Log EC50 Research Articles

The effects of local over-expression of VEGF on the uterine arteries and uterine artery endothelial cells of pregnant sheep

IntroductionImpaired utero-placental perfusion leads to fetal growth restriction. The authors have shown a significant increase in blood flow in the uterine arteries (UAs) of pregnant sheep and a significantly reduced...

Modeling the interaction of binary and ternary mixtures of estradiol with bisphenol A and bisphenol AF in an in vitro estrogen-mediated transcriptional activation assay (T47D-KBluc).

Exposure to xenoestrogens occurs against a backdrop to physiological levels of endogenous estrogens. Endogenous estrogen levels vary from low levels in early childhood to high levels during pregnancy and in young women. However, few studies have addressed how xenoestrogens interact with endogenous estrogens. The current study was designed to characterize the individual dose-response curves of estradiol-17beta (E(2)), bisphenol A (BPA), tetrabromo-bisphenol A (TBBPA), and bisphenol AF (BPAF, 4,4'-hexafluoroisopropylidene diphenol) on estrogen-dependent luciferase expression in T47D-KBluc cells and to determine how binary (8 x 8 factorial) and ternary (4 x 4 x 4 factorial) mixtures of an endogenous estrogen (E(2)) interact with BPA and/or BPAF. Log EC(50) and hillslope values with SEs, respectively, for individual compounds were as follows: E(2), -12.10M +/- 0.06071, 0.7702 +/- 0.1739; BPA, -6.679M +/- 0.08505, 1.194 +/- 0.2137; and BPAF, -7.648M +/- 0.05527, 1.273 +/- 0.1739. TBBPA was not evaluated in mixture studies because of its minimally estrogenic response at 3 x10(-5)M and elicited cytotoxicity at higher concentrations. Both the binary mixtures of E(2) with BPA and BPAF and the ternary mixture of E(2), BPA, and BPAF behaved in an additive manner. For binary mixtures, as E(2) concentration increased, higher concentrations of BPA and BPAF were necessary to induce a significant increase in the estrogenic response. Understanding the behavior of mixture interactions of xenoestrogens, like BPA and BPAF, with endogenous estrogens will allow a better assessment of the potential risk associated with exposure to these chemicals, individually or as mixtures.

W1778 Epidermal Growth Factor Effect on NHE8 in the Intestine

G A A b st ra ct s acetylcholine (ACh)-induced vasodilation in pressurized mesenteric arteries (diameter: 170250 microns).Methods: Isobaric preparations of 4th-order mesenteric arteries from SpragueDawley rats were used. The arterial diameter was continuously recorded using video microscopy. The effect of Chrm1-4, COX, NOS and potassium channel inhibitors on ACh-induced vasodilation was investigated. The role of Chrm on pressure-induced vascular tone (20-100 mm Hg) and phenylephrine sensitivity was also determined. Only arteries that developed myogenic tone were used. Results: Arteries spontaneously developed >20% reduction in passive diameter. ACh-induced dose-dependent (0.001-10 μM) vasodilation (ANOVA: P<0.05; log EC50: -6.49) was blocked by atropine (non-selective Chrm inhibitor;1 μM) and 4-DAMP (Chrm3-selective inhibitor; 0.1 μM; log EC50: -3.98) (P<0.05 for both). Pirenzepine (Chrm1-selective inhibitor; 1 μM; log EC50: -5.78), methoctramine (Chrm2-selective inhibitor; 1 μM; log EC50: -6.21) and tropicamide (Chrm4-selective inhibitor; 1 μM; log EC50: -5.73) had no effect on ACh-mediated vasodilation. ACh also stimilated dose-dependent vasodilation in arteries incubated with indomethacin (INDO) and L-NAME (COX and NOS inhibitors; 10 and 300 μM, respectively; log EC50: -6.53; ANOVA: P<0.05). Glibenclamide (KATP blocker; 10 μM), 4-aminopyridine (KV blocker; 1 mM), BaCl (KIR blocker; 30 μM) had no effect while tetraethylammonium (KCa blocker; 10 mM) attenuated INDOand LNAME-resistant vasodilation (P<0.05). Intraluminal application of apamin plus charybdotoxin (SKCa and IKCa blockers respectively; 50 nM each) completely abolished INDOand L-NAME-resistant ACh-mediated vasodilation (P<0.05). Additionally, 4-DAMP blocked INDOand L-NAME-resistant ACh-mediated vasodilation (P<0.05) but had no effect on pressure-induced myogenic tone or phenylephrine dose-response. Conclusions:These data indicate that, in small mesenteric arteries in rats, (i) Chrm3 plays a key role in ACh-mediated vasodilation but has no role in regulating basal vascular tone; (ii) Non-prostaglandin and non-NO mechanisms play an important role in Chrm3-mediated vasodilation that is likely induced by EDHF. We conclude that Chrm3 activation induces EDHF-mediated vasodilation in rat small mesenteric arteries.

