http://http://dspace.unitus.it:80 The DSpace digital repository system captures, stores, indexes, preserves, and distributes digital research material. Sun, 26 May 2013 05:18:13 GMT 2013-05-26T05:18:13Z http://hdl.handle.net/2067/1429 Title: Expression, localization and functional coupling of the somatostatin receptor subtype 2 in a mouse model of oxygen-induced retinopathy Authors: Dal Monte, Massimo; Ristori, Chiara; Videau, Catherine; Loudes, Catherine; Martini, Davide; Casini, Giovanni; Epelbaum, Jacques; Bagnoli, Paola Abstract: PURPOSE. In the mouse model of oxygen-induced retinopathy (OIR) somatostatin (SRIF) acting at the SRIF receptor subtype 2 (sst2) inhibits angiogenic responses to hypoxia through a downregulation of vascular endothelial growth factor. Information on the sites where SRIF-sst2 interactions take place is lacking, and downstream effectors mediating SRIF-sst2 antiangiogenic actions are unknown. METHODS. In the OIR model, retinal expression of SRIF was evaluated with RT-PCR and RIA. The bindings of [125I]LTT-SRIF-28 and [125I]Tyr3-octreotide were measured in coronal sections of the eye. With Western blot we evaluated the levels of sst2A as well as the expression and the activity of the Signal Transducer and Activator of Transcription (STAT)3. The analysis of STAT3 was performed in hypoxic mice treated with the sst2 agonist octreotide or with the sst2 antagonist D-Tyr8 cyanamid 154806 (CYN). Retinal localization of sst2A was assessed by single and double immunohistochemistry with an endothelial cell marker. RESULTS. In the hypoxic retina, both SRIF and sst2 levels as well as [125I]Tyr3-octreotide binding were downregulated. In addition, sst2A immunostaining was decreased in the neuroretina, but was increased in capillaries. Hypoxia increased both expression and activity of STAT3. This increase was inhibited by octreotide, while was strengthened by CYN. CONCLUSIONS. These data suggest that i. sst2 expressed by capillaries may be responsible of the antiangiogenic effects of SRIF and ii. downstream effectors in this action include the transcription factor STAT3. These results support the possibility of using sst2- selective ligands in the treatment of proliferative retinopathies and indicate STAT3 as an additional target for novel therapeutic approach. Description: L'articolo è disponibile sul sito dell'editore http://www.arvo.org/eweb/StartPage.aspx?Site=arvo2 Thu, 31 Dec 2009 23:00:00 GMT http://hdl.handle.net/2067/1429 2009-12-31T23:00:00Z http://hdl.handle.net/2067/1448 Title: Adenylyl cyclase/cAMP system involvement in the antiangiogenic effect of somatostatin in the retina. Results from transgenic mice. Authors: Ristori, Chiara; Ferretti, Maria Enrica; Pavan, Barbara; Cervellati, Franco; Casini, Giovanni; Catalani, Elisabetta; Dal Monte, Massimo; Biondi, Carla Abstract: Neoangiogenesis is a response to retinal hypoxia that is inhibited by somatostatin (SRIF) through its subtype 2 receptor (sst2). Using a mouse model of hypoxia-induced retinopathy, we investigated the possibility that inhibition of adenylyl cyclase (AC) is involved in SRIF anti-angiogenic actions. Hypoxia increased AC responsiveness in wild type (WT) retinas and in retinas lacking sst2, but not in sst2-overexpressing retinas. Hypoxia also altered AC isoform expression, but with different patterns depending on sst2 expression level. Among the nine AC isoforms, AC VII isoform mRNA and protein resulted the most affected. Indeed, in hypoxia AC VII expression was significantly enhanced in WT retinas and it was further increased in sst2-lacking retinas, but not in retinas overexpressing sst2. These data suggest an involvement of AC/cAMP in mediating both hypoxia-evoked retinal neoangiogenesis and SRIF protective actions. The AC VII isoform is a candidate to a main role in these mechanisms. Description: L'articolo è disponibile sul sito dell'editore http://www.springerlink.com/ Mon, 31 Dec 2007 23:00:00 GMT http://hdl.handle.net/2067/1448 2007-12-31T23:00:00Z http://hdl.handle.net/2067/1486 Title: The cyclooxygenase-2/prostaglandin E2 pathway is involved in the somatostatin-induced decrease of epileptiform bursting in the mouse hippocampus Authors: Ristori, Chiara; Cammalleri, Maurizio; Martini, Davide; Pavan, Barbara; Casini, Giovanni; Cervia, Davide; Bagnoli, Paola Abstract: The neuromodulatory peptide somatostatin-14 (SRIF) plays an important inhibitory role in epilepsy, but little is known on the signalling mechanisms coupled to this effect of SRIF. We have previously demonstrated that SRIF induces reduction of epileptiform