Biomedical subjects

Paolo Mannella

Publications and source records attributed to Paolo Mannella.

16 recordsLinked to original sources

PubMed2006

Estrogen receptor protein interaction with phosphatidylinositol 3-kinase leads to activation of phosphorylated Akt and extracellular signal-regulated kinase 1/2 in the same population of cortical neurons: a unified mechanism of estrogen action.

17Beta-estradiol (E2)-induced neuroprotection is dependent on mitogen-activated protein kinase (MAPK) and phosphatidylinositol-3-kinase (PI3K) signaling cascades. We sought to determine whether E2 neuroprotective mechanisms are mediated by a unified signaling cascade activated by estrogen receptor (ER)-PI3K interaction within the same population of neurons or whether E2 activation of extracellular signal-regulated kinase 1/2 (ERK1/2) and Akt are independent signaling events in different neuronal populations. Immunoprecipitation of E2-treated cortical neurons was conducted to determine a protein-protein interaction between ER and the PI3K regulatory subunit p85. Subsequently, cortical neurons were treated with E2 alone or in presence of MAPK inhibitors or PI3K inhibitors. Results of these analyses indicated a protein-protein interaction between ER and p85 that was time-dependent and consistent with the temporal profile for generation of Akt (pAkt) and ERK1/2 phosphorylation (pERK1/2). E2-induced phosphorylation of Akt, was first apparent at 10 min and maximal at 30 min. Simultaneously, E2-induced pERK1/2 was first apparent at 5-10 min and maximal at 30 min. Inhibition of PI3K completely blocked E2 activation of pAkt at 10 and 30 min and blocked E2 activation of ERK1/2 at 10 min, which revealed a PI3K-independent activation of ERK at 30 min. Double immunocytochemical labeling for pERK1/2 and pAkt demonstrated that E2 induced both signaling pathways in the same neurons. These results indicate a unified signaling mechanism for rapid E2 action that leads to the coordinated activation of both pERK1/2 and pAkt in the same population of neurons. Implications of these results for understanding estrogen mechanism of action in neurons and therapeutic development are considered.

Alzheimer Disease↗

PubMed2006

Effects of dydrogesterone and of its stable metabolite, 20-alpha-dihydrodydrogesterone, on nitric oxide synthesis in human endothelial cells.

OBJECTIVE: To investigate the effects of P, medroxyprogesterone acetate (MPA), and dydrogesterone (DYD) and its metabolite, 20-alpha-dihydrodydrogesterone (DHD) on endothelial synthesis of nitric oxide (NO) and characterize the signaling events recruited by these compounds. The Women's Health Initiative trial reports an excess of heart disease in postmenopausal women receiving MPA. DESIGN: Cell culture. SETTING: Research laboratory. PATIENT(S): Human endothelial cells from umbilical vein. INTERVENTION(S): Treatments with P, MPA, DYD, or DHD. MAIN OUTCOME MEASURE(S): Measure of NO release, endothelial nitric oxide synthase (eNOS) activity and expression, and activation of ERK 1/2 and Akt. RESULT(S): The administration of DYD alone or in combination with estrogen to endothelial cells results in neutral effects on NO synthesis and on the activity and expression of eNOS. In parallel, the stable metabolite DHD acts similarly to natural P, enhancing the expression of eNOS and inducing rapid activation of the enzyme through the regulation of the ERK 1/2 mitogen-activated protein kinase cascade. 20-Alpha-dihydrodydrogesterone and P also potentiate eNOS induction by E2. On the contrary, MPA does not trigger eNOS enzymatic activation and decreases the extent of eNOS induction by E2. CONCLUSION(S): These findings support the concept that synthetic progestins act differently on vascular cells and that hormonal preparations may differ as to their cardiovascular effects.

Cells, Cultured↗

PubMed2006

Estrogen receptor alpha interacts with Galpha13 to drive actin remodeling and endothelial cell migration via the RhoA/Rho kinase/moesin pathway.