20-HETE Mediates Ozone-Induced, Neutrophil-Independent Airway Hyper-Responsiveness in Mice

BackgroundOzone, a pollutant known to induce airway hyper-responsiveness (AHR), increases morbidity and mortality in patients with obstructive airway diseases and asthma. We postulate oxidized lipids mediate in vivo ozone-induced AHR in murine airways.Methodology/Principal FindingsMale BALB/c mice were exposed to ozone (3 or 6 ppm) or filtered air (controls) for 2 h. Precision cut lung slices (PCLS; 250 µm thickness) containing an intrapulmonary airway (∼0.01 mm2 lumen area) were prepared immediately after exposure or 16 h later. After 24 h, airways were contracted to carbachol (CCh). Log EC50 and Emax values were then calculated by measuring the airway lumen area with respect to baseline. In parallel studies, dexamethasone (2.5 mg/kg), or 1-aminobenzotriazol (ABT) (50 mg/kg) were given intraperitoneal injection to naïve mice 18 h prior to ozone exposure. Indomethacin (10 mg/kg) was administered 2 h prior. Cell counts, cytokine levels and liquid chromatography-mass spectrometry (LC-MS) for lipid analysis were assessed in bronchoalveolar lavage (BAL) fluid from ozone exposed and control mice. Ozone acutely induced AHR to CCh. Dexamethasone or indomethacin had little effect on the ozone-induced AHR; while, ABT, a cytochrome P450 inhibitor, markedly attenuated airway sensitivity. BAL fluid from ozone exposed animals, which did not contain an increase in neutrophils or interleukin (IL)-6 levels, increased airway sensitivity following in vitro incubation with a naïve PCLS. In parallel, significant increases in oxidized lipids were also identified using LC-MS with increases of 20-HETE that were decreased following ABT treatment.Conclusions/SignificanceThese data show that ozone acutely induces AHR to CCh independent of inflammation and is insensitive to steroid treatment or cyclooxygenase (COX) inhibition. BAL fluid from ozone exposed mice mimicked the effects of in vivo ozone exposure that were associated with marked increases in oxidized lipids. 20-HETE plays a pivotal role in mediating acute ozone-induced AHR.

Endothelium-dependent and -independent vasorelaxation induced by CIJ-3-2F, a novel benzyl-furoquinoline with antiarrhythmic action, in rat aorta

Aims This study was designed to examine the mechanism of relaxation induced by CIJ-3-2F, a benzyl-furoquinoline antiarrhythmic agent, in rat thoracic aorta at the tissue and cellular levels. Main methods Isometric tension of rat aortic ring was measured in response to drugs. Ionic channel activities in freshly dissociated aortic vascular smooth muscle cells (VSMCs) were investigated using a whole-cell patch-clamp technique. Key findings CIJ-3-2F relaxed both phenylephrine (PE) and high KCl (60 mM)-induced contractions with respective pEC 50 (-log EC 50) values of 6.91 ± 0.07 and 6.32 ± 0.06. Removal of endothelium or pretreatment with nitric oxide (NO)-pathway inhibitors N ω-nitro- l-arginine methyl ester (L-NAME), N G-monomethyl- l-arginine (L-NMMA), N 5-(1-iminoethyl)- l-ornithine (L-NIO), hemoglobin, methylene blue or 1 H-[1,2,4]oxadiazolo[4,2-α]quinoxalin-1-one (ODQ) reduced the relaxant effect of CIJ-3-2F. Relaxation to CIJ-3-2F was also attenuated by K + channel blockers tetraethylammonium (TEA) or 4-aminopyridine (4-AP), but not by charybdotoxin plus apamin, iberiotoxin, glibenclamide, or BaCl 2. CIJ-3-2F non-competitively antagonized the contractions induced by PE, Ca 2+, and Bay K8644 in endothelium-denuded rings. In addition, CIJ-3-2F inhibited both the phasic and tonic contractions induced by PE but did not affect the transient contraction induced by caffeine. CIJ-3-2F reduced the Ba 2+ inward current through L-type Ca 2+ channel (IC 50 = 4.1 μM) and enhanced the voltage-dependent K + (K v) current in aortic VSMCs. Significance These results suggest that CIJ-3-2F induced both endothelium-dependent and -independent vasorelaxation; the former is likely mediated by the NO/cGMP pathway whereas the latter is probably mediated through inhibition of Ca 2+ influx or inositol 1,4,5-triphosphate (IP 3)-sensitive intracellular Ca 2+ release, or through activation of K v channels.

