leucocytes' adhesion to endothelial cells. In addition, NO inhibits platelet aggregation and facilitates the dissolution of small platelet aggregates. However, the regulatory action of NO on blood cells is most likely limited to the luminal surface of endothelial cells since NO is rapidly scavenged by hemoglobin in erythrocytes and inactivated by oxygen-derived radicals such as superoxide anions. NO can also affect the fibrinolytic activity by regulating the release of tissue-type plasminogen activator and plasminogen activator inhibitor-1. The crucial role of vascular NO in the control of blood fluidity has been demonstrated by the regulation of the bleeding time in humans.

Animals↗

PubMed1999

Growth-inhibitory effect of cyclic GMP- and cyclic AMP-dependent vasodilators on rat vascular smooth muscle cells: effect on cell cycle and cyclin expression.

1. The possibility that the antiproliferative effect of cyclic GMP- and cyclic AMP-dependent vasodilators involves an impaired progression of vascular smooth muscle cells (VSMC) through the cell cycle and expression of cyclins, which in association with the cyclin-dependent kinases control the transition between the distinct phases of the cell cycle, was examined. 2. FCS (10%) stimulated the transition of quiescent VSMC from the G0/G1 to the S phase (maximum within 18-24 h and then to the G2/M phase (maximum within 22-28 h). Sodium nitroprusside and 8-Br-cyclic GMP, as well as forskolin and 8-Br-cyclic AMP markedly reduced the percentage of cells in the S phase after FCS stimulation. 3. FCS stimulated the low basal protein expression of cyclin D1 (maximum within 8-24 h) and E (maximum within 8-38 h) and of cyclin A (maximum within 14-30 h). The stimulatory effect of FCS on cyclin D1 and A expression was inhibited, but that of cyclin E was only minimally affected by the vasodilators. 4. FCS increased the low basal level of cyclin D1 mRNA after a lag phase of 2 h and that of cyclin A after 12 h. The vasodilators significantly reduced the FCS-stimulated expression of cyclin D1 and A mRNA. 5. These findings indicate that cyclic GMP- and cyclic AMP-dependent vasodilators inhibit the proliferation of VSMC by preventing the progression of the cell cycle from the G0/G1 into the S phase, an effect which can be attributed to the impaired expression of cyclin D1 and A.

Animals↗

PubMed1999

Vascular endothelial growth factor activates nuclear factor-kappaB and induces monocyte chemoattractant protein-1 in bovine retinal endothelial cells.

Vascular endothelial growth factor (VEGF) has been suggested to play a role in the pathogenesis of diabetic vascular complications. In the present study, we investigated whether expression of monocyte chemoattractant protein-1 (MCP-1), a chemokine that has been proposed to recruit leukocytes to sites of inflammation, neovascularization, and vascular injury, can be modulated by VEGF in bovine retinal microvascular endothelial cells (BRECs). VEGF induced expression of MCP-1 mRNA in BRECs in a concentration- and time-dependent manner. Secretion of MCP-1 into the culture medium of BRECs treated with VEGF for 24 h was increased by 2.2-fold compared with the control. Inhibitors of transcription factor NF-kappaB, N-alpha-tosyl-L-lysine chloromethylketone (TLCK) and N-acetylcysteine (NAC), as well as an inhibitor of the extracellular signal-regulated kinase (ERK) pathway, PD 98059, attenuated VEGF-induced expression of MCP-1 mRNA. Using electrophoretic gel mobility shift assay, we observed that VEGF stimulated binding activity of NF-kappaB. VEGF-induced NF-kappaB activation was inhibited by TLCK and NAC, but not by PD 98059. Binding activity of transcription factor AP-1, which is suggested to regulate induction of the MCP-1 gene together with NF-kappaB, was also stimulated by VEGF. PD 98059 inhibited the VEGF-induced activation of AP-1. These results indicate that VEGF induces MCP-1 expression in BRECs most likely by activating NF-kappaB and AP-1 via ERK-independent and -dependent pathways. Activation of NF-kappaB and induction of MCP-1 by VEGF in microvascular endothelial cells may contribute to the development of diabetic vascular complications.