Sex steroids control cell movement and tissue organization; however, little is known of the involved mechanisms. This report describes the ongoing dynamic regulation by estrogen of the actin cytoskeleton and cell movement in human vascular endothelial cells that depends on rapid activation of the actin-regulatory protein moesin. Moesin activation is triggered by the interaction of the C-terminal portion of cell membrane estrogen receptor alpha with the G protein Galpha(13), leading to activation of the small GTPase RhoA and of the downstream effector Rho-associated kinase. The resulting phosphorylation of moesin on Thr(558) is the means of moesin's binding to actin and the remodeling of the actin cytoskeleton. This cascade of events ensues within minutes of estradiol administration and results in changes in cell morphology and to the development of specialized cell membrane structures such as ruffles and pseudopodia that are necessary for cell movement. These findings expand our knowledge of the basis of estrogen's effects on human cells, including the regulation of actin assembly, cell movement and migration. They highlight novel pathways of signal transduction of estrogen receptor alpha through nontranscriptional mechanisms. Furthermore, exposure of this estrogen receptor-dependent, nongenomic action of estrogen on human vascular endothelial cells is especially relevant to the present interest in the role of estrogen in cardiovascular protection.

Actins↗

PubMed2005

Differential estrogen signaling in endothelial cells upon pulsed or continuous administration.

UNLABELLED: While experimental evidence demonstrates that estrogen protects vascular cells, clinical trials on hormone replacement therapies (HRT) fail to report cardiovascular benefits. This discrepancy may indicate that estrogen signaling during HRT may not be fully effective in vascular cells, possibly due to the way of delivering estrogens to vascular tissues. We therefore, tested whether a different kinetics of exposure of endothelial cells to estrogens may alter the balance between transcriptional and non-transcriptional signaling. METHODS AND RESULTS: Cultured human umbilical vein endothelial cells (HUVEC) were exposed to equal amounts of 17beta-estradiol administered transiently (7 nM for 1 h) or continuously (0.29 nM for 24 h), using endothelial nitric oxide synthase (eNOS) expression and activity as functional endpoints. Transient and continuous administrations equally increased eNOS expression, implying that a short contact between hormone and cells is sufficient to trigger genomic pathways. In agreement, in ovariectomized rats pulse-like changes in plasma estradiol or stable levels equally increased aortic eNOS. More importantly, we extensively show that transient estradiol administration results in preferential recruitment of non-genomic pathways, leading to more vivid activation of MAPK and phosphatidylinositol-3 kinase (PI3K) cascades. CONCLUSIONS: Our data suggest that the kinetics of contact of estrogen with the cells could represent a sensor for estrogen receptor (ER) signaling towards non-genomic or genomic pathways. These findings deepen our understanding of estrogen signaling, and have important implications for the design of safer and more effective HRT.

Animals↗

PubMed2004

Differential signal transduction of progesterone and medroxyprogesterone acetate in human endothelial cells.

The conjugated equine estrogens-only arm of the Women's Health Initiative trial, showing a trend toward protection from heart disease as opposed to women receiving also medroxyprogesterone acetate (MPA), strengthens the debate on the cardiovascular effects of progestins. We compared the effects of progesterone (P) or MPA on the synthesis of nitric oxide and on the expression of leukocyte adhesion molecules, characterizing the signaling events recruited by these compounds. Although P significantly increases nitric oxide synthesis via transcriptional and nontranscriptional mechanisms, MPA is devoid of such effects. Moreover, when used together with physiological estradiol (E2) concentrations, P potentiates E2 effects, whereas MPA impairs E2 signaling. These findings are observed both in isolated human endothelial cells as well as in vivo, in ovariectomized rat aortas. A marked difference in the recruitment of MAPK and phosphatidylinositol-3 kinase explains the divergent effects of the two gestagens. In addition, both P and MPA decrease the adhesiveness of endothelial cells for leukocytes when given alone or with estrogen. MPA is more potent than P in inhibiting the expression of vascular cell adhesion molecule-1 and intercellular adhesion molecule-1. However, when administered together with physiological amounts of glucocorticoids, MPA (which also binds glucocorticoid receptor) markedly interferes with the hydrocortisone-dependent stabilization of the transcription factor nuclear factor kappaB and with the expression of adhesion molecules, acting as a partial glucocorticoid receptor antagonist. Our findings show significant differences in the signal transduction pathways recruited by P and MPA in endothelial cells, which may have relevant clinical implications.

Cell Communication↗

PubMed2004

Genomic and non-genomic effects of estrogens on endothelial cells.

