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F Pugliese

Publications and source records attributed to F Pugliese.

At least 55 records · Page 3Linked to original sources
PubMed1994

Calcium release-activated calcium influx in cultured human mesangial cells.

Ca2+ influx is a major component of the response of cultured human mesangial cells (HMC) to vasoconstrictors. Activators of phospholipase C such as angiotensin II (Ang II) release Ca2+ from intracellular stores and enhance Ca2+ influx, which in turn is modulated by Na+/Ca2+ exchange. By microfluorometry we studied the mechanisms of Ca2+ entry in resting and stimulated fura-2-loaded monolayers or single HMC. Addition of 1 to 10 mM extracellular Ca2+ to cells equilibrated in Ca(2+)-free media resulted in a rapid, persistent elevation of free cytosolic Ca2+ ([Ca2+]i), from 52 +/- 5 to 113 +/- 18 and 226 +/- 37 nM, respectively. Ca2+ influx was blocked by lanthanum or chelation with EGTA, while it was only partially inhibited by voltage-operated Ca2+ channel (VOC) blockers, such as nifedipine or verapamil. The rise of [Ca2+]i at high external [Ca2+] was not due to a Ca(2+)-sensing mechanism with release of intracellular stored Ca2+, since it was prolonged, and it was not seen in cells maintained in normal 1.25 mM [Ca2+] media. Moreover, it was not abolished by prior depletion of Ca2+ stores with 0.5 microM thapsigargin or 5 microM ionomycin in Ca(2+)-free media, which transiently increased [Ca2+]i (to 281 +/- 39 and 380 +/- 51 nM, respectively). On the contrary, both agents markedly potentiated Ca2+ influx upon addition of 1 to 10 mM [Ca2+]e, (to a maximum of 686 +/- 111 and 633 +/- 150 nM, P < 0.05 vs. control).(ABSTRACT TRUNCATED AT 250 WORDS)

Angiotensin II↗
PubMed1994

Amlodipine versus ramipril in the treatment of mild to moderate hypertension: evaluation by 24-hour ambulatory blood pressure monitoring.

This study was designed to compare the efficacy and tolerability of amlodipine (AML) and ramipril (RAM) administered once a day in patients affected by mild to moderate primary systemic hypertension. Twenty outpatients, 12 men and 8 women (age range 35-64 years), were enrolled. The patients received single-blind placebo for 2 weeks and thereafter in a double-blind, randomized crossover sequence AML (10 mg) and RAM (5 mg), both for 4 weeks. At the end of each period, the patients underwent 24-hour noninvasive blood pressure monitoring with readings taken every 10 min during daytime (from 07.00 to 23.00 h) and 20 min during nighttime (from 23.00 to 07.00 h). Both AML and RAM induced a highly significant (p < 0.0001) decrease in blood pressure from 162/103 +/- 7/3 to 132/82 +/- 6/6 and 135/83 +/- 6/5 mm Hg, respectively. The mean blood pressure decreased from 122 +/- 5 to 99 +/- 6 (AML; p < 0.0001) and 100 +/- 5 mm Hg (RAM; p < 0.0001). No significant differences in heart rate were noted during drug administrations. Treatment did not have to be discontinued in any patient because of adverse reactions. In conclusion, both AML and RAM reduced the blood pressure markedly, even if AML proved to be significantly more effective than RAM.

Adult↗
PubMed1994

Mechanisms of glucose-enhanced extracellular matrix accumulation in rat glomerular mesangial cells.

