INSIGHTS ON AROMATASE IMMUNOHISTOCHEMISTRY: VARIATIONS BETWEEN INTRINSIC MOLECULAR SUBTYPES OF BREAST CANCERS.

CONTEXT: Aromatase is a key enzyme in local estrogen production by androgen conversion, especially in women post-menopause. There have been controversies concerning aromatase localization in breast carcinomas and its association with current histopathological variables. MATERIAL AND METHODS: Using polyclonal antibody immunohistochemistry we assessed (by intensity and percentage scores) the immunolocalization of aromatase in 70 tissue samples, and described particularities within the molecular subtypes of breast cancer. RESULTS: Aromatase was found in all tissue compartments: tumor (95.7%), stroma (58.6%) and adipose tissue (94.3%). Aromatase expression in tumor cells correlated inversely with tumor grading (p=-0.361, p=0.027), and positively with estrogen receptor status (ER, p=0.143, p<0.001). Dividing the study group by intrinsic subtypes, a strongly inversely association between tumor aromatase and grading (p=-0.486, p<0.001), and between stromal aromatase and Ki67-index (p=-0.448, p=0.048) was observed in luminal A breast cancer. Tumor aromatase and ER percentage scores had stronger correlations in luminal B HER2 negative (p=0.632, p=0.002), and positive (p=0.324, p=0.026) tumors. In contrast, in triple negative tumors, a positive association stromal aromatase and Ki67 index (p=-0.359, p=0.007) was observed. CONCLUSION: Local aromatase was linked to better tumor differentiation and proliferation in luminal breast subtypes, and not in triple negative cases, suggesting a potential prognostic role of aromatase in breast carcinomas.

[Relevance of L-type calcium channels for the non-genomic effects of estradiol and aldosterone upon contractile activity in isolated arteries]. / Relevanta canalelor de calciu de tip L pentru efectele non-genomice ale estradiolului si aldosteronului asupra activitatii contractile în artere izolate.

UNLABELLED: Non-genomic vascular effects of steroids are incompletely understood, despite progress made regarding some aspects, such as the mechanism of endothelium-dependent relaxation by estrogens. AIM: To investigate the involvement of certain mechanisms in the rapid, non-genomic effects of estradiol (EST) and aldosterone (ALD) on endothelium-dependent and -independent vasomotor responses. MATERIAL AND METHODS: Isometric myography of rings from aorta, mesenteric arch, and first order mesenteric branches isolated from male Wistar rats was used. RESULTS: We found that L-type calcium channels (Cav1.2) are important for endothelium-independent relaxation induced by EST, while ALD reduces the involvement of Cav1.2 in phenylephrine-induced contraction and potentates both NO- and EDHF-mediated endothelium-dependent relaxation. To further examine the relevance of Cav1.2 for the vascular effects of EST and ALD, we were using rings with and without functional endothelium, precontracted by direct activation of Cav1.2 (Bay K 8644), high extracellular K+, phenylephrine, and under complete Cav1.2 block (nifedipine). Data suggest that EST, which directly inhibits Cav1.2 in transfected HEK cells, uses mainly this path to induce endothelium-independent relaxation, and that ALD may induce a rightward shift in the voltage-dependence of Cav1.2.

The importance of EDHF in endothelium-dependent relaxation increases distally in mesenteric arteries depending upon the contracting agent.

AIM: The importance of endothelium-derived hyperpolarizing factor (EDHF) in endothelium-dependent relaxation (EDR) is influenced by multiple factors, including vascular territory and caliber, pre-existing tone and its determining factors. METHOD: Using isometric myography we noticed that in rat mesenteric resistance arteries (RMA2; 2nd order branches) EDHF-mediated relaxation is increased when precontraction is induced by prostaglandin F2 (PGF) compared to phenylephrine (PHE) and we investigated the participation of certain K channels. Here we extend the study on more proximal vascular fragments; from mesenteric arcade and from 1st order branches. RESULTS: The EDHF component of EDR is stronger distally only when precontraction is induced by PHE. Moreover, morphometric analysis shows a strong inverse correlation between the magnitude of EDHF response and arterial caliber. CONCLUSIONS: Other authors have shown that EDHF increases in relative importance distally, but we show that this change in EDR profile depends upon the contracting agent used, with implications regarding the physiological relevance of accumulated data refering to EDHF and nitric oxide as mediators of EDR in resistance arteries.

