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Biomedical subjects

G Egidy

Publications and source records attributed to G Egidy.

16 recordsLinked to original sources
PubMed2005

The endothelin system and renin in human fetal membranes.

Term human fetal membranes express prorenin, a key enzyme within the renin-angiotensin system. High levels of another vasoactive peptide, endothelin-1 (ET-1), are found in human amniotic fluid. To address the question of the relationship between these two vasoactive systems, we analyzed the expression of the components of the ET-1 system in fetal membranes in which cell types had been identified using different markers. Immunohistochemistry was performed with antibodies raised against the human proteins of the ET system. Term fetal membranes displayed ubiquitous labeling of endothelin-converting enzyme-1 (ECE-1) and ET-1. ETA receptors were detected in the chorionic connective tissue and the attached decidua; ETB receptors were localized to chorionic trophoblast cells and decidua. The localization of the ET-1 receptor subtype was confirmed by in-situ receptor binding. Renin immunoreactivity was detected in the chorionic connective tissue and the decidua. These findings suggest that ET-1 is produced ubiquitously in human fetal membranes, and its targets may be, trophoblast cells following ETB receptor activation, vascular structures and fibroblasts in the connective tissue and decidua via ETA and ETB receptors. It appears possible that renin and ET may contribute to the pathophysiological changes associated with premature labor and preeclampsia.

Binding Sites↗
PubMed2001

Pulmonary endothelinergic system in experimental congestive heart failure.

OBJECTIVES: Endothelin-1 (ET-1), plays an important role in the pathophysiology of CHF and the pulmonary endothelium is an early hemodynamic target in diastolic left ventricular dysfunction. Therefore we hypothesized that the lung is a main source of humoral endothelin in CHF and that its secretion is proportional to the degree of heart failure. METHODS AND RESULTS: We used rats with coronary artery ligation as an experimental model of either compensated or decompensated heart failure, depending on infarct size. Reverse transcriptase polymerase chain reaction (RT-PCR) revealed that in the lung, the expression of preproET-1 mRNA was higher in decompensated HF than in control and compensated HF rats (P<0.001). Run-on assay demonstrated that ET-1 overexpression is regulated at a transcriptional level (P<0.01). In contrast, there was no change in ET-1 mRNA expression in aortae, left ventricular myocardium and skeletal muscle. The expression of endothelin-converting enzyme (ECE)-1 mRNA was not modified and the expression of ET(B) receptor mRNA in the congestive lung was significantly lower than in control and compensated HF rats (P<0.0001), while the expression of ET(A) receptor mRNA did not differ between groups. The lung and plasma ET-1 peptide levels were respectively 4.2 and 9 fold higher in the rats with decompensated HF than in control rats (P<0.05; P<0.0001). Organoculture experiments showed that the lung ET-1 peptide secretion level in rats with decompensated HF was higher than that in control rats (P<0.01). In contrast, there was no change in ET-1 peptide secretion by the left ventricular myocardium and skeletal muscle. In plasma of rats with decompensated HF, the rate of bigET-1 conversion to ET-1 was 22%. ET-1 peptide was also present in the pleural effusion of decompensated heart failure. Plasma ET-1 concentration was significantly correlated with upstream markers of left ventricular diastolic dysfunction, with the expression of preproET-1 mRNA in the lung, with lung and pleural ET-1 concentration and with the expression ratio of ET-1/ET(B) receptor mRNA. CONCLUSION: Taken together, these data suggest that overexpression of ET-1 and down-regulation of ET(B) receptors in the lung are determinants of circulating endothelin in CHF. As a corollary, increased plasma endothelin may provide evidence of pulmonary endothelial dysfunction in CHF.

Analysis of Variance↗
PubMed2001

Increased AT(1) receptor expression and mRNA in kidney glomeruli of AT(2) receptor gene-disrupted mice.