In view of the importance of mesangial extracellular matrix (ECM) accumulation in the pathogenesis of diabetic glomerulosclerosis, we investigated 1) the effects of high glucose on ECM production by rat glomerular mesangial cells in culture (study A) and 2) the mechanisms underlying these effects, particularly the role of high sugar levels irrespective of intracellular metabolism (study B1) and of excess glucose disposal via the polyol pathway and associated biochemical alterations (study B2). Cells were cultured for 4 weeks, through six to eight passages, under the experimental conditions indicated below and, at each passage, the levels of fibronectin (FN), laminin (LAM), and collagen types I (C-I), III (C-III), IV (C-IV), and VI (C-VI) in media and cell extracts were quantified by an enzyme immunoassay. In study A, medium and cell content of matrix were assessed, together with [3H]leucine and [3H]thymidine incorporation into monolayers, polyol, fructose, and myo-inositol levels and the cytosolic redox state, in cells grown in high (30 mM) D-glucose or iso-osmolar mannitol versus cells cultured in normal (5.5 mM) D-glucose. FN, LAM, C-IV, and C-VI accumulation, but not C-I and C-III accumulation, was increased by 30 mM glucose, but not by iso-osmolar mannitol, when compared with 5.5 mM glucose, starting at week 2 and, except for C-VI, persisting throughout the remaining 2 weeks, whereas no change was observed in the measured indexes of total protein synthesis and DNA synthesis/cell proliferation. At any time point, polyol levels were increased, whereas myo-inositol was reduced by high glucose; in cells grown under elevated glucose concentrations, the lactate/pyruvate (L/P) ratio, an index of the cytosolic redox state, progressively increased. In study B1, the effects of high D-glucose were compared with those of iso-osmolar concentrations of sugars that are partly or not metabolized but are capable of inducing nonenzymatic glycosylation, such as D-galactose and L-glucose, and of mannitol, which does not enter the cell. Both D-galactose and L-glucose, but not mannitol, partly mimicked D-glucose-induced ECM overproduction. Although D-galactose is metabolized via the polyol pathway and alters the cytosolic redox state, ECM changes induced by high galactose were not prevented by the use of an aldose reductase inhibitor (ARI), Alcon 1576 (14 microM).(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1994

[Isradipine versus diltiazem in the treatment of stable effort angina pectoris: ergometric evaluation in a crossover double-blind study].

OBJECTIVES: To evaluate the effects of isradipine (ISR) and diltiazem (DIL) on exercise tolerance and ischemic ST depression in patients with stable effort angina. METHODS: Fourteen out-patients, 9 males and 5 females, aged 46-65 years (mean +/- SD = 57 +/- 8), with ischemic heart disease and reproducible ST-segment depression on two consecutive exercise stress tests in baseline conditions, underwent a study consisting of 4 periods: 1 and 3 placebo, 2 and 4 at random ISR (5 mg b.i.d.) and DIL (120 mg b.i.d.). At the end of each period a multistage treadmill exercise stress test (Bruce protocol) was performed. RESULTS: Both drugs significantly (p < 0.001) increased ischemia time (IT) (0.1 mV ST depression) as compared to placebo, from 438 +/- 132 s. to 620 +/- 164 s. (ISR) and 583 +/- 147 s. (DIL) without statistical difference between two drugs (p = 0.2), and significantly reduced (p < 0.002) the maximal ST depression, from -0.20 +/- 0.11 mV to -0.07 +/- 0.07 mV (ISR) and -0.09 +/- 0.11 mV (DIL). At the IT, systolic blood pressure increased (p = 0.02), from 180 +/- 19 mm Hg to 187 +/- 15 mm Hg (ISR) and 191 +/- 15 mm Hg (DIL); similarly, heart rate increased from 133 +/- 24 bpm to 144 +/- 18 bpm (ISR: p = 0.002) and 140 +/- 17 bpm (DIL: p = NS). CONCLUSIONS: ISR and DIL, at the above dosage have showed an important and significant anti-ischemic effect (IT = +41.5% during ISR and +33.1% during DIL).

Aged↗
PubMed1993

High glucose inhibits cytosolic calcium signaling in cultured rat mesangial cells.

