Biomedical subjects

M Muratori

Publications and source records attributed to M Muratori.

At least 19 recordsLinked to original sources

PubMed2006

Antineoplastic effects of rosiglitazone and PPARgamma transactivation in neuroblastoma cells.

Neuroblastoma (NB) is the most common extracranial solid tumour in infants. Unfortunately, most children present with advanced disease and have a poor prognosis. In the present study, we evaluated the role of the peroxisome proliferator-activated receptor gamma (PPARgamma) agonist rosiglitazone (RGZ) in two NB cell lines (SK-N-AS and SH-SY5Y), which express PPARgamma. Rosiglitazone decreased cell proliferation and viability to a greater extent in SK-N-AS than in SH-SY5Y. Furthermore, 20 microM RGZ significantly inhibited cell adhesion, invasiveness and apoptosis in SK-N-AS, but not in SH-SY5Y. Because of the different response of SK-N-AS and SH-SY5Y cells to RGZ, the function of PPARgamma as a transcriptional activator was assessed. Noticeably, transient transcription experiments with a PPARgamma responsive element showed that RGZ induced a three-fold increase of the reporter activity in SK-N-AS, whereas no effect was observed in SH-SY5Y. The different PPARgamma activity may be likely due to the markedly lower amount of phopshorylated (i.e. inactive) protein observed in SK-N-AS. To our knowledge, this is the first demonstration that the differential response of NB cells to RGZ may be related to differences in PPARgamma transactivation. This finding indicates that PPARgamma activity may be useful to select those patients, for whom PPARgamma agonists may have a beneficial therapeutic effect.

Antineoplastic Agents↗

PubMed2004

The androgen receptor associates with the epidermal growth factor receptor in androgen-sensitive prostate cancer cells.

Many recent evidences indicate that androgen-sensitive prostate cancer cells have a lower malignant phenotype that is in particular characterized by a reduced migration and invasion. We previously demonstrated that expression of androgen receptor (AR) by transfection of the androgen-independent prostate cancer cell line PC3 decreases invasion and adhesion of these cells (PC3-AR) through modulation of alpha6beta4 integrin expression. The treatment with the synthetic androgen R1881 further reduced invasion of the cells without, however, modifying alpha6beta4 expression on the cell surface, suggesting an interference with the invasion process in response to EGF. We investigated whether the presence of the AR could affect EGF receptor (EGFR)-mediated signaling in response to EGF by evaluating autotransphosphorylation of the receptor as well as activation of downstream signalling pathways. Immunoprecipitation studies demonstrated a reduction of EGF-induced tyrosine phosphorylation of EGFR in PC3-AR cells. In addition, EGF-stimulated PI3K activity, a key signalling pathway for invasion of these cells, was decreased in PC3-AR cells and further reduced by treatment with R1881, indicating decreased functionality of EGFR. An interaction between EGFR and AR has been demonstrated by immunoconfocal and co-immunoprecipitation analysis in PC3-AR cells, suggesting a possible interference of AR on EGFR signalling by interaction of the two proteins. In conclusion, our results suggest that the expression of AR by transfection in PC3 cells confers a less malignant phenotype by interfering with EGFR autophosphorylation and signalling in response to EGF leading to invasion through a mechanism involving an interaction between AR and EGFR.

Androgens↗

PubMed2004

Signaling mechanisms that mediate invasion in prostate cancer cells.