Estrogen receptors act via the regulation of transcriptional processes, involving nuclear translocation and binding on specific response elements, thus leading to regulation of target gene expression. However, novel non-transcriptional mechanisms of signal transduction through steroid hormone receptors have been identified. These so-called "non-genomic" effects are independent by gene transcription or protein synthesis and involve steroid-induced modulation of cytoplasmic or of cell membrane-bound regulatory proteins. Relevant biological actions of steroids have been associated with this signaling in different tissues. Ubiquitary regulatory cascades such as mitogen-activated protein kinases (MAPK), the phosphatidylinositol 3-OH kinase (PI3K) and tyrosine kinases are modulated through non-transcriptional mechanisms by steroid hormones. Furthermore, steroid hormone receptors modulation of cell membrane-associated molecules such as ion channels and G-protein-coupled receptors has been shown in diverse tissues. The vascular wall is a site where non-genomic steroid hormones actions are particularly prominent. For instance, estrogens and glucocorticoids trigger rapid vasodilatation due to rapid induction of nitric oxide synthesis in endothelial cells via the estrogen receptor-dependent activation of MAPK and PI3K, leading to relevant pathophysiological consequences, in vitro and in vivo. The growing amount of evidence collected in the last years claims that non-transcriptional signaling mechanisms play a primary role in the generation of the effects of steroids on endothelial cells, which may turn out to be of relevance for clinical purposes.

Animals↗

PubMed2004

Tibolone activates nitric oxide synthesis in human endothelial cells.

After the unexpected findings of the Women's Health Initiative trial, indicating that traditional cardiovascular risk markers fail to predict the effects of hormone replacement therapy, it is of interest to characterize how steroids act on vascular cells. This is particularly important for tissue-specific drugs such as tibolone, whose actions may differ from other preparations. Because nitric oxide (NO) is a key regulator of vascular tone and atherogenesis, we studied its regulation by tibolone and its metabolites on human endothelial cells. Tibolone and its estrogenic metabolites (3alpha- and 3beta-OH tibolone) activate NO synthesis by recruiting functional estrogen receptors, whereas the progestogenic/androgenic metabolite (Delta(4) isomer) has no effect. During prolonged exposures, tibolone and the estrogenic compounds enhance the expression of endothelial NO synthase (eNOS). In addition, tibolone is able to induce rapid activation of eNOS, leading to rapid increases in the release of NO. Relevant for its clinical effects, the sulfated metabolites of tibolone are also effective in activating eNOS. Different from estrogen, rapid activation of eNOS does not rely on recruitment of phosphatidylinositol-3 kinase but rather on MAPK-dependent cascades. These results help to understand the mechanisms of action of tibolone on the cardiovascular system and have relevant clinical implications.

Cells, Cultured↗

PubMed2003

In vitro effects of progesterone and progestins on vascular cells.

The impact of progesterone on the cardiovascular system is relevant, but not as well characterized as the effects of estrogens. The recent early interruption of the conjugated equine estrogens (CEE)-medroxyprogesterone acetate (MPA) arm of the Women's Health Initiative trial, but not of the parallel CEE-only treatment arm, suggesting the possibility of harmful cardiovascular effects of the progestins, boosts the debate on the role of progesterone and progestins on the vascular tree. The data available up to now show the presence of important regulatory effects of progestagens on vascular cells. Additionally, the presence of a progestagen results in diverse modifications of the effects of estrogens, sometimes acting synergically, others being neutral or antagonizing estrogens' effects. Notwithstanding the availability of consistent observations on the functional effects of progestins on the cardiovascular system, the molecular mechanisms of progestins actions on vascular cells have been up to now only scarcely characterized. Novel mechanisms of signal transduction are being discovered for progesterone receptors in different tissues, some of which are independent of gene transcription regulation, and are therefore indicated as "nongenomic." Furthermore, the contribution to signal transduction of co-activators is currently widely investigated, in order to understand the ways to tissue-specificity and to engineer new progesterone receptor modulators. The understanding of the molecular basis of progesterone receptor signaling in vascular tissue is therefore of paramount importance for the development of hormonal agents with an optimal cardiovascular profile.

Blood Pressure↗

PubMed2003

Dehydroepiandrosterone modulates endothelial nitric oxide synthesis via direct genomic and nongenomic mechanisms.