Acetylcysteine↗

PubMed1999

Dual effects of insulin-like growth factor-I on the constitutive and inducible nitric oxide (NO) synthase-dependent formation of NO in vascular cells.

The free radical nitric oxide (NO) has emerged as an important signal effector molecule that controls critical functions in the mammalian cardiovascular system in both healthy and diseased states. In normal blood vessels, NO is synthesized from L-arginine by a constitutively expressed NO synthase (NOS III) in endothelial cells. The endothelial formation of NO can be increased by both pharmacological and physiological agonists (e.g. bradykinin) and this effect is dependent on the interaction of calcium/calmodulin with NOS III. Recent observations suggest that NO is a pivotal mediator of insulin-like growth factor (IGF)-I-induced vasodilatation in humans and experimental animals. Administration of IGF-I in the human brachial artery increased blood flow in the forearm, an effect which was abolished by N(G)-monomethyl-L-arginine (L-NMMA), an inhibitor of NOS. Inhibitors of NOS also prevented the IGF-I-induced renal vasodilatation in anesthetized rats and in in vitro blood-perfused preparations of rat juxtamedullary nephrons. In addition, IGF-I caused concentration-dependent relaxation of precontracted isolated arteries that required the presence of a functional endothelium, and this effect was abolished by L-NMMA. Moreover, the rapid formation of NO in response to IGF-I is detected in cultured endothelial cells by an amperometric NO sensor. The signalling of NO formation is independent of changes in intracellular Ca2+ and involves tyrosine kinase and phosphatidylinositol-3-kinase. At sites of vascular injury, proinflammatory mediators can stimulate the expression of an inducible NOS (NOS II) that generates large amounts of NO for prolonged periods of time. IGF-I has been shown to inhibit the interleukin-1beta-induced formation of NO in cultured vascular smooth muscle cells by preventing the induction of NOS II. In conclusion, IGF-I may be an important regulator of vascular tone in part by modulating the formation of NO by NOS III and NOS II in the vascular wall.

Animals↗

PubMed1998

Thrombin receptor expression is increased by angiotensin II in cultured and native vascular smooth muscle cells.

OBJECTIVE: The factors involved in restenosis after balloon angioplasty are poorly characterized but the local concentration of the potent mitogens angiotensin II (AII) and thrombin is known to be increased at sites of vascular injury. We investigated the possibility of a synergistic interaction between AII and thrombin by studying the effects of AII on the expression of the thrombin receptor in rat aortic smooth muscle cells (VMSC). METHODS: Thrombin receptor mRNA expression was studied by Northern blot analysis and RT-PCR using total RNA extracted from VMSC or from endothelium-denuded rat aortae. As a measure of thrombin receptor protein expression, we assessed either the thrombin-stimulated release of 6-keto-prostaglandin F1 alpha from VMSC or the contraction of endothelium-denuded rat aortic rings. RESULTS: The thrombin receptor mRNA was expressed at a low level in both cultured and native VMSC. AII concentration- and time-dependently increased expression of thrombin receptor mRNA in VMSC and augmented the thrombin-induced release of 6-keto-prostaglandin F1 alpha as well as the thrombin induced contraction. Blockade of the angiotensin subtype 1 (AT1) receptor by EXP3174 or D8731 prevented the AII-mediated increase in thrombin receptor expression. The effect of AII on the increase in thrombin receptor mRNA expression was enhanced by the protein kinase C inhibitor Ro 31-8220, but was unaffected by prolonged incubation with phorbol myristate acetate or the tyrosine kinase inhibitors genistein and erbstatin A. CONCLUSION: These results demonstrate that AII enhances the expression of thrombin receptor in cultured and native VMSC. In cultured cells, this effect is mediated by the activation of the AT1 receptor subtype. This synergistic effect between AII and the thrombin receptor may promote the extensive proliferation of smooth muscle cells in response to vascular injury.

6-Ketoprostaglandin F1 alpha↗