Recent evidence indicates that androgen-sensitive prostate cancer cells have a less malignant phenotype characterized by reduced migration and invasion. We investigated whether the presence of the androgen receptor could affect EGFR-mediated signaling by evaluating autotransphosphorylation of the receptor as well as activation of the downstream signaling pathway PI3K/AKT. Immunoprecipitation studies demonstrated a reduction of EGF-induced tyrosine phosphorylation of EGFR in PC3-AR cells. In addition, EGF-stimulated PI3K activity, a key signaling pathway for invasion of these cells, was decreased in PC3-AR cells and further reduced by treatment with R1881, indicating decreased functionality of EGFR. Our results suggest that the expression of androgen receptors by transfection in PC3 cells confers a less malignant phenotype by interfering with EGFR autophosphorylation and signaling leading to invasion in response to EGF. We used the selective tyrosine kinase inhibitor of the EGFR gefitinib (also known as Iressa or ZD1839) to further investigate the role of EGFR in the invasion and growth of PC cells. We demonstrate that in the androgen-insensitive cell lines PC3 and DU145 this compound was able to decrease in vitro invasion of Matrigel by inhibiting EGFR autotransphosphorylation and subsequent PI3K activation. Gefitinib may be useful in the treatment of androgen-independent prostate cancer to limit not only the proliferation but also the invasion of these tumors.

Androgens↗

PubMed2003

Pharmacological profile of MEN 11066, a novel potent and selective aromatase inhibitor.

MEN 11066 is a new non-steroidal compound which potently inhibits human placenta (K(i)=0.5 nM) and rat ovarian (K(i)=0.2 nM) aromatase in vitro. In vivo, a single oral dose of 0.3 mgkg(-1) significantly decreased uterus weight in immature rats after stimulation of uterus growth by androstenedione. MEN 11066 reduced in a dose-dependent manner plasma estradiol levels in adult female rats treated with pregnant mare serum gonadotropin (PMSG). After 2 weeks of repeated daily treatment in adult rats, a significant decrease in uterine weight was observed together with a 65% decrease in plasma estradiol, whereas plasma levels of testosterone, progesterone, aldosterone, corticosterone, cholesterol, LH and FSH were not affected. The lack of any effect by MEN 11066 on adrenal steroids was confirmed by the unchanged plasma corticosterone and aldosterone levels in immature rats and also in adult rats when the repeated treatment with MEN 11066 (15 days) was followed by the administration of a synthetic ACTH analogue. No change in 11beta-hydroxylase or 21-hydroxylase activities was produced in vitro by the addition of 10 microM MEN 11066. Fifteen-day treatment with MEN 11066 did not produce changes in several rat hepatic enzymatic activities involved in the metabolism of xenobiotics. These results demonstrated that MEN 11066 is a potent inhibitor of aromatase which does not interfere with the cytochrome P450 involved in the synthesis of other steroids or in the metabolism of xenobiotics.

Administration, Oral↗

PubMed2003

Inhibition of spontaneous and androgen-induced prostate growth by a nonhypercalcemic calcitriol analog.

We have recently found that analog V (BXL-353, a calcitriol analog) inhibits growth factor (GF)-stimulated human benign prostate hyperplasia (BPH) cell proliferation by disrupting signal transduction, reducing Bcl-2 expression, and inducing apoptosis. We now report that BXL-353 blocks in vitro and in vivo testosterone (T) activity. BPH cells responded to T and dihydrotestosterone (DHT) with dose-dependent growth and reduced apoptosis. Exposure of BPH cells to BXL-353 significantly antagonized both T- and DHT-induced proliferation and induced apoptosis, even in the presence of T. To verify whether BXL-353 reduced prostate growth in vivo, we administered it orally to either intact or castrated rats, supplemented with T enanthate. Nonhypercalcemic doses of BXL-353 time- and dose-dependently reduced the androgen effect on ventral prostate weight, similarly to finasteride. Comparable results were obtained after chronic administration of BXL-353 to intact rats. Clusterin (an atrophy marker) gene and protein were up-regulated by BXL-353 in rat prostate, and nuclear fragmentation was widely present. The antiandrogenic properties of BXL-353 did not interfere with pituitary and testis function, as assessed by serum determination of rat LH and T. BXL-353 did not compete for androgen binding to BPH homogenates and failed to inhibit 5alpha-reductase type 1 and type 2 activities. In conclusion, BXL-353 blocks in vitro and in vivo androgen-stimulated prostate cell growth, probably acting downstream from the androgen receptor, without affecting calcemia or sex hormone secretion. BXL-353 and other vitamin D(3) analogs might thus represent an interesting class of compounds for treating patients with BPH.

