In vitro effects of selected endocrine disruptors (DEHP, PCB118, BPA) on narrow-clawed crayfish (Astacus leptodactylus) primary cells.

Environmental pollutants with endocrine-disrupting effect are of global importance due to their contribution to the aethiologies of variety of complex diseases. These lipophilic pollutants are persistent in the environment and able to bioaccummulate in nontarget organisms. BPA, DEHP and PCB118 (dioxin-like PCB) are associated with endocrine disruption effects, while information on their effects on aquatic invertebrates are limited. In the current study, the effects of these compounds, which are ubiqutous and present at low concentrations in the environment, are studied in the primary hepatopancreas, muscle, gill, intestine and gonadal cultures of narrow-clawed crayfish (Astacus leptodactylus Eschscholtz, 1823), a widely distributed freshwater crayfish in Turkey with high economic importance. IC50 values following MTT assay ranged 0.27-12.61 nM; when compared with other tissues, the gonads were more affected with lower IC50 values. PCB118 induced higher cytotoxicity, while DEHP was the least toxic compound. This is the first study on the primary culture of A. leptodactylus¸ and the toxic effects of these compounds in this organism providing mechanistic insights on the responses and detoxification capacity of the organs. This study provides basis to unravel the mechanism of action of the tested EDCs in crayfish and improvement of cell culture conditions for ecotoxicity and screening assays.

The impact of occlusal plane cant along with gingival display on smile attractiveness.

OBJECTIVES: To evaluate the influence of occlusal plane cant in conjunction with maxillary gingival display on perception of smile attractiveness by orthodontists, dentists, and laypersons. SETTING AND SAMPLE POPULATION: Faculty of Dentistry at Baskent University. A total of 204 raters for smile attractiveness. MATERIALS AND METHODS: A frontal intra-oral photograph of aligned teeth was modified using image processing software. Six different occlusal lines representing 0° to 5° cants were obtained by tilting the photographs. Each occlusal cant was adjusted in five manners resulting in five different gingival display amounts. Attractiveness of the 30 different smiles was evaluated by 204 raters divided into three groups (n = 68 in each group). RESULTS: Both occlusal cant (p < 0.001) and gingival display amount (p < 0.001) had a statistically significant influence on smile attractiveness. Smile attractiveness scores with reference to amount of gingival display showed a significant difference between rater groups (p < 0.001). Orthodontists preferred 1-mm coverage of upper central incisors by the upper lip, whereas dentists and laypersons preferred 2 mm. Significant (p < 0.001) interaction was observed between occlusal cant and gingival display amount, which influenced smile attractiveness. CONCLUSION: Increase in both occlusal plane cant and gingival display negatively influences smile attractiveness. The influence of occlusal plane cant becomes less when gingival display increases, whereas the influence of gingival display decreases when occlusal cant increases. Dentists are more generous than orthodontists, while laypersons are the most generous regarding smile attractiveness scores.

Osteoblastoma originating from frontoethmoidal sinus causing personality disorders and superior gaze palsy.

Osteoblastoma is a rare, solitary benign tumor that is usually situated in axial skeleton mainly in vertebra. It is rarely seen in ethmoid and frontal sinuses. A 40-year-old man who had osteoblastoma originated from frontal and ethmoidal sinuses that extends up to frontal lobe and gave rise to personality disorders by compressing the frontal lobe, and caused superior gaze palsy by compressing the superior rectus muscle. We present this rare case with clinical, radiological and histopathological findings.

Quercetin both partially attenuates hydrogen peroxide-induced toxicity and decreases viability of rat glial cells.

Quercetin is one of the most ubiquitous flavonoids in foods of plant origin. Although quercetin is generally considered to provide protection against oxidative injury, recent studies have shown to be cytotoxic to many cell types. We intended here to determine whether quercetin protects against H2O2-induced toxicity and/or affects viability of rat mixed glial cells. The cells were obtained from 1-3 day olds rat brains and incubated in a humidified atmosphere of 5% CO2, at 37 °C in flasks. In the quercetin groups, different quercetin concentrations (1, 10, 50, 75 or 100 µM) were applied alone for 24 h. For H2O2 cytotoxicity group, the glial cells were treated for 3 h with 100 µM H2O2 which induced 75% cell death. In another group, the cells were treated with 100 µM H2O2 plus respective quercetin concentrations simultaneously for 3 h, the medium was removed and refed for 24 h. MTT test was then applied and statistical significance was ascertained by one way analysis of variance, followed by Tukey's multiple comparison test. Quercetin starting from 50 µM decreased the glia survival significantly. In H2O2 plus quercetin co-treated groups, both 75 and 100 µM quercetin attenuated toxic effect of H2O2 by 15%. In conclusion, quercetin both partially protects H2O2-induced gliotoxicity and decreases rat glial cell viability in primary culture.

