Biomedical subjects

P Pignatelli

Publications and source records attributed to P Pignatelli.

33 records · Page 2Linked to original sources

PubMed1999

Platelet hyperactivity in hypertensive older patients is controlled by lowering blood pressure.

OBJECTIVE: Patients with hypertension tend to have a high prevalence of atherothrombotic accidents. Platelet hyperactivity is frequently associated with hypertension. Because the vascular disease associated with hypertension evolves over the years, we investigated platelet activity parameters in a population of older hypertensive patients with no other risk factors for cardiovascular disease. PARTICIPANTS: We studied 34 older, nonsmoking patients (mean age 74 +/- 5 years) with uncomplicated hypertension before and after the normalization of blood pressure (BP) was achieved with the angiotensin-converting enzyme inhibitor quinapril alone or in combination with the Ca2+ antagonist nifedipine. MEASUREMENTS: Platelet aggregation, P-selectin (CD62) expression on the platelet surface, serum levels of Interleukin-1beta (IL-1beta) and of Interleukin-6 (IL-6), as well as plasma levels of soluble P-selectin and Endothelin-1 (ET-1), were analyzed. RESULTS: All platelet hyperactivity parameters were reduced significantly with the normalization of BP at the end of antihypertensive drug treatment (systolic/diastolic: 186.2 +/- 2.7/103.4 +/- 1.1 mm Hg vs 135.0 +/- 1.3/85.9 +/- 1.9 mm Hg; P < .001). Those factors more strictly associated with endothelium injury, such as ET-1 and IL-6, did not show variations. A significant correlation (Spearman Rank test) was observed among all platelet function parameters and blood pressure values. CONCLUSIONS: This study demonstrated that even in a population of older hypertensive patients with no other risk factor for atherogenic disease, normalization of blood pressure induces a significant reduction of the parameters of enhanced platelet hyperactivity independent of the action exerted, at the platelet level, by the antihypertensive drugs.

Aged↗

PubMed1999

Vitamin E inhibits collagen-induced platelet activation by blunting hydrogen peroxide.

In this study, we investigated whether vitamin E at concentrations achievable in blood after supplementation inhibits platelet function in humans. Gel-filtered platelets were incubated 30 minutes with scalar concentrations (50 to 250 mmol/L) of vitamin E and then stimulated with collagen. Compared with controls, vitamin E inhibited collagen-induced platelet aggregation and thromboxane A2 formation in a dose-dependent manner. Furthermore, vitamin E inhibited, in a dose-dependent manner, Ca(2+) mobilization and formation of inositol 1,4,5-triphosphate. Because it was previously shown that hydrogen peroxide formation mediates arachidonic acid metabolism and phospholipase C activation in collagen-induced platelet activation, we investigated whether vitamin E was able to blunt hydrogen peroxide. In experiments performed in unstimulated platelets supplemented with hydrogen peroxide and in collagen-stimulated platelets, vitamin E was able to blunt hydrogen peroxide. In 6 healthy subjects given vitamin E for 2 weeks (600 mg/d), we found a significant decrease of collagen-induced H(2)O(2) formation, platelet aggregation, and calcium mobilization. This study demonstrated in vitro and ex vivo that vitamin E inhibits collagen-induced platelet activation by blunting hydrogen peroxide formation.

Adult↗

PubMed1998

Concomitant activation of Gi and Gq protein-coupled receptors does not require an increase in cytosolic calcium for platelet aggregation.

U46619 is a potent platelet agonist, its binding to the thromboxane A2 receptor resulting in Gq-binding protein-mediated responses; nevertheless, it is unable to cause platelet aggregation, unless released ADP is present. In this study we demonstrate that Gi activation is the step U46619 lacks to cause platelet aggregation; in fact, when platelets were treated with an ADP scavenger system, the response to U46619 was restored by the addition of epinephrine, which activates platelets via a Gi protein. The concomitant activation of Gi and Gq proteins does not require increased cytosolic calcium to cause aggregation, as assessed by the fact that platelets treated with the intracellular calcium chelator BAPTA were able to respond to U46619 provided ADP or epinephrine was present. Moreover, as the calcium ionophore ionomycin, at low concentrations, potentiated the response to U46619 but not to epinephrine, we may conclude that calcium influx preferentially activates a Gi downstream signalling pathway.