3-Oxo-5-alpha-Steroid 4-Dehydrogenase↗

PubMed2003

Expression of functional estrogen receptors in human fetal male external genitalia.

It is generally assumed that male genital development is determined by androgens on a default program leading to female genitalia. Female genitalia virilization is due to high levels of androgens, whereas feminization is linked to reduction or lack of fetal androgen. Excess androgen determines sex reversion in female, whereas excess estrogen does not cause male feminization. In the present study, we investigate the presence of androgen receptors (AR) and estrogen receptors (ER) in human fetal penile tissue and in a cellular model of human fetal penile smooth muscle cells (hfPSMC). By immunohistochemistry, we showed the presence of ER and AR in the developing penile tissue of male fetuses. Besides the presence of AR, hfPSMC showed ERalpha/beta as demonstrated by RT-PCR, Western blot, and binding techniques. These receptors are functionally active because cell stimulation with 17beta-estradiol increased progesterone receptor B expression and inhibited hfPSMC growth, both effects being reversed by tamoxifen. Conversely, cell proliferation was stimulated by R1881 and testosterone, an effect enhanced by letrozole. These findings are the first demonstration of the presence of functional ER in differentiating male external genitalia and indicate a possible novel inhibitory role of estrogens in the regulation of the development of these sex structures.

Aromatase Inhibitors↗

PubMed2001

Transesophageal echocardiography as predictor of mitral valve repair.

BACKGROUND AND AIM OF THE STUDY: Mitral valve repair has recently emerged as the treatment of choice in patients presenting with insufficiency due to valve prolapse. The study aims were to evaluate: (i) the clinical presentation in a consecutive series of patients with mitral valve prolapse undergoing surgical repair; (ii) the correlation between pre- and intraoperative echocardiographic features and surgical findings in these patients; and (iii) whether clinical and echocardiographic data may predict surgical outcome. METHODS: Between March 1997 and May 2000, 152 patients (110 men, 42 women; mean age 59+/-13 years) were recruited into the study. All patients had myxomatous mitral valve disease causing severe regurgitation and underwent systematic examination by transesophageal echocardiography (TEE) for clear delineation of the three scallops of the posterior leaflet and juxtaposed segments of the anterior leaflet. RESULTS: In 119 patients (78%) a flail valve was documented by TEE and confirmed on surgical inspection; an anterior leaflet chordal rupture was not visualized by TEE in one case. In 15 cases (10%) there was flail of the anterior leaflet, and in 105 cases (69%) flail of the posterior leaflet. A bileaflet complex prolapse without chordal rupture was found in 32 cases. On the basis of TEE evaluation, mitral valve replacement was performed electively in 10 patients (7%); the other 142 (93%) underwent mitral valve repair. Adequate repair was obtained in 93% of cases; residual mitral regurgitation (eight cases; grade 3+) and mitral stenosis (one case) were documented by intraoperative TEE, and nine patients (6%) underwent valve replacement. CONCLUSION: The majority of patients with myxomatous mitral valve prolapse and severe regurgitation undergoing valve repair have chordal rupture of the posterior mitral leaflet, a condition in which results of valve repair are excellent. TEE provides a powerful means to define the mechanisms of mitral regurgitation and to identify the suitability of patients for valvuloplasty.

Adult↗

PubMed2000

Effect of the aromatase inhibitor, MEN 11066, on growth of two different MCF-7 sublines.