The effect of adrenomedullin, amylin fragment 8-37 and calcitonin gene-related peptide on contractile force, heart rate and coronary perfusion pressure in isolated rat hearts.

The effect of human adrenomedullin, human amylin fragment 8-37 (amylin 8-37) and rat calcitonin gene-related peptide (CGRP) on contractile force, heart rate and coronary perfusion pressure has been investigated in the isolated perfused rat hearts. Adrenomedullin (2x10(-10), 2x10(-9) and 2x10(-8) M) produced a significant decrease in contractile force and perfusion pressure, but only the peptide caused a decline in heart rate at the highest dose. Amylin (10(-9), 10(-8) and 10(-7) M) significantly increased and then decreased contractile force. Two doses of amylin (10(-8) and 10(-7) M) induced a significant increase in heart rate, however amylin did not change perfusion pressure in all the doses used. Rat alpha CGRP (10(-8), 10(-7) and 10(-6) M) evoked a slight decline in contractile force following a significant increase in contractile force induced by the peptide. CGRP in all the doses raised heart rate and lowered perfusion pressure. Our results suggest that adrenomedullin has negative inotropic, negative chronotropic and coronary vasodilator actions. Amylin produces a biphasic inotropic effect and evokes a positive chronotropy. CGRP causes positive inotropic, positive chronotropic and vasodilatory effects in isolated rat hearts.

The mitogenic activity of endothelin-1 on megakaryocytopoiesis in vitro.

Endothelin (ET)-1 induces proliferation of various cells including smooth muscle cells, fibroblasts, glomerular mesangial cells, endothelial cells and osteoblasts. ET-1 also stimulates synthesis of interleukin (IL)-6 in endothelial and bone marrow stromal cells of rat. It is well known that IL-6 modulates megakaryocytopoiesis. Some studies have indicated that megakaryocytes express both ET receptors and they are targets for ET. Therefore we planned to examine the effects of ET-1 on the growth of normal megakaryocytic cells in rat bone marrow primary cell culture. Bone marrow cells were cultured at 37 degrees C, in an incubator atmosphere of 5% CO2 in air and 95% relative humidity for nine days. ET-1 at 10(-7), 10(-8 ) and 10(-11) M, and control with saline were added at the beginning of the experiment protocol. At each day, plasma clots were stained using direct-coloring thiocholin method for acetylcholinesterase activity. Although 10(-7) M ET-1 did not change the proliferation of megakaryocytic cells, this could be due to the presence of over crowded fibroblasts in the same environment. 10(-8) M ET-1 stimulated megakaryocytic cell growth to 234% over the control on the fifth day. ET-1 at a concentration of 10(-11) M also rised the megakaryocytic cell number significantly reaching up to 86% at the sixth day. Our results indicate that ET-1 may modulate the growth of megakaryocytic cells by an autocrine and/or paracrine action.

Antiproliferative properties of the lazaroids U-83836E and U-74389G on glioma cells in vitro.

The 21-aminosteroids (lazaroids) are a new family of steroid compounds that inhibit lipid peroxidation reactions. They are novel antioxidant agents, which have been shown to have antiproliferative properties on cancer cells and also are thought to prevent free radical-mediated blood-brain barrier damage. In order to understand the effect of lazaroids on glioma, we tested U-83836E and U-74389G at doses ranging between 0.1 100 m mM on primary cultures of glioblastoma multiforme from three patients, rat C6 glioma cell line, and 5 th subculture established from one of the patients. The effects of both compounds on cell proliferation were determined using 3-(4,5-dimethyl thiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) colorimetric assay. U-83836E in the primary cultures was found to have 50% inhibitory concentrations (IC50 ) of 6.30, 6.75 and 6.50 m mM, respectively. The IC50 value of U-74389G was calculated as 91 m mM in only one of the patients. On C6 glioma cells, while the IC50 of U-83836E was 45 m mM, U-74389G showed no cytotoxic effect. On the 5 th subculture, U-83836E had an IC50 of 37.5 m mM, but the cytotoxic effects of U-74389G was less than in that of the primary culture. In conclusion, these compounds were found to be more cytotoxic in primary culture than the cell lines and there were also differences between their members in the inhibition of cell survival.