Adenosine Diphosphate↗

PubMed1998

Hydrogen peroxide is involved in collagen-induced platelet activation.

In this study, we investigated whether (1) collagen-induced platelet aggregation is associated with a burst of H2O2, (2) this oxidant species is involved in the activation of platelets, and (3) the pathways of platelet activation are stimulated by H2O2. Collagen-induced platelet aggregation was associated with production of H2O2, which was abolished by catalase, an enzyme that destroys H2O2. H2O2 production was not observed when ADP or thrombin were used as agonists. Catalase inhibited dose-dependently thromboxane A2 production, release of arachidonic acid from platelet membrane, and Inositol 1,4,5P3 (IP3) formation. In aspirin-treated platelets stimulated with high concentrations of collagen, catalase inhibited platelet aggregation, calcium mobilization, and IP3 production. This study suggests that collagen-induced platelet aggregation is associated with a burst of H2O2 that acts as a second messenger by stimulating the arachidonic acid metabolism and phospholipase C pathway.

Blood Platelets↗

PubMed1998

Effects on platelet function of combination etoposide and carboplatin chemotherapy in pediatric oncology patients.

The effects of a therapeutic course of the combination of carboplatin and etoposide on platelet function have been evaluated in 10 pediatric patients with brain tumors. Platelet count, in vitro aggregation tests, P-selectin expression and agonist-induced ATP release were evaluated before, and 7 and 15 days after one cycle of chemotherapy. The analysis of the results demonstrated the presence of an in vitro platelet aggregation defect in response to collagen and arachidonic acid in all patients 7 days after therapy. A concomitant decrease of collagen- and arachidonic acid-induced ATP release was also observed. Both platelet aggregation and ATP release returned to baseline values 15 days after chemotherapy administration. Conversely, in vitro platelet aggregation and secretion induced by ADP and epinephrine were unaltered by carboplatin and etoposide administration. Furthermore, P-selectin expression was negative at baseline and did not change after chemotherapy. These results support the hypothesis that combination etoposide and carboplatin chemotherapy in pediatric patients is responsible for possible disturbances in biochemical pathways required for platelet secretion and aggregation.

Journal Article↗

PubMed1998

Nickel enhances collagen-induced platelet activation acting by increasing the organization of the cytoskeleton.

In keratinocytes, osteoclasts and enterocytes, Ni2+ acts as an agonist working through selective activation of the polyvalent cation-sensing receptor. We report here that while Ni2+ alone had no direct ability to induce platelet aggregation or secretion, Ni2+ pretreatment produced these responses when platelets were stimulated with subthreshold concentrations of collagen. In addition, pretreatment with Ni2+ significantly enhanced collagen-induced phospholipase C activation and calcium mobilization. Platelet adhesion to collagen was increased and the inhibition of collagen-induced adhesion normally seen after cytochalasin D treatment was significantly diminished. When Ni2+ was added to platelets alone, tyrosine phosphorylation of p60src was increased. Moreover, Ni2+ enhanced the amount of protein, especially actin, found in the low-speed Triton X-100 insoluble cytoskeleton. Our results indicate that nickel, possibly acting via a platelet cation sensing receptor analogous to that which has been described in other cell types, may cause a rapid tyrosine kinase-dependent cytoskeleton reorganization leading to enhanced adhesion of platelets to collagen and increasing collagen-dependent responses.

Blood Platelets↗

PubMed1998

Platelet activation and cytokine production during hypothermic cardiopulmonary bypass--a possible correlation?

