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M Prosdocimi

Publications and source records attributed to M Prosdocimi.

At least 55 records · Page 3Linked to original sources

Cystometric changes in alloxan diabetic rats: evidence for functional and structural correlates of diabetic autonomic neuropathy.

Autonomic neuropathy and urinary bladder function were compared in Sprague-Dawley rats with alloxan-diabetes of 3 months duration, rats fed sucrose for 8 weeks, and rats examined 8 weeks after pelvic nerve surgical axotomy; normal age-matched rats were used as controls. All experimental interventions induced bladder hypertrophy with increased bladder weight. In diabetic and sucrose-fed animals, water intake and urinary output increased. Cystometric recordings of normal rats in vivo showed rhythmic contractions (1.25 +/- 0.25 contr/min) with threshold volume for micturition reflex at 0.51 +/- 0.04 ml. In diabetic rats, bladder contractions were irregular and of lower frequency (0.60 +/- 0.04 contr/min), while threshold volume was significantly higher (1.00 +/- 0.11 ml). Bladder contractions were normal in sucrose-fed animals, though threshold volume was markedly augmented (1.27 +/- 0.19 ml). Pelvic nerve surgical ablation abolished micturition reflex. In bladder strips excised post-mortem, contractile response to field stimulation was reduced in diabetic rats compared to control and sucrose-fed animals. Morphological examination of pelvic and hypogastric nerves revealed abnormalities characteristic of diabetic neuropathy only in diabetic rats. These data suggest that in alloxan-induced diabetes the decrease in the rate of bladder contraction is the result of autonomic neuropathy; while bladder hypertrophy in sucrose-fed rats appears to be an organ adaptation to hyperdiuresis.

Alloxan↗

Inhibition of platelet function by polymorphonuclear leukocytes.

Human polymorphonuclear leukocytes (PMNs) were evaluated for their ability to modulate platelet response induced by collagen, thrombin, platelet-activating factor and the stable analog of cyclic endoperoxides U46619. Platelet aggregation was first evaluated in whole blood and in leukocyte-depleted whole blood by the impedance method. This novel approach highlighted the inhibitory role of leukocytes on platelet aggregation in whole blood. The inhibitory role of PMNs on platelet function was subsequently evaluated on washed cells. PMN inhibition of platelet aggregation and beta-thromboglobulin release was more evident with threshold concentrations of stimuli. The inhibition also depended on the number of PMNs incubated in mixed cellular suspensions. Higher concentrations of stimuli may overcome the PMN-dependent inhibition. Under this condition, preincubation of cells with N-formyl-methionyl-leucyl-phenylalanine (a specific PMN agonist) restored the inhibitory effect of PMNs on platelet aggregation in whole blood and in mixed cellular suspensions. Not only PMNs, but also PMN-derived supernatants, dose-dependently inhibited U46619-induced platelet aggregation, suggesting that the inhibition observed may be exerted by chemically stable compound(s). Cytoplasmic Ca2+ movement was measured in aequorin-loaded platelets exposed to thrombin or U46619 to see whether cytoplasmic Ca2+ levels were affected by PMN. Ca2+ levels were similar in the presence or absence of PMNs, suggesting that inhibition may be related to a subsequent platelet response step. A series of bioassay experiments showed that PMNs were able to remove and/or convert adenosine diphosphate available for platelet aggregation but not to reduce U46619 availability. Our findings suggest that (1) unstimulated PMNs may release factor(s) that inhibit platelet aggregation and beta-thromboglobulin release; (2) this in itself is sufficient to block the platelet response to a threshold concentration of stimuli; (3) release of the same or other inhibitory mediators from stimulated PMNs may have to be greater to inhibit platelet response to higher concentrations of stimuli. Data presented here suggest that adenosine diphosphatase activity and chemically stable, as yet unidentified, compounds besides previously well-characterized labile compounds such as nitric oxide and arachidonic acid metabolites are responsible for the PMN-dependent mechanism of inhibition of platelet response that could be relevant in physiopathologic conditions.

Adenosine Diphosphate↗

Cholesterol diet followed by normal diet alters functional and metabolic responses in rabbit aorta.