[Essential hypertension in children--risk factors and target organ damage]. / Hipertensiunea arteriala esentiala la copil--factori de risc si afectarea organelor tinta.

UNLABELLED: Regarding that systemic hypertension is an important condition in childhood, we try to clarify existing data about this pathology. MATERIAL AND METHODS: The aim of our study was to evaluate the incident cases of essential hypertension in children admitted during a four years period in the Children Hospital "Sf. Maria" Iasi, Romania. The study included 60 children with essential hypertension. RESULTS: 10 (16,7%) of children were in prehypertension group, 38 (63,3%) have stage 1 hypertension and 12 (20%) have stage 2 hypertension. We objectivate a different gender distribution - boy/girl: 1,5/1. The frequency of essential hypertension increased with age, the incidence was higher over 14 years. We established a statistically significant relationship between the essential hypertension body mass index and prematurity (gestation age under 37 weeks). Left ventricular hypertrophy was objectivated in 26,7% of cases. CONCLUSIONS: Essential hypertension in children is a pathology with increasing prevalence nowadays. This is due to synergic action of multiple risk factors involved. That is why is important to know and to prevent as much as possible this disease.

Relaxation and polarization of precontracted rat aortic smooth muscle by sodium nitrite and an L-type channel blocker.

UNLABELLED: Membrane potential (MP) is essential for smooth muscle (SM) contraction, mainly by determining the activity of L-type Ca2+ channels (Ca(L)). Although a widely used SM model, there are few electrophysiology studies in rat aorta. AIM: We investigated the mechanism of SM relaxation induced by NaNO2 in comparison with a Ca(L) blocker. METHOD: The dynamic MP-force relation was studied in de-endothelised rat aorta rings during contraction by 40 mM K+ or 0.01 mM phenylephrine (PHE) and relaxation by 0.01 mM methoxy-verapamil (D600) or 0.1 mM NaNO2. RESULTS: We confirm data on resting MP and depolarization by K+ or PHE. MP and Ca(L) are essential for K+ contraction and have little involvement in PHE contraction. NaNO2 polarization is not positively correlated with the relaxing effect. CONCLUSIONS: Our studies on the MP-force relation provide novel information regarding the excitation-contraction coupling in SM. Polarization seems to be just an additional mechanism within nitrite-induced relaxation and may not involve K+ channel activation.

[Evaluation of umbilical artery resistance index and plasma angiotensin II in pregnancy induced hypertension]. / Evaluarea indicelui de rezistenta al arterei ombilicale si nivelului plasmatic de angiotensina II in hipertensiunea indusa de sarcina.

UNLABELLED: It has already been proven that the renin-angiotensin system has deep implications during the normal pregnancy and along with pregnancy induced hypertension. MATERIAL AND METHOD: We evaluated 14 third trimester pregnancies (8 normotensive and 6 hypertensive women) for body mass index, parity, age, blood pressure (BP), resistance index of the umbilical artery (RIUA), plasma angiotensin II (AngII) level. RESULTS: We identified statistical significant correlations for BP with RIUA, AngII. The covariance analysis showed that the correlation was not influenced by the gestational age. CONCLUSION: AngII and RIUA have statistical significant correlations with BP but no interdependence between them was identified.

Relaxing effects of adrenomedullin in rat mesenteric resistance arteries.

