Altered anxiety-related behavior in nociceptin/orphanin FQ receptor gene knockout mice

Studies showed that nociceptin/orphanin FQ (N/OFQ) peptide receptor (NOP) agonists produce anxiolytic-like actions, while little is known about the effects of blockade of NOP receptor signaling in anxiety. To this aim, we investigated the behavioral phenotype of NOP receptor gene knockout mice (NOP(-/-)) in different assays. In the elevated plus-maze and light-dark box, NOP(-/-) mice displayed increased anxiety-related behavior. In the novelty-suppressed feeding behavior and elevated T-maze, NOP(-/-) mice showed anxiolytic-like phenotype, while no differences were found in the open-field, hole-board, marble-burying, and stress-induced hyperthermia. Altogether, these findings suggest that the N/OFQ-NOP receptor system modulates anxiety-related behavior in a complex manner.     

The domestication of artichoke and cardoon: from Roman times to the genomic age

BACKGROUND: The history of domestication of artichoke and leafy cardoon is not yet fully understood and when and where it occurred remains unknown. Evidence supports the hypothesis that wild cardoon is the wild progenitor of both these crops. Selection for large, non-spiny heads resulted in artichoke and selection for non-spiny, large stalked tender leaves resulted in leafy cardoon. The two crops differ in their reproductive system: artichoke is mostly vegetatively propagated and perennial, while leafy cardoon is seed propagated and mostly grown as an annual plant. Here, new trends in artichoke cultivation are analysed, while the consequences of these tendencies on the conservation of artichoke genetic resources are highlighted. SCOPE: The historical and artistic records, together with recent literature on genetics and biosystematics, are examined with the aim of achieving a better understanding of the present-day knowledge on the domestication of these two crops. CONCLUSIONS: Historical, linguistic and artistic records are consistent with genetic and biosystematic data and indicate that the domestication of artichoke and cardoon diverged at different times and in different places. Apparently, artichoke was domesticated in Roman times, possibly in Sicily, and spread by the Arabs during early Middle Ages. The cardoon was probably domesticated in the western Mediterranean in a later period.     

The late Messinian Lago-Mare episode in the Mediterranean Basin: Preliminary report on the occurrence of Paratethyan ostracod fauna from central Crete (Greece)

In the Crete Island, late Messinian Lago-Mare facies are not well known. At present, the occurrence in Crete of the uppermost Messinian post evaporitic deposits is a matter of debate. According to several authors, the well-known late Messinian Lago-Mare facies does not occur in Crete. In this paper the preliminary results obtained from the biostratigraphical analysis of some sections sampled in the Messarà Plain will be shown. Nearby Faneromeni and Ano Akria villages, the Miocene/Pliocene boundary is well exposed. There, gypsum-bearing clay, laminated microcrystalline gypsum and gypsum-rudites characterize the evaporitic deposits of the Messinian stage. In these areas, above the Messinian evaporite, post-evaporitic fine-laminated polychrome clays, with intercalations of sandstones and conglomerates, have been found. In both the Faneromeni and Ano Akria area, the Pliocene grey clays and conglomerates rest unconformably on the uppermost Messinian post-evaporitic deposits. A 20 cm-spaced sampling has been performed in both the sections, for more than 100 samples collected. The results of the micropaleontological analysis performed on the Faneromeni and Ano Akria sections point to the occurrence of ostracod assemblages containing: Loxocauda limata (Schneider in Agalarova et al.), Loxocauda sp., Cytherura pyrama Schneider, Cyprideis anlavauxensis Carbonnel, Cyprideis agrigentina Decima, Amnicythere palimpsesta (Livental), Amnicythere propinqua (Livental), Amnicythere accicularia (Olteanu), Amnicythere costata (Olteanu), Amnicythere multituberculata (Livental), Amnicythere sp. D (Miculan in Bassetti et al.), Amnicythere sp. 2 Gliozzi and Grossi, Amnicythere sp., Euxinocythere (Maeotocythere) praebaquana (Livental in Agalarova et al.), Mediocytherideini indet., Pontoniella pontica (Agalarova), Camptocypria sp. 1 Gliozzi and Grossi, Caspiocypris sp., Zalanyiella venusta (Zalanyi), Tyrrhenocythere sp., Loxoconcha rhombovalis Pokorny, Loxoconcha eichwaldi Livental, Loxoconcha sp. A (Miculan in Bassetti et al.), Loxocorniculina djafarovi (Schneider in Suzin). In the analysed samples, reworked planktonic foraminifers and well-preserved charophyte gyrogonites have been also found. The ostracod assemblages found in the Messarà Plain belong to the Loxocorniculina djafarovi Zone (sensu Carbonnel, 1978), which characterizes the uppermost Messinian deposits of the whole Mediterranean Basin. At that time, the well-known Lago-Mare biofacies was also widespread on the Crete Island. The presence of Paratethyan ostracods in the post-evaporitic Messinian deposits of both Faneromeni and Ano Akria sections suggests that in the Crete Island the latest Messinian sedimentation took place in brackish water palaeoenvironments.     