Cardiopulmonary bypass (CPB) is associated with impaired platelet function and a systemic inflammatory response. The present study was designed to evaluate whether any correlation between platelet activation and inflammatory response during CPB exists. The results obtained from 8 patients undergoing hypothermic CPB for cardiac surgery showed the occurrence of a moderate degree of platelet activation during CPB, demonstrated by an increase of platelet CD62P expression in correlation with an increase of beta-thromboglobulin levels, with a concomitant decrease of in vitro platelet response. Plasma IL-1beta levels significantly increased during CPB, with a peak between 1 and 4 h after CPB. Similarly, IL-6 levels were elevated 30 min from CPB starting, peaked at 4 h, and remained elevated after 24 h. A direct correlation was found between plasma IL-1beta and IL-6 levels. A significant correlation between plasma IL-1beta and beta-thromboglobulin levels was also found. In turn, plasma beta-thromboglobulin levels correlated with CD62P expression on activated platelets. An inverse correlation was found between in vitro platelet aggregation and plasma IL-1beta or IL-6 levels. From the present results it may be speculated that platelet activation during CPB may contribute, through the release of IL-1beta, to activation of endothelial cells and subsequent release of other cytokines with chemotactic and pro-inflammatory properties, thus playing an important role in the inflammatory response associated with CPB.

Aged↗

PubMed1997

Platelet activation by superoxide anion and hydroxyl radicals intrinsically generated by platelets that had undergone anoxia and then reoxygenated.

BACKGROUND: Platelet activation has been demonstrated in experimental and clinical models of ischemia-reperfusion, but the underlying mechanism is still unclear. We mimicked the ischemia-reperfusion model in vitro by exposing platelets to anoxia-reoxygenation (A-R) and evaluated the role of oxygen free radicals (OFRs), which are usually produced during the reperfusion phase, in inducing platelet activation. METHODS AND RESULTS: Human platelets were exposed to 15 and 30 minutes of anoxia and then reoxygenated. Compared with control platelets kept in atmospheric conditions, platelets exposed to A-R showed spontaneous platelet aggregation (SPA), which was maximal after 30 minutes of anoxia. Superoxide dismutase (SOD) (-74%, P < .005), catalase (-67%. P < .005). SOD plus catalase (-82%, P < .005), and the hydroxyl radical (OH0) scavengers mannitol (-66%, P < .005) and deoxyribose (-55%, P < .005) inhibited SPA. Platelets that had undergone A-R released superoxide anion (0-2), as detected by lucigenin chemiluminescence. Also, platelets exposed to A-R and incubated with salicylic acid generated 2.3- and 2,5-dihydroxybenzoates, which derive from salicylic acid reaction with OH0. SPA was significantly inhibited by the cyclooxygenase enzyme inhibitors aspirin and indomethacin: by SQ29548, a thromboxane (Tx) A2 receptor antagonist; by diphenyliodonium an inhibitor of flavoprotein-dependent enzymes: and by arachidonyl trifluoromethyl ketone, a selective inhibitor of cytosolic phospholipase A2. Platelets exposed to A-R markedly generated inositol 1,3,4-trisphosphate and TxA2, which were inhibited by incubation of platelets with SOD plus catalase. CONCLUSIONS: This study shows that platelets exposed to A-R intrinsically generated 0-2 and OH0, which in turn activate arachidonic acid metabolism via phospholipases A2 and C, and provides further support for the use of antioxidant agents as inhibitors of platelet function in ischemia-reperfusion models.

Acridines↗

PubMed1997

Mechanism of the persisting TxA2 receptor antagonism by picotamide.