UNLABELLED: Exogenous adrenomedullin (ADM) is hypotensive in very small doses, through direct vasodilation of resistance arteries. In different region this is variably mediated by ADM and CGRP receptors. Mechanisms involve ADM effects upon smooth muscle to increase cAMP and to activate K+ channels, plus stimulation of endothelial NO release and inhibition of sympathetic noradrenaline release. AIM: We studied the mechanism of the vasodilator effect of ADM in vitro. METHOD: Rat mesenteric resistance arteries (RMRA) in isometric conditions, were stimulated using four contracting agents: K+ 40 microM, phenylephrine 10 mM, PGF2alpha 10 microM, angiotensin II 1 microM. We used methylene blue 10 microM to examine the involvement of soluble guanylate cyclase (GC) in the relaxation induced by ADM. RESULTS: We found that relaxation induced by ADM is endothelium-independent, similar regardless of the contracting agent (including high K+), and partially sensitive to methylene blue. CONCLUSIONS: K+ channel activation by ADM is not essential for relaxation. Opposite to other authors, we suggest that ADM-induced vasodilation may involve endothelium-independent activation of soluble GC.

Effects of novel leukotriene receptor antagonists in the isolated rat aorta.

We screened 14 novel antagonists of the LTB(4) and LTD(4) receptors (also inhibitors of LTB(4) synthesis) for their vasoactive properties in the isolated rat aorta. The compounds belong to three classes, e.g. quinoline (Q), phenetylamido (P), and resatophenone (R) derivatives. They are effectiveless in resting conditions and generally display a weak relaxing ability against contraction by either high K(+) or alpha(1) adrenoceptor activation, either in the presence or absence of a functional endothelium. There is little overlap with the generally lower concentration range where their anti-LT properties are already manifested. We could not find any correlation between any of the anti-LT properties and the vasorelaxant effects. Concerning the non-specific vasoactive properties, choice compounds of the examined groups could be further tested regarding the mechanisms of their relaxing effects. Given the many uncertainties concerning LT and vascular physiology, it may be worthy to proceed with this line of investigation.

[L-type calcium channels involvement in aortic smooth muscle contraction as revealed by membrane potential and active force dynamics]. / Implicatii ale canalelor de calciu de tip L în contractia muschiului neted aortic relevate de dinamica potentialului membranar si fortei active.

Membrane potential (MP) is essential in smooth muscle (SM) contractile activity, mainly by its effect upon L-type Ca2+ channels. We simultaneously recorded SM isometric tension and MP in de-endothelised rat aorta rings and examined their submaximal activation by K+, norepinephrine (NE) or phenylephrine (PHE) and the influence of methoxyverapamil (D600). K+ -induced contraction strictly correlated with depolarization, while faster contractions induced by NE or PHE started and peaked with a less depolarized membrane. D600 completely relaxed K+ or NE contracted rings, time-correlated with full repolarization, but partially relaxed PHE-contracted rings, with partial repolarization, which did not precede relaxation. The observed MP and force dynamics support known mechanisms of action of the drugs used. L-type channels participate in the depolarizing and contractile effect of NE, as opposite to their minor involvement in the effects of PHE.

[Cytosolic calcium dynamics and smooth muscle contraction.III Plasmalemmal calcium fluxes]. / Dinamica citosolica a calciului si contractia muschiului neted. III. Fluxuri de calciu plasmalemale.

Smooth muscle contractile activity depends upon cytosolic Ca2+, the Ca(2+)-sensitivity of actin-myosin interaction and several auxiliary mechanisms. This section presents the plasmalemmal Ca2+ fluxes in relation with the functional structure of their supportive proteins and the contractile impact. Summaries of classical data are accompanied by examples of recent advances. Ca2+ influx occurs mainly via the L and T types of voltage-operated Ca2+ channels, the store-operated Ca2+ channels and the non-selective cation channels (operated by membrane receptors or mechanical stimuli). The plasmalemmal Ca2+ ATPase and Na/Ca antiport function to limit increases in cytosolic Ca2+; Na/Ca effect is opposite when driven to operate in the reverse mode.

[Cytosolic calcium dynamics and smooth muscle contraction(II): Reticular calcium fluxes]. / Dinamica citosolica a calciului si contractia muschiului neted (II): Fluxuri de calciu reticulare.