beta1-Adrenoreceptor activation contributes to ischemia-reperfusion damage as well as playing a role in ischemic preconditioning

Protein kinase A (PKA) activation has been implicated in early-phase ischemic preconditioning. We recently found that during ischemia PKA activation causes inactivation of cytochrome-c oxidase (CcO) and contributes to myocardial damage due to ischemia-reperfusion. It may be that beta-adrenergic stimulation during ischemia via endogenous catecholamine release activates PKA. Thus beta-adrenergic stimulation may mediate both myocardial protection and damage during ischemia. The present studies were designed to determine the role of the beta(1)-adrenergic receptor (beta(1)-AR) in myocardial ischemic damage and ischemic preconditioning. Langendorff-perfused rabbit hearts underwent 30-min ischemia by anterior coronary artery ligation followed by 2-h reperfusion. Occlusion-reperfusion damage was evaluated by delineating the nonperfused volume of myocardium at risk and volume of myocardial necrosis after 2-h reperfusion. In some hearts ischemic preconditioning was accomplished by two 5-min episodes of global low-flow ischemia separated by 10 min before coronary occlusion-reperfusion. Orthogonal electrocardiograms were recorded, and coronary flow was monitored by a drip count. Three hearts from each experimental group were used to determine mitochondrial CcO and aconitase activities. Two-hour reperfusion after occlusion caused an additional decrease in CcO activity vs. that after 30-min occlusion alone. Blocking the beta(1)-AR during occlusion-reperfusion reversed CcO activity depression and preserved myocardium at risk for necrosis. Similarly, mitochondrial aconitase activity exhibited a parallel response after occlusion-reperfusion as well as for the other interventions. Furthermore, classic ischemic preconditioning had no effect on CcO depression. However, blocking the beta(1)-AR during preconditioning eliminated the cardioprotection. If the beta(1)-AR was blocked after preconditioning, the myocardium was preserved. Interestingly, in both of the latter cases the depression in CcO activity was reversed. Thus the beta(1)-AR plays a dual role in myocardial ischemic damage. Our findings may lead to therapeutic strategies for preserving myocardium at risk for infarction, especially in coronary reperfusion intervention.     

Helper-dependent adenovirus for the gene therapy of proliferative retinopathies: stable gene transfer, regulated gene expression and therapeutic efficacy

BACKGROUND: Ocular neovascular disorders, such as diabetic retinopathy and age-related macular degeneration, are the principal causes of blindness in developed countries. Current treatments are of limited efficacy, whereas a therapy based on intraocular gene transfer of angiostatic factors represents a promising alternative. For the first time we have explored the potential of helper-dependent adenovirus (HD-Ad), the last generation of Ad vectors, in the therapy of retinal neovascularization. METHODS: We first analyzed efficiency and stability of intraretinal gene transfer following intravitreous injection in mice. A HD-Ad vector expressing green fluorescent protein (GFP) under the control of the cytomegalovirus (CMV) promoter (HD-Ad/GFP) was compared with a first-generation (E1/E3-deleted) Ad vector carrying an identical GFP expression cassette (FG-Ad/GFP). We also constructed HD-Ad vectors expressing a soluble form of the VEGF receptor (sFlt-1) in a constitutive (HD-Ad/sFlt-1) or doxycycline (dox)-inducible (HD-Ad/S-M2/sFlt-1) manner and tested their therapeutic efficacy upon intravitreous delivery in a rat model of oxygen-induced retinopathy (OIR). RESULTS: HD-Ad/GFP promoted long-lasting (up to 1 year) transgene expression in retinal Müller cells, in marked contrast with the short-term expression observed with FG-Ad/GFP. Intravitreous injection of HD-Ad vectors expressing sFlt-1 resulted in detectable levels of sFlt-1 and inhibited retinal neovascularization by more than 60% in a rat model of OIR. Notably, the therapeutic efficacy of the inducible vector HD-Ad/S-M2/sFlt-1 was strictly dox-dependent. CONCLUSIONS: HD-Ad vectors enable stable gene transfer and regulated expression of angiostatic factors following intravitreous injection and thus are attractive vehicles for the gene therapy of neovascular diseases of the retina.     

