Lack of iNOS impairs endothelial function in endothelin-1 transgenic mice.

BACKGROUND: Endothelin-1 (ET-1) is one of the most potent biologic vasoconstrictors. Nevertheless, transgenic mice overexpressing ET-1 exhibit normal blood pressure. We hypothesized that in states of ET-1 overproduction, the lack of counterregulatory mediators such as nitric oxide (NO), produced by the inducible NO synthase (iNOS), may critically impair endothelial function and may result in blood pressure elevation. METHODS: We generated crossbred animals of ET transgenic mice (ET+/+) and iNOS knockout mice (iNOS-/-) and evaluated blood pressure and endothelial function in these animals. Endothelium-dependent and -independent vascular function was assessed as relaxation/contraction of isolated preconstricted aortic rings to acetylcholine, sodium nitroprusside, and ET-1, alone or in the presence of BQ123 or BQ788. RESULTS: Systolic blood pressure was similar in ET+/+, iNOS-/- and wild-type mice, but was significantly elevated in ET+/+ iNOS-/- crossbred animals versus ET+/+ mice. Maximum endothelium-dependent relaxation was enhanced in ET+/+ mice (95 +/- 5 vs. 78 +/- 5% of preconstriction in wild-type littermates; p < 0.05). Additional knockout of iNOS led to a significant decrease of endothelium-dependent relaxation in combined ET+/+ iNOS-/- animals (75 +/- 6%; p < 0.05 vs. ET+/+ mice). Endothelium-independent relaxation was comparable among all groups. Maximum vascular contraction to ET-1 was reduced in ET+/+ mice (33 +/- 4%), iNOS-/- mice (38 +/- 5%) and ET+/+ iNOS-/- mice (44 +/- 4%) to a similar extent as compared with wild-type littermates (66 +/- 4%; p < 0.05). CONCLUSIONS: Our data show for the first time that in transgenic mice overexpressing human ET-1, additional knockout of iNOS results in impaired endothelium-dependent vasodilatation thus contributing to elevated blood pressure in ET+/+ iNOS-/- animals.

The estrogen receptor-alpha agonist 16alpha-LE2 inhibits cardiac hypertrophy and improves hemodynamic function in estrogen-deficient spontaneously hypertensive rats.

OBJECTIVE: Cardiac mass increases with age and with declining estradiol serum levels in postmenopausal women. Although the non-selective estrogen receptor-alpha and -beta agonist 17beta-estradiol attenuates cardiac hypertrophy in animal models and in observational studies, it remains unknown whether activation of a specific estrogen receptor subtype (ERalpha or ERbeta) might give similar or divergent results. Therefore, we analyzed myocardial hypertrophy as well as cardiac function and gene expression in ovariectomized, spontaneously hypertensive rats (SHR) treated with the subtype-selective ERalpha agonist 16alpha-LE2 or 17beta-estradiol. METHODS AND RESULTS: Long-term administration of 16alpha-LE2 or 17beta-estradiol did not affect elevated blood pressure, but both agonists efficiently attenuated cardiac hypertrophy and increased cardiac output, left ventricular stroke volume, papillary muscle strip contractility, and cardiac alpha-myosin heavy chain expression. The observed effects of E2 and 16alpha-LE2 were abrogated by the ER antagonist ZM-182780. Improved left ventricular function upon 16alpha-LE2 treatment was also observed in cardiac MRI studies. In contrast to estradiol and 16alpha-LE2, tamoxifen inhibited cardiac hypertrophy but failed to increase alpha-myosin heavy chain expression and cardiac output. CONCLUSIONS: These results support the hypothesis that activation of ERalpha favorably affects cardiac hypertrophy, myocardial contractility, and gene expression in ovariectomized SHR. Further studies are required to determine whether activation ERbeta mediates redundant or divergent effects.

Effect of additional temporary glycoprotein IIb/IIIa receptor inhibition on troponin release in elective percutaneous coronary interventions after pretreatment with aspirin and clopidogrel (TOPSTAR trial).

