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E Baldi

Publications and source records attributed to E Baldi.

At least 73 records · Page 4Linked to original sources
PubMed1995

Identification and characterization of functional angiotensin II receptors in human neuroblastoma cells.

The presence of specific AII receptors in 6 different human neuroblastoma cell lines was investigated using binding, cAMP and [Ca2+]i studies. We found high affinity (0.1 nM), low capacity ((1-2).10(3) sites/cell) binding sites for [125I](Sar-1,Ile-8)AII in one half of the cell lines studied. In the positive cell lines mathematical modeling of multiple competition curves among AII and analogs strongly indicated the presence of a homogenous class of sites, i.e., AT1 receptors. The presence of AT1 receptors was further substantiated by AII-induced inhibition of VIP-stimulated cAMP levels and by AII-evoked [Ca2+]i transient. The density of AT1 receptors in neuroblastoma cells was not affected by treatment with pertussis toxin and retinoic acid but was significantly increased by subacute treatment with VIP. In neuroblastoma cells, AII does not stimulate DNA synthesis, suggesting other roles rather than mitogenesis. Neuroblastoma cells represents an interesting model to investigate the function of AII in neural crest derived tissues.

1-Sarcosine-8-Isoleucine Angiotensin II↗
PubMed1995

Stimulation of protein tyrosine phosphorylation by platelet-activating factor and progesterone in human spermatozoa.

Tyrosine phosphorylation of proteins is involved in several sperm functions, including capacitation, motility, and acrosome reaction of spermatozoa. This study was undertaken to determine changes of tyrosine phosphorylation during 'in vitro' capacitation as well as the ability of platelet-activating factor (PAF) and progesterone (P), two known activators of sperm functions, to stimulate tyrosine phosphorylation of human sperm proteins. Spermatozoa were capacitated in BSA-containing medium and incubated with PAF (10-1000 nM) and progesterone (0.1-1 microgram/ml). After SDS-PAGE, sperm proteins were transferred to nitrocellulose and tyrosine phosphorylated proteins immunodetected by reacting with anti-phosphotyrosine antibody. The antibody mainly reacted with two proteins of approximately 97 and 75 kDa. The level of phosphorylation increased in these two proteins as a function of capacitation time, with a maximum between 120 and 180 min. In addition, phosphorylation in these two proteins was increased in capacitated spermatozoa by treatment with progesterone and PAF and was greatly reduced by pre-incubation with the tryosine kinase inhibitor erbstatin. Furthermore, pre-incubation with the two tyrosine kinase inhibitors erbstatin and genistein inhibited the induction of acrosome reaction by progesterone and, partially, by PAF. Our results suggest a role for tyrosine kinase(s) in the mechanism of capacitation and activation of human spermatozoa by PAF and progesterone.

Acrosome↗
PubMed1995

Time-dependent deficits of rat's memory consolidation induced by tetrodotoxin injections into the caudate-putamen, nucleus accumbens, and globus pallidus.

In order to better define corpus striatum involvement in memory processing, tetrodotoxin functional ablation was employed to assess the respective roles of the caudate-putamen as a whole and of its anterior, median, and posterior regions, of the nucleus accumbens, and of the globus pallidus during early passive avoidance response consolidation of the rat. Under general ketamine anesthesia (100 mg/kg) by means of microinjections, tetrodotoxin (10 ng/microliter) was bilaterally administered at the appropriate stereotaxic locations of the chosen sites and subsites at different postacquisition delays (0.25, 1.5, and 6 h). Saline solution was administered to control subjects in the same sites. Rats always underwent retrieval testing 48 h after treatment, i.e., when tetrodotoxin blockade had completely disappeared. The results show that both total caudate-putamen and median caudate-putamen blockades caused passive avoidance responding impairment when performed at 0.25- and 1.5-h postacquisition delays, that caudate-putamen anterior and caudate-putamen posterior blockades were never followed by passive avoidance impairment, and that both nucleus accumbens and globus pallidus blockades were followed by passive avoidance impairment when performed at 0.25- and 1.5-h postacquisition delays. In all structures, tetrodotoxin administration caused no impairment at the 6-h delay. The results are discussed with reference both to previous results obtained after reversible or irreversible lesions of the same structures and to the functional mnemonic relationships of the corpus striatum and its components with other subcortical structures, particularly the substantia nigra.

Animals↗
PubMed1995

Actions of progesterone on human sperm: a model of non-genomic effects of steroids.

