Coronary artery reperfusion: The ADP receptor P2Y<Subscript>1</Subscript> mediates early reactive hyperemia <Emphasis Type="BoldItalic">in vivo</Emphasis> in pigs

The physiological mechanisms that regulate reactive hyperemia are not fully understood. We postulated that the endothelial P2Y₁ receptor that release vasodilatory factors in response to ADP might play a vital role in the regulation of coronary flow. Intracoronary flow was measured with a Doppler flow-wire in a porcine model. 2-MeSADP (10^–5 M), ATP (10^–4 M) or UTP (10^–4 M) alone or as co-infusion with a selective P2Y₁ receptor blocker, MRS 2179 (10^–3 M) was locally delivered through the tip of a coronary angioplasty balloon. In separate pigs the coronary artery was occluded with the balloon for 10 min. During the first and tenth minutes of coronary ischemia, 2.5 ml of MRS 2179 (10^–3 M) was delivered distal to the occlusion in 8 pigs, 10 pigs were used as controls. MRS 2179 fully inhibited the 2-MeSADP-mediated coronary flow increase (P < 0.05) with no effect on UTP, indicating selective P2Y₁ inhibition. ATP-mediated flow increase was significantly inhibited by MRS 2179. During reactive hyperemia following coronary occlusion, flow increased by nearly sevenfold. MRS 2179, however, reduced the post-ischemic hyperemia by a mean of 46% during the period 1–2.5 min following balloon deflation (P < 0.05), which corresponds to peak velocity flow during reperfusion. In conclusion, MRS 2179, a selective P2Y₁ receptor blocker, significantly reduces the increased coronary flow caused both by 2-MeSADP and reactive hyperemia in coronary arteries. Thus, ADP acting on the endothelial P2Y₁ receptor may play a major role in coronary flow during post-ischemic hyperemia.     

Morphological features and isoenzyme characterization of endosymbiotic algae from green hydra

Symbiotic associations are of a wide significance in evolution and biodiversity. Green hydra is a typical example of endosymbiosis. In its gastrodermal myoepithelial cells, it harbors individuals of unicellular green algae. Morphological characteristics of isolated algae determined by light and electron microscopy are presented. Cytological morphometric parameters (cell area, cell radius, chloroplast area) of isolated algae from green hydra (Cx), as well as from reference species Chlorella kessleri (Ck) and Chlorella vulgaris (Cv), revealed similarity between the isolated endosymbiont and C. kessleri. Isoenzyme patterns of esterase (EST), peroxidase (POX), and catalase (CAT) were used for the investigation of genetic variability in endosymbiotic algae isolated from green hydra. Out of 14 EST isoenzymes observed in Cx species, 9 were expressed in the Cx sample. Results of the EST isoenzyme analysis indicated a higher degree of similarity between Cx and Cv than between Cx and Ck. Due to much higher heterogeneity, EST isoenzymes seem to be more suitable genetic markers for identification of different Chlorella species than CAT and POX isoenzymes. Results obtained suggest that symbiogenesis in green hydra has probably not been terminated yet.     

Recombination between the linear plasmid pPZG101 and the linear chromosome of Streptomyces rimosus can lead to exchange of ends

The 387 kb linear plasmid pPZG101 of Streptomyces rimosus R6 can integrate into the chromosome or form a prime plasmid carrying the oxytetracycline biosynthesis cluster. The integration of plasmid pPZG101 into the linear chromosome of S. rimosus R6-501 in mutant MV25 was shown to be due to a single cross-over at a 4 bp common sequence. pPZG101 had integrated into a 250 kb DNA sequence that was reiterated at a low level. This sequence includes the oxytetracycline biosynthesis cluster, so that homologous recombination generated a mixed population carrying different copy numbers of the region. The 1 Mb linear plasmid pPZG103 in mutant MV17 had also arisen from a cross-over between pPZG101 and the chromosome, so that one end of pPZG103 consists of c . 850 kb of chromosomal sequence including the oxytetracycline biosynthesis cluster. The plasmid pPZG101 was shown to consist of a unique central region of about 30 kb flanked by terminal inverted repeats of about 180 kb. Analysis of a presumed ancestor plasmid pPZG102 suggested that the long terminal repeats had arisen by a recombination event during the strain development programme.     

Complementary chimeric isoforms reveal Dscam1 binding specificity in vivo

Dscam1 potentially encodes 19,008 ectodomains of a cell recognition molecule of the immunoglobulin (Ig) superfamily through alternative splicing. Each ectodomain, comprising a unique combination of three variable (Ig) domains, exhibits isoform-specific homophilic binding in?vitro. Although we have proposed that the ability of Dscam1 isoforms to distinguish between one another is crucial for neural circuit assembly, via a process called self-avoidance, whether recognition specificity is essential in?vivo has not been addressed. Here we tackle this issue by assessing the function of Dscam1 isoforms with altered binding specificities. We generated pairs of chimeric isoforms that bind to each other (heterophilic) but not to themselves (homophilic). These isoforms failed to support self-avoidance or did so poorly. By contrast, coexpression of complementary isoforms within the same neuron restored self-avoidance. These data establish that recognition between Dscam1 isoforms on neurites of the same cell provides the molecular basis for self-avoidance.     

