Toxic effects of natural piperine and its derivatives on epimastigotes and amastigotes of Trypanosoma cruzi

We describe herein an evaluation of trypanocidal effects of the natural alkaloid piperine and twelve synthetic derivatives against epimastigote and amastigote forms of the protozoan parasite Trypanosoma cruzi, the causative agent of the incurable human disease, Chagas' disease. The results obtained point to piperine as a suitable template for the development of new drugs with trypanocidal activity.     

Daily rhythms of serum leptin in ewes: effects of feeding, pregnancy and lactation

The aim of the present study was to test whether serum concentrations of leptin in ewes vary with a daily rhythm. For this purpose, we examined 24 h serum leptin profiles of ewes exposed to natural photoperiodic conditions and subjected to two different feeding schedules (regular feeding and fasting). The results show for the first time the existence of daily rhythm of plasma leptin in regularly fed ewes, with a minimum during the light phase and a peak during the dark phase. Daily rhythms of serum leptin persisted after 50 h of fasting, although fasting shifted the peak of the rhythm to the beginning of the light phase and significantly reduced daily leptin production. To gain a better understanding of the role of leptin in the temporal organization of physiological events related to pregnancy and lactation, we measured serum leptin profiles throughout 24 h in ewes either during pregnancy or lactation. Daily leptin rhythms were found to persist during pregnancy and lactation, but both physiological conditions altered leptin concentrations. Maternal serum leptin concentration rose between early and mid pregnancy, then decreased in the late pregnancy and during lactation. Daily serum leptin concentration was significantly lower in nonpregnant, nonlactating ewes, compared either to lactating or to early pregnant ewes.     

Structure at 1.3 A resolution of Rhodothermus marinus caa(3) cytochrome c domain

The cytochrome c domain of subunit II from the Rhodothermus marinus caa(3) HiPIP:oxygen oxidoreductase, a member of the superfamily of heme-copper-containing terminal oxidases, was produced in Escherichia coli and characterised. The recombinant protein, which shows the same optical absorption and redox properties as the corresponding domain in the holo enzyme, was crystallized and its structure was determined to a resolution of 1.3 A by the multiwavelength anomalous dispersion (MAD) technique using the anomalous dispersion of the heme iron atom. The model was refined to final R(cryst) and R(free) values of 13.9% and 16.7%, respectively. The structure reveals the insertion of two short antiparallel beta-strands forming a small beta-sheet, an interesting variation of the classical all alpha-helical cytochrome c fold. This modification appears to be common to all known caa(3)-type terminal oxidases, as judged by comparative modelling and by analyses of the available amino acid sequences for these enzymes. This is the first high-resolution crystal structure reported for a cytochrome c domain of a caa(3)-type terminal oxidase. The R.marinus caa(3) uses HiPIP as the redox partner. The calculation of the electrostatic potential at the molecular surface of this extra C-terminal domain provides insights into the binding to its redox partner on one side and its interaction with the remaining subunit II on the other side.     

Controlled nitric oxide photo-release from nitro ruthenium complexes: The vasodilator response produced by UV light irradiation

Preliminary pharmacological studies of various nitric oxide (NO) photo-releasing agents are reported based on the flash-photolysis studies of the nitro ruthenium complexes cis-[Ru^II(NO₂)L(bpy)₂]⁺ (bpy = 2,2′-bipyridine and L = pyridine, 4-picoline and pyrazine) and [Ru^II(NO₂)(bpy)(terpy)]⁺ (terpy = terpyridine) in physiological medium. The net photoreactions under these conditions are two primary photoproducts, in (I) there is Ru^II-NO₂ photoaquation, where the photoproducts are Ru^II-H₂O plus and (II) homolytic dissociation of NO from a coordinated nitrito to derive the Ru^II-OH₂ specie and NO. Based on photochemical processes, the nitro ruthenium complexes were incorporated in water in oil (W/O) microemulsion and used in the vasorelaxation induced experiment. Denuded rat aortas were contracted with KCl and nitro ruthenium complexes in microemulsion were added. Perfusion pressures were recorded while arteries were irradiated at 355 nm The time to reach maximum relaxation was longer for [Ru^II(NO₂)(bpy)(terpy)]⁺ complex (ca. 50 min, n = 6) than for cis-[Ru(NO₂)L(bpy)₂]⁺ with L = py and 4-pic complex (ca. 28 min, n = 6) and cis-[Ru(NO₂)(bpy)₂ (pz)]²⁺ complex (ca. 24 min, n = 5).     

Light-induced protein-matrix uncoupling and protein relaxation in dry samples of trehalose-coated MbCO at room temperature

In humid samples of trehalose-coated carboxy-myoglobin (MbCO), thermally driven conformational relaxation takes place after photodissociation of the carbon monoxide (CO) molecule at room temperature. In such samples, because of the extreme viscosity of the external matrix, photodissociated CO cannot diffuse out of the protein and explores the whole (proximal and distal side) heme pocket, experiencing averaged protein heme pocket structures, as a results of the presence of Brownian motions. At variance, in very dry samples, a lower portion of the photodissociated CO diffuses from the distal to the proximal heme pocket side probing in nonaveraged structures. We revisit here the flash photolysis data by Librizzi et al. (2002) and report on new, room temperature experiments in MbCO-trehalose samples, shortly illuminated prior the laser pulse. In dry samples, pre-illumination increased the diffusion of CO from the distal to the proximal heme pocket side, which resulted in less structure than in non-pre-illuminated samples. Such an effect, which is absent in humid samples, stems from a decoupling of the protein internal degrees of freedom from those of the external water-sugar matrix. We suggest that such a decoupling can be brought about by the continuous attempts performed by the protein during pre-illumination to undergo relaxation toward the photodissociated deoxy state. This, in turn, causes a collapse in the hydrogen bond network, which connects the protein surface to the water-sugar matrix, as reported by Cottone et al. (2002) and Giuffrida et al. (2003). In the conclusion section, we discuss the possible involvement of the processes invoked to rationalize the present data, in the function of macromolecules and interactions in living cells.     

Cellular and functional defects in a mouse model of heart failure

Heart failure and dilated cardiomyopathy develop in mice that lack the muscle LIM protein (MLP) gene (MLP(-/-)). The character and extent of the heart failure that occurs in MLP(-/-) mice were investigated using echocardiography and in vivo pressure-volume (P-V) loop measurements. P-V loop data were obtained with a new method for mice (sonomicrometry) using two pairs of orthogonal piezoelectric crystals implanted in the endocardial wall. Sonomicrometry revealed right-shifted P-V loops in MLP(-/-) mice, depressed systolic contractility, and additional evidence of heart failure. Cellular changes in MLP(-/-) mice were examined in isolated single cells using patch-clamp and confocal Ca(2+) concentration ([Ca(2+)]) imaging techniques. This cellular investigation revealed unchanged Ca(2+) currents and Ca(2+) spark characteristics but decreased intracellular [Ca(2+)] transients and contractile responses and a defect in excitation-contraction coupling. Normal cellular and whole heart function was restored in MLP(-/-) mice that express a cardiac-targeted transgene, which blocks the function of beta-adrenergic receptor (beta-AR) kinase-1 (betaARK1). These data suggest that, despite the persistent stimulus to develop heart failure in MLP(-/-) mice (i.e., loss of the structural protein MLP), downregulation and desensitization of the beta-ARs may play a pivotal role in the pathogenesis. Furthermore, this work suggests that the inhibition of betaARK1 action may prove an effective therapy for heart failure.