The proposed feedback between angiotensin II AT(2) and AT(1) receptors prompted us to study AT(1) receptor expression in kidneys of male AT(2) receptor-gene disrupted mice (agtr2 -/y). In wild-type (agtr2 +/y) mice, AT(1) receptor binding and mRNA is abundant in glomeruli, and AT(1) receptor binding is also high in the inner stripe of the outer medulla. AT(2) receptors are scarce, primarily associated to cortical vascular structures. In agtr2 -/y mice, AT(1) receptor binding and mRNA were increased in the kidney glomeruli, and AT(1) receptor binding was higher in the rest of the cortex and outer stripe of the outer medulla, but not in its inner stripe, indicating different cellular regulation. Although AT(2) receptor expression is very low in male agtr 2 +/y mice, their gene disruption alters AT(1) receptor expression. AT(1) upregulation alone may explain the AT(2) gene-disrupted mice phenotype such as increased blood pressure, higher sensitivity to angiotensin II, and altered renal function. The indirect AT(1)/AT(2) receptor feedback could have clinical significance because AT(1) antagonists are widely used in medical practice.

Angiotensin II↗
PubMed2001

Localization of the endothelin system in aldosterone-producing adenomas.

Endothelin-1 (ET-1) could play a role in the regulation of aldosterone secretion of the human adrenal gland. The presence of the endothelin-converting enzyme 1 (ECE-1) and ET-1 suggests that there is a local ET system in the adrenal cortex, but the in situ synthesis of ET-1 remains to be confirmed. The cellular distribution of the whole ET system was evaluated in 20 cases of aldosterone-producing adenomas. Polymerase chain reaction studies gave strong signals for ECE-1 mRNA and the mRNAs for endothelin type A (ET(A)) and B (ET(B)) receptors and faint signals for prepro-ET-1 mRNA. In situ hybridization showed ET(A) receptors scattered throughout the adenoma, in both secretory cells and vascular structures (score, +). There were more ET(B) receptors (score, ++), but they were restricted mainly to the endothelium. ECE-1 mRNA and protein were ubiquitous and abundant in secretory cells (score, +++) and vascular structures (score, ++); the enzyme was active on big ET-1. There was no prepro-ET-1 mRNA in the cortex, except in the thickened precapillary arterioles present in only 30% of the aldosterone-producing adenomas studied. ET-1 immunoreactivity was detected in vascular structures (score, +), probably bound to receptors, suggesting that ET-1 has an endocrine action. The low concentrations of ET-1 could also indicate that it acts in a paracrine-autocrine fashion to control adrenal blood flow. The discrepancy between the concentrations of ECE-1 and its substrate suggests that ECE-1 has another role in the adrenal secretory cells. Our data indicate that ET probably is not a primary cause of the development or maintenance of the adenoma.

Adrenal Cortex↗
PubMed2000

Modulation of human colon tumor-stromal interactions by the endothelin system.

Tumor neovascularization is considered to be a critical step in the development of a malignant tumor. Endothelin (ET)-1 is a powerful vasoconstrictor and mitogenic peptide that is produced by many cancer cell lines. The cellular distribution of the ET components was evaluated in human colon tumors and compared to normal colon. There was more of the ET components (preproET-1, endothelin-converting enzyme-1, and ETA and ETB receptors) in adenomas and adenocarcinomas than in the normal colon. There was overproduction of preproET-1 and endothelin-converting enzyme-1 in carcinoma cells and stromal vessels, suggesting that they are a local source of ET-1. ETA receptors were present in stromal myofibroblasts of neoplastic tissue, and there were large amounts of ETB receptors in the endothelium and myofibroblasts. There was also a redistribution of alpha-smooth muscle actin-positive cells in the vascular structures of tumors. An experimental rat model of induced colon cancer treated for 30 days with bosentan, a mixed antagonist of both ET receptors, confirmed the morphological changes observed during the tumor vascularization. Our data suggest that ET-1 and its receptor play a role in colon cancer progression, with ET-1 functioning as a negative modulator of the stromal response.

Adenocarcinoma↗
PubMed2000

The endothelin system in human glioblastoma.