Glomerular vasodilatation in the early stages of type I diabetes mellitus apparently results from arteriolar insensitivity to vasoconstrictors. Since cytosolic free calcium ([Ca2+]i) is a major signaling mechanism for smooth muscle contraction, we studied whether growth of smooth muscle-like rat glomerular mesangial cells in media with high glucose concentration affects [Ca2+]i responses to vasoconstrictors. In cells grown for five days in 22 mM glucose, we observed blunted responsiveness to three structurally unrelated vasoconstrictors that elevate [Ca2+]i via a phospholipase C-dependent mechanism, angiotensin II, prostaglandin F2 alpha, and arginine vasopressin. Inhibition of [Ca2+]i responses was not due to an osmotic effect of high glucose, since it was not mimicked by hypertonic mannitol. While the size of intracellular Ca2+ pools was unaffected by elevated glucose, Na+/Ca2+ exchange was markedly inhibited, thus ruling out both impaired filling of Ca2+ stores and enhanced counter-regulatory mechanisms. Impaired myoinositol transport or intracellular sorbitol accumulation were not responsible for the effects of high glucose, since supplementation of media with myo-inositol or with the aldose reductase inhibitor. Alcon 1576, failed to reverse insensitivity to vasoconstrictors. On the other hand, down-regulation or pharmacological inhibition of protein kinase C completely reversed the effects of high glucose, thus indicating involvement of this signal transduction pathway. These data suggest a possible intracellular mechanism for the impaired vascular sensitivity underlying early renal hemodynamic changes in diabetes mellitus.

Animals↗
PubMed1993

Urinary styrene in the biological monitoring of styrene exposure.

The urinary excretion of styrene represents a promising indicator of exposure to this solvent. Nevertheless extensive research under field conditions is scant. In this investigation 214 styrene-exposed workers from 10 fiberglass-reinforced plastics factories were studied. Environmental monitoring was performed by personal passive sampling. Blood styrene and the urinary excretion of styrene and its main metabolites, mandelic acid (MA) and phenylglyoxylic acid (PGA), were measured. The correlation coefficient between the time-weighted average of environmental styrene and the mean urinary excretion of styrene was 0.88 (0.91 after logarithmic transformation), compared with the 0.82 and 0.78 of the end-of-shift MA and PGA values, respectively. A high correlation (0.86) was also found between styrene in the blood and urine. The results, obtained under field conditions with a large group of exposed workers, confirm the usefulness of the urinary excretion of styrene as an exposure index for the biological monitoring of styrene exposure.

Adolescent↗
PubMed1993

[An ergometric cross-over evaluation of the anti-ischemic efficacy of amlodipine and gallopamil in stable angina of effort].

In the setting of stable effort angina a single-blind, randomized, cross-over study to evaluate the effects of gallopamil (GAL) and amlodipine (AML) on exercise tolerance and ischemic ST depression was conducted. Fifteen outpatients, 12 males and 3 females, aged 40-65 years (57 +/- 9), with documented coronary atherosclerosis and reproducible ST-segment depression on 2 consecutive baseline exercise stress tests, completed the study, which consisted of 4 periods: 1 and 3 placebo, 2 and 4 at random GAL (50 mg tid) and AML (10 mg/daily). At the end of each period a multistage treadmill exercise stress test (Bruce protocol) was performed. Both drugs significantly (p = 0.0001) increased the ischemia time (IT) (0.1 mV ST depression) as compared to placebo, from 416 +/- 165 s to 635 +/- 161 s (GAL) and 607 +/- 152 s (AML) with significant difference (p = 0.2) between the 2 drugs, and reduced significantly (p = 0.001) the maximal ST depression from -0.25 +/- 0.09 mV to -0.11 +/- 0.08 mV (GAL) and -0.12 +/- 0.09 mV (AML). At the IT, the systolic blood pressure increased from 178 +/- 23 mmHg to 185 +/- 20 mmHg (GAL) and remained unchanged during AML treatment (178 +/- 15 mmHg); similarly, the heart rate increased from 126 +/- 22 b/min to 139 +/- 21 b/min (GAL) and 138 +/- 19 b/min (AML). In conclusion, both GAL and AML showed a good anti-ischemic effect (IT = +52.6% during GAL and +45.9% during AML), even if GAL proved to be significantly more effective than AML.

Adult↗
PubMed1992

Signal transduction in mesangial cells.