Male New Zealand rabbits were fed a high cholesterol (1%) diet for 8 weeks followed by a normal diet for 8 additional weeks. Rabbits on normal diet were used as controls. Serum cholesterol, elevated at the end of the cholesterol feeding period, returned to normal at the time of observation. Aortic wall lesions were severe and vessel wall cholesterol in the thoracic aorta was approximately 10 times the level in control animals. In vitro, thoracic aortic rings were used to measure contractile response to noradrenaline (NOR); no difference in response was found between normal and treated rabbits. Rings pre-contracted with NOR, were exposed to acetylcholine (Ach), ATP and NaNO2. Proximal and distal rings from normal animals showed a dose-dependent relaxation response to all agents, though maximal dilation occurred at an intermediate concentration of Ach. The rings from treated rabbits relaxed normally in response to NaNO2, but ATP relaxation was reduced in proximal and distal rings and Ach induced a contractile response. HPLC analysis of tissue extracts from aortic arch and distal thoracic aorta showed reduced ATP, ADP, GTP and GDP, and increased AMP. These changes resulted in a reduced energy charge and a reduction in total adenine nucleotides. The data indicate that cholesterol feeding, followed by normal diet, causes severe alterations of aortic vessel wall and that at an advanced stage of the diet-induced experimental disease energy metabolism and endothelium dependent vasodilation remain impaired even in the presence of normal serum lipids.

Acetylcholine↗

Improved method for purification of human platelet factor 4 by affinity and ion-exchange chromatography.

A simple method for the reproducible purification of human platelet factor 4 (PF4) is described. PF4 is obtained in a highly purified form from platelet concentrate by utilizing a combination of affinity and FPLC ion-exchange chromatography. In every instance, after elution from heparin affinity and cation exchange chromatography, SDS gel electrophoresis reveals a single band attributable to PF4. Moreover, the amino acid composition of PF4 isolated by this method is compatible with that described for a PF4 cDNA clone and with other reported PF4 analysis. The purified protein is used to study in vitro the affinity of PF4 for several glycosaminoglycans (GAGs), by measuring the fluorescence of each PF4-GAG complex bound to fluorescamine. PF4 affinity for GAGs is as follows: heparin greater than heparan sulphate much greater than dermatan sulphate.

Amino Acids↗

Molecular aspects of cloricromene (AD6) distribution in human platelets and its pharmacological effects.

Cloricromene (AD6) is an investigational drug which inhibits platelet aggregation and release reaction. We studied the relationship between its action and its distribution and metabolism in platelets. Incubation of anticoagulated whole blood or platelet-rich plasma (PRP) without exogenous aggregating agents resulted in a progressive decrease of platelet count with a concomitant increase of beta-thromboglobulin (BTG) release. AD6 (20-50 mumols/l), but not acetylsalicylic acid (ASA), incubated with whole blood or PRP, prevented the fall in platelet count and the release of BTG for at least 150 min. Moreover, incubation of PRP with AD6 (50 mumols/l) and subsequent stimulation by ADP at threshold concentrations resulted in a significant reduction (about 30%) in aggregation for at least 90 min. AD6 (20 mumols/l) added to PRP was rapidly metabolized by hydrolysis of an ester bond to AD6 acid, a stable catabolite pharmacologically inactive in platelets. Significant amounts of AD6 acid (up to 13.26 +/- 2.80 pmol/10(6) platelets) were associated with the platelets after incubation either at 37 degrees C or 4 degrees C. The amount of AD6 acid in the platelet pellet was proportional to AD6 concentration (2 to 100 mumols/l). PRP incubation with AD6 acid (20 mumols/l) resulted in very low levels (less than 1 pmol/10(6) platelets) of the same compound in the platelet pellet after 1, 5 or 30 min. These data suggest that AD6 is taken up as an ester and converted to its acid catabolite with a consequent long-lasting inhibition of platelet function.

Adenosine Diphosphate↗

In vitro inhibition of human polymorphonuclear cell function by cloricromene.

Cloricromene, a coumarin derivative with antiaggregating and vasodilating properties, was tested in vitro on polymorphonuclear cell (PMN) adhesion to the endothelium, superoxide anion generation and chemotaxis. PMN adhesion was measured using cultured human umbilical vein endothelial cells (EC) either untreated or previously activated with interleukin-1 (IL-1). Cloricromene (5-50 microM) induced dose-related inhibition of PMN adhesion to untreated and IL-1 treated EC. Cloricromene also inhibited PMN superoxide generation induced by the tumor promoter 12-O-tetradecanoyl-phorbol-13-acetate (TPA) or by N-formyl-methionyl-leucyl-phenylalanine (FMLP). PMN and monocyte chemotaxis was evaluated by a modification of the Boyden chamber technique. Cloricromene inhibited both types of cell motility induced by FMLP in a concentration-dependent fashion. The major cloricromene metabolite (cloricromene acid) had no effect on any of the biological parameters studied up to a concentration of 500 microM. HPLC measurement showed that cloricromene accumulated in PMN within a few minutes and levels of the drug were still high after 60 min. In contrast its acid metabolite was not taken up in a significant amount during incubation periods up to 60 min. We conclude that cloricromene inhibits a series of PMN activities in vitro. This effect might be of pharmacological interest in view of the role of PMN activation in different pathophysiological conditions.