Non-genomic actions of steroids have been extensively studied in the last few years. Among these actions, the non-genomic effect of progesterone (P) on human spermatozoa appears to be very promising, in view of the dramatic effect of this steroid on intracellular calcium, activation of tyrosine kinase, and induction of acrosome reaction. We have shown that the ability of spermatozoa to respond to P increases during the process of capacitation and is not counteracted by the P-receptor antagonist RU486 nor by the GABAA antagonists bicuculline and picrotoxin. We have also shown that P increases tyrosine phosphorylation of a sperm protein of about 97 kDa, suggesting activation of tyrosine kinase(s). In addition, we found that P induces a perturbation of sperm membrane phospholipid metabolism resulting in an increase of synthesis of platelet-activating factor and liberation of arachidonic acid. Results of these biochemical studies indicate that P is able to stimulate several signal transduction pathways in human sperm. We have also investigated responsiveness to P in sperm of oligozoospermic subjects as well as of men undergoing an in vitro fertilization (IVF) program. Our results show that the percentage increases of intracellular calcium and acrosome reaction in response to P is significantly reduced in oligozoospermic men as well as in subjects with reduced fertilization rate. Moreover, in the latter subjects response to P is highly significant correlated to fertilization rate of oocytes. These studies indicate that a biochemical alteration of sperm in their capacity to respond to P might be responsible for reduced fertilizing ability.

Acrosome↗
PubMed1995

Intracellular calcium increase and acrosome reaction in response to progesterone in human spermatozoa are correlated with in-vitro fertilization.

In this study we have investigated responsiveness to progesterone in spermatozoa from a group of unselected male partners of couples undergoing in-vitro fertilization (IVF). We evaluated progesterone-stimulated intracellular Ca2+ ([Ca2+]i) and percentage increase in acrosome reaction in the same sperm sample used for oocyte inseminations. [Ca2+]i was measured with a fluorimetric method, while the acrosome reaction was assessed using a fluorescent probe (fluorescein isothiocyanate-labelled peanut lectin). The average percentage [Ca2+]i as well as the rate of increase in the frequency of acrosome reaction following progesterone challenge were significantly lower (P < 0.005) in the group of patients with a fertilization rate < 50%. In addition, significant correlations between the fertilization rate and the progesterone-stimulated [Ca2+]i and acrosome reaction increases (r = 0.78 and r = 0.79 respectively) were observed. Furthermore, in cases of fertilization failure, no increase of [Ca2+]i or acrosome reaction was observed in response to progesterone with the exception of one case. Our results indicate that [Ca2+]i and acrosome reaction increases in response to progesterone can be of value in the prediction of sperm fertilizing ability. As the two parameters were significantly correlated to each other (r = 0.86), the two assays have similar IVF predictive value and might be used interchangeably as a diagnostic tool in the assignment of male patients to the different kinds of assisted fertilization techniques.

Acrosome↗
PubMed1995

Transitory effects of intracerebral administration of protein synthesis inhibitors on rat's spontaneous behavior.

In order to evaluate whether the effects of the local administration of protein synthesis inhibitors are transient or permanent, small volumes of cycloheximide, 2-deoxy-d-galactose, tetrodotoxin, and saline solution were stereotaxically administered in the amygdala, substantia nigra, and nucleus basalis magnocellularis of naive male adult Wistar rats. Three spontaneous activities (feeding, drinking, locomotion) were continuously recorded. 12 groups of animals were housed in activity cages for 6 consecutive days with a light/dark schedule (LD 12:12). The active compounds at doses sufficient to cause only limited effects were administered at the beginning of the light period of the fourth day. During the initial three days spontaneous activities remained constant, following the normal circadian distribution of rodents. Cycloheximide and tetrodotoxin administrations were followed by transient site-specific and activity-specific changes, which occurred only during the fourth and fifth day. The results show that protein synthesis inhibitors can be locally administered into the CNS at doses affecting only limited subcortical structures. The observed modifications of the three spontaneous activities were all transient and were always followed by complete recovery within 48 hours, thus confirming previous biochemical data. Therefore, the local injection of protein synthesis inhibitors, like tetrodotoxin, lidocaine or other compounds, can be employed to induce transient functional effects on well delimited brain structures in order to study the time-course of the mechanism of mnemonic trace formation.

Amygdala↗
PubMed1995

Energy expenditure (EE) and substrate utilization (SU) in the perioperative period in orthotopic liver transplantation (OLTX).