Endothelin-1 (ET-1) is a powerful mitogenic and/or anti-apoptotic peptide produced by many cancer cells. To evaluate the potential role of the endothelin system in glioblastoma we first determined the cellular distribution of the mRNA and proteins of the components of the endothelin system, preproendothelin-1 (PPET-1), endothelin-converting enzyme-1 (ECE-1), and ET(A) and ET(B) receptors in human glioblastoma tissue and glioblastoma cell lines. PPET-1, ECE-1, and ET(A) receptor were highly expressed in glioblastoma vessels and in some scattered glioblastoma areas whereas ET(B) receptor was mainly found in cancer cells. This suggests that glioblastoma vessels constitute an important source of ET-1 that acts on cancer cells via the ET(B) receptor. Four human glioblastoma cell lines expressed mRNA for all of the components of the ET-1 pathway. Bosentan, a mixed ET(A) and ET(B) receptor antagonist, induced apoptosis in these cell lines in a dose-dependent manner. Apoptosis was potentiated by Fas Ligand (APO-1L, CD95L), a pro-apoptotic peptide, only in LNZ308 cells, corresponding to the known functional Fas expression in these cell lines. LNZ308 cells also expressed the long and short forms of the cellular FLICE/caspase-8 inhibitory protein (FLIP). Bosentan and a protein kinase C inhibitor down-regulated short FLIP in these cells. ET-1 induced transient phosphorylation of extracellular signal-regulated kinase but did not induce long-term thymidine incorporation in LNZ308 glioblastoma cells. These results suggest that, in glioblastoma cells, ET-1, mainly acting via the ET(B) receptor, is a survival/antiapoptotic factor produced by tumor vasculature, but not a proliferation factor, involving protein kinase C and extracellular signal-regulated kinase pathways, and stabilization of the short form of FLIP.

Base Sequence↗
PubMed2000

Endothelin receptor blockade potentiates FasL-induced apoptosis in colon carcinoma cells via the protein kinase C-pathway.

An imbalance between proliferation and apoptosis is important in tumor progression. Endothelin-1 (ET-1) has vasoconstricting and mitogenic activities and may be involved in apoptosis regulation. We found that ET-1 and FasL systems were colocalized in human colon tumors and that ET-1 was secreted by human (HT-29, SW480) and rat (PROb, REGb) colon carcinoma cell lines. Bosentan, a mixed endothelin-A- and -B- (ET(A)/ET(B)) receptor antagonist, potentiated FasL- (APO-1, CD95) induced apoptosis in these cells. The specific inhibition of enzymes involved in ceramide production did not restore survival of cells exposed to FasL and bosentan. Inhibition of PKC with bisindolylmaleimide IX enhanced FasL-induced apoptosis in HT-29, PROb and REGb cells in the absence of bosentan. These results suggest that ET-1 is an autocrine survival factor able to protect colon carcinoma cells against FasL-induced apoptosis, involving the protein kinase C (PKC) but not the sphingomyelin-ceramide signaling transduction pathways.

Animals↗
PubMed2000

The endothelin system in normal human colon.

Endothelin (ET)-1 is a potent vasoconstrictor and mitogenic peptide that has a variety of biological effects in noncardiovascular tissues. The precise cellular distribution of the ET-1 system in the wall of the normal human colon was studied to identify the physiological role of ET in the gut. In situ hybridization revealed ET-converting enzyme-1 (ECE-1) mRNA in all vessels, the colon epithelium, and macrophages. Prepro-ET-1 (PPET-1) mRNA had a similar distribution except for a scattered signal in mucosal microvessels. ET(A) and ET(B) receptor mRNAs were mainly in the lamina propria, pericryptal myofibroblasts, microvessels, and mononuclear cells, with ET(A) mRNA more abundant than ET(B) mRNA. (125)I-ET-1 binding showed ET(B) along the crypts and in nerve fibers descending from the ganglionic plexus that contained PPET-1, ECE-1, and ET(B) transcripts, whereas glia contained ET(A) receptors. The finding of the entire ET system in the normal mucosa suggests its implication in some characteristic functions of the colon and its secretion as both a neuroactive and a vasoactive peptide.