Peripheral microvascular responses to norepinephrine in women with orthostatic intolerance

We determined the effects of estradiol (E2) and progesterone (P4) on the cutaneous microvasculature during adrenergic (norepinephrine, NE) stimulation in eight orthostatic tolerant (OT) and eight intolerant (OI) women. We hypothesized that 1) OI have a blunted vasoconstrictor response to NE compared to OT; and 2) that female sex hormones modulate vasoconstriction. A cumulative stress index (CSI) during maximal lower body negative pressure determined orthostatic tolerance. Endogenous E2 and P4 was suppressed with daily administration of a GnRH antagonist (GnRH) for 16 days; we gave E2 (0.2 mg/day, patch) on days 4–16, and P4 (200 mg/day, pills) on days 13–16. We measured skin blood flow (laser Doppler flowmetry) during graded NE infusions (skin microdialysis) during GnRH, E2, and E2+P4. CSI was 871±86 and 397±65 for OT and OI respectively (P &lt; 0.05). The vasoconstrictor response to NE during GnRH was similar between groups (log EC50: OT −3.9, CI −4.9 to −3.1; OI −4.4, CI −5.4 to −3.4). Combined E2+P4 enhanced the vasoconstrictor response in OT (EC50 −4.8, CI −5.6 to −4.1) but was reversed with Ketorolac (−3.8, −4.6 to −3.0). Hormones did not alter vasoconstriction in OI (EC50 −4.3, CI −5.0 to −3.7; −4.9, CI −5.5 to −4.4). Adrenergic response in skin microvasculature was similar between OT and OI. Although OI were insensitive to hormone treatments, P4 augments cutaneous adrenergic responses in OT. HL071159.

Pharmacological characterization of KR-30988, a novel non-peptide AT1 receptor antagonist, in rat, rabbit and dog.

The pharmacological profile of KR-30988, a non-peptide AT1-selective angiotensin receptor antagonist, has been investigated by use of a variety of experimental models in-vitro and in-vivo. KR-30988 inhibited the specific binding of [125I][Sar1, Ile8]-angiotensin II to the recombinant AT1 receptor from man with a potency similar to that of losartan (IC50 values, the concentrations of drugs displacing 50% of specific binding, 13.6 and 12.3 nM, respectively), but did not inhibit the binding of [125I]CGP 42112A to recombinant AT2 receptor from man (IC50 >10 microM for both drugs). Scatchard analysis showed that KR-30988 interacted competitively with recombinant AT1 receptor from man in the same manner as losartan. In functional studies with rat and rabbit aorta, KR-30988 noncompetitively inhibited the contractile response to angiotensin II (pD2, = -log EC50 (where EC50 is the dose resulting in 50% of a reference contraction), 8.64 and 7.73, respectively) with a 20-85% decrease in the maximum contractile responses, unlike losartan. In pithed rats intravenous KR-30988 resulted in a non-parallel shift to the right of the dose-pressor response curve to angiotensin II (ID50 value, the dose inhibiting the pressor response to angiotensin II by 50%, 0.09 mg kg(-1)) with a dose-dependent reduction in the maximum responses; in this antagonistic effect KR-30988 was 20 times (approx.) more potent than losartan (ID50 1-74 mg kg(-1)). In conscious renal hypertensive rats oral administration of KR-30988 produced a dose-dependent and long-lasting (>24 h) anti-hypertensive effect; the potency was six times that of losartan (ED30 values, the dose reducing mean arterial blood pressure by 30 mmHg, 0.48 and 2.97 mg kg(-1), respectively). In conscious furosemide-treated dogs oral administration of KR-30988 produced a dose-dependent and long-lasting (>8 h) hypotensive effect with a rapid onset of action (time to Emax, the maximum effect, 1-2 h); KR-30988 was eight times more potent than losartan (ED20, the dose reducing mean arterial blood pressure by 20 mm Hg, 1.04 and 7.96 mg kg(-1), respectively). These results suggest that KR-30988 is a potent, orally active selective AT1 receptor antagonist with a mode of insurmountable antagonism.