The effects of anticancer drugs in combination with nimodipine and verapamil on cultured cells of glioblastoma multiforme.

The presence of the cellular multidrug resistance (MDR1) gene and its product, P-glycoprotein (Pgp), is thought to be a mechanism for the failure of chemotherapy in cancer patients. Calcium channel blockers have been shown to sensitise cancer cells to anticancer drugs by reversing Pgp expression in cell lines. The interactions between anticancer drugs such as carmustine (BCNU), vincristine (VCR) and procarbazine (PCB) and calcium channel blockers such as nimodipine and verapamil on cultured cells of glioblastoma from eight patients were therefore tested. Pgp expression was examined immunohistochemically using C219 monoclonal antibody in cytospin preparation. The cytotoxicity of the drugs was screened using microculture tetrazolium assay. The cells from five patients showed positive immunoreaction for Pgp. Nimodipine showed growth-inhibitory activity against glioblastoma cells at a rate of 16.55-26.88% (P < 0.05), but a similar effect was not observed with verapamil. While antiproliferative effects of BCNU were around 20.91-45.09% (P < 0.05) on the cells from seven patients, VCR was the most effective agent in inhibition of cell growth at a rate of 26.43-48.47% (P < 0.05). The response of the cells from five patients to PCB was from 11.98 to 16.32% (P < 0.05). When used together, nimodipine further enriched cytotoxicity of the anticancer drugs up to 11.14-40.85% (P < 0.05) without relation to Pgp expression. In conclusion, the enhancement of cytotoxicity of anticancer drugs by nimodipine suggests that there might be a synergy between anticancer drugs and nimodipine in the inhibition of glioma cell growth.

Luteinizing hormone releasing hormone increases proliferation of meningioma cells in vitro.

The fact that meningioma shows at least a 2:1 predilection for women over men is considered to be due to endocrinological and paracrine regulation of the development of this tumour. The presence of receptors for the luteinizing hormone releasing hormone (LHRH) in gynaecological cancer permits the use of LHRH agonistic or antagonistic analogues with a direct effect or by the gonado-pituitary axis suppression in the treatment of these tumours. Therefore, the effect of LHRH on meningioma cells is tested in this study. Meningioma cells from three female patients were cultured and LHRH (50 ng/ml) was added to the growth medium daily, for fourteen days. At the end of this period the cells were counted by means of a Coulter Counter. The stimulating effects of LHRH on the increase of the amount of cells in the meningioma monolayer culture were 146% (p < 0.01), 134% (p < 0.05) and 141% (p < 0.05) of the control, respectively, for the three patients.

Prostaglandin-independent decrease of sheep ureter contractility induced by bradykinin.

The mechanisms that mediate the actions of bradykinin on ureteral motility are poorly defined and mediation via prostaglandins has not been examined. Therefore, the effects of bradykinin on contractility and the possible mediator role of prostaglandins have been investigated in sheep ureter. At the concentrations of 10(-8), 10(-7) and 10(-6) M, bradykinin elicited marked reductions in contractile force. When ureteral strips were treated separately with 10(-6) M indomethacin, 2 x 10(-6) M sodium salicylate and 10(-5) M aspirin, each drug produced a significant decrease in contractile force. In strips in which prostaglandin synthesis was inhibited by the above concentrations of indomethacin, sodium salicylate and aspirin, 10(-7) M bradykinin significantly decreased the contractility. From these data, we concluded that in ureter bradykinin decreases contractility via a mechanism not involving prostaglandin generation.

Effects of rat platelet extracts and human platelet-derived growth factor on megakaryocytopoiesis in vitro.

An improved plasma clot culture system was developed for the culture of rat megakaryocyte colonies in vitro. Greater stimulation of megakaryocyte colony growth was provided when both serum and plasma were added to the culture medium than when plasma was added alone. Freeze-thawed extracts prepared from 900 x 10(6) platelets/ml saline and also 9 ng/ml of human platelet-derived growth factor (PDGF) caused maximum increases over the controls of 71% and 42%, respectively, in the numbers of megakaryocyte colonies. The increase in the numbers of megakaryocyte colonies induced by platelet extracts and by PDGF implied that substance(s) released by platelets might play an important role in the control of megakaryocytopoiesis.