Viktor Drvota

Effects of intramyocardial injection of phVEGF‐A165 as sole therapy in patients with refractory coronary artery disease – 12‐month follow‐up: Angiogenic gene therapy

Abstract. Sarkar N, Rück A, Källner G, Y‐Hassan S, Blomberg P, Islam KB, van der Linden J, Lindblom D, Nygren AT, Lind B, Brodin L‐Å, Drvota V, Sylvén C (Karolinska Institute, Huddinge University Hospital, Novum, Stockholm, Sweden). Effects of intramyocardial injection of phVEGF‐A165 as sole therapy in patients with refractory coronary artery disease: 12‐month follow‐up. Angiogenic gene therapy. J Intern Med 2001; 250: 373–381.

Amiodarone Is a Dose-Dependent Noncompetitive and Competitive Inhibitor of T3 Binding to Thyroid Hormone Receptor Subtype β1, Whereas Disopyramide, Lignocaine, Propafenone, Metoprolol, dl-Sotalol, and Verapamil Have No Inhibitory Effect

The cardiovascular and electrophysiological effects of amiodarone resemble those of hypothyroidism. The drug has a structural resemblance to thyroid hormone (T3). Previous studies indicate that amiodarone exerts its major effect through antagonism of T3, probably as a result of inhibition of ligand binding to the thyroid hormone receptor (ThR). There are five subtypes of ThR, of which the β1 is the most prominent in the human heart. Our first aim was to investigate whether ThR is involved in a general antiarrhythmic mechanism for antiarrhythmic drugs or whether this action is specific for amiodarone. Therefore, we studied the affinity of one antiarrhythmic drug from every Vaughan-Williams group on T3 binding to human ThRβ1 (hThRβ1). Second, we wished to investigate whether amiodarone is a competitive or noncompetitive inhibitor. hThRβ1, expressed in insect cells using a recombinant baculovirus, was used in regular binding competition assays. Disopyramide, lignocaine, propafenone, metoprolol, dl-sotalol, and verapamil had no effect on T3 binding to hThRβ1. Amiodarone showed a noncompetitive binding pattern at low concentrations (0.25–2 μM) and a competitive binding at high concentrations (2–8 μM). Among the antiarrhythmics tested, only amiodarone had affinity for hThRβ1.This may represent a novel type of antiarrhythmic mechanism. The finding that amiodarone, in concentrations corresponding to therapeutic range in plasma, shifts from a noncompetitive to a competitive inhibitor, is of clinical interest in comparisons of low- and high-dose treatment.

Desethylamiodarone Prolongation of Cardiac Repolarization is Dependent on Gene Expression: A Novel Antiarrhythmic Mechanism

Desethylamiodarone (DEA) is the major metabolite of amiodarone and has similar electrophysiologic effects with prolongation of the repolarization that is reversed by thyroid hormone (T3). Some of the electrophysiologic effects are probably due to antagonism of T3 at the receptor level. Such effects of T3 are mediated by modulation of gene transcription. The aim of this study was to investigate whether cycloheximide (Cy), an inhibitor of protein synthesis, and actinomycin D (ActD), a RNA-synthesis inhibitor, block DEA-induced prolongation of the repolarization and whether DEA takes part in the autoregulation of the nuclear thyroid hormone-receptor subtypes (ThR). Corrected monophasic action potentials (MAPc) and QTc were measured in Langendorff-perfused guinea pig hearts for 1 h. The hearts were continuously perfused with (a) vehicle, (b) 7.5 microM Cy, (c) 5 microM DEA, (d) 5 microM DEA + 7.5 microM Cy, (e) 1 microM T3, (f) 5 microM DEA + 1 microM T3, (g) 1.5 microM ActD, and (h) ActD + DEA. A potassium channel blocker with class III antiarrhythmic effects, 0.5 microM almokalant, was used as a control, separately and together with Cy. Western blot analysis for the ThR subtypes alpha, beta1, and beta2 was performed on vehicle- and DEA-treated hearts. DEA increased MAPc by 19% (p < 0.0005) and QTc by 18% (p < 0.0005). There was no effect on MAPc or QTc when Cy, ActD, or T3 was added with DEA. Almokalant increased MAPc by 14% (p < 0.005) and QTc by 13% (p < 0.0005). When Cy was present, almokalant still induced a similar prolongation of MAPc by 14% (p < 0.005) and QTc by 17% (p < 0.0005). Western blot analysis revealed no change in the expression of the ThR protein. In conclusion, the prolongation of the cardiac repolarization by DEA, but not almokalant, can be totally blocked by Cy and ActD. This indicates that the class III action of DEA is at least in part dependent on transcription rather than a direct effect on cell-membrane channels or receptors. The action of DEA could be reversed by T3, indicating an antagonism between DEA and T3. These results suggest a new antiarrhythmic mechanism dependent on gene expression.