Quantitative structure–activity relationships (QSARs) to estimate ionic liquids ecotoxicity EC50 ( Vibrio fischeri)

Many ionic liquids are soluble in water and their impact on the aquatic environment has to be evaluated. However, due to the large number of ionic liquids and the lack of experimental data, it is necessary to develop estimation procedures in order to reduce the materials and time consumption. Quantitative structure–activity relationships (QSARs) are models that can be used to estimate the physicochemical and toxicological properties of molecules from the molecular structure or properties of similar compounds whose activities have already been assessed. In this work, a novel QSAR based on multiple linear regression is applied in order to estimate the ecotoxicity of ionic liquids, expressed as EC 50 ( Vibrio fischeri), involving 9 kind of cations and 17 anions. The range of log EC 50 values covered by the novel QSAR is from -0.23 to 5.00. From the results, the influence of cations, anions and substitutions on the ecotoxicity of ionic liquids is established.

Estimating the toxicities of organic chemicals in activated sludge process

The experimental log EC 50 toxicity values of 104 compounds causing bioluminescent repression of the bacterium strain Pseudomonas isolated from an industrial wastewater were studied. Using the Best Multilinear Regression method implemented in CODESSA PRO, models with up to 8 theoretical descriptors were obtained. Utilizing a rigorous descriptor selection and validation procedure a reliable QSAR model with four parameters was selected as best. The proposed model emphasizes the importance of the halogen atoms presented in each compound, the possibility of H-bond formation and the flexibility and degree of branching of the molecules. As pointed out by many researchers, the contribution of the octanol–water partition coefficient to the explanation of the toxicity effect was also found to be significant. In addition, the model currently proposed was compared to those reported earlier and its advantages were discussed in detail.

Effects of abnormal cannabidiol on oxytocin-induced myometrial contractility

The objective of this study was to investigate the effects of abnormal cannabidiol (abn-cbd) on oxytocin-induced myometrial contractility occurring during pregnancy. Isometric tension recordings were performed in isolated myometrial strips from biopsies obtained at elective cesarean section. The effects of cumulative doses of abn-cbd (10(-9)-10(-5) M) on oxytocin-induced myometrial contractions alone, and on those following pre-incubation with SR 144528, AM 251, methylene blue, and iberiotoxin were measured, and dose-response curves were constructed. The pD(2) (-log EC(50)) values and the maximal inhibitory (MMI) values that were achieved were compared for each tissue type. Abn-cbd exerted a potent relaxant effect on oxytocin-induced myometrial contractions in vitro. Pre-incubation with the guanylate cyclase inhibitor, methylene blue, and the BK(Ca) channel antagonist, iberiotoxin, significantly attenuated this effect (for pD(2), P<0.01; for MMI, P<0.01). Abn-cbd exerts a potent inhibitory effect on human uterine contractility. This effect is partially mediated through modulation of guanylate cyclase and activation of BK(Ca) channel activity. These findings have implications for physiologic regulation of myometrial quiescence.