The racemate compound MEN 11066 (1-[(benzofuran-2-yl)(4'-cyanophenyl)methyl]-1H-1,2,4-triazole) and its enantiomers, (+)-MEN 11623 and (-)-MEN 11622, showed potent and selective aromatase activity on human placental microsomes. In addition, to better evaluate their potency as anticancer drugs, the compounds were assayed on testosterone-induced cell proliferation to measure their ability in inhibiting oestrogen-dependent tumour growth. Two different sublines originated from the human breast carcinoma MCF-7 were used. One, named MCF-7(tumour aromatase) (TA), that had maintained its intrinsic aromatase activity, was more sensitive to estradiol or testosterone-induced growth than the second subline named MCF-7(human placental aromatase) (hPA). The latter had been transfected with the human placental aromatase cDNA, after recognizing that the parental cells had aromatase activity reduced to undetectable levels. The MEN compounds completely reverted the testosterone-induced proliferation in both MCF-7(TA) and MCF-7(hPA) cells, while they did not affect the estradiol-triggered proliferation as a proof of their specificity for aromatase enzyme. Interestingly, MCF-7(TA) cells were more susceptible to the effects of aromatase inhibitors than the MCF-7(hPA) cell. These data suggest the efficacy of aromatase inhibitors in breast cancer when the growth dependency from oestrogen is high and a relatively low aromatase activity may be extremely important for tumour development.

Androstenedione↗

PubMed2000

Uteroglobin reverts the transformed phenotype in the endometrial adenocarcinoma cell line HEC-1A by disrupting the metabolic pathways generating platelet-activating factor.

Uteroglobin, originally named blastokinin, is a protein synthesized and secreted by most epithelia, including the endometrium. Uteroglobin has strong anti-inflammatory properties that appear to be due, at least in part, to its inhibitory effect on the activity of the enzyme phospholipase A(2). In addition, recent experimental evidence indicates that uteroglobin exerts antiproliferative and antimetastatic effects in different cancer cells via a membrane receptor. The human endometrial adenocarcinoma cell line HEC-1A does not express uteroglobin. Thus, we transfected HEC-1A cells with human uteroglobin cDNA. The transfectants showed a markedly reduced proliferative potential as assessed by impaired plating efficiency as well as by reduced growth in soft agar. Cytofluorimetric analysis clearly indicated that in uteroglobin-transfected cells the time for completion of the cell cycle was increased. We previously demonstrated that HEC-1A cells actively synthesize platelet-activating factor, one of the products of phospholipase A(2) activity. In addition, we demonstrated that platelet-activating factor stimulates the proliferation of these cells through an autocrine loop. In uteroglobin transfectants, the activity of phospholipase A(2) and platelet-activating factor acetyl-transferase, which are involved in the synthesis of platelet-activating factor, was significantly reduced compared with wild-type and vector-transfected cells (p < 0.05). Our results indicate that enforced expression of uteroglobin in HEC-1A cells markedly reduced their growth potential and significantly impaired the synthesis of platelet-activating factor, an autocrine growth factor for these cells. These data suggest that one possible mechanism for the recently observed antineoplastic properties of uteroglobin may be the inhibition of the synthesis of platelet-activating factor.

Acetyltransferases↗

PubMed2000

Intracellular events and signaling pathways involved in sperm acquisition of fertilizing capacity and acrosome reaction.

Two processes, namely capacitation and acrosome reaction, are of fundamental importance in the fertilization of oocyte by spermatozoon. Physiologically occurring in the female genital tract, capacitation is a complex process, which renders the sperm cell capable for specific interaction with the oocyte. During capacitation, modification of membrane characteristics, enzyme activity and motility properties of spermatozoa render these cells able to penetrate oocyte investments and responsive to stimuli that induce acrosome reaction prior to fertilization. Physiological acrosome reaction occurs upon interaction of the spermatozoon with the zona pellucida protein ZP3. This is followed by liberation of several acrosomal enzymes and other constituents that facilitate penetration of the zona and expose molecules on the sperm equatorial segment that allows fusion of sperm membrane with the oolemma. The molecular mechanisms and the signal transduction pathways mediating the processes of capacitation and acrosome reaction have been partially defined, and appear to involve modifications of intracellular calcium and other ions, lipid transfer and phospholipid remodeling in sperm plasma membrane as well as changes in protein phosphorylation. Some of the kinases and phosphorylated proteins that are involved in the processes of capacitation and acrosome reaction have been now characterized. Characterization of sperm receptors to physiological inducers of acrosome reaction is in progress. This review summarizes the main signal transduction pathways involved in capacitation and acrosome reaction.Furthermore, the mechanisms underlying sperm DNA fragmentation are also briefly reviewed.