Phenotype, growth, and functional characteristics of glomerular mesangial "myofibroblasts" are under the control of multiple hormones, vasoactive agents, autacoids, and cytokines. Several parallel signal transduction pathways couple receptor occupancy with functional changes, including phospholipases C, A2, and D breakdown of membrane phospholipids, and adenylate/guanylate cyclase activation. Changes of cytosolic ion concentrations, cyclic nucleotide accumulation, and eicosanoid biosynthesis are currently interpreted as intracellular signals for protein kinase activation. Phosphorylation of multiple substrates by serine/threonine kinases C, A, and G or by tyrosine kinases directly coupled to receptors, is a final step in cell activation. Cross-talk between signal transduction pathways, along with the release of eicosanoids and cytokines acting as intercellular mediators, provides the potential for interactive regulation of glomerular cell functions.

Adenylyl Cyclases↗
PubMed1992

Regulation of cultured human mesangial cell growth by ionized macromolecules.

We evaluated the importance of the net charge of polyionic macromolecules in the regulation of cultured human mesangial cell growth. Structurally unrelated polyanionic compounds, i.e., heparin, suramin, poly-L-aspartic acid, and poly-L-glutamic acid, strongly inhibited 10% fetal bovine serum-stimulated cell proliferation. On the other hand, two polycations, protamin sulfate and poly-L-lysine, were equally effective in inhibiting cell growth. The antiproliferative activity of each compound was neutralized by molecules with opposite net charge. These data indicate that both anionic and cationic macromolecules exert an antimitogenic effect on cultured human mesangial cells. This inhibitory effect is dependent upon charge density rather than on the net electric charge of each compound.

Cell Division↗
PubMed1992

Thromboxane A2 regulates protein synthesis of cultured human mesangial cells.

Cultured human glomerular mesangial cells express functional receptors for vasoconstrictor metabolites of arachidonate, such as thromboxane A2. Binding of thromboxane A2 analogs triggers phosphatidylinositol breakdown and a rise of intracellular free Ca2+, followed by complex effects on cell growth. We have evaluated the effects of the thromboxane A2 mimetic, U-46619, on the biosynthesis of human mesangial cell proteins. After 4 to 24 hours of incubation with U-46619, cells were metabolically labeled with tritiated leucine or sulfur 35-methionine and analyzed by one-dimensional and two-dimensional sodium dodecylsulfate polyacrylamide gel electrophoresis and fluorography. U-46619 modestly stimulated total protein synthesis in both quiescent and cycling cells to a maximum of 12% and 23% above control at 1 mumol/L after 24 hours, respectively. The eicosanoid selectively enhanced the labeling of crude membrane and cytosolic proteins of molecular weights of approximately 38 to 53 kd and 125 to 200 kd in the presence or absence of serum, respectively. Cellular tubulin and collagen type IV, but not actin, were markedly stimulated, as determined by immunoblotting of cell-associated proteins. On the contrary, U-46619 potently inhibited labeling of soluble proteins that were released into the media, to a maximum of 49% after 24 hours. Inhibition was confirmed by immunoblotting of collagen type IV. Intraglomerular accumulation of thromboxane A2 during chronic inflammation may modify the functional characteristics of the mesangium by regulation of the biosynthesis of structural proteins.

15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5↗
PubMed1991

Regulation of Na(+)-Ca2+ exchange in cultured human mesangial cells.

Na(+)-Ca2+ exchange contributes to regulation of cytosolic free Ca2+ levels ([Ca2+]i) of cultured human mesangial cells following phospholipase C stimulation, as shown by larger responses to vasoconstrictors such as angiotensin II (ANG II) or endothelin 1 in Na(+)-free media. In turn, previous activation of phospholipase C by vasoconstrictors significantly enhances the amplitude of the [Ca2+]i elevation induced by Na+ withdrawal. We studied the mechanisms of upregulation in monolayer cultures loaded with the fluorescent Ca(2+)-sensitive probe fura-2. The exchanger was stimulated by insulin and inhibited by chronic exposure to serum. A rise of [Ca2+]i was not sufficient per se to enhance exchange activity, as prior elevation of [Ca2+]i with the ionophores ionomycin or 8-bromo-A23187 failed to augment the response to Na+ withdrawal. Protein kinase C (PKC) activation by phorbol 12-myristate-13-acetate (PMA), alone or in combination with a rise of [Ca2+]i, potently inhibited basal and vasoconstrictor-enhanced Na(+)-Ca2+ exchange. Suppression of the effects of ANG II was not due to frustrated phospholipase C activation by PMA, because addition of PMA after ANG II also inhibited Na(+)-Ca2+ exchange. PKC downregulation by 24-h pretreatment with PMA or inhibition with 1-(5-isoquinolinesulfonyl)-2-methylpiperazine or staurosporine did not prevent activation by ANG II. The exchanger was markedly potentiated by Na+ loading the cells with gramicidin D or reducing extracellular K+. ANG II failed to stimulate Na(+)-Ca2+ exchange when added in the absence of extracellular Na+. Therefore vasoconstrictors promote Na(+)-Ca2+ exchange by a mechanism independent of [Ca2+]i and PKC while presumably linked to Na+ influx.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed1991