Regulation of Cerebromicrovascular Permeability by Lysophosphatidic Acid

Lysophosphatidic acid (LPA) increases permeability of cerebral endothelium in culture, but it has been suggested that histamine release is required in vivo. Cerebral venular permeability was measured by using the single-vessel micro-occlusion technique, and fura-2 ratios were used to track changes in endothelial [Ca(2+)]. Topical acute LPA application dose-dependently increased permeability (log EC(50)-9.4; similar to the K(d) of the LPA1 receptor). The calcium response to LPA was similar to histamine, but the permeability response was unaffected by H(2)-histamine receptor antagonism, and was blocked by Ki16425, a LPA1 receptor antagonist. The permeability response was blocked by nitric oxide synthase and free radical scavenging, which were carried out together, but not separately. Intravascular LPA bolus injection increased permeability. Whole serum albumin, or plasma albumin co-applied with LPA, increased permeability, but less potently than LPA itself (log EC(50) 5.1 and 6.1, respectively). Tachyphylaxis of the LPA1 receptor was demonstrated by LPA application for 10 minutes, which resulted in suppression of the response to subsequent applications for the following 15 minutes. Lysophosphatidic acid increases cerebrovascular permeability by acting directly on the endothelium and utilizes both nitric oxide and free radical signaling pathways.

A quantum-chemical-based guide to analyze/quantify the cytotoxicity of ionic liquids

A COSMO-RS descriptor (Sσ-profile) has been used in quantitative structure–activity relationship studies (QSARs) based on a neural network for the prediction of the toxicological effect of ionic liquids (ILs) on a leukemia rat cell line (LogEC50 IPC-81) for a wide variety of compounds including imidazolium, pyridinium, ammonium, phosphonium, pyrrolidinium and quinolinium ILs. Sσ-profile is a two-dimensional quantum-chemical parameter capable of characterising the electronic structure and molecular size of cations and anions. By using a COSMO-RS descriptor for a training set of 105 compounds (96 ILs and 9 closely related salts) with known biological activities (experimental LogEC50 IPC-81 values), a reliable neural network was designed for the systematic analysis of the influence of structural IL elements (cation side chain, head group, anion type and the presence of functional groups) on the cytotoxicity of ∼450 IL compounds. The Quantitative Structure–Activity Map (QSAM), a new concept developed here, was proposed as a valuable tool for (i) the molecular understanding of IL toxicity, by relating Log EC50 IPC-81 parameters to the electronic structure of compounds given by quantum-chemical calculations; and (ii) the sustainable design of IL products with low toxicity, by linking the chemical structure of counterions to the predictions of IL cytotoxicity in handy contour plots. As a principal contribution, quantum-chemical-based QSAM guides allow the analysis/quantification of the non-linear mixture effects of the toxicophores constituting the IL structures. Based on these favorable results, the QSAR model was applied to estimate IL cytotoxicities in order to screen commercially available compounds with comparatively low toxicities.

Abstract 5226: Endothelial Nox4 NADPH Oxidase Enhances Vasodilatation via Hydrogen Peroxide-induced Hyperpolarization and Reduces Blood Pressure

Introduction NADPH oxidases (Noxs) are reactive oxygen species-generating enzymes implicated in cardiovascular disease. Nox4 is the most abundantly expressed isoform in endothelial cells but its function is unclear. We investigated the role of endothelial Nox4 on vascular function in vivo . Methods and Results We generated transgenic mice with endothelium-specific overexpression of Nox4 (Nox4TG) and studied the effects on endothelial function (aortic rings ex vivo) and blood pressure (BP, telemetry). Nox4 protein levels were 2-fold higher in Nox4TG aorta compared to wild-type littermates (WT), with no changes in expression of other Nox isoforms. Nox4TG had enhanced relaxation to acetylcholine (ACh) than WT (−log EC50 7.76±0.07 vs. 7.20±0.05; n=12; p&lt;0.001) but similar relaxation to sodium nitroprusside. The ACh response in Nox4TG and WT was identical in the presence of catalase (1500U/ml) but remained greater in Nox4TG in the presence of inhibitors of nitric oxide synthesis (L-NMMA, 100 μ M; −log EC50 7.44±0.22 vs. 6.93±0.22; n=6; p&lt;0.01), soluble guanylate cyclase (ODQ, 5 μ M; −log EC50 6.92±0.21 vs. 6.78±0.30; n=6; p&lt;0.001) or protein kinase G (KT5823, 2 μ M; −log EC50 7.77±0.11 vs. 7.27±0.12; n=6; p&lt;0.001). However, the difference between Nox4TG and WT was abolished in the presence of high extracellular potassium (30mM). Nox4TG also had significantly lower BP than WT (mean BP 102.5±1.8 vs. 109.5±2.0mmHg; n=10; P&lt;0.05), which was abolished after chronic treatment with N-acetylcysteine or an SOD/catalase mimetic, EUK-8. Plasma nitrite/nitrate levels and aortic levels of phosphorylated VASP were identical and acute iv treatment with L-NMMA (10mg/kg) increased BP to a similar extent in Nox4TG and WT. The hypertensive response to chronic 14-day angiotensin II infusion (1.1mg/kg/d) was lower in Nox4TG compared to WT (mean BP 116.7±4.7 vs. 129.4±3.5mmHg; n=10; P&lt;0.05). Conclusions Nox4TG had significantly enhanced ACh-induced vasodilatation compared to WT as a result of H 2 O 2 -induced hyperpolarization. Nox4TG also had a lower BP, which was not attributable to altered nitric oxide bioactivity but was normalized by chronic antioxidant treatment. These results suggest that endothelial Nox4 has potentially beneficial effects on vascular tone and BP.