Acrosome Reaction↗

PubMed2000

A novel functional estrogen receptor on human sperm membrane interferes with progesterone effects.

We have identified an estrogen receptor of approximately 29 kDa apparent molecular weight in human sperm membranes by ligand and Western blot analysis, respectively, using peroxidase-conjugated E(2) and an antibody directed against the ligand binding region of the genomic receptor (alphaH222). Such receptor is functional since 17betaE(2) induces a rapid and sustained increase of intracellular calcium concentrations ([Ca(2+)](i)) which is completely eliminated by preincubation with alphaH222. 17betaE(2) effects on calcium are clearly mediated by a membrane receptor, as they are reproduced by the membrane-impermeable conjugate of the hormone BSA-E(2). Dose-response curve for this effect is biphasic with EC(50)s in the nanomolar and micromolar range. In addition to calcium increase, 17betaE(2) stimulates tyrosine phosphorylation of several sperm proteins including the 29-kDa protein band. Preincubation of human sperm with 17betaE(2) inhibits calcium and acrosome reaction increases in response to progesterone. We conclude that estrogens may play a role in the modulation of non-genomic action of progesterone (P) in human sperm during the process of fertilization.

Acrosome Reaction↗

PubMed2000

Role of cine-fluoroscopy, transthoracic, and transesophageal echocardiography in patients with suspected prosthetic heart valve thrombosis.

Prosthetic heart valve thrombosis (PVT) is a rare but potentially life-threatening complication of heart valve replacement. An effective, quick, and easy diagnostic method is highly desirable. We evaluated the diagnostic efficacy of cine-fluoroscopy (CF), transthoracic (TTE), and transesophageal (TEE) echocardiography in 82 consecutive patients with mechanical valves and suspected PVT. Criteria for PVT were: leaflet(s) motion restriction at CF, increased Doppler gradients at TTE, and evidence of thrombi at TEE. Patients were divided in 4 groups (A, B, C, and D) according to results of CF and TTE. Group A was composed of 24 patients with positive CF and TTE. Thrombi were detected by TEE in all cases, suggesting that when both are positive, CF and TTE correctly identified PVT in all patients so that TEE may be deferred. Group B was composed of 12 patients with positive CF and negative TTE; TEE showed PVT in 4 patients (33%). These patients had very slight leaflet motion restriction as in the case of initial PVT. This suggests that CF compared with Doppler may identify patients with "hemodynamically significant" PVT. The remaining 8 patients in this group had monocuspid prostheses with negative TEE, suggesting that abnormal leaflet motion at CF may be due to functional changes. Therefore, TEE should always be performed in case of monocuspid prostheses with isolated CF abnormalities. Group C was composed of 18 asymptomatic patients with small-sized aortic prostheses and very high Doppler gradients on routine TTE. CF showed normal leaflet motion and TEE ruled out PVT in all cases outlining the diagnostic role of CF in this particular subset. Finally, group D was composed of 28 patients with negative CF and TTE. TEE did not show thrombi in 24 of 28 patients (86%), confirming that, when both yield negative results, CF and TTE are reliable methods to rule out valve thrombosis in most cases. However, in 4 of 28 patients (14%) TEE showed "nonobstructive" prosthetic thrombosis: these patients had mitral prostheses, chronic atrial fibrillation, and 3 of 4 had systemic embolisms. Thus, TEE should be performed in selected patients despite negative CF and TTE results. Sensitivity, specificity, and positive and negative predictive values were 87%, 78%, 80%, and 91% for CF and 75%, 64%, 57%, and 78% for TTE, respectively. CF and TTE correctly identified PVT in 70 of 82 patients (85%). TEE was actually required in 15% of the cases. Thus, CF and TTE are quick, effective, and complementary diagnostic tools to diagnose PVT in most patients. TEE still remains the gold standard technique in selected cases.