OBJECTIVES: The Troponin in Planned PTCA/Stent Implantation With or Without Administration of the Glycoprotein IIb/IIIa Receptor Antagonist Tirofiban (TOPSTAR) trial investigated: 1) the amount of troponin T (TnT) release after nonacute, elective percutaneous coronary intervention (PCI) in patients pretreated with aspirin and clopidogrel; and 2) the effect of additional glycoprotein (GP) IIb/IIIa receptor inhibiton on postinterventional TnT release. BACKGROUND: No data are available yet as to whether additional administration of a GP IIb/IIIa receptor antagonist might be beneficial in patients undergoing elective PCI already pretreated with aspirin and clopidogrel. METHODS: After bolus application of the study medication (tirofiban [T] or placebo [P]), PCI was performed followed by an 18-h continuous infusion of T/P. Primary end point of the study was incidence and amount of TnT release after elective PCI after 24 h. RESULTS: A total of 12 h after PCI troponin release was detected in 63% of the patients receiving P and in 40% of the patients receiving T (p < 0.05), after 24 h in 69% (P) and 48% (T) (p < 0.05) and after 48 h in 74% (P) versus 58% (T) (p < 0.08) of the patients. No differences were observed regarding major bleeding, intracranial bleeding or nonhemorrhagic strokes. After nine months a reduction of combined death/myocardial infarction/target vessel revascularization could be observed in the tirofiban group ([T] 2.3% vs. [P] 13.04%, p < 0.05). CONCLUSIONS: Troponin T release occurs after successful intervention in 74% of the patients undergoing elective PCI after 48 h even after pretreatment with aspirin and clopidogrel. The GP IIb/IIIa receptor antagonist tirofiban is able to decrease the incidence of troponin release significantly in this patient population.

Administration of testosterone is associated with a reduced susceptibility to myocardial ischemia.

This study investigated the impact of testosterone on myocardial ischemia-reperfusion injury and corresponding intracellular calcium ([Ca2+]i) metabolism. Nonorchiectomized mature male Wistar rats were randomly assigned to placebo, a single dose of testosterone undecanoate, or 5alpha-dihydrotestosterone. In a further series, orchiectomized rats were treated with placebo. After 2 wk of treatment, the hearts were removed and placed in a Langendorff setup. The isolated, buffer-perfused hearts were subjected to 30 min of no-flow ischemia and 30 min of reperfusion. Recovery of myocardial function was measured by analyzing pre- and postischemic left ventricular (LV) systolic/diastolic pressure and coronary perfusion pressure simultaneously, together with [Ca2+]i handling (aequorin luminescence). Calcium regulatory proteins were analyzed by Western blotting. LV weight/body weight ratio was increased after administration of testosterone vs. orchectomized rats. The recovery of contractile function was improved in testosterone-treated rats: at the end of the reperfusion, LV systolic pressure was higher and end-diastolic pressure was lower in testosterone-treated rats. End-ischemic [Ca2+]i and [Ca2+]i overload upon reperfusion was significantly lower in testosterone vs. orchiectomized rats, too. However, levels of calcium regulatory proteins remained unaffected. In conclusion, administration of testosterone significantly improves recovery from global ischemia. These beneficial effects are associated with an attenuation of reperfusion induced [Ca2+]i overload.

Cytokine response after percutaneous coronary intervention in stable angina: effect of selective glycoprotein IIb/IIIa receptor antagonism.

BACKGROUND: It is unclear whether modulation of inflammatory markers by glycoprotein IIb/IIIa receptor inhibition after percutaneous coronary intervention (PCI) is caused by an interaction with the alpha(v)beta3 and alpha(M)beta2 receptor or it correlates with ischemic events during PCI. This study investigates the inflammatory profile after elective, nonacute PCI and whether and how administration of the glycoprotein IIb/IIIa receptor antagonist tirofiban modulates the postinterventional inflammatory myocardial response. METHODS: The time course of inflammatory parameters (C-reactive protein [CRP], interleukin-1 [IL-1], interleukin-6 [IL-6], and tumor necrosis factor alpha [TNF-alpha]) of patients receiving peri- and postinterventional placebo (n = 46) or tirofiban infusion (n = 50) was analyzed by use of enzyme-linked immuno assays. Samples were collected before and 30 minutes, 2.5 hours, 6.5 hours, 12 hours, 24 hours, and 48 hours after elective PCI. RESULTS: Among the inflammatory markers analyzed, TNF-alpha, IL-6, and CRP levels increased significantly. However, the latter markers followed individual time courses in patients given placebo and patients treated with tirofiban after PCI, compared with pre-PCI levels (P <.01), with no significant differences between the placebo and tirofiban-treated groups. However, by subgroup analysis, significant differences were revealed in TNF-alpha, IL-6, and CRP levels of patients who were troponin T-positive versus patients who were troponin T-negative after PCI. CONCLUSIONS: The administration of the selective glycoprotein IIb/IIIa receptor antagonist tirofiban has no direct impact on the inflammatory profile after elective PCI. Change of the inflammatory profile was only related to the presence or absence of postinterventional troponin.