Serotonin and the glomerular mesangium. Mechanisms of intracellular signaling.

Serotonin, a release product of activated platelets, stimulates proliferation and prostaglandin synthesis in cultured smooth muscle-like glomerular rat mesangial cells by activation of phospholipase and protein kinase C. To further characterize the signaling mechanisms used by serotonin, we monitored its effects on intracellular free Ca2+, pH, and membrane potential of cultured rat mesangial cells with sensitive fluorometric techniques. Activation of a 5-HT2 receptor, blocked by the specific receptor antagonists ketanserin and ritanserin, triggered immediate discharge of intracellular Ca2+ stores. The resulting rise of cytosolic free Ca2+ was accompanied by simultaneous membrane depolarization and followed within 30-60 seconds by prolonged cytosolic alkalinization. Depolarization and cytosolic free Ca2+ elevation were persistent in the continued presence of serotonin and were rapidly reversed by competitive receptor displacement with ketanserin or ritanserin. Depolarization is secondary to enhanced Cl- conductance, whereas it is relatively independent of Na+, K+, and Ca2+ fluxes. The putative Cl- channel is regulated by Ca2+ since ionomycin and other stimuli of cytosolic free Ca2+ mimic the effects of serotonin on membrane potential, whereas serotonin-induced depolarization is blunted in cells pretreated with the intracellular Ca2+ chelator BAPTA. Cytosolic alkalinization occurs in HCO3(-)-free solutions resulting from enhanced activity of a Na(+)-H+ exchanger and blocked by extracellular Na+ removal or amiloride. In the presence of HCO3-, serotonin elicits a persistent acidification, revealing simultaneous enhancement of a Na(+)-independent Cl(-)-HCO3- countertransport. These findings indicate multiple pathways for contraction and long-term functional changes induced by serotonin in mesangial cells, with potential relevance to glomerular and systemic hypertension.

Animals↗
PubMed1990

Identification and characteristics of a Na+/Ca2+ exchanger in cultured human mesangial cells.

Excitable cells express Na+/Ca2+ exchange activity among other mechanisms modulating rapid fluctuations of cytosolic free Ca2+ ([Ca2+]i). We studied functions and regulation of a Na+/Ca2+ exchanger in cultured human glomerular mesangial cells. Fura-2-loaded confluent monolayers reacted to removal of ambient Na+ with an immediate, transient elevation of [Ca2+]i, assessed by single/dual wavelength fluorometry. Peak [Ca2+]i was inversely correlated with the extracellular Na+ concentration. Ca2+ influx was the sole mechanism implicated, as the [Ca2+]i rise was prevented by EGTA. The process was inhibited by 1 mM amiloride, but not by blockers of voltage-operated Ca2+ channels. Re-addition of Na+ resulted in a rapid decrease of [Ca2+]i, indicating bimodal operation of the exchanger. Na(+)-loading the cells with monensin and ouabain enhanced Ca2+ uptake. Prior stimulation of [Ca2+]i with the thromboxane A2 mimetic, U-46619, or angiotensin II also increased Ca2+ uptake upon subsequent Na+ removal, suggesting induction of the exchanger by vasoconstrictors. Moreover, the magnitude of agonist-induced [Ca2+]i transients was amplified by Na+ removal, indicating that the exchanger modulates the effects of vasoconstrictors. These results demonstrate that an inducible Na+/Ca2+ antiporter is operative in resting and stimulated human mesangial cells, further confirming their smooth muscle origin and potential regulatory role on glomerular hemodynamics.