Aged↗

PubMed2000

How the left and right sides of the heart, as well as pulmonary venous drainage, adapt to an increasing degree of head-up tilting in hypertrophic cardiomyopathy: differences from the normal heart.

OBJECTIVES: We aimed to assess the differences in the adaptive response of patients with hypertrophic cardiomyopathy (HCM) compared with normal subjects, as well as any association with increased susceptibility to the test. BACKGROUND: Diastolic function contributes importantly in the adaptation of the normal heart to head-up tilting. This mechanism may be disturbed by an impaired relaxation in HCM. METHODS: Twenty-one male patients with HCM (46 +/- 6 years old) and 22 healthy men (44 +/- 8 years) were studied using Doppler echocardiography after 1 and 10 min of head-up tilting at 20 degrees, 40 degrees and 60 degrees. RESULTS: In control subjects, tilting was associated with 1) a predominance of diastolic pulmonary venous flow and early left ventricular (LV) filling (atrium functioning as an open conduit); 2) right ventricular (RV) shrinkage; and 3) no LV dimensional variations. In patients with HCM, tilting was associated with 1) a prevalence of systolic pulmonary venous flow (atrium functioning as a reservoir in which filling depends on atrial relaxation and compliance) and late diastolic transmitral flow (atrium working as a booster pump); 2) LV shrinkage; and 3) no RV dimension variations. These mechanisms did not prevent stroke volume (SV) from decreasing at 40 degrees and 60 degrees in both groups. Because of a lower increase in heart rate (HR), a reduction in cardiac output (CO) was greater in patients with HCM. The responses were similar after 1 and 10 min of tilting in control subjects, whereas in patients, blood pressure (BP), SV and LV dimension fell more after 10 min. CONCLUSIONS: Adaptation of the normal heart to tilting is based on a ventricular interaction and LV diastolic properties; HCM relies on left atrial diastolic and systolic functions. An inadequate HR reaction to a fall in BP and SV in HCM (depressed reflexogenic activity) contributes to making CO more vulnerable by greater and more prolonged displacements.

Adaptation, Physiological↗

PubMed2000

Exercise echocardiography versus exercise electrocardiography in the diagnosis of coronary artery disease in hypertension.

In hypertension, coronary artery disease (CAD) can be overestimated by stress electrocardiography (ECG) and scintigraphy due to frequent false-positive results. Exercise tests are also limited by an excessive blood pressure increase, and pharmacologic pressure normalization decreases the accuracy of the test. The aim of this study was to assess the accuracy of exercise echocardiography as an alternative test for CAD detection in hypertension, both before and after adequate blood pressure control. We studied 59 hypertensive and 59 normotensive patients undergoing coronary angiography for chest pain. Upright bicycle exercise ECG and echocardiographic tests were performed in each group in the absence of therapy; in hypertensives, the tests were repeated a day apart after blood pressure normalization with sublingual nifedipine. Significant CAD (lumen narrowing >50%) was detected in 22 hypertensive and 41 normotensive patients. In the two groups, sensitivity, specificity, and diagnostic accuracy of exercise echocardiography performed before treatment were not statistically different (95%, 94%, 94% in hypertensives and 82%, 77%, 83% in normotensives, respectively), but were significantly higher than for the exercise ECG test (68%, 70%, and 69%, respectively). After blood pressure lowering, exercise echocardiography sensitivity slightly decreased (91%), whereas specificity (100%) and diagnostic accuracy (96%) did not vary; on the contrary, exercise ECG sensitivity decreased to 45%. Therefore, according to our data, exercise echocardiography can be an accurate test and more reliable than exercise ECG to detect CAD in normotensives as well as in hypertensives. Normalization of blood pressure with nifedipine does not affect its accuracy, but markedly reduces the sensitivity of exercise ECG.