Abstract 5222: Genetic Deficiency of Arginase II Reduces Endothelial Dysfunction and Atherosclerosis in ApoE-Knockout Mice

Introduction: Arginase II is known to be one of the key mediator in vascular pathophysiology of cholesterol-medicated endothelial dysfunction by reciprocal regulation of NOS and NO production through substrate competition. We aimed to investigate whether the transgenic Arginase II inhibition could reduce the endothelial dysfunction and atherosclerosis in apolipoprotein E-knockout (ApoE-KO) mice. Methods and Results: ApoE-KO mice were bred on an arginase II KO (ArgII-KO) background. Mice that were homozygous deleted at both alleles were used in the study. Male 8 – 12 wk old Arg-KO mice, ApoE-KO mice, Arg/ApoE-KO (Dbl-KO) mice fed high cholesterol diet for 16 weeks, and age-matched wild type (WT) mice fed normal diet were enrolled. Arginase activity was significantly higher in the aorta of ApoE-KO mice than compared to WT mice (1612.2±445.9 pmol Urea/mg protein/min vs. 836.3±12.2 pmol Urea/mg protein/min, n=5, p &lt; 0.01), but not detected in Arg-KO and Dbl-KO mice. We measured NO an ROS in vascular strips pinned down endothelial side up using the fluorescent probes DAF-FM diacetate and DHE. Dbl-KO mice showed significantly higher NO production (0.0737±0.0056 intensity/sec vs. 0.0256±0.0029 intensity/sec, n=6, p &lt; 0.05) and lower ROS production than ApoE-KO mice (0.0676±0.0065 intensity/sec vs. 0.1147±0.01619 intensity/sec, n=6, p &lt; 0.05). We tested endothelial dependent vasorelaxation in aortic rings in organ chambers. The vasorelaxation response to acetylcholine (Ach) in Dbl-KO mice was greater than in ApoE-KO mice (log EC 50 , −5.68 vs. −5.13, p &lt;0.05), although it was less than in Arg-KO and WT mice. N G -nitro-L-arginine methyl ester (L-NAME) incubation didn’t only impair Ach response, but also abolished the inter-group difference. The aortic atheromatous plaque burden as measured by Sudan IV staining was significantly reduced in Dbl-KO group compared to ApoE-KO group (38.4±3.8% vs. 52.0±2.8%, n=6, p &lt;0.01), No significant plaque was detectable in Arg-KO and WT mice. Conclusion: The genetic inhibition of Arginase II enhanced NO production, reduced ROS produnction restored endothelial function and lowered atheromatous plaque burden. Therefore, Arginase II represents a crtical novel target for atherosclerosis therapy.