The contractile status of smooth muscle depends upon cytosolic Ca2+, the Ca(2+)-sensitivity of actin-myosin interaction and various calcium-independent mechanisms. This second part of our overview is devoted to the complex involvement of endoplasmic reticulum in the cytosolic Ca2+ signals related to smooth muscle contractile activity, with a focus on the functional structure of reticular membrane proteins that ensure the respective Ca2+ fluxes. Ca2+ release is activated by cytosolic Ca2+, involving reticular channels called inositol triphosphate receptors and ryanodine receptors. Beside calcium and inositol triphosphate, cyclic ADP-ribose and nicotinic acid adenine dinucleotide phosphate have recently emerged as intracellular signals that activate Ca2+ release. The reticular Ca2+ pump is essential both for the control of cytosolic Ca2+ and for the preservation of reticular stores.

Implications of imidazolines and imidazoline receptors role at the vascular level.

There are both post- and pre-synaptic vascular imidazoline (IM) binding sites. The importance of direct IM actions and that of peripheral imidazoline receptors (IRs) are shadowed by the central effects of IMs and by their interaction with alpha 2 adrenoceptors. Since the discovery of clonidine the many studies on IMs have been focused on their hypotensive effect, with rilmenidine and moxonidine as representative drugs. Formerly called IM preferring alpha 2 or IM/guanidium sites, the IRs (idazoxan-sensitive) are the plasmalemmal I1 (clonidine-sensitive) and the various I2 (one structure identified as MAO). I1 signaling includes activation of phosphatidylcholine-selective phospholipase C and inhibition of some ligand-gated channels. Inhibitory IRs on postganglionic sympathetic terminals, are not alpha 2, H3, I1 or I2. Some IMs directly affect CaL, while others inhibit K+ efflux. Clonidine-displacing substances including agmatine are endogenous ligands at IRs and alpha 2 and may participate in arterial pressure control. Beside few speculations, the roles of vascular IRs are largely unknown.

Characteristics and possible mechanisms of low-Na+ induced contractions in rat aorta.

The influence of reducing external Na+ concentration ([Na+]ex) upon vascular smooth muscle contractility was investigated using the rat isolated aorta. NaCl from the physiological saline solution (PSS) was replaced with either choline-Cl, sucrose, or LiCl to give the following [Na+]ex (mM): 115, 85, 55, and 25 (115NaPSS to 25NaPSS). Small reductions in [Na+]ex (115NaPSS) induced a biphasic contraction, comparable in amplitude with the control one induced by phenylephrine 10(-6) M. Elimination of the endogenous catecholamine participation using either phentolamine 10(-5) M or guanethidine 3.10(-6) M similarly reduces these contractions to 25% (sucrose replacement). A similar relaxing effect was obtained with D600 10(-5) M, an antagonist of the voltage operated Ca2+ channels (25-30% residual tension for all the substitutes). Large reductions in [Na+]ex (25NaPSS) induced contractions comparable in amplitude and shape, but less sensitive to phentolamine and guanethidine (residual tension 65-75%, sucrose replacement) and insensitive to D600 (all the substitutes). The Na+/K+ ATPase inhibitor ouabain (10(-4) M) elicited slowly developing contractions, the amplitude being 115% of the phenylephrine 10(-6) M control. Phenylephrine further contracted the 115NaPSS precontracted preparations, but was significantly less effective in 25NaPSS, although the precontraction levels were similar for the same substitute used. The amplitude of the superimposed phenylephrine contractions exhibited [Na+]ex dependence. Phenylephrine 10(-6) M failed to further contract the ouabain 10(-4) M precontracted rings. We conclude that relatively small reductions in [Na+]ex are able to induce contractions of rat aorta primarily through release of endogenous catecholamines, probably through neural Na+/Ca2+ exchange. Larger reductions in [Na+]ex appear to cause contraction through muscular Na+/Ca2+ exchange.

Effects of liposome-entrapped adenosine in the isolated rat aorta.