Picotamide is a dual TxA2 receptor antagonist/TxA2 synthetase inhibitor. As the picotamide effect is maximum only after 5-10 min of incubation, aim of the present work was to study whether the effect of picotamide could be observed even after wash out of the drug from the external buffer. Platelet aggregation, serotonin release and changes in intracellular calcium were analyzed in platelets treated with picotamide and then gel-filtered, in comparison with gel-filtered platelets treated with picotamide. To exclude the possibility that the inhibitory effect of picotamide is due to its intracytosolic storage, serotonin release in digitonin-permeabilized platelets was measured. Platelet aggregation and serotonin release in response either to U46619, a synthetic agonist of the thromboxane A2 receptor, or arachidonic acid or collagen, as well as the changes in intracellular calcium concentration after U46619 or arachidonic acid stimulation, were inhibited by picotamide even when it had been washed out by gel-filtration. Both inhibitions were very similar to that observed in experiments in which picotamide was present in the medium. As the serotonin release in digitonin permeabilized platelets presented the same inhibition it may be excluded that picotamide effect is consequent upon the cytosolic storage of the drug. Since our results clearly indicate that the action of picotamide persists even after washing out of the drug from the medium, the idea that this antagonistic effect may be dependent on its binding to platelet plasma membrane rather than on its cytosolic concentration, is strongly substantiated.

Blood Platelets↗

PubMed1996

Effects of storage on in vitro platelet responses: comparison of ACD and Na citrate anticoagulated samples.

The present study was designed to evaluate whether the use of acid citrate dextrose (ACD) Formula A may enhance the survival of platelets during storage, thus allowing the continuance of platelet studies over the period of 2-3 hours usually recommended. For this purpose the effects of time on in vitro platelet response to several agonists have been investigated in platelet-rich plasma (PRP) obtained from blood samples anticoagulated with either Na citrate or ACD Formula A. The analysis of the data obtained in in vitro platelet aggregation studies using various parameters and at different time points demonstrated that storage of PRP obtained from citrated samples caused a marked reduction of platelet responses. This reduction was already evident after 6 hours, and a strong decrease was observed after 8 hours with all the agonists used. On the other hand, storage of ACD anticoagulated blood did not cause any significant decrease of platelet responsiveness up to 6 hours. A reduction of platelet aggregation became evident only after 8 hours, but not to the same extent as the one observed in citrated samples. Therefore, it may be concluded that the use of ACD Formula A as anticoagulant is capable of maintaining a normal platelet responsiveness up to 6-8 hours, thus permitting the investigation of platelet function for periods of time over those commonly recommended.

Adenosine Diphosphate↗

PubMed1995

Acid citrate dextrose (ACD) formula A as a new anticoagulant in the measurement of in vitro platelet aggregation.

To evaluate whether the use of ACD Formula A may affect in vitro platelet function, blood samples were obtained from 21 healthy blood donors and anticoagulated in ACD (acid-citrate dextrose, NIH Formula A), Na citrate 3.8%, and K3EDTA. Platelet count, mean platelet volume, and in vitro platelet aggregation were evaluated on each sample. No significant difference was observed in platelet count and mean platelet volume among the different samples. Conversely, the ACD treated platelets showed a higher reactivity to the agonists as demonstrated by a significant increase of the maximum percentages of aggregation induced by ADP, epinephrine, and collagen, as well as a significant decrease of secondary aggregation thresholds to ADP and epinephrine. In conclusion, it may be speculated that ACD Formula A is capable of better maintaining the intraplatelet signal transduction mechanisms during PRP preparation, thus improving the overall responsiveness of platelets.

Adult↗

PubMed1995

Use of a rapid and simple method to extract proviral DNA in the identification of HIV-1 by PCR.