Involvement of IL-13 in Tobacco Smoke–Induced Changes in the Structure and Function of Rat Intrapulmonary Airways

Chronic obstructive pulmonary disease (COPD) involves disease of small airways with an increase in airway smooth muscle sensitivity to spasmogens and with structural changes described as airway remodeling. We investigated the effect of tobacco smoke (TS) exposure on the structure and function of small airways in rats and the role of IL-13 in this response. Precision-cut lung slices (230-280 microm) were prepared from male Sprague-Dawley rats after acute (3 d) or chronic (8 or 16 wk) daily exposure to TS or air. Carbachol (CCh) and 5-hydroxytryptamine (5HT) concentration responses were performed on airways (50-400 microm diameter). The effect of IL-13 in vitro on small airway sensitivity to CCh and 5HT was also determined. Acute exposure to TS did not affect the sensitivity of the intrapulmonary airways to either spasmogen. After 8 weeks of TS exposure, airway hyperresponsiveness (AHR) to CCh was evident (log EC(50) CCh: air = 0.22 microM; TS = -0.12 microM; P = 0.019); AHR to 5HT was also observed after the 16-week exposure to TS (air = -0.85 microM; TS = -1.06 microM; P = 0.038). Chronic TS exposure increased airway wall SMA content, which correlated with increased expression of IL-13 and transforming growth factor (TGF)-beta(1) in the lung tissues. In vitro incubation with IL-13, but not TGF-beta(1), induced changes in small airway sensitivity to CCh and 5HT. Chronic TS exposure induces increased responsiveness in intrapulmonary airways of rats that may be mediated in part by an increase in IL-13.

Primary rat hepatocytes in chemical testing and QSAR predictive applicability

Primary rat hepatocytes were used to test acute toxicities of 16 neutral aliphatic alcohols, ketones and esters. Their effects on cell viability and metabolic function (ureogenesis, i.e. biotransformation of ornithine to urea) were measured and expressed as EC50 values. Log EC50 values from both tests correlated with the log partition coefficients for the chemicals between n-octanol and water and log P ow-based QSAR models were derived. Log EC50 (viability) tightly correlates with log EC50 (ureogenesis): log EC50 (viability) = 0.91 log EC50 (ureogenesis) + 0.06. Each of these toxic indices can be substituted by the other one. The toxic indices for both cell viability and metabolic disorder can be estimated using log EC50 for movement inhibition in the oligochaete Tubifex tubifex and the respective QSAR equation. It eliminates a usage of rats. Their correlations were proved and justified.

Acute toxicity assessment of perfluorinated carboxylic acids towards the Baltic microalgae

The presence of high-energy carbon–fluorine bonds in perfluoro compounds lends them great stability and causes them to be environmentally persistent. Relatively little is known about the acute toxicity of perfluorinated carboxylic acids (PFCAs) to ecotoxicological markers such as aquatic plants and animals. This study tested the toxicity of these compounds to the green alga Chlorella vulgaris, the diatom Skeletonema marinoi and the blue-green alga Geitlerinema amphibium, which are species representative of the algal flora of the Baltic Sea. The EC 50 values obtained range from 0.28 mM to 12.84 mM. A distinct relationship between hydrophobicity and toxicity is demonstrated. For every extra perfluoromethylene group in the alkyl chain, the toxicity increases twofold. Log EC 50 values are very well correlated linearly with both the number of carbon atoms in the perfluoroalkyl chain and the partition coefficients. The results also indicate that there are clear differences between the responses of particular taxonomic groups of algae: blue-green algae and diatoms are far more sensitive to PFCAs than green algae, probably because of differences in cell wall structure.

Pharmacological characterization of the serotonin receptor mediating contraction in the rat jugular vein