Dehydroepiandrosterone (DHEA) and its sulfate ester (DHEAS) are the major circulating steroid hormones in humans, and their levels progressively decline with age. Epidemiological studies suggest that DHEA/DHEAS concentrations may be inversely related to cardiovascular risk, but disagreement exists on this issue. Preliminary studies show that DHEA regulates vascular function, but few data have been published on the mechanisms. We show that DHEA administration to human endothelial cells triggers nitric oxide synthesis, due to enhanced expression and stabilization of endothelial nitric oxide synthase (eNOS). Additionally, DHEA rapidly activates eNOS, through a nontranscriptional mechanism that depends on ERK1/2 MAPK, but not on phosphatidylinositol 3-kinase/Akt. DHEA is not converted to estrogens or androgens by endothelial cells, and its genomic and nongenomic effects are not blocked by antagonists of the estrogen, progesterone, glucocorticoid, or androgen receptors, suggesting that DHEA acts through a specific receptor. Oral DHEA administration to ovariectomized Wistar rats dose-dependently restores aortic eNOS levels and eNOS activity, confirming the effects of DHEA in vivo. Our present data suggest that DHEA may have direct genomic and nongenomic effects on the vascular wall that are not mediated by other steroid hormone receptors, leading to eNOS activation and induction.

Androgen Receptor Antagonists↗

PubMed2003

[Cardiovascular effects of selective estrogen receptor modulators. Current perspectives].

The use of hormone replacement therapy (HRT) after the menopause for the prevention of the long-term complications of estrogen deprivation has recently been questioned after the publication of large clinical trials that failed to show benefits for postmenopausal women. Although these trials risk to dump the widespread opinion of the cardioprotective effects of long-term estrogen use, they have many pitfalls that prevent a direct clinical application of these negative results. Furthermore, the large amount of epidemiological and experimental evidence indicating estrogens as protective on the vascular system cannot be ignored, and efforts should be devoted to understand the reasons for the discrepancy of results of these recent large trials. In the meanwhile, different molecules should be studied in depth as for the actions on the cardiovascular system, and their specific mechanisms of actions should be elucidated. Selective estrogen receptor modulators (SERM) are a promising family of molecules and some of these compounds have positive effects on cardiovascular risk parameters as well as on vascular cells. Large trials are ongoing to study the impact of these substances on cardiovascular risk, and the near future should provide us with answers on the possible use of SERM as possible safer alternatives to HRT for the long-term prevention of cardiovascular disease in postmenopausal women.

Aged↗

PubMed2002

Novel non-transcriptional mechanisms for estrogen receptor signaling in the cardiovascular system. Interaction of estrogen receptor alpha with phosphatidylinositol 3-OH kinase.

Estrogen receptor (ER) signaling has been, for a long time, associated with transcriptional processes involving nuclear translocation and binding on specific response elements, leading to regulation of target gene expression. However, rapid, non-transcriptional mechanisms of signal transduction through steroid hormone receptors have been identified. These so-called 'non-genomic' effects are independent from gene transcription or protein synthesis and involve steroid-induced modulation of cytoplasmic or cell membrane-bound regulatory proteins. Several biological actions of estrogen have been associated with this type of signaling, and intracellular regulatory cascades such as extracellular signal-regulated kinase/mitogen-activated protein kinases (ERK/MAPK) and tyrosine kinases or the modulation of G-protein-coupled receptors have been shown to be non-transcriptionally recruited by estrogen in diverse tissues. The vascular wall is one of these sites, where estrogen triggers rapid vasodilatation mainly due to increased nitric oxide (NO) release. We have recently described a novel, non-transcriptional mechanism for ER signaling in human as well as in animal endothelial cells, showing that ER alpha can physically and functionally couple to the lipid kinase phosphatidylinositol 3-OH kinase (PI3K). This interaction leads to activation of PI3K signaling cascade to Ser/Thr kinase Akt, which mediates several PI3K-dependent intracellular effects, including endothelial isoform of NO synthase (eNOS) phosphorylation and activation. This original non-transcriptional mechanism for ER signaling may play an important role in the generation of some of the rapid 'non-genomic' effects of estrogen.

Cardiovascular System↗

PubMed2002

Genomic and nongenomic mechanisms of nitric oxide synthesis induction in human endothelial cells by a fourth-generation selective estrogen receptor modulator.