DNA extraction is a critical step in PCR analysis and is closely related to its sensitivity. Traditional methods, based on phenol-chloroform extraction, require more time and the use of toxic reagents. GeneReleaser (Bio Ventures Inc.) is a commercial product which releases DNA from whole blood, cell cultures, bacterial colonies and the like. Cells lysis and DNA extraction are accomplished directly in the amplification tube on a thermocycler. We used GeneReleaser in the identification of HIV-1 proviral DNA by PCR on whole blood samples. All samples arrived at our laboratory for HIV-1 detection were treated with two different procedures. The classical one was based on the lysis of separated lymphocytes by proteinase K, while the other consisted in DNA extraction by GeneReleaser from 5 microliters of whole blood in sodium citrate. All samples were amplified for HIV-1 GAG region; to prevent carry-over contamination Uracil N-glycosylase (UNG) sterilization was performed. Amplified sequences were revealed using the DEIA commercial system (Sorin Biomedica, Italy). To verify the suitability both of cell lysates and GeneReleaser DNA-extracted samples for PCR, we amplified a specific sequence of HLA-DQ-alpha gene. Initial data indicate that this new method might reduce the performance time of PCR (DNA extraction time was around 15 minutes) and improve PCR sensitivity.

Adult↗

PubMed1994

Effect of picotamide on the calcium mobilization and phospholipase C activation in human platelets.

The effect of picotamide (G137 or N,N'-bis-3-picolyl-4-methoxyisophthalamide), a dual thromboxane A2 (TxA2) synthetase inhibitor/TxA2 endoperoxide receptor antagonist, on the phospholipase C (PLC) activation and calcium mobilization in human platelets stimulated by arachidonic acid (AA) and TxA2 receptor synthetic agonist U46619, has been studied. Preincubation with picotamide (10(-4) M) for 1 and 3 min significantly reduced (p < 0.03 and p < 0.005 respectively) the calcium concentration changes induced in gel-filtered platelets (GFPs) by U46619 125 nM and 250 nM. Picotamide also reduced the calcium concentration changes induced in GFPs by AA 75 and 150 microM and by ADP 5 and 10 microM. In thrombin degranulated platelets picotamide inhibited the effect of U46619 up to 500 nM. The PLC activation, as indicated by inositol-1,3,4 P3 (Ins 1,3,4 P3) formation in response to U46619 250 nM and AA 150 microM was also inhibited by picotamide. These results may suggest a dual effect of picotamide on the receptor/effector systems through which TxA2 mediates platelet activation.

15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5↗

PubMed1984

[Intravenous glucose tolerance test during muscular exercise].

We have valued in 8 individuals subjected to muscular moderate work the degree of use of glucose in the body tissues, measured by means of K-Conard, and the role that holds the insulin hormone in this particular condition. The IVGTT was carried out in 8 individuals, at first, in condition of relax. In the same individuals, the day after, the IVGTT was carried out after 5 minutes from the beginning of muscular work to cycloergometer set a 50 watts for a term of 30 minutes. The study demonstrates that, during the muscular moderate work, we have an improvement, statistically significant, of the use of glucose, measured by means of K-Conard (t = 2.42; P less than 0.05). Then, it demonstrates that the muscular moderate work has not any influence on plasmatic concentrations of insulin.

Adult↗

PubMedDate not supplied

Metformin decreases platelet superoxide anion production in diabetic patients.

BACKGROUND: Patients with type 2 diabetes mellitus are usually treated with oral antidiabetic agents but it is still not known whether these drugs have antioxidant effects in humans. METHODS: We studied 60 patients with type 2 diabetes mellitus, divided into three groups on the basis of hypoglycaemic treatment (Group A: metformin, Group B: glibenclamide, Group C: diet). All patients were followed for at least 1 year. The three subgroups had similar clinical characteristics. Twenty healthy subjects, of comparable sex and age, were enrolled as controls. In each subject, platelet production of superoxide anion (O(2)(-)) elicited by collagen, was determined by lucigenin assay. RESULTS: Patients with diabetes had higher platelet O(2)(-) production than controls; no correlation was observed between blood glucose and platelet O(2)(-) production. Group A patients had platelet O(2)(-) production similar to that of healthy subjects but lower than Group B and Group C patients. CONCLUSION: The present findings suggest an in vivo antioxidant activity of metformin and warrant prospective studies to further explore this hypothesis.

Antioxidants↗