Cell Adhesion↗

Endothelium-dependent relaxation, cholesterol content and high energy metabolite balance in Watanabe hyperlipemic rabbit aorta.

Functional and metabolic parameters of thoracic aorta from Watanabe heritable hyperlipemic (WHHL) rabbits (aged 11-14 months) were investigated in vitro. The aortic preparations, normally responsive to noradrenaline, showed a diminished response to the endothelium-dependent agent, acetylcholine, in comparison with control preparations from age-matched New Zealand rabbits (maximal relaxation: 33 +/- 4% in WHHL vs. 52 +/- 2% in controls: P less than 0.005). ATP relaxant effect (only partially endothelium-dependent) was unimpaired in WHHL aorta, and it was much higher than in controls (maximal response: 63 +/- 6% vs. 37 +/- 3%, respectively; P less than 0.005). The response to NaNO2, an endothelium-independent relaxant, was unchanged in WHHL aortas. Acetylcholine-induced response was found to be inversely related to the degree of total cholesterol infiltration in aorta (r = -0.62, P less than 0.05). No correlation was observed between either total serum cholesterol or triglycerides and ACh-induced response. Furthermore, the concentration of adenine nucleotides and nucleosides in the aortic tissue of WHHL rabbits was lower than in controls, indicating a loss of energy balance. The results indicate a functional damage induced by genetic hyperlipidemia on endothelium-dependent relaxation and an impairment of energy-rich phosphate metabolism of the aortic wall. The relationship between functional and metabolic parameters is not yet clarified.

Adenosine Triphosphate↗

Autonomic neuropathy in BB rats and alterations in bladder function.

In vivo urinary bladder function was examined in BB rats after 4 and 6 mo of diabetes, and the data were correlated with morphometric changes in the pelvic and hypogastric nerves, which constitute the micturition reflex arc. After controlled bladder distension, diabetic animals revealed irregular bladder contractions at frequencies that were reduced to 33% of normal values and with significantly increased amplitudes. The abnormal micturition in diabetic animals was elicited at moderately elevated threshold volumes. These functional abnormalities of the diabetic bladder were associated with a progressive axonopathy of afferent myelinated sensory fibers and later-occurring axonal atrophy of unmyelinated efferent preganglionic fibers. These data suggest that diabetic urinary bladder dysfunction is initiated by a visceral sensory neuropathy involving the afferent limb of the micturition reflex arc.

Animals↗

Cloricromene improves survival rate and peritoneal macrophage function in splanchnic artery occlusion shock in rats.

Splanchnic artery occlusion (SAO) shock, induced by a transient occlusion of splanchnic arteries for 45 min, was performed in male rats, treated with vehicle or cloricromene, a coumarin derivative, 15 min before surgery. Survival rate, plasma levels of myocardial depressant factor (MDF), macrophage phagocytosis and killing of Candida albicans, and thromboxane B2 (TxB2) synthesis by peritoneal macrophages were evaluated. Of the SAO-shocked animals, 10% survived for 6 hr after the release of the occlusion of the splanchnic arteries, whereas none of the sham-shocked rats died. Peritoneal macrophages of shocked animals exhibited decreased phagocytosis (24.7 +/- 2.7%) and killing (8.0 +/- 2.1%) and increased TxB2 levels (3.23 +/- 0.27 ng/ml) with respect to those collected from sham-shocked animals (phagocytosis 48.8 +/- 3.0%; killing 16.5 +/- 2.4%; TxB2 0.30 +/- 0.18 ng/ml). MDF was also increased (114.3 +/- 21.5 U/ml) compared with sham-shocked animals (31.5 +/- 3.7 U/ml). Cloricromene, given intravenously (i.v.) at doses of 1, 2, and 4 mg/kg, significantly increased survival rate and lowered MDF in shocked rats. Lower doses (0.25 and 0.5 mg/kg/i.v.) were without effect. Doses that were able to reduce mortality partially reverted shock-induced macrophage impairment of phagocytosis, killing of C. albicans, and TxB2 synthesis. In addition, cloricromene (5, 10, and 25 microM) added in vitro to peritoneal macrophages, collected from shocked rats, significantly enhanced their phagocytic activity depressed by shock.