bursting in a model of interictal-like activity in mouse hippocampal slices. In this same model, we investigated whether the cyclooxygenase 2 (COX-2)/prostaglandin E2 (PGE2) pathway is part of those signalling mechanisms mediating SRIF anti-epileptic actions. Both the expression of COX-2 (mRNA and protein) and the endogenous release of PGE2 increased in concomitance with epileptiform bursting. In particular, COX-2 protein increased in CA1/CA3 pyramidal layer and in the granular layer of the dentate gyrus. In addition, the selective inhibition of COX-2 by NS-398 markedly decreased endogenous PGE2 release induced by epileptiform bursting and the epileptiform bursting itself. Similar effects on epileptiform bursting were obtained with another COX-2 inhibitor, i.e., meloxicam. SRIF application counteracted the increase of both COX-2 expression and PGE2 release which occurred in concomitance with epileptiform bursting. Interestingly, SRIF and NS-398 comparably reduced epileptiform bursting in a non-additive manner and PGE2 abolished the inhibitory effect of SRIF on epileptiform bursting. These results demonstrate that: i) the COX-2/PGE2 pathway facilitates epileptiform bursting; and ii) SRIF exerts an anti-epileptic role by coupling to the COX-2/PGE2 pathway. In conclusion, we have identified a key set of signalling events that underlie anti-convulsant effects of SRIF in a mouse model of hippocampal bursting, thus providing useful data not only to identify alternative intervention points for the modulation of SRIF function, but also to exploit new chemical space for drug-like molecules. Description: L'articolo è disponibile sul sito dell'editore http://www.sciencedirect.com/ Mon, 31 Dec 2007 23:00:00 GMT http://hdl.handle.net/2067/1486 2007-12-31T23:00:00Z http://hdl.handle.net/2067/1485 Title: Involvement of the cAMP-dependent pathway in the reduction of epileptiform bursting caused by somatostatin in the mouse hippocampus Authors: Ristori, Chiara; Cammalleri, Maurizio; Martini, Davide; Pavan, Barbara; Liu, Yanqiang; Casini, Giovanni; Dal Monte, Massimo; Bagnoli, Paola Abstract: The cAMP pathway is major signal transduction system involved in hippocampal neurotransmission. Recently, the peptide somatostatin-14 (SRIF) has emerged as a key signal that, by activating its receptors, inhibits epileptiform bursting in the mouse hippocampus. Little is known on transduction mechanisms which may mediate SRIF function in native cell/tissues. Using a well established model of epileptiform activity induced by Mg2+-free medium with 4-aminopyridine (0 Mg2+⁄4-AP) in mouse hippocampal slices, we demonstrated that PKA-related signaling is upregulated by hippocampal bursting and that treatment with SRIF normalizes this upregulation. We also demonstrated that the SRIF-induced inhibition of PKA impairs phosphorylation of the NMDA receptor subunit NR1. Extracellular recordings of the 0 Mg2+⁄4-AP-induced hippocampal discharge from the CA3 region demonstrated that treating slices with compounds which interfere with PKA activity prevent SRIF inhibition of epileptiform bursting. Our results suggest that SRIF modulation of hippocampal activity may involve PKA-related signaling. Description: L'articolo è disponibile sul sito dell'editore http://www.springerlink.com Mon, 31 Dec 2007 23:00:00 GMT http://hdl.handle.net/2067/1485 2007-12-31T23:00:00Z http://hdl.handle.net/2067/1469 Title: Action Mechanisms of the Secondary Metabolite Euplotin C: Signaling and Functional Role in Euplotes Authors: Trielli, Francesca; Cervia, Davide; Di Giuseppe, Graziano; Ristori, Chiara; Kruppel, Thomas; Burlando, Bruno; Guella, Graziano; Viarengo, Aldo; Bagnoli, Paola; Delmonte Corrado, Maria Umberta; Dini, Fernando Abstract: Among secondary metabolites, the acetylated hemiacetal sesquiterpene euplotin C has been isolated from the marine, ciliated protist Euplotes crassus, and provides an effective mechanism for reducing populations of potential competitors through its cytotoxic properties. However, intracellular signaling mechanisms and their functional correlates mediating the ecological role of euplotin C are largely unknown. We report here that, in E. vannus (an Euplotes morphospecies which does not produce euplotin C and shares with E. crasssus the same interstitial habitat), euplotin C rapidly increases the intracellular concentration of both Ca2+ and Na+, suggesting a generalized effect of this metabolite on cation transport systems. In addition, euplotin C does not induce oxidative stress, but modulates the electrical properties of E. vannus through an increase