12 patients were investigated with IC (Datex, Deltatrac) preoperatively and during the surgical procedure of OLTX, VO2, VCO2 were continuously measured and RQ, EE and SU were calculated considering the different periods of the procedure: preoperative resting EE: (PREE), anaesthesiological procedures (ANEE), liver preparation EE (LPEEE), liver removal EE (LREE), anhepatic phase EE (APEE), reperfusion, EE (RPEE) and end of operation EE (EOEE). EE were expressed as % respect value calculated with H.B. (Harris-Benedict) formula. Data were analyzed with Student T-test and p < 0.01** or < 0.05* PREE is typical in end stage liver disease with low RQ values and increased EE. Energy production depends on lipid utilization since liver gluconeogenesis and glycogen stores are impaired. Anesthesia reduces energy needs and production up to 50% of the preoperative values, reducing VO2 more than VCO2 and therefore an augmented RQ value over 1.0. SU analysis indicates an increased glucose and aminoacid utilization coupled with high nitrogen catabolism that continues in the postoperative period (from 0.08 0.01 gN/kg b.w. to 0.20 0.06 gN/kg b.w.). When the new liver is reperfused, VO2 increases more than VCO2 indicating the risk of reperfusion injury.

Calorimetry, Indirect↗
PubMed1994

Platelet-activating factor mediates an autocrine proliferative loop in the endometrial adenocarcinoma cell line HEC-1A.

We investigated the synthesis and biological effects of platelet-activating factor (PAF) in the human endometrial cancer cell line HEC-1A. We found that HEC-1A cells actively synthesize and release PAF, as demonstrated by both [3H]acetate incorporation into PAF and gas chromatography-mass spectrometry studies. HEC-1A cells not only synthesize but also respond to PAF. Indeed, in fura-2-loaded cells, PAF stimulates [Ca2+]i increase with a median effective concentration of 5.6 nM. Furthermore, PAF induces a time-dependent expression increase of the nuclear protooncogene c-fos with a median effective concentration of 130 nM and stimulates DNA synthesis (median effective concentration, 700 nM). All of these effects are inhibited by the PAF receptor antagonist L659,989. Radioligand binding studies indicated the presence of two populations of PAF receptors with affinity constants in the nanomolar and micromolar range. Since the PAF antagonist per se inhibits DNA synthesis and cell proliferation, we suggest that PAF supports an autocrine growth circuit in HEC-1A cells. On the contrary, in the uterine leiomyosarcoma cell line SK-UT-1, which does not express specific binding sites for PAF, neither this phospholipid nor its receptor antagonist affect DNA synthesis. Our results provide evidence for the existence of an autocrine proliferative loop involving PAF in the endometrial cancer cell line HEC-1A.

Acetyl-CoA C-Acetyltransferase↗
PubMed1994

Post-training nucleus basalis magnocellularis functional tetrodotoxin blockade effects on passive avoidance consolidation in the rat.

The tetrodotoxin (TTX) functional ablation technique was employed in order to evaluate the temporal coordinates of the rat's nucleus basalis magnocellularis (NBM) involvement in memory trace processing. Under ketamine general anesthesia, TTX (10 ng in 1 microliter saline) was stereotaxically administered to rats, either in one or both NBMs. TTX was injected to different groups of rats, respectively 15 min, 6, 24, 48, 96 h after passive avoidance acquisition testing. The rats underwent retrieval testing 48 h later, i.e. after full recovery from TTX effects. Results show that: (1) monolateral TTX blockade significantly impairs PAR conditioned responding if induced up to 6 h but not 24 h after acquisition testing; (2) bilateral TTX blockade dramatically impairs passive avoidance responding up to a 48-h delay but not 96 h after acquisition testing. The results indicate a very profound involvement of NBM in passive avoidance response consolidation. The experimental evidence is discussed together with previous functional ablation findings concerning amygdala, parabrachial nuclei and neocortex.

Amygdala↗
PubMed1994

Identification, characterization, and biological activity of somatostatin receptors in human neuroblastoma cell lines.

To investigate the presence of biologically active somatostatin (SS) receptors in neural crest-derived tumors, radioligand binding studies, cyclic AMP accumulation, intracellular calcium, and growth assays were performed in eight human neuroblastoma (NB) cell lines. Mathematical modeling of binding experiments strongly indicates the presence of heterogeneity of sites. The first site (SSR1) is present in 40% of the NB cell lines and binds with low capacity (0.5 pmol/mg protein) and high affinity (0.1-1 nM) SS14, SS28, and analogues. The second site (SSR2) is a high capacity site (200 pmol/mg protein), widely distributed in all of the cell lines investigated, that shows relative selectivity yet low affinity (100 nM) for SS14, SS28, and [D-Trp8]SS14 without any apparent biological activity. SSR1 is coupled to a pertussis toxin-sensitive G protein, inhibits forskolin- or VIP-stimulated adenylate cyclase activity, decreases intracellular free calcium, and mediates inhibition (30%) of both DNA synthesis and cell growth. Analysis of cell cycle distribution in aphidicolin-synchronized SSR1-positive NB cells indicated that this inhibitory effect is partially mediated by a transient accumulation in G0-G1. Our data indicate high affinity binding sites for SS14, and analogues are present and biologically active in a subset of NB cells.