Calcium↗
PubMed1990

Glomerular polyanion and control of cell function.

The glomerular polyanion comprises all anionic sites of glomerular cell surfaces, basement membranes and extracellular matrix. Charged structures may play a critical pathophysiologic role within the glomerular microcirculation, as loss of charges results in altered permselectivity of the filtration barrier and proteinuria. Neutralization of cell surface negative charges of cultured glomerular epithelial and mesangial cells by the polycation poly-L-lysine (PL) is accompanied by increased prostanoid synthesis. Recent studies have shown that PL enhances mesangial cell proliferation in culture. Conversely, the polyanion heparin prevents the effect of PL, and inhibits the growth-stimulatory effect of serum. Thus, our data suggest that the glomerular polyanion, in addition to maintaining the integrity of the filtration barrier, regulates key cell functions such as eicosanoid biosynthesis and proliferation.

Animals↗
PubMed1990

Cellular basis of hormonal actions in the glomerulus.

Mesangial cells are contractile pericytes of the kidney glomerulus. Mesangial contraction/relaxation contributes to the regulation of glomerular hemodynamics. Additionally, mesangial cells process filtered macromolecules, synthesize extracellular matrix, respond to and release a number of cytokines and vasoactive mediators. Cultured mesangial cells express receptors for circulating and local agents that affect glomerular function. These receptors are coupled to distinct signaling pathways, namely phospholipase C and A2, transducing vasoconstrictor stimuli, and adenylate/guanylate cyclase, transducing vasodilators. Early intracellular signals include changes of cytosolic ions and cyclic nucleotides. They translate into short-term responses, such as cell depolarization and contraction, and later events, such as prostanoid synthesis and cell proliferation. Studies of mesangial cell behavior in culture may largely enhance our current understanding of glomerular pathophysiology.

Animals↗
PubMed1989

Fractional conversion of thromboxane B2 to urinary 11-dehydrothromboxane B2 in man.

Thromboxane (TX) B2, the chemically stable hydration product of pro-aggregatory TXA2, undergoes two major pathways of metabolism in man, resulting in the formation of 2.3-dinor-TXB2 and 11-dehydro-TXB2, respectively. We have measured the excretion of the latter during the infusion of exogenous TXB2 over a 50-fold dose range in order to examine the fractional conversion of TXB2 to urinary 11-dehydro-TXB2 and to re-assess the rate of entry of endogenous TXB2 into the circulation. Four healthy male volunteers received 6-h intravenous infusions of the vehicle alone and TXB2 at 0.1, 1.0 and 5.0 ng.kg-1.min-1 in random order. They were pretreated with aspirin 325 mg/d in order to suppress endogenous TXB2 production. Urinary 11-dehydro-TXB2 and 2,3-dinor-TXB2 were measured before, during and up to 24 h after the infusions and in aspirin-free periods, by means of NICI-GC/MS-validated radioimmunoassays. Aspirin treatment suppressed urinary 11-dehydro-TXB2 by 91%. The fractional elimination of 11-dehydro-TXB2 was independent of the rate of TXB2 infusion and averaged 6.8 +/- 0.7%, as compared to 6.4 +/- 0.9% for 2,3-dinor-TXB2. Interpolation of 11-dehydro-TXB2 values obtained in aspirin-free periods onto the linear relationship between the quantities of infused TXB2 and the amount of metabolite excreted in excess of control values (y = 0.0058x, r = 0.94, P less than 0.001) permitted calculation of the mean rate of entry of endogenous TXB2 into the circulation as 0.12 ng.kg-1.min-1. We conclude that: (a) urinary 11-dehydro-TXB2 is at least as abundant a conversion product of exogenously infused TXB2 as 2,3-dinor-TXB2; (b) its excretion increases linearly as a function of the rate of entry of TXB2 into the circulation up to approx. 40-fold the calculated rate of secretion of endogenous TXB2; (c) the latter is consistent with previous estimates based on monitoring of the beta-oxidation pathway of TXB2 metabolism.

Adult↗