Animals↗

Segmental impairment of endothelium-mediated relaxation in thoracic aortas from atherosclerotic rabbits. Comparison to cholesterol infiltration and energy metabolism.

Three sets of parameters, (i) relaxation to acetylcholine (Ach), ATP and NaNO2, (ii) cholesterol content in aortic tissue, and (iii) energy metabolism were compared in normal and atherosclerotic rabbits, fed 1% cholesterol for eight weeks. A special protocol was envisaged to permit a strict comparison between Ach, ATP and NaN O2 at different levels of thoracic aorta, in each rabbit. A gradual impairment of the endothelium-dependent relaxation to Ach and ATP was found at different levels of the thoracic aorta from hypercholesteromic rabbits. By contrast, NaNO2--endothelium-independent--maintained its relaxing power quite normally at all aortic levels. A close correlation was evident between the impairment of aorta relaxation to Ach and the cholesterol infiltration in the vessel wall, being the correlation coefficient -0.85 (P less than 0.001). A correlation was also evident for ATP, but to a lower degree, being the correlation coefficient -0.61 (P less than 0.01). Energy metabolism and related parameters (ATP, ADP, AMP, adenosine, inosine, GTP, GDP, guanosine, NAD, NADP, total adenylate nucleotides and adenylate energy charge) were not modified by the cholesterol diet. These data show that the gradual impairment of endothelium-dependent relaxation, decreasing down the thoracic aorta of hypercholesterolemic rabbits, and correlated with cholesterol content in the aortic wall, may be considered as an index of a very early atherogenic damage, prior to variation in the parameters of energy metabolism and purine turnover.

Acetylcholine↗

Prostacyclin release in the coronary circulation during sustained stimulation in in vitro and in in vivo experimental systems.

Prostacyclin release from rat isolated perfused hearts and from dog coronary circulation was studied by measuring immunoreactive 6-keto-PGF1 alpha (6-keto-PGF1a) in heart perfusate and in plasma obtained from the great cardiac vein respectively. Continuous infusion of arachidonic acid at constant concentration in isolated perfused hearts induced an increased prostacyclin release. This release showed a rapid peak within 10 min and a subsequent decrease. Low-flow ischemia induced an increased perfusate concentration of 6-keto-PGF1a but, considering the decreased flow, prostacyclin release was actually reduced. During the whole period of ischemia (60 min) prostacyclin release was constant. In open-chest anesthetized dogs 6-keto-PGF1a concentration in the great cardiac vein was increased after ligation of the left anterior descending coronary artery. A prolonged period of coronary occlusion (4.5 hours) resulted in a progressive rise of prostacyclin release. 6-keto-PGF1a determinations in the femoral vein and in the aorta did not show relevant variations during the observation period.

Animals↗

Effect of AD6 (8-monochloro-3-beta-diethylamino-ethyl-4-methyl-7-ethoxycarbonylme tho xy coumarin) on cyclic nucleotide phosphodiesterases in human platelets.

The effect of AD6 (8-monochloro-3-beta-diethylamino-ethyl-4-methyl-7-ethoxycarbonylmeth oxy coumarin), an inhibitor of platelet aggregation, on cyclic nucleotide metabolism was investigated. AD6 inhibited selectively human platelet cyclic GMP phosphodiesterase, which was separated from cyclic AMP phosphodiesterase by DEAE-cellulose chromatography. Addition of AD6 to washed platelets increased cyclic GMP levels significantly in two minutes. These results could be useful in elucidating the action of AD6 on intact platelets.

3',5'-Cyclic-GMP Phosphodiesterases↗

Stenosis and vascular damage as a cause of thrombosis in the dog femoral artery.