The purpose of this study was to characterize pharmacologically the serotonin (5‐hydroxytryptamine; 5‐HT) receptors mediating contraction in the rat jugular vein (RJV). We hypothesized that 5‐HT2A receptor is not the only receptor mediating contraction in RJV. Endothelium‐intact rings of RJV were placed in a tissue bath for measurement of isometric contractile force induced by cumulative addition of agonists (1nM to 10 μM). The contractions induced by the 5‐HT1A/1B and 5‐HT1A receptor agonists, RU 24969 (pD2=‐log EC50 [M]=6.4±0.05) and 8‐OH‐DPAT (pD2=5.5±0.07), respectively, were not as robust as that induced by 5‐HT (pD2=6.8±0.17). The 5‐HT1B/1D/1F agonist sumatriptan failed to induce contraction. The 5‐HT2A receptor agonist DOI (pD2=7.78±0.12) was more potent, whereas the 5‐HT2B receptor agonist BW723C86 was less potent (pD2=6.17±0.12) when compared to the contraction induced by 5‐HT. The antagonists with high affinity for the 5‐HT2A and 5‐HT2B receptors, ketanserin (30nM) and LY266097 (10nM), respectively, caused a right‐shift of 5‐HT‐induced contraction. The 5‐HT1B/1D receptor antagonist GR127935 (10nM) had no effect on 5‐HT‐induced contraction. We conclude that 5‐HT‐induced contraction of RJV is mediated primarily by 5‐HT2A receptor. The involvement of 5‐HT2B and 5‐HT1A receptor subtypes can not be ruled out and deserve further investigation. By identifying receptor mechanisms, we can better understand how the newly discovered serotonergic system in veins may locally modify contraction.

Kinetic Glutathione Chemoassay To Quantify Thiol Reactivity of Organic Electrophiles—Application to α,β-Unsaturated Ketones, Acrylates, and Propiolates

Glutathione (GSH) is a soft nucleophile and, as such, can be used to sense the reactivity of electrophilic agents toward the thiol group and other electron-rich sites of molecular structures. A new kinetic GSH chemoassay is introduced that employs a photometric method to quantify GSH loss and enables an efficient determination of second-order rate constants, k(GSH), of the reaction between electrophilic substances and GSH. Comparison with an existing 2 h static assay shows that the new kinetic variant is superior with respect to the detectable range of electrophilic reactivity and to confounding factors such as additional GSH loss due to oxidation. Analysis of the chemoassay degradation kinetics provides insight into the characteristic reaction times and the contributions of GSH-electrophile Michael addition and GSH oxidation to the overall GSH loss. For 15 alpha,beta-unsaturated ketones, nine acrylates, and two propiolates acting as Michael acceptors, the measured k(GSH) values span ca. 5 orders of magnitude. Moreover, log k(GSH) correlates with the compounds' toxicity toward the ciliates Tetrahymena pyriformis in terms of 48 h log EC(50) (50% growth inhibition) values, yielding a squared correlation coefficient (r(2)) of 0.91 and a root-mean-square error of 0.30 log units. It shows that for these and related compounds, aquatic toxicity is driven by electrophilic reactivity. The findings demonstrate that the kinetic GSH chemoassay can be used as an efficient tool to analyze, interpret, and predict correspondingly reactive toxicity in the context of qualitative and quantitative structure-activity relationship studies and as a nonanimal tool of integrated testing strategies for REACH to screen compounds for excess toxicity.

Phosphodiesterases inhibition unmask a positive inotropic effect mediated by β 2-adrenoceptors in rat ventricular myocardium

The effects of salbutamol on contractility and cAMP levels were investigated in rat right ventricular myocardium. Salbutamol (1–300 μM), produced a concentration-dependent positive inotropic effect which was not affected by ICI 118551 (50 nM), a β 2-adrenoceptor antagonist but was abolished by CGP 20712A (1 μM) a β 1-adrenoceptor antagonist. However, in rats pretreated with pertussis toxin (30 μg/kg intraperitoneal injection) salbutamol increases contractility ( E max = 9.8 ± 1.8%, − log EC 50 = 6.25 ± 0.07, n = 5). The combination of salbutamol + CGP 20712A, also produces a concentration-dependent enhancement of contractility ( E max = 43.0 ± 7.5%, − log EC 50 = 6.3 ± 0.04, n = 6), in the presence of 30 μM of the non selective phosphodiesterase (PDE) inhibitor 3-isobutylmethylxantine (IBMX) which was prevented by ICI 118551 (50 nM). Also, salbutamol + CGP 20712A fail to increase cAMP tissue levels but enhance them in the presence of IBMX. This effect was also prevented by ICI 118551. These results indicate that PDEs blunt contractility and cAMP production mediated by β 2-adrenoceptors in rat ventricular myocardium. Gi protein, although less efficiently than PDEs, also limits inotropic effects of salbutamol mediated by β 2-adrenoceptors in this tissue.