Adult↗
PubMed1999

A fourth isoform of endothelin-converting enzyme (ECE-1) is generated from an additional promoter molecular cloning and characterization.

Human endothelin-converting enzyme (ECE-1) has been shown to exist as three isoforms (ECE-1a, ECE-1b and ECE-1c) diverging in their N-terminal sequence and displaying different patterns of subcellular localization. We report here the cloning of ECE-1d, a novel isoform of 767 amino acids, which is generated from the same gene via the existence of an additional promoter located upstream from the third exon of the ECE-1 gene. ECE-1d converting activity is comparable to that of the other three isoenzymes. In contrast to ECE-1b, ECE-1d is expressed at the cell surface, although less strongly than ECE-1a. We have also shown, by identifying ECE-1b and ECE-1d in rat, that the ECE-1 diversity is conserved between human and rodent, suggesting its physiological relevance. The mRNA levels of the four isoforms were assessed in the two species in various cell types, revealing some differences. In particular, the ECE-1a isoform, strongly expressed at the plasma membrane, was found to be highly expressed in primary cultures of endothelial cells but absent from primary cultures of smooth muscle cells.

Amino Acid Sequence↗
PubMed1997

Construction, expression and characterization of a soluble form of human endothelin-converting-enzyme-1.

Endothelin-converting-enzyme-1 (ECE-1) belongs to the family of zinc metallopeptidases and is responsible for generating endothelin (ET) peptides from their inactive precursors the big endothelins (bigET). The enzyme is a type II integral membrane protein consisting of a short amino-terminal cytosolic domain of 56 amino acids, a single transmembrane domain and a large putative extracellular domain containing the catalytic site. Recombinant and native ECE-1 are expressed as a dimer. We have constructed a soluble form of ECE, named sECE*, by fusing the cleavable signal peptide of pro-opiomelanocortin in frame to the complete extracellular domain of human ECE-1. Stable expression of this construct in CHO cells resulted in the secretion of a fully active enzyme. In contrast to membrane-bound ECE, sECE* was expressed as a monomer, highly glycosylated, as assessed by gel filtration and Western blot. However, recombinant sECE* converted bigET-1 with similar specific activity as ECE-1a. This activity was completely inhibited by phosphoramidon, but not by thiorphan and captopril. sECE* was active in a broad range of pH, showing an optimum of 6.6-6.8 for bigET-1. Thus, the extracellular domain alone is sufficient for conferring full ECE-1 activity, inhibitors recognition and substrate specificity.

Animals↗
PubMed1997

Angiotensin II AT1A receptor mRNA expression is induced by estrogen-progesterone in dopaminergic neurons of the female rat arcuate nucleus.

Brain angiotensin II (Ang II) inhibits pituitary prolactin release by an indirect mechanism requiring stimulation of dopamine formation and release. We report that [125I]Sar1-Ang II binding to AT1 receptors and AT1A receptor mRNA expression increase selectively in the dorsomedial arcuate nucleus of 17beta-estradiol-primed ovariectomized rats after treatment with progesterone. In hormone-treated rats, arcuate nucleus AT1A receptor mRNA expression is associated with tyrosine hydroxylase-positive neurons. No AT1A receptor mRNA was detected in tyrosine hydroxylase-positive cells of the arcuate nucleus of intact male rats. Conversely, in the anterior pituitary, where local or circulating Ang II stimulates prolactin release, [125I]Sar1-Ang II binding to AT1 receptors and AT1B receptor mRNA expression are decreased in 17beta-estradiol/progesterone-treated ovariectomized rats. Thus, AT1A receptors in the dorsal arcuate nucleus and AT1B receptors in the anterior pituitary are regulated inversely by estrogen/progesterone treatment, supporting the hypothesis of a dual role for brain and pituitary Ang II on prolactin release. The colocalization of AT1A receptor mRNA and tyrosine hydroxylase in neurons of the arcuate nucleus furthermore indicates that within this area central Ang II acts directly on dopaminergic neurons. These results support the hypothesis that central Ang II inhibits pituitary prolactin release indirectly via modulation of dopaminergic activity in the arcuate nucleus.