Cardiovascular disease is the leading cause of morbidity and mortality in postmenopausal women. EM-652 (acolbifene) is a fourth-generation selective ER modulator (SERM) exerting complete antiestrogenic effects on the breast and uterus. EM-652 potently inhibits bone resorption and induces positive lipid modifications in estrogen-deficient animals. As most of the cardioprotective actions of estrogen are exerted directly at the vascular level, we studied the effects of EM-652 on endothelial production of nitric oxide (NO) in vitro and in vivo. EM-652 triggers NO release by human umbilical vein endothelial cells through nongenomic mechanisms, rapidly activating endothelial nitric oxide synthase (eNOS) via an ER-dependent sequential activation of MAPKs and PI3K/Akt pathways independently from gene transcription or protein synthesis. Moreover, EM-652 increases eNOS protein levels during prolonged treatments. Upon pharmacological comparison, EM-652 is markedly more potent than the SERMs raloxifene and tamoxifen in increasing NO synthesis from endothelial cells. In ovariectomized and fertile rats, EM-652 increases aortic eNOS expression and enzymatic activity at low, but not at higher, dosages. The present data show that EM-652 (acolbifene) has estrogen-like activity on the vascular wall, directly increasing NO production through genomic and nongenomic mechanisms in vitro and in vivo.

Animals↗

PubMedDate not supplied

Activation of nitric oxide synthesis in human endothelial cells by red clover extracts.

OBJECTIVE: The unexpected findings of the Women's Health Initiative trial, where surrogate cardiovascular risk markers have failed to predict the cardiovascular performance of hormone therapy, showing no reduction of cardiovascular disease in postmenopausal women receiving hormonal preparations inducing a favorable lipid profile, raise the interest on how molecules with hormone-like activity used for the treatment of menopausal symptoms act on vascular cells. This is particularly important for estrogen-like compounds such as phytoestrogens, whose mechanisms of action may significantly differ from those of other estrogenic compounds. DESIGN: Because endothelial-derived nitric oxide (NO) is a key regulator of vascular tone and atherogenesis as well as a well-characterized estrogen-regulated molecule, we studied the regulation of NO synthesis in cultured human endothelial cells by phytoestrogens contained in red clover extracts. RESULTS: We show that red clover extracts activate NO synthesis in endothelial cells by recruiting transcriptional pathways but are not capable of inducing rapid NO synthesis through nongenomic mechanisms. During prolonged exposures, red clover extracts enhance the expression as well as the activity of endothelial nitric oxide synthase. These effects are mediated by a recruitment of estrogen receptor-beta. Moreover, we show that red clover-derived isoflavones synergize with 17beta-estradiol in increasing endothelial nitric oxide synthase activity and expression, therefore being devoid of antiestrogenic effects in human endothelial cells. CONCLUSIONS: These results help to understand the mechanisms of action of phytoestrogens on the cardiovascular system and have relevant clinical implications.

Cells, Cultured↗

PubMedDate not supplied

Diagnostic approach to a persistent anterior knee pain.

The case of a 67-year-old female patient with persistent pain in the right anterior knee appeared after she had an accidental fall, is discussed. Laboratory tests and imaging were negative. Persistent pain required MRI. Based on MRI findings the diagnosis of patellar fracture responsible for knee pain could be established.

Accidental Falls↗

PubMedDate not supplied

Obstructive jaundice.

The case of a 67-year-old male patient affected by ingravescent obstructive jaundice, calcific choledocholithiasis and cardiopathic hypertension, is presented. On sonography, marked dilatation of intra- and extrahepatic bile ducts with a hyperechoic round neoformation was visualized. CT was performed to ascertain the nature of the lesion. However, it did not confirm the lithiasic nature of the stenosis. Sampling for ERCP was inadequate. Cholangiopancreatography was therefore necessary to establish a correct diagnosis and the subsequent therapeutic approach. Distal choledochal obstruction was evidenced, suspicious for lithiasis obstruction. A definitive diagnosis could not be established on exclusive imaging findings. Surgery was performed with the diagnosis of cholangiocarcinoma with lithiasis.

Aged↗

PubMedDate not supplied

Perception and reality in a case of suspected cervical fracture.

The case of a 36-year-old female, who following a road accident, had a cervical whiplash, is discussed. A radiographic examination of the affected region was performed at the Emergency Department. It showed a transverse radiolucent line at the basal odontoid process of the axis. A definitive interpretation was not possible and fracture of the odontoid process of the axis was suggested as differential diagnosis. On coronal CT reconstruction the presence of this fracture was ruled out. The line seen on the film was actually the product of a psychophysiologic visual illusion known as "Mach band". The use of conventional radiography correlated with digital CT images shows that the practical application of Mach band theory allows a better differentiation between normal and pathologic anatomy.

Accidents, Traffic↗