Long-term reduction of mortality in the 4-year follow up of tirofiban therapy in elective percutaneous coronary interventions (TOPSTAR) trial.

BACKGROUND: TOPSTAR was a randomized, placebo-controlled trial studying the effects of adding the glycoprotein IIb/IIIa inhibitor tirofiban to conventional treatment with aspirin and clopidogrel in patients undergoing elective percutaneous coronary interventions (PCI). TOPSTAR demonstrated a lower periprocedural troponin release and a reduced 6-month mortality risk following PCI. The present study analyzed the corresponding long-term effects. METHODS: All 96 patients who were initially included were followed for a minimum of 4 years (median follow-up time, 4.3 years). The prespecified endpoints were: 1) all-cause mortality and 2) the combined endpoint of all-cause death, myocardial infarction and target vessel revascularization by intention-to-treat analysis in patients randomly assigned to elective PCI. Survival analyses were carried out using Kaplan-Meier analysis and Cox proportional hazard regression. RESULTS: After 4 years of follow up, no differences were observed between the two groups with respect to medical therapy, NYHA classification and number of reinterventions and target vessel revascularizations. All-cause mortality was still higher in the placebo group (10.9%; 5/46) compared with the tirofiban group (0%; 0/50; Kaplan-Meier log rank = 0.017). The combined endpoint occurred in 21.7% (10/46) in the placebo group versus 8.0% (4/50) in the tirofiban group (Kaplan-Meier log rank = 0.056). CONCLUSIONS: The reduced 6-month mortality risk after elective PCI in the TOPSTAR trial persisted after 4 years of follow up. Even in this relatively small study, periprocedural effective platelet inhibition had a sustained impact on long-term mortality risk.

Functional properties and [Ca(2+)](i) metabolism of creatine kinase--KO mice myocardium.

One major function of the creatine kinase system is to maintain energy demand of myofibrillar contraction processes. Loss of the CK-system led to adaptations in skeletal muscle. To analyze the impact on myocardial function contractile parameters and intracellular calcium metabolism of transgenic mice lacking mitochondrial CK (ScCKmit(-/-)) alone or both mitochondrial and cytoplasmic ScCK (CK(-/-)) were investigated compared to wild type at various workload conditions using isolated intact muscle fibers. Force development at baseline conditions, force-frequency relationship (60-600/min), and rapid frequency switch (60-600/min) were unaltered in myocardium of transgenic mice compared to wild type. Intracellular calcium metabolism revealed unchanged amplitude of the intracellular calcium transients (ICT), refilling of the sarcoplasmic reticulum (calcium reuptake, post-rest behavior) in the ScCKmit(-/-) and CK(-/-) mice. The results demonstrate the effectiveness of myocardial energy-recruiting compensatory mechanisms at baseline as well as under stress conditions in CK depleted myocardium of transgenic mice.

Increased myocardial oxygen consumption by TNF-alpha is mediated by a sphingosine signaling pathway.

The present study investigated the effect of tumor necrosis factor (TNF)-alpha on myocardial energy metabolism as estimated by myocardial oxygen consumption (MVo(2)). MVo(2) of electrically stimulated isolated trabeculae of right ventricular Wistar rat myocardium was analyzed using a Clark-type oxygen probe. After the initial data collection in the absence of TNF-alpha, measurements were repeated after TNF-alpha stimulation. In separate experiments, pretreatment with the nitric oxide (NO) synthase inhibitor N(G)-nitro-l-arginine methyl ester (l-NAME) or the ceramidase inhibitor n-oleoylethanolamine (NOE) was done to investigate NO/sphingosine-related effects. TNF-alpha impaired myocardial economy at increasing stimulation frequencies without altering baseline MVo(2). Incubation with TNF-alpha in the presence of l-NAME further impaired myocardial economy. NOE preincubation abrogated the TNF-alpha effect on myocardial economy. Moreover, the negative inotropic effect of TNF-alpha was observed in NOE-pretreated but not l-NAME-pretreated muscle fibers. Exogenous sphingosine mimicked the TNF-alpha effect on mechanics and energetics. We conclude that TNF-alpha impairs the economy of chemomechanical energy transduction primarily through a sphingosine-mediated pathway.