Angiotensin II↗
PubMed1997

CGP-42112 partially activates human monocytes and reduces their stimulation by lipopolysaccharides.

CGP 42112, a high-affinity ligand for angiotensin II AT2 receptors, binds to rat macrophage/microglia lacking AT2 receptors. Here we report that CGP-42112 binds to human monocytes and exerts specific effects. Binding studies revealed a single site, highly specific for CGP-42112, not displaceable by angiotensin II, angiotensin fragments, tumor necrosis factor-alpha (TNF-alpha), interleukin (IL)-4, IL-10, transforming growth factor-beta, or lipopolysaccharide (LPS). Incubation of purified human monocytes in serum-free medium with CGP-42112 enhanced, in a dose-dependent manner, cell attachment to fibronectin and collagen-coated dishes as well as matrix metalloproteinase-9 secretion. CGP-42112 did not promote cytokine secretion. In contrast, when added in the presence of low doses of LPS, CGP-42112 reduced the LPS-stimulated secretion of TNF-alpha, IL-1 alpha, IL-1 beta, and IL-6 without affecting IL-10 and decreased the LPS-stimulated matrix metalloproteinase-9 activity. Additionally, CGP-42112 inhibited the increase in protein kinase A activity produced by LPS. Our results indicate that CGP-42112 may modulate monocyte activation through binding to a novel receptor.

Animals↗
PubMed1991

Activation of the cAMP cascade by steroidogenic hormones and glucagon in the pathogenic fungus Candida albicans.

Incubation of Candida albicans yeast cells with human luteinizing hormone (hLH), human chorionic gonadotrophin (hCG) or glucagon produced a significant rise in cAMP total levels. The effect of these hormones in permeabilized cells of the fungus produced a 2-3 fold increase in the Mg2+, GTP-dependent adenylyl cyclase activity as well as full activation of the cAMP-dependent protein kinase (PKA) activity. These results indicate that the interaction of the mammalian hormones with the fungus triggered the cAMP activation cascade in a similar way to that found in higher eukaryotic organisms.

Adenylyl Cyclases↗
PubMed1990

A guanine nucleotide-sensitive, glucagon-stimulated adenylyl cyclase in Candida albicans: effect of glucagon on cell morphology.

GTP, GTP-gamma-S and Gpp(NH)p produced a significant activation of Mg2(+)-dependent adenylyl cyclase in permeabilized cells of Candida albicans. This activation was inhibited by GDP-beta-S. Maximal stimulation (4-6 fold) was obtained when Mg2+ and GTP-gamma-S were added during permeabilization. The guanine nucleotide-stimulated activity could be further activated by glucagon in a dose dependent manner being the maximal stimulation (4-5 fold) attained at 10(-6) M glucagon. Addition of the hormone to yeast cells incubated under conditions conducive to produce germ tubes blocked hyphal development and promoted multibudded cell morphology.

Adenylyl Cyclases↗
PubMed1990

cAMP levels and in situ measurement of cAMP related enzymes during yeast-to-hyphae transition in Candida albicans.

Intracellular levels of cAMP and specific activities of adenylate cyclase, cAMP phosphodiesterase and cAMP-dependent protein kinase were measured during filamentation in the dimorphic fungus Candida albicans. Enzymatic assays were performed in permeabilized cells under conditions prevented endogenous proteolysis. The variations observed in cAMP levels were mainly accounted for by variations in the specific activities of adenylate cyclase and cAMP phosphodiesterase at different stages during germ tube formation. cAMP-dependent protein kinase, measured with kemptide as exogenous substrate, was developmental regulated. Some properties of the enzymatic activities from cell-free extracts are described.

3',5'-Cyclic-AMP Phosphodiesterases↗