Aged↗

PubMed2000

Androgen receptor expression in prostate carcinoma cells suppresses alpha6beta4 integrin-mediated invasive phenotype.

Prostate cancer cells may lose androgen-sensitivity after androgen ablation therapy, becoming highly invasive and metastatic. The biological mechanisms responsible for higher tumurogenicity of androgen-independent prostate carcinomas are not entirely known. We demonstrate that androgen receptor regulation of adhesion and invasion of prostate cancer cells through modulation of alpha6beta4 integrin expression may be one of the molecular mechanisms responsible of this phenomenon. We found that protein and gene expressions of alpha6 and beta4 subunits were strongly reduced in the androgen-sensitive cell line LNCaP respect to the androgen-independent PC3 and that transfection of PC3 cells with a full-length androgen receptor expression vector resulted in a decreased expression of alpha6beta4 integrin, reduced adhesion on laminin, and suppressed Matrigel invasion. Growth in soft agar was also suppressed in androgen receptor-positive PC3 clones. Treatment of androgen receptor positive clones with the synthetic androgen R1881 further reduced alpha6 and beta4 messenger RNA expression as well as adhesion on laminin and Matrigel invasion. Our results indicate that androgens regulate cell-extracellular matrix adhesion and invasion by modulation of integrin expression and function, thus keeping a low invasive phenotype of prostate cancer cells.

Antigens, Surface↗

PubMed1999

Identification and characterization of a novel functional estrogen receptor on human sperm membrane that interferes with progesterone effects.

The presence of a novel functional estrogen receptor on the human sperm surface has been demonstrated by using different experimental approaches. Ligand blot analysis of sperm lysates, using peroxidase-conjugated estradiol as probe, identified a specific estradiol-binding protein of approximately 29-kDa apparent molecular mass. The same protein band was also revealed by using alphaH222 antibody, which is directed against the steroid binding domain of the genomic estrogen receptor. The biological effects of estrogen receptor were investigated by analyzing calcium fluxes, tyrosine phosphorylation, and acrosome reaction (AR) in response to 17beta-estradiol (17betaE2) and by measuring the steroid influence on calcium and AR in responses to progesterone (P), a well-known physiological stimulus for human spermatozoa. Our results demonstrate that 17betaE2 induces a rapid and sustained increase of intracellular calcium concentrations ([Ca2+]i). This effect is totally dependent on the presence of extracellular calcium, because it is completely abolished in a calcium-depleted medium. The dose-response curve for calcium increase to 17betaE2 is biphasic with a first component in the nanomolar range (effective concentration 50 = 0.60 +/- 0.12 nmol/L) and a second component in the micromolar range (EC50 = 3.80 +/- 0.26 micromol/L). 17BetaE2 stimulates tyrosine phosphorylation of several sperm proteins, including the 29-kDa protein band, and determines a reduction of calcium response to P, finally resulting in inhibition of P-stimulated sperm AR. Conversely, no direct effect of 17betaE2 is observed on AR. 17BetaE2 effects on calcium are clearly mediated by a membrane receptor, because they are reproduced by the membrane-impermeable conjugate of the hormone BSA-E2 and reduced by sperm preincubation with alphaH222 antibody. Taken together, our results clearly show the presence of a functional surface estrogen receptor, of 29 kDa, on human spermatozoa. This receptor may play a role in the modulation of nongenomic action of P in these cells during the process of fertilization.