Endogenously nitrated proteins in mouse brain: links to neurodegenerative disease

Increased abundance of nitrotyrosine modifications of proteins have been documented in multiple pathologies in a variety of tissue types and play a role in the redox regulation of normal metabolism. To identify proteins sensitive to nitrating conditions in vivo, a comprehensive proteomic data set identifying 7792 proteins from a whole mouse brain, generated by LC/LC-MS/MS analyses, was used to identify nitrated proteins. This analysis resulted in the identification of 31 unique nitrotyrosine sites within 29 different proteins. More than half of the nitrated proteins that have been identified are involved in Parkinson's disease, Alzheimer's disease, or other neurodegenerative disorders. Similarly, nitrotyrosine immunoblots of whole brain homogenates show that treatment of mice with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), an experimental model of Parkinson's disease, induces an increased level of nitration of the same protein bands observed to be nitrated in brains of untreated animals. Comparing sequences and available high-resolution structures around nitrated tyrosines with those of unmodified sites indicates a preference of nitration in vivo for surface accessible tyrosines in loops, a characteristic consistent with peroxynitrite-induced tyrosine modification. In addition, most sequences contain cysteines or methionines proximal to nitrotyrosines, contrary to suggestions that these amino acid side chains prevent tyrosine nitration. More striking is the presence of a positively charged moiety near the sites of nitration, which is not observed for non-nitrated tyrosines. Together, these observations suggest a predictive tool of functionally important sites of nitration and that cellular nitrating conditions play a role in neurodegenerative changes in the brain.     

Ultrasound evaluation of extracranial carotid artery lesions in Parkinsonian patients

The purpose of this investigation was to determine the atherosclerotic changes in patients with vascular parkinsonism and in patients with idiopathic Parkinson's disease, in order to evaluate the possible influence of the extracranial pathology of carotid arteries in developing lacunar cerebral infarcts. Degree of stenosis and plaque morphology of the extracranial part of carotids in both group of patients were evaluated by color Doppler flow imaging ultrasound investigation and the results were compared. We selected two matched groups of patients with parkinsonism: 22 patients with vascular parkinsonism, and 28 with idiopathic Parkinson's disease.The atherosclerotic changes found in patients with Parkinson's disease showed mild carotid lesions with mostly stable calcified plaques and lesser risk for embolic cerebral intravascular events contrary to the higher degree of carotid stenosis found in patients with vascular parkinsonism with mostly mixed plaques prone to embolization. Therefore, we suggest performing ultrasonographic examination of the extracranial part of carotid arteries in all patients with parkinsonism to assess risk of vascular accidents originating from carotid lesions. That would enable adequate treatment of parkinsonism and prevent further occurrence of intracranial vascular changes.     

Contrasting the excited-state dynamics of the photoactive yellow protein chromophore: protein versus solvent environments

Wavelength- and time-resolved fluorescence experiments have been performed on the photoactive yellow protein, the E46Q mutant, the hybrids of these proteins containing a nonisomerizing “locked” chromophore, and the native and locked chromophores in aqueous solution. The ultrafast dynamics of these six systems is compared and spectral signatures of isomerization and solvation are discussed. We find that the ultrafast red-shifting of fluorescence is associated mostly with solvation dynamics, whereas isomerization manifests itself as quenching of fluorescence. The observed multiexponential quenching of the protein samples differs from the single-exponential lifetimes of the chromophores in solution. The locked chromophore in the protein environment decays faster than in solution. This is due to additional channels of excited-state energy dissipation via the covalent and hydrogen bonds with the protein environment. The observed large dispersion of quenching timescales observed in the protein samples that contain the native pigment favors both an inhomogeneous model and an excited-state barrier for isomerization.     

C-H. . .pi interactions in the metal-porphyrin complexes with chelate ring as the H acceptor

Specific C-H. . .pi interactions with the pi-system of porphyrinato chelate ring were found in crystal structures of transition metal complexes from the Cambridge Structural Database and statistical analysis of geometrical parameters for intramolecular and intermolecular interactions was done. By density functional theory calculations on a model system it was evaluated that an interaction energy is above 1.5 kcal/mol and that the strongest interaction occurs when the distance between hydrogen atom and the center of the chelate ring is 2.6 A. This prediction is in good agreement with the distances for intermolecular interactions found in the crystal structures. In many cases the intramolecular interaction distances are much shorter than 2.6 A, and these short distances are caused by geometrical constrains. The C-H. . .pi interactions with chelate ring of porphyrinato ligand can influence the structure, contribute to its stability, and play some role in the function of biomolecules with metalo porphyrins.     

Cytotoxicity of some platinum(IV) complexes with ethylenediamine-N,N'-di-3-propionato ligand

The cytotoxicities of two platinum(IV) complexes of formula [PtX2(eddp)].nH2O (eddp=ethylenediamine-N,N'-di-3-propionate, X=chloro [I] or bromo [II], n=1 or 1.24) are reported. The complexes have been obtained by direct reaction of potassium hexahaloplatinate(IV) with H2eddp.2HCl followed by addition of a base (LiOH). The crystal and molecular structure has confirmed that the complex with bromo ligands, similarly to the complex with chloro ligands previously reported, has trans configuration of the halogens. In both the chloro and bromo complexes there appear to be intramolecular N-H...X interactions which account for a narrowing of the corresponding X-Pt-N angles below 90degrees. The trans isomer (configuration index OC-6-13, two nitrogens and two oxygens of eddp bound in the equatorial plane) is the only one obtained in the reaction of hexahaloplatinate(IV) with the eddp ligand while a similar reaction performed with ethylenediamine-N,N'-diacetate (edda) affords exclusively the symmetrical cis-isomer (configuration index OC-6-33, equatorial nitrogen and axial oxygen atoms of edda). The longer chain of the propionato groups (as compared to the acetato ones) is responsible for such a change in preferred configuration. NMR data have revealed a very large diastereotopic splitting of the propionato methylene protons to the nitrogens (0.88 ppm). The trans disposition of the halogen ligands in the compounds with eddp leads to deactivation of platinum(IV) complexes in comparison to those with edda having cis disposition of the leaving chlorides (human ovarian cancer cell line A2780, IC50 [muM] of 92.6 +/- 12 and 30.3 +/- 7.5 for [I] and [II], respectively).