Reprint of "Structural and mechanistic aspects of Amt/Rh proteins" [J. Struct. Biol. 158 (2007) 472-481

Amt/Rh proteins, which mediate movement of ammonium across cell membranes, are spread throughout the three kingdoms of life. Most functional studies on various members of the family have been performed using cellular assays in heterologous expression systems, which are, however, not very well suited for detailed mechanistic studies. Although now generally considered to be ammonia conducting channels, based on a number of experimental studies and structural insights, the possibility remains that some plant Amts facilitate net ammonium ion transport. The Escherichia coli channel AmtB has become the model system of choice for analysis of the mechanism of ammonia conductance, increasingly also through molecular dynamics simulations. Further progress in a more detailed mechanistic understanding of these proteins requires a reliable in vitro assay using purified protein, allowing quantitative kinetic measurements under a variety of experimental conditions for different Amt/Rh proteins, including mutants. Here, we critically review the existing functional data in the context of the most interesting and unresolved mechanistic questions and we present our results, obtained using an in vitro assay set up with the purified E. coli channel AmtB.     

Molecular and karyological aspects of Batoidea (Chondrichthyes, Elasmobranchi) phylogeny

Although considerable progress has been made in elucidating the relationships within the Chondrichthyes, there is no agreement as it concerns the systematics of Batoidea, the most derived superorder among cartilaginous fishes, and many different interpretations exist. Our investigation provides the first assessment of relationships among the described batoid species using sequences from both mtDNA and nuclear genes as well as karyological morphology. Our work consists primarily in reconstructing the phylogenetic relationships of Batoidea by examining the mtDNA (16S) and nuclear gene (18S) sequences from 11 batoid species. The three analytical methods (NJ, MP and Bayesian analysis) grouped Rajiformes, Myliobatiformes and Rhinobatiformes. In these trees the two torpedoes diverge from the other batoid fishes. We also compare the molecular data with the available karyological evidence, which consist of the diploid number and the karyotype morphology of eight species belonging to the four orders examined. The results show that the karyological structure in the different species is generally consistent with the various phylogenetical trees, and that Torpediniformes confirm their unique genome organization.     

Association of body iron stores with low molecular weight iron and oxidant damage of human atherosclerotic plaques

The association between iron, an oxidant catalyst, and atherosclerosis is controversial. In particular, it is unknown whether: (1) stored iron, namely serum ferritin, is correlated with catalytic iron and oxidant damage of human atherosclerotic plaques; (2) catalytic iron is related to oxidative injury within such plaques; (3) plaque oxidant burden is associated with the severity of atherosclerosis. Thus, we assessed low molecular weight iron (LMWI), which represents the metal catalytically active form, together with fluorescent damage products of lipid peroxidation (FDPL) and lipid hydroperoxides (LOOH), in 38 atherosclerotic plaques surgically removed from 38 patients who had undergone selective carotid endarterectomy. In each patient, the levels of serum ferritin were measured and correlated with those of plaque LMWI and lipoperoxides by the Spearman rank correlation test with Spearman rank correlation coefficient (r(S)) calculation. Moreover, in patients selected from the same study population, we compared plaque analyte levels between two groups with different severity of atherosclerotic carotid stenosis, i.e., <90% (group A, n = 25) or > or =90% (group B, n = 13), and between another two groups without (group C, n = 27) and with (group D, n = 11) associated contralateral carotid stenosis > or =50%, indicative of "extensive" and more severe atherosclerotic disease. In group A patients, serum ferritin was directly and significantly correlated with plaque LMWI (r(S) = 0.46, P < 0.025) and FDPL (r(S) = 0.58, P < 0.005), while its correlation with plaque LOOH, albeit direct, did not attain statistical significance. Moreover, a direct and significant relationship was evident between the plaque content of LMWI and that of both FDPL (r(S) = 0.61, P < 0.0025) and LOOH (r(S) = 0.51, P < 0.025), suggesting a prooxidant role of catalytic iron within human atherosclerotic plaques. Considering the 13 patients of group B, a positive and significant correlation was observed between the levels of serum ferritin and those of plaque LMWI (r(S) = 0.83, P < 0.0001); on the other hand, serum ferritin, as well as plaque LMWI, showed no significant correlation with either plaque FDPL or LOOH, conceivably reflecting the small number of patients belonging to group B. Finally, plaque LMWI, FDPL, and LOOH content was significantly higher in group B than in group A, and in group D than in group C. These data suggest a role for catalytic iron in atherosclerotic plaque oxidation and in the severity of atherosclerosis, which appears indeed associated with plaque oxidant burden.