U. Deniz Dincer

Impaired capsaicin-induced relaxation of coronary arteries in a porcine model of the metabolic syndrome

Recent studies implicate channels of the transient receptor potential vanilloid family (e.g., TRPV1) in regulating vascular tone; however, little is known about these channels in the coronary circulation. Furthermore, it is unclear whether metabolic syndrome alters the function and/or expression of TRPV1. We tested the hypothesis that TRPV1 mediates coronary vasodilation through endothelium-dependent mechanisms that are impaired by the metabolic syndrome. Studies were conducted on coronary arteries from lean and obese male Ossabaw miniature swine. In lean pigs, capsaicin, a TRPV1 agonist, relaxed arteries in a dose-dependent manner (EC50 = 116 +/- 41 nM). Capsaicin-induced relaxation was blocked by the TRPV1 antagonist capsazepine, endothelial denudation, inhibition of nitric oxide synthase, and K+ channel antagonists. Capsaicin-induced relaxation was impaired in rings from pigs with metabolic syndrome (91 +/- 4% vs. 51 +/- 10% relaxation at 100 microM). TRPV1 immunoreactivity was prominent in coronary endothelial cells. TRPV1 protein expression was decreased 40 +/- 11% in obese pigs. Capsaicin (100 microM) elicited divalent cation influx that was abolished in endothelial cells from obese pigs. These data indicate that TRPV1 channels are functionally expressed in the coronary circulation and mediate endothelium-dependent vasodilation through a mechanism involving nitric oxide and K+ channels. Impaired capsaicin-induced vasodilation in the metabolic syndrome is associated with decreased expression of TRPV1 and cation influx.

Impaired function of coronary BKCa channels in metabolic syndrome

The role of large-conductance Ca(2+)-activated K(+) (BK(Ca)) channels in regulation of coronary microvascular function is widely appreciated, but molecular and functional changes underlying the deleterious influence of metabolic syndrome (MetS) have not been determined. Male Ossabaw miniature swine consumed for 3-6 mo a normal diet (11% kcal from fat) or an excess-calorie atherogenic diet that induces MetS (45% kcal from fat, 2% cholesterol, 20% kcal from fructose). MetS significantly impaired coronary vasodilation to the BK(Ca) opener NS-1619 in vivo (30-100 microg) and reduced the contribution of these channels to adenosine-induced microvascular vasodilation in vitro (1-100 microM). MetS reduced whole cell penitrem A (1 microM)-sensitive K(+) current and NS-1619-activated (10 microM) current in isolated coronary vascular smooth muscle cells. MetS increased the concentration of free intracellular Ca(2+) and augmented coronary vasoconstriction to the L-type Ca(2+) channel agonist BAY K 8644 (10 pM-10 nM). BK(Ca) channel alpha and beta(1) protein expression was increased in coronary arteries from MetS swine. Coronary vascular dysfunction in MetS is related to impaired BK(Ca) channel function and is accompanied by significant increases in L-type Ca(2+) channel-mediated coronary vasoconstriction.

Mechanisms of Coronary Dysfunction in Obesity and Insulin Resistance

ABSTRACT

Periadventitial adipose tissue impairs coronary endothelial function via PKC-β-dependent phosphorylation of nitric oxide synthase

Endogenous periadventitial adipose-derived factors have been shown to contribute to coronary vascular regulation by impairing endothelial function through a direct inhibition of endothelial nitric oxide synthase (eNOS). However, our understanding of the underlying mechanisms remains uncertain. Accordingly, this study was designed to test the hypothesis that periadventitial adipose tissue releases agents that attenuate coronary endothelial nitric oxide production via a protein kinase C (PKC)-beta-dependent mechanism. Isometric tension studies were conducted on isolated canine circumflex coronary arteries with and without natural amounts of periadventitial adipose tissue. Adipose tissue significantly diminished coronary endothelial-dependent vasodilation and nitric oxide production in response to bradykinin and acetylcholine. The selective inhibition of endothelial PKC-beta with ruboxistaurin (1 microM) abolished the adipose-induced impairment of bradykinin-mediated coronary vasodilation and the endothelial production of nitric oxide. Western blot analysis revealed a significant increase in eNOS phosphorylation at the inhibitory residue Thr(495) in arteries exposed to periadventitial adipose tissue. This site-specific phosphorylation of eNOS was prevented by the inhibition of PKC-beta. These data demonstrate that periadventitial adipose-derived factors impair coronary endothelial nitric oxide production via a PKC-beta-dependent, site-specific phosphorylation of eNOS at Thr(495).

Voltage-dependent K+ channels regulate the duration of reactive hyperemia in the canine coronary circulation

We previously demonstrated a role for voltage-dependent K(+) (K(V)) channels in coronary vasodilation elicited by myocardial metabolism and exogenous H(2)O(2), as responses were attenuated by the K(V) channel blocker 4-aminopyridine (4-AP). Here we tested the hypothesis that K(V) channels participate in coronary reactive hyperemia and examined the role of K(V) channels in responses to nitric oxide (NO) and adenosine, two putative mediators. Reactive hyperemia (30-s occlusion) was measured in open-chest dogs before and during 4-AP treatment [intracoronary (ic), plasma concentration 0.3 mM]. 4-AP reduced baseline flow 34 +/- 5% and inhibited hyperemic volume 32 +/- 5%. Administration of 8-phenyltheophylline (8-PT; 0.3 mM ic or 5 mg/kg iv) or N(G)-nitro-L-arginine methyl ester (L-NAME; 1 mg/min ic) inhibited early and late portions of hyperemic flow, supporting roles for adenosine and NO. 4-AP further inhibited hyperemia in the presence of 8-PT or L-NAME. Adenosine-induced blood flow responses were attenuated by 4-AP (52 +/- 6% block at 9 microg/min). Dilation of arterioles to adenosine was attenuated by 0.3 mM 4-AP and 1 microM correolide, a selective K(V)1 antagonist (76 +/- 7% and 47 +/- 2% block, respectively, at 1 microM). Dilation in response to sodium nitroprusside, an NO donor, was attenuated by 4-AP in vivo (41 +/- 6% block at 10 microg/min) and by correolide in vitro (29 +/- 4% block at 1 microM). K(V) current in smooth muscle cells was inhibited by 4-AP (IC(50) 1.1 +/- 0.1 mM) and virtually eliminated by correolide. Expression of mRNA for K(V)1 family members was detected in coronary arteries. Our data indicate that K(V) channels play an important role in regulating resting coronary blood flow, determining duration of reactive hyperemia, and mediating adenosine- and NO-induced vasodilation.