of the amplitude of graded action potentials. These events parallel the disassembling of the ciliary structures, the inhibition of cell motility, the occurrence of aberrant cytoplasmic vacuoles, and the rapid inhibition of phagocytic activity. Euplotin C also increases lysosomal pH and decreases lysosomal membrane stability of E. vannus. These results suggest that euplotin C exerts a marked disruption of those homeostatic mechanisms whose efficiency represents the essential prerequisite to face the challenges of the interstitial environmental. Description: L'articolo è disponibile sul sito dell'editore http://onlinelibrary.wiley.com/ Mon, 31 Dec 2007 23:00:00 GMT http://hdl.handle.net/2067/1469 2007-12-31T23:00:00Z http://hdl.handle.net/2067/1470 Title: Modulation of the neuronal response to ischemia by somatostatin analogues in wild-type and knock-out mouse retinas Authors: Cervia, Davide; Martini, Davide; Ristori, Chiara; Catalani, Elisabetta; Timperio, Anna Maria; Bagnoli, Paola; Casini, Giovanni Abstract: Somatostatin acts at five G protein-coupled receptors, sst1-sst5. In mouse ischemic retinas, the over-expression of sst2 (as in sst1 knock-out mice) results in reduction of cell death and glutamate release. Here, we reported that, in wild-type retinas, somatostatin, the multireceptor ligand pasireotide and the sst2 agonist octreotide decreased ischemia-induced cell death and that octreotide also decreased glutamate release. In contrast, cell death was increased by blocking sst2 with cyanamide. In sst2 over-expressing ischemic retinas, somatostatin analogues increased cell death, and octreotide also increased glutamate release. To explain this reversal of the anti-ischemic effect of somatostatin agonists in the presence of sst2 over-expression, we tested sst2 desensitisation due to internalisation or altered receptor function. We observed that: i) sst2 was not internalised, ii) among G protein-coupled receptor kinases (GRKs) and regulators of G protein signalling (RGSs), GRK1 and RGS1 expression increased following ischemia, iii) both GRK1 and RGS1 were down-regulated by octreotide in wild-type ischemic retinas, iv) octreotide down-regulated GRK1 but not RGS1 in sst2 over-expressing ischemic retinas. These results demonstrate that sst2 activation protects against retinal ischemia. However, in the presence of sst2 over-expression sst2 is functionally desensitised by agonists, possibly due to sustained RGS1 levels. Description: L'articolo è disponibile sul sito dell'editore http://onlinelibrary.wiley.com/ Mon, 31 Dec 2007 23:00:00 GMT http://hdl.handle.net/2067/1470 2007-12-31T23:00:00Z http://hdl.handle.net/2067/1474 Title: The secondary metabolite euplotin c induces apoptosis-like death in the marine ciliated protist euplotes vannus Authors: Cervia, Davide; Di Giuseppe, Graziano; Ristori, Chiara; Martini, Davide; Gambellini, Gabriella; Bagnoli, Paola; Dini, Fernando Abstract: The sesquiterpenoid euplotin C is a secondary metabolite produced by the ciliated protist Euplotes crassus and provides a mechanism for damping populations of potential competitors. Indeed, E. crassus is virtually resistant to its own product while different non-producer species representing an unbiased sample of the marine, interstitial, ciliate diversity are sensitive. For instance, euplotin C exerts a marked disruption of different homeostatic mechanisms in Euplotes vannus. We demonstrate by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay that euplotin C quickly decreases viability and mitochondrial function of E. vannus with a very high efficacy and at micromolar potency. In addition, euplotin C induces apoptosis in E. vannus as 4,6-diamino-2-phenylindole and Terminal Transferase dUTP Nick End Labeling staining show the rapid condensation and fragmentation of nuclear material in cells treated with euplotin C. These effects occur without detectable permeabilisation or rupture of cell membranes and with no major changes in the overall morphology, although some traits, such as vacuolisation and disorganised microtubules, can be observed by transmission electron microscopy. In particular, E. vannus show profound changes of the mitochondrial ultrastructure. Finally, we also show that caspase activity in E. vannus is increased by euplotin C. These data elucidate the pro-apoptotic role of euplotin C and suggest a mechanism for its impact on natural selection. Description: L'articolo è disponibile sul sito dell'editore http://onlinelibrary.wiley.com/ Wed, 31 Dec 2008 23:00:00 GMT http://hdl.handle.net/2067/1474 2008-12-31T23:00:00Z