We describe here an experimental model of peripheral arterial thrombosis and the effect of several drugs which are known to affect vessel and platelet biological functions. A similar method has been previously applied by us and others on dog coronary arteries. Male Beagle dogs, under pentobarbital anesthesia, were instrumented to measure arterial pressure, heart rate, ECG, femoral blood flow and expired CO2. A segment of the femoral artery was squeezed with forceps to damage the endothelium, and a plastic cylinder was placed around the vessel in the area of the damage. The cylinders had a length of 2 mm and an internal diameter of 1.6-1.8 mm. Under these circumstances blood flow in the stenosed artery was reduced by about 60-70% from control value and showed cyclic blood flow variations (CBFV). CBFV eventually led either to a total occlusion of the vessel (documented by blood flow measurement and by angiographic analysis), or to a spontaneous partial restoration of flow, followed by another decrease, in a repetitive fashion. Drug effect was monitored by observing the changes in frequency and amplitude of CBFV. Ketanserin (0.25 mg/kg), dazmegrel (0.5 mg/kg), and chlorpromazine (0.5 mg/kg), abolished or greatly reduced CBFV in all the experiments, while acetylsalycilic acid (ASA, 10 mg/kg) reduced or abolished CBFV in 60% of the treated dogs. Heparin (50 I.U./kg), dipyridamole (1.0 mg/kg) and prazosin (0.1 mg/kg) did not change CBFV. These results emphasize the importance of serotonin and thromboxane as mediators of vascular occlusion in this particular experimental model. This approach provides a reproducible in vivo preparation to study the pharmacological control of peripheral arterial thrombosis.

Angiography↗

General toxicity and peripheral nerve alterations induced by chronic vincristine treatment in the rabbit.

The effects of five 0.3 mg/kg intravenous administrations of vincristine (VCR) at weekly intervals were studied in the rabbit. Body weight gain was impaired starting from the first injection, while gross signs of motor paralysis and hair loss initiated from the third week. At the end of the observation period blood analysis revealed normocytic normochromic anemia, elevated serum creatine kinase, and low serum alkaline phosphatase, whereas all the tested parameters related to liver and kidney functions where within normal limits. The decreased number of red blood cells was the consequence of a complete, although reversible, blockade of staminal hematopoietic activity. Two important indexes of peripheral nerve function were clearly altered at the end of the treatment: (i) the sciatic nerve conduction velocity in vitro was 27% reduced and (ii) the latency between sciatic nerve stimulation and extensor digitorum longus (EDL) twitch in vivo was 34% prolonged. The usefulness of the rabbit as an animal model to study side-effects of VCR treatment is discussed.

Alkaline Phosphatase↗

Platelet activating factor induces dopamine release in PC-12 cell line.

The ability of platelet activating factor (PAF) to stimulate dopamine release and modify calcium homeostasis in PC-12 cell line was studied. PAF-induced dopamine release is related to its molecular form, with only the R-form steric configuration [(R)PAF], but not its S-form or its 2-lyso derivative, effective at being active. In addition, PAF acts at very low concentrations in a dose-dependent manner (0.1-30 nM). Preincubation with PAF receptor antagonists (CV-3988 and BN52021) as well as the specific desensitization of PC-12 cells to (R)PAF abolish the (R)PAF-induced dopamine release. Several lines of evidence suggest that dopamine release is dependent on a (R)PAF-induced calcium influx and efflux modulation. Dopamine release by PC-12 cells challenged with (R)PAF is associated with a rapid 45Ca influx and efflux and a rise in cytoplasmic calcium concentrations ([Ca2+]i) evaluated by using the calcium indicators fura-2 and quin2. At 30 nM (R)PAF, the absence of extracellular calcium inhibits the dopamine release but not the rise of [Ca2+]i from the internal stores, suggesting the importance of calcium influx in (R)PAF-induced dopamine release. PAF, which has been reported to be synthesized by stimulated neuronal cells (J. Biol. Chem. 261: 16502-16508, 1986) may thus have a physiological modulatory role on cells with neurosecretory properties.

Adrenal Gland Neoplasms↗

AD6 (8-monochloro-3-beta-diethylamino-ethyl-4-methyl-7-ethoxycarbonyl-meth oxy coumarin) inhibits the release of arachidonic acid in human platelets stimulated by thrombin.

The coumarin derivative AD6 is known to inhibit platelet aggregation and release and it possesses vasodilatory properties on coronary arteries of laboratory animals. Furthermore, the inhibition of the production of TxB2 from endogenous substrates after stimulation of human platelets with collagen has been demonstrated. The present report demonstrates that AD6 inhibits the production of labeled arachidonic acid and diglycerides from phospholipids of platelets stimulated with thrombin. This effect is dose-dependent and is already evident at a concentration of the drug (25 microM) which is unable to prevent the aggregation. Apparently, AD6 inhibits the release of arachidonic acid from phosphatidylinositol and choline phosphoglycerides which are the main sources of the substrate for the synthesis of prostaglandins and thromboxanes.

Arachidonic Acid↗