Sensitization of Coronary α -Adrenoceptor Vasoconstriction in the Prediabetic Metabolic Syndrome

Objective: This study tested whether α ‐adrenoceptor‐mediated coronary vasoconstriction is augmented in the metabolic syndrome and is accompanied by the alteration of specific α1‐ and α2‐coronary adrenoceptors.

Coronary Vasomotor Reactivity to Endothelin-1 in the Prediabetic Metabolic Syndrome

Objective:The purpose of the present investigation was to test the hypothesis that coronary vasoconstrictor responses to endothelin‐1 are augmented in the prediabetic metabolic syndrome.

Diabetes decreases mRNA levels of calcium-release channels in human atrial appendage

Patients with chronic diabetes mellitus usually develop reductions in rate and force of cardiac contractions. Since calcium-release channels (ryanodine receptors (RyRs) and inositol 1,4,5-trisphosphate receptors (IP3Rs)) play integral roles in effecting these processes, we rationalize that alterations in their expression may underlie these defects. To test this hypothesis, right atrial appendages were obtained from diabetic (65.0 ± 4.5 years) and nondiabetic (56.2 ± 2.6 years) patients undergoing coronary arterial by-pass grafting and reverse transcription-polymerase chain reactions were used to compare steady state levels of mRNA encoding the three major isoforms of RyRs and IP3Rs. In this study we did not detect either RyR1 or RyR3 in human atrial appendage. When compared with nondiabetic patients, mRNA encoding RyR2 from diabetic patients decreased by 74.2 ± 6.2% (p< 0.01). Diabetes also significantly decreased steady-state levels of mRNA encoding the IP3Rs in human atrial appendage. IP3R1 decreased by 24.2 ± 4.6%, IP3R2 decreased by 63.0 ± 4.6% and IP3R3 decreased by 55.5 ± 6.5%. Since a reduction in steady-state mRNA is usually indicative of a decrease in protein levels, these data suggest that the decrease in chronotropy and inotropy seen in chronic diabetic patients may be due in part to a decrease in expression of calcium-release channels. (Mol Cell Biochem 263: 143-150, 2004).Patients with chronic diabetes mellitus usually develop reductions in rate and force of cardiac contractions. Since calcium-release channels (ryanodine receptors (RyRs) and inositol 1,4,5-trisphosphate receptors (IP3Rs)) play integral roles in effecting these processes, we rationalize that alterations in their expression may underlie these defects. To test this hypothesis, right atrial appendages were obtained from diabetic (65.0 ± 4.5 years) and nondiabetic (56.2 ± 2.6 years) patients undergoing coronary arterial by-pass grafting and reverse transcription-polymerase chain reactions were used to compare steady state levels of mRNA encoding the three major isoforms of RyRs and IP3Rs. In this study we did not detect either RyR1 or RyR3 in human atrial appendage. When compared with nondiabetic patients, mRNA encoding RyR2 from diabetic patients decreased by 74.2 ± 6.2% (p< 0.01). Diabetes also significantly decreased steady-state levels of mRNA encoding the IP3Rs in human atrial appendage. IP3R1 decreased by 24.2 ± 4.6%, IP3R2 decreased by 63.0 ± 4.6% and IP3R3 decreased by 55.5 ± 6.5%. Since a reduction in steady-state mRNA is usually indicative of a decrease in protein levels, these data suggest that the decrease in chronotropy and inotropy seen in chronic diabetic patients may be due in part to a decrease in expression of calcium-release channels. (Mol Cell Biochem 263: 143–150, 2004)

Decreased expression of β1- and β2-adrenoceptors in human diabetic atrial appendage

BackgroundUsing the streptozotocin-induced diabetic rat model, we have recently showed that the expression and function of β1-adrenoreceptor were decreased in the diabetic rat heart. However, the effect of diabetes on expression of β-adrenoreceptors in human cardiac tissue remains undefined. Therefore, the focus of the present study was to investigate the effect of diabetes on mRNA encoding β1- and β2-ARs in human atrial tissues.MethodsRight atrial appendages from five diabetic (mean age 65 ± 4.5; 4 female, 1 male) and five nondiabetic patients (mean age 56.2 ± 2.8; 4 male, 1 female) undergoing coronary artery bypass grafting were collected and assayed using reverse transcriptase-polymerase chain reaction (RT-PCR) for their mRNA content. No patient from these two groups suffered from acute myocardial infarction and/or failure. All diabetic patients received insulin for at least two years and had been diagnosed as diabetics for at least five years.ResultsWhen compared with levels in nondiabetics, steady state levels of mRNA encoding β1-adrenoreceptor decreased by 69.2 ± 7.6 % in diabetic patients while β2-adrenoreceptor mRNA decreased by 32.2 ± 5.5 % (p < 0.001).ConclusionsOur findings show a decreased expression of β1- and β2-adrenoreceptors in human diabetic atrial appendage.