Binding Sites↗
PubMed1994

Platelet-activating factor in human endometrium.

Platelet-activating factor (PAF) is a phospholipid actively produced by human endometrium and deeply involved in the processes of ovoimplantation and labor. We recently found that PAF represents a new autocrine growth factor for a human adenocarcinoma cell line, HEC-1A. Indeed, biologically active PAF is synthesized by HEC-1A cells, under progesterone control. In HEC-1A cells, PAF regulates intracellular calcium concentration ([Ca2+]), DNA synthesis and expression of early oncogenes. All these effects are blocked by the receptor antagonist L659,989. However, while nanomolar concentrations of PAF mobilize [Ca2+], only micromolar concentrations affect cell growth, suggesting heterogeneity of PAF receptors or signaling. Two distinct populations of PAF receptors are present in HEC-1A cells, which bind PAF in nanomolar and micromolar concentrations, respectively. Since HEC-1A cells are producing elevated concentrations of PAF and micromolar concentrations of the PAF antagonist L659,989 inhibit cell proliferation, an autocrine role for PAF is suggested in HEC-1A cells.

Adenocarcinoma↗
PubMed1994

Effect of platelet-activating factor on motility and acrosome reaction of human spermatozoa.

The effect of platelet-activating factor (PAF) on motility parameters and induction of the acrosome reaction in human spermatozoa was investigated in 36 unselected men with different degrees of initial sperm motility. The characteristics of sperm movement were assessed by computer-assisted sperm analysis (Hamilton-Thorn Motility Analyser) and the percentage of acrosome-reacted spermatozoa was evaluated after 1 h incubation with PAF (10 nM) and staining with fluorescent peanut lectin. We found that short-term (4 h max) incubation with PAF significantly enhanced total and progressive sperm motility as well as acrosome reaction. An increase of sperm motility in response to PAF was present in 16 out of the 25 subjects studied (defined as responders) and was inversely correlated with basal motility. In the 11 samples (six responders and five non-responders) where the incubation with PAF was prolonged overnight, an increase of sperm motility was present in all the subjects studied. Similarly, an increase in numbers of acrosome reactions in response to 10 nM PAF was present in 20 out of the 26 subjects examined, and was inhibited by the PAF receptor antagonist L659 989. Our results indicate a possible physiological role for PAF in fertilization and suggest a potential use of PAF in in-vitro fertilization techniques in cases of reduced sperm motility.

Acrosome↗
PubMed1994

Endothelin stimulates phosphatidylcholine hydrolysis through both PLC and PLD pathways in mesangial cells.

Endothelin (ET) is a recently characterized vasoconstrictor hormone that has potent effects on glomerular function. Many vasoconstrictors, like ET, that stimulate phospholipase C (PLC) hydrolysis of polyphosphoinositides also stimulate phosphatidylcholine (PtdCho) hydrolysis via both PLC and phospholipase D (PLD) pathways. We have previously reported that ET stimulates a protein kinase C (PKC)-regulated, intracellular calcium-insensitive PLD activity that forms phosphatidic acid (PA) in rat mesangial cells (MC). We now ask whether ET-induced diglyceride (DG) production is also, in part, a result of either PLC- or PLD-induced hydrolysis of PtdCho. ET induced both a time- and dose-dependent stimulation in DG as measured by radioflux and mass assays. ET-stimulated DG production was still elevated even at time points where inositol polyphosphates had returned to basal levels. In addition, using [3H]choline-labeled cells, ET stimulated [3H]phosphocholine accumulation, suggesting a PLC-mediated hydrolysis of PtdCho. Stimulation of DG was unaffected by the presence of ethanol or propranolol, suggesting that ET-stimulated DG were not a result of a sequential PLD/PA phosphohydrolase activity. We further dissociated PtdCho-dependent PLC and PLD activities because, in contrast to ET-induced stimulation of PLD, the effect of ET on DG formation was mimicked with ionomycin and was inhibited with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid but not ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid. ET stimulation of DG could not be mimicked by phorbol myristate acetate and was not blocked by PKC inhibition or depletion. Together, these data suggest that ET stimulates multiple signaling pathways in MC that hydrolyze PtdCho via separate PLC and PLD mechanisms.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

Antagonists for the human oxytocin receptor: an in vitro study.