Thyroid hormone α1 and β1 receptor mRNA are downregulated by amiodarone in mouse myocardium

Amiodarone, a powerful antiarrhythmic drug, may exert its effect by antagonism of the thyroid hormone, probably at the receptor level. The aim of this study was to investigate whether amiodarone affects the levels of thyroid hormone receptor (TR) messenger RNA (mRNA) subtypes in mouse hearts. Mice were treated with 10, 25, and 50 mg/kg body weight (BW) amiodarone or vehicle (propyleneglycol) intraperitoneally, daily for 14 days. The heart rate dose-dependently decreased in the 25 mg/kg BW (p < 0.05) and 50 mg/kg BW (p < 0.005) amiodarone-treated mice compared with control. Serum T3 levels were significantly decreased by 25% (4.2 +/- 0.7 pM) in the 50 mg/kg BW amiodarone group in comparison to control (5.6 +/- 1.4 pM; p < 0.05). The serum T4 levels were 1.3 times higher in 50 mg/kg BW amiodarone-treated mice (13.2 +/-1.6 pM) compared with the control (10.3 +/- 1.3 pM; p < 0.005). Determination of TRalpha1, alpha2, beta1, and beta2 mRNA in the heart were performed by reverse transcriptase-polymerase chain reaction (RT-PCR)/enzyme-linked immunosorbent assay (ELISA). Both in treated and untreated mice, TRalpha2 mRNA had the highest density in mouse heart, whereas TRbeta2 mRNA had the lowest density. Amiodarone dose-dependently downregulated the levels of TRalpha1 and beta1 mRNA in comparison to the control. There were, however, no differences in the TRalpha2 and TRbeta2 mRNA levels in the mice heart treated with different doses of amiodarone in comparison with the control group. In conclusion, this study shows that amiodarone subtype selectively downregulates the TR mRNA levels in mouse myocardium in a dose-dependent manner. These results support a thyroid hormone-dependent action of amiodarone.

Protein and Angiogenic Dose-Response Expression of phVEGF-A165 Gene in Rat Myocardium

Therapeutic myocardial angiogenesis by means of transient overexpression of angiogenic growth factors is a potential treatment modality for severe ischemic heart disease. This study was undertaken in the rat to examine effects of phVEGF-A165 myocardial transfection in terms of dose-response as regards the number of hVEGF-A expressing cells on one hand and on the other angiogenesis. Non-surgical echocardiography-guided intramyocardial injection of phVEGF-A165 was done into normoxic or hypoxic (10[emsp4 ]% O2) rats. Cardiomyocytes expressing VEGF-A protein, capillary morphology and density were determined after 5 days. VEGF protein expression was seen in rat cardiomyocytes located around the tip of the injection scar and increased dose-dependently (p<0.05). Microvessel density also increased dose-dependently with phVEGF165 (p<0.05) and with hypoxia (p<0.05). No vascular tumours were observed. In conclusion, direct intramyocardial injection of phVEGF-A165 in the rat results in a dose-dependent increase both in transfected hVEGF-A protein producing cells and in angiogenesis.