This study examined the effects of adenosine- and adenosine deaminase-loaded liposomes upon the contractile activity of the vascular smooth muscle, using the isolated, de-endothelised rat aorta ring as in vitro model. While control liposomes had no effect, intraliposomal adenosine (5 x 10(-3) M) induced contraction of the preparation. Intraliposomal adenosine deaminase induced partial relaxation of high K(+)-precontracted rings. The adenosine-induced contraction seems to involve Ca2+ influx through L-type channels as an essential component, but protein kinase C may also have a modulatory role.

Effects of liposome-entrapped D-myo-inositol 1,4,5-trisphosphate and D-myo-inositol 1,3,4,5-tetrakisphosphate in the isolated rat aorta.

This study examined the effects of D-myo-inositol 1,4,5-trisphosphate (Ins(1,4,5)P3)- and D-myo-inositol 1,3,4,5-tetrakisphosphate (Ins(1,3,4,5)P4)-loaded liposomes upon the contractile activity of vascular smooth muscle, using the isolated (endothelium removed) rat aortic ring as in vitro model. While control liposomes had no effect, the administration of Ins(1,4,5)P3-containing liposomes contracted the smooth muscle preparation. Furthermore, a similar effect was seen with the administration of Ins(1,3,4,5)P4-filled liposomes but, in this case, the rings developed a significantly higher level of active tension. Pretreatment of the aortic preparation with heparin-loaded liposomes blocked the contractions induced by both Ins(1,4,5)P3- and Ins(1,3,4,5)P4-containing liposomes.

Adenosine alters the vascular effects of a diacylglycerol analogue and polymyxin B.

Endothelium-denuded rat aorta rings were used to study the possible relationship between protein kinase C and the mechanism of adenosine-induced smooth muscle relaxation. Adenosine (5 x 10(-4) M) partially relaxed the aortic rings contracted by either a depolarising amount of KCl (4 x 10(-2) M) or activation of protein kinase C with 1-oleoyl-2-acetyl-sn-glycerol (10(-6) M). The same amount of adenosine blocked the further relaxation obtained in the presence of polymyxin B (5 x 10(-5) M), a protein kinase C blocking agent. These results suggest a possible interaction in vascular smooth muscle between adenosine and protein kinase C.

Adenosine effects upon the spontaneous quantal transmitter release at the frog neuromuscular junction in the presence of protein kinase C-blocking and -activating agents.

This paper gives experimental evidence involving protein kinase C (PKC) in the inhibitory effects of adenosine (ADO) upon the spontaneous transmitter release at the frog neuromuscular junction. In the presence of two PKC inhibitors--polymyxin B (5 x 10(-6) mol/l) and H-7 (10(-5) mol/l), both adenosine (5 x 10(-5) mol/l) and its stable analogue 1-PIA (5 x 10(-8) mol/l), significantly increased the rate of the spontaneous release of acetylcholine quanta. Even when PKC was activated with OAG (5 x 10(-6) mol/l) or TPA (162 x 10(-9) mol/l) and quantal release was increased greatly, ADO still inhibited release. ADO deaminase increased the PKC-induced activation of the transmitter release significantly.

Short-time adenosine deaminase interventions upon underperfused rat hearts in vitro--a test for the role of adenosine in the coronary flow control.

Spontaneously beating rat heart Langendorff preparations, perfused with Tyrode solution under low flow conditions (1.62 +/- 0.47 ml/min at 55 mmHg), were used to investigate the role of adenosine in the coronary flow control, by means of adenosine-deaminase injections into the perfusion system. Adenosine deaminase did not influence the control coronary flow, but significantly reduced autoregulation, hypoxic vasodilation, reactive hyperemia and functional adrenaline-induced hyperemia. All these effects, excepting the hypoxic vasodilation reduction, dependent upon the moment of enzyme injection during the experiment. Under the mentioned conditions adenosine seems to be responsible for more than half of the autoregulation of the coronary flow. Adenosine deaminase completely abolished reactive hyperemia during control perfusion but only delayed it under adrenaline perfusion.