Acrosome Reaction↗

PubMed1997

Protein tyrosine kinase, mitogen-activated protein kinase and protein kinase C are involved in the mitogenic signaling of platelet-activating factor (PAF) in HEC-1A cells.

We have recently demonstrated that the phospholipid platelet-activating factor (PAF) mediates an autocrine proliferative loop in the endometrial adenocarcinoma cell line HEC-1A. In the present study we investigated the signaling pathways involved in PAF-mediated increase of proliferation in these cells. In particular, we studied the effect of PAF on protein tyrosine phosphorylation and mitogen-activated protein kinase (MAPK) activity, as well as the effect of protein tyrosine kinase (PTK) and protein kinase C (PKC) inhibition on PAF-induced increase of c-fos gene expression and thymidine incorporation in HEC-1A cells. We found that PAF induced a rapid, time- and dose-dependent increase of tyrosine phosphorylation of a subset of proteins of 42, 44, 78, 99, and 150 kDa molecular weight. We also found that PAF increased tyrosine phosphorylation and activity of p42 MAPK, suggesting the involvement of this important intermediary enzyme in the proliferative effect of PAF. The effect of PAF on c-fos gene expression was not prevented by pre incubation with the PTK inhibitors genistein or methyl-2,5-dihydroxycinnamate, whereas was strongly affected by PKC down regulation after long term incubation with PMA or by PKC inhibition with sangivamycin. We also found that genistein and methyl 2,5-dihydroxycinnamate decreased both basal and PAF-stimulated [3H]thymidine uptake in these cells. Similar results were obtained with PD 098059, a specific inhibitor of MAP kinase cascade. PAF-stimulated [3H]thymidine uptake was also prevented by PKC down regulation after long term exposure to PMA and PKC inhibition with the two inhibitors sangivamycin and bis-indolylmaleimide. In conclusion, our results indicate that PAF-induced mitogenesis in HEC-1A cells is mediated by the activation of multiple signaling pathways, involving PTK, MAPK, and PKC activation.

Adenocarcinoma↗

PubMed1997

Use of cabergoline in the long-term treatment of hyperprolactinemic and acromegalic patients.

Cabergoline (Cab), a very potent and long-lasting dopaminergic compound, was administered to 26 women with pituitary microprolactinoma [mean serum PRL levels: 124.8 +/- 11.3 micrograms/l (+/- SE), range 62-300 micrograms/l] and 3 patients with GH-secreting pituitary adenoma (2 with associated PRL hypersecretion) for 12 and 24 months, respectively. In microprolactinomas, a stable normoprolactinemia was achieved in 96.1% of cases: in 13 women (50%) with the lowest dose of the drug (0.5 mg/week), and in other 12 patients (46.1%) with increasing doses up to 3 mg/week. All the oligomenorrheic/amenorrheic women, except one, restored regular and ovulatory menses. Two patients became pregnant. Pituitary abnormalities at high resolution-CT (HR-CT) scan disappeared in 13 of 19 patients (68.4%) after 12 months of therapy and this feature persisted in 8/13 cases (61.5%) 12 months after drug withdrawal. During Cab discontinuation (range: 3-60 months), mean serum PRL levels remained significantly lower than the basal ones. Six of 25 women are still without therapy. In 2 patients, normoprolactinemia persisted up to 38 and 60 months, respectively. Cab treatment was re-instituted in 13 patients because of the recurrence of hyperprolactinemia. Five patients were lost at follow up. In all the acromegalic patients, Cab (1-3 mg/week) normalized serum GH, IGF-I and PRL levels. A clear improvement in clinical symptoms was observed in all patients, but neuroradiological improvement in only one. Cab therapy was very well tolerated, as only seven patients complained of mild and transient side-effects and none had to stop treatment. In conclusion, Cab is an effective, safe, and well tolerated dopaminergic compound for the treatment of hyperprolactinemic disorders and the control of the clinical and hormonal features of dopamine-sensitive acromegalic patients.

Acromegaly↗