The oxytocin antagonist [Mpa1, D-Tyr(Et)2, Thr4, Orn8]-oxytocin has been successfully used for treating premature labour. The interactions of this antagonist with neurohypophysialhormone receptors in the human myometrium were investigated. Competition curves among [3H]oxytocin, [3H]arginine vasopressin, [3H][1-(beta-mercapto-beta,beta-cyclopentamethylenepropionic acid)2-(O-methyl)-tyrosine, 8-arginine] vasopressin, the corresponding unlabelled peptides and a series of oxytocin antagonists including [Mpa1,D-Tyr(Et)2,Thr4,Orn8]-oxytocin were constructed from results taken from the myometrium of pregnant women and rabbits, and were analysed simultaneously using the computer program LIGAND. The biological activity of [Mpa1,D-Tyr(Et)2,Thr4,Orn8]-oxytocin in the human uterus was investigated by studying its effect on oxytocin-induced intracellular Ca2+ mobilization in human myometrial cells in culture that were expressing high concentrations of oxytocin receptors. The results indicate that [Mpa1,D-Tyr(Et)2,Thr4,Orn8]-oxytocin and related antagonists are selective for the oxytocin receptor in the myometrium of pregnant rabbits but not of pregnant women. In women, they bind with high affinity to the V1 vasopressin receptor. In myometrial cells [Mpa1,D-Tyr(Et)2,Thr4,Orn8]- oxytocin inhibits the oxytocin-induced increase in intracellular Ca2+ concentration in a dose-dependent fashion, with an IC50 value of 5 nmol l-1. The uterine relaxant effect of this antagonist might result not only from the block of the oxytocin receptor, but also from interaction with the V1 vasopressin receptor.

Animals↗
PubMed1994

Endothelin in the human uterus during pregnancy.

In this study we report the immunolocalization, binding and biological activity of endothelins in the human uterus. Since, in previous studies in the rabbit, sex steroids greatly affected uterine endothelin-1 (ET-1) immunolocalization and binding, we sought to compare results obtained in a relatively steroid-deprived uterus (postmenopausal women) with those obtained in late pregnancy. Two classes of ET receptors were identified in human pregnant and non-pregnant myometrium. One site (ETB) was a low capacity site (0.3 pM/mg protein) that bound with high affinity (0.1 nM), yet no selectivity, ET-1, ET-2, ET-3, sarafotoxin (SRTX) and vasoactive intestinal contractor (VIC). The second site (ETA) was six fold more concentrated than the former (1.9 pM/mg protein) and was relatively selective for ET-1, ET-2 and VIC but showed lower affinity for ET-3 and SRTX. Studies with human myometrial cells indicated that the ETA receptor mediates an increase in intracellular calcium, while the physiological function of the ETB receptor is still unclear. Homologous competition curves for ET-1 were used in order to study the ET receptor density (ETA+ETB) in individual myometrial samples. We found that the concentration of ET receptors did not change during different stages of labour or in postmenopausal women. We identified cells with intense positivity for ET-1 in human decidua. Similar cells were also present in pregnant myometrium, intimately associated with smooth muscle cells. Conversely, no staining for ET-1 was observed in non-pregnant myometrium. A paracrine role for ET-1 in the human uterus is suggested.

Adult↗
PubMed1994

Passive avoidance response distribution by post-training substantia nigra functional tetrodotoxin inactivation in the rat.

The tetrodotoxin (TTX) functional ablation technique was employed to assess the temporal coordinates of rat's substantia nigra (SN) in memory processing. TTX (10 ng in 1 microliter saline) was stereotaxically administered to rats under general ketamine anesthesia, either bilaterally or unilaterally. TTX was injected in different groups of rats respectively 0.25, 6, 24, and 48 hours after passive avoidance acquisition testing. Rats always underwent retrieval testing 48 hours later, after full recovery from TTX effects. The results show that: i) unilateral TTX blockade significantly impairs PAR only up to 0.25 h and not 6 h after acquisition testing, and ii) bilateral TTX blockade dramatically disrupts passive avoidance responding up to 24 but not 48 hours after acquisition testing. The results indicate a much more important SN role in memory processing than was previously assessed. The experimental evidence is discussed both in relation to previous TTX functional ablation findings (amygdala, parabrachial nuclei, nucleus basalis magnocellularis) and in relation to SN anatomical and functional connections with other subcortical structures.

Animals↗