Badania interakcji chloropiryfosu i cypermetryny

Praca wykonana w: Katedra i Zakład Toksykologii

Synthetic Pyrethroids Exposure and Embryological Outcomes: A Cohort Study in Women from Fertility Clinic

Pyrethroids exposure has been associated with adverse reproductive outcome. However, there is no study that explores the effect of environmental exposure and embryological outcomes. This question was addressed in a prospective cohort of couples undergoing fertility treatment. The study aims to assess the association between urinary metabolites of synthetic pyrethroids and embryological outcomes (MII oocyte count, top quality embryo, fertilization and implantation rate). We included 450 women aged 25–45 undergoing assisted reproductive technology (ART) cycle at Infertility Clinic in Poland. Urine samples were collected at the time of fertility procedure(s) to assess four urinary synthetic pyrethroids concentrations (3-phenoxybenzoic acid (3PBA), cis-3-(2,2-dichlorovinyl)-2,2-dimethylcyclopropane carboxylic acid (cis-DCCA), trans-3-(2,2-dichlorovinyl)-2,2-dimethylcyclopropane carboxylic acid (trans-DCCA), cis-2,2-dibromovinyl-2,2-dimethylocyclopropane-1-carboxylic acid (DBCA)) using validated gas chromatography ion-tap mass spectrometry and calculated for each cycle-specific metabolite. To evaluate the effect of environmental exposure to synthetic pyrethroids and embryological outcomes (methaphase II (MII) oocyte yield, top quality embryo, fertilization rate, implantation rate), multivariable generalized linear mixed analyses with random intercepts were prepared. Urinary 3-PBA concentrations decrease MII oocyte count (p = 0.007) in the fourth quartile (>75 percentile) compared to women in the first quartile (≤25 percentile). Additionally, when 3-PBA was treated as continuous variable, the negative association between exposure to pyrethroids and MII oocyte count was also observed (p = 0.012). Exposure to other pyrethroid metabolities (CDCCA, TDCCA, DBCA) was not related to any of the examined embryological outcomes. Exposure to synthetic pyrethroids may be associated with poorer embryological outcome among couples seeking fertility treatments. As this is the first study on this topic, the results need to be confirmed in further studies

New Insights into Butyrylcholinesterase Activity Assay: Serum Dilution Factor as a Crucial Parameter.

Butyrylcholinesterase (BChE) activity assay and inhibitor phenotyping can help to identify patients at risk of prolonged paralysis following the administration of neuromuscular blocking agents. The assay plays an important role in clinical chemistry as a good diagnostic marker for intoxication with pesticides and nerve agents. Furthermore, the assay is also commonly used for in vitro characterization of cholinesterases, their toxins and drugs. There is still lack of standardized procedure for measurement of BChE activity and many laboratories use different substrates at various concentrations. The purpose of this study was to validate the BChE activity assay to determine the best dilution of human serum and the most optimal concentration of substrates and inhibitors. Serum BChE activity was measured using modified Ellman's method applicable for a microplate reader. We present our experience and new insights into the protocol for high-throughput routine assays of human plasma cholinesterase activities adapted to a microplate reader. During our routine assays used for the determination of BChE activity, we have observed that serum dilution factor influences the results obtained. We show that a 400-fold dilution of serum and 5mM S-butyrylthiocholine iodide can be successfully used for the accurate measurement of BChE activity in human serum. We also discuss usage of various concentrations of dibucaine and fluoride in BChE phenotyping. This study indicates that some factors of such a multicomponent clinical material like serum can influence kinetic parameters of the BChE. The observed inhibitory effect is dependent on serum dilution factor used in the assay

Urinary Bisphenol A Concentrations and Parameters of Ovarian Reserve among Women from a Fertility Clinic

Background: Human exposure to environmentally widespread endocrine disruptors, especially bisphenