Virtual histology assessment of cardiac allograft vasculopathy following introduction of everolimus--results of a multicenter trial.

In this 12-month multicenter Scandinavian study, 78 maintenance heart transplant (HTx) recipients randomized to everolimus with reduced calcineurin inhibitor (CNI) exposure or continued standard CNI-therapy underwent matched virtual histology (VH) examination to evaluate morphological progression of cardiac allograft vasculopathy (CAV). Parallel measurement of a range of inflammatory markers was also performed. A similar rate of quantitative CAV progression was observed in the everolimus (n = 30) and standard CNI group (n = 48) (plaque index 1.9 ± 3.8% and 1.6 ± 3.9%, respectively; p = 0.65). However, VH analysis revealed a significant increase in calcified (2.4 ± 4.0 vs. 0.3 ± 3.1%; p = 0.02) and necrotic component (6.5 ± 8.5 vs. 1.1 ± 8.6%; p = 0.01) among everolimus patients compared to controls. The increase in necrotic and calcified components was most prominent in everolimus patients with time since HTx >5.1 years and was accompanied by a significant increase in levels of von Willebrand (vWF) factor (p = 0.04) and vascular cell adhesion molecule (VCAM) (p = 0.03). Conversion to everolimus and reduced CNI is associated with a significant increase in calcified and necrotic intimal components and is more prominent in patients with a longer time since HTx. A significant increase in vWF and VCAM accompanied these qualitative changes and the prognostic implication of these findings requires further investigation.

Survival in patients with pulmonary arterial hypertension associated with systemic sclerosis from a Swedish single centre: prognosis still poor and prediction difficult.

OBJECTIVES: To describe the survival rate in a cohort of systemic sclerosis (SSc) patients with pulmonary arterial hypertension (PAH) and to evaluate possible predictors for SSc-PAH in a cohort of SSc patients. METHODS: Thirty patients with SSc-PAH and 150 SSc patients without PAH were included. Survival and survival on therapy were calculated. Clinical features at baseline were correlated to the risk for development of PAH during follow-up. RESULTS: The 1-, 2-, 3-, and 4-year survival rates were 86, 59, 39, and 22%, respectively, from diagnosis of PAH. The hazard ratio for total mortality in the SSc-PAH group was 3.2 [95% confidence interval (CI) 1.8-5.7] compared to SSc without PAH (p < 0.001). Risk factors at baseline for the development of PAH were: limited skin involvement, low diffusing capacity of the lung for carbon monoxide (DL(CO)), high N-terminal pro-brain natriuretic peptide (NTProBNP), increased estimated systolic pulmonary arterial pressure (ESPAP), and the presence of teleangiectases. Severe peripheral vascular disease requiring iloprost treatment during follow-up was associated with an eightfold increased risk of PAH. CONCLUSION: Despite modern treatment and yearly screening by echocardiography, the survival in SSc-PAH is still low in our cohort. The identified risk factors should be assessed to select patients eligible for right heart catheterization (RHC) to make an earlier diagnosis.

Electrical and mechanical responses to inhibition of cell respiration in vascular smooth muscle of the rat portal vein.

Metabolic regulation of contractility in vascular smooth muscle was studied in the spontaneously active rat portal vein using respiratory depression by cyanide (0.2-2.0 mM) as a model for tissue hypoxia. Intracellular recordings of electrical activity were done with concomitant registration of force development. Average membrane potential in the absence of cyanide was -61 +/- 1 mV (n = 27). Addition of cyanide to normal Krebs solution resulted in a reduction of force amplitude and the number of action potentials per burst, with a relatively more pronounced effect on the mechanical activity. At moderate levels of inhibition of force amplitude the frequency of spontaneous bursts of action potentials transiently increased concomitant with a slight depolarization, but after prolonged (15-20 min) exposure to cyanide the membrane repolarized to the level prior to cyanide addition and the burst frequency decreased to be equal to or lower than that in the absence of cyanide. Higher concentrations of cyanide totally inhibited spontaneous mechanical and electrical activity. In contrast to the results with glucose, it was found that when beta-hydroxybutyrate was used as substrate the addition of 2 mM cyanide led to a marked hyperpolarization (13 +/- 1 mV) after total inhibition of spontaneous activity. The hyperpolarization was not prevented by administration of 4-aminopyridine (2.5 mM) or tetraethylammonium (4-6 mM) prior to the addition of cyanide. To investigate the effects of increased metabolic demand on the relation between force and membrane potential in cyanide-treated muscle, high-K+ (40 mM) contractures were studied. Contractures were associated with depolarization of 34 +/- 3 mV (n = 5). 1 mM cyanide reduced the amplitude of the contractures to about 9% of control with a moderate reduction in the amount of depolarization (28 +/- 1 mV, n = 5). It is concluded that the decrease of mechanical activity during respiratory inhibition may partly reflect a reduction in the number of spikes per burst but that other mechanisms, independent of membrane activity, also contribute to the inhibition. The increase of glycolysis during respiratory inhibition seems to prevent more pronounced changes in membrane potential.

Contractile and metabolic characteristics of creatine-depleted vascular smooth muscle of the rat portal vein.

The functional consequences of phosphocreatine (PCr) depletion for mechanical properties, O2 consumption, and lactate production of the rat portal vein were investigated. After feeding rats for 8-9 weeks on a diet containing 2% beta-guanidino propionic acid (BGPA), PCr of the portal vein was reduced to 14% of control, whereas ATP was unchanged. No significant change was found in the level of spontaneous contractile activity or the force developed in a high-K+ contracture. Lactate production and the relationship between contractile force and O2 consumption were uninfluenced by BGPA treatment. The force-velocity relation of electrically stimulated portal veins showed no influence of BGPA treatment on Vmax. To investigate whether decrease in PCr influenced the response to metabolic stress, portal veins were exposed to graded concentrations (0.1-0.5 mM) of cyanide to depress cellular respiration. Veins from control and BGPA-treated rats showed the same relative decrease of contractile activity and O2 consumption, and the same increase in lactate production. Cyanide treatment resulting in a reduction of electrically stimulated force to 70-80% of the original gave a reduction of Vmax to 85-90%. The relative degree of reduction was uninfluenced by BGPA treatment. Reduction of PCr content thus does not affect the functional properties of metabolism or contractility under normoxic conditions. Furthermore, it can be inferred that the PCr reduction known to occur in smooth muscle exposed to hypoxia (Lövgren & Hellstrand 1985) is not in itself the major factor causing hypoxic inhibition of mechanical activity.

Shortening velocity, myosin light chain phosphorylation and Ca2+ dependence of force during metabolic inhibition in smooth muscle of rat portal vein.

The concentration-response relation for Ca2+ (0.2-5.0 mM) of high-K+ contractures (40 mM) in the rat portal vein during respiratory inhibition by 0.2 mM cyanide was investigated. A reduction of force in the presence of cyanide to about 30% of control was associated with a leftward shift of the normalized concentration-response relation. When force at the plateau of high-K+ contractures (at about 2 min) was reduced to 65 +/- 2% due to the addition of cyanide, the maximal shortening velocity (Vmax) was 94 +/- 5% of control (n = 6). In electrically (AC) stimulated preparations giving short tetanic contractions, a reduction of active force to 58 +/- 2% of control in the presence of cyanide was associated with a reduction of Vmax to 83 +/- 5% (n = 7). Phosphorylation of the 20-kDa regulatory light chains (LC20) of the myosin molecule was studied in the relaxed state and at the plateau of high-K+ contractures for comparison with the mechanical data. Both control and cyanide-treated preparations showed 9% LC20 phosphorylation in nominally Ca2+-free solution (n = 6). After activation the level of phosphorylation increased to 30 +/- 3% (n = 9) in the control veins. In cyanide-treated veins, where force was reduced to 42 +/- 6% compared to a preceding control period, the phosphorylation level was 17 +/- 2% (n = 7). The study suggests that the mechanical changes caused by inhibition of cellular respiration may involve the combined effect of several metabolic alterations, including decreased LC20 phosphorylation during contraction, but apparently not decreased intracellular Ca2+ concentration or sensitivity of the contractile system to Ca2+.

Demonstration of voltage-dependent and TTX-sensitive Na(+)-channels in human melanocytes.

The electrophysiological properties of cultured human melanocytes were investigated using the whole-cell configuration of the patch-clamp technique. Depolarizations to membrane potentials more positive than -30 mV resulted in the rapid development ( < 1 ms to peak) of an inward current. The maximum peak current was observed at +10 mV and reached an average amplitude of about 270 pA. During the depolarizations, the current inactivated with a time constant of about 2 ms. The current was abolished by the addition of 0.3 microM tetrodotoxin, a blocker of voltage-gated Na(+)-channels, and disappeared when Na+ was omitted from the extracellular medium. In addition, the melanocytes contain at least two types of outward K(+)-current. The first type, observed in every cell, was highly sensitive (Ki 1 mM) to the K(+)-channel blocker TEA, required depolarizations beyond zero to be activated and did not inactivate. The second type was less regularly observed (10% of the cells). This current activated at more negative voltages (-20 mV), was resistant to TEA (20 mM) but was blocked by 2 mM 4-aminopyridine and inactivated rapidly during depolarizations. We conclude that human melanocytes are equipped with voltage-dependent Na(+)-channels, a delayed rectifying K(+)-current and a K(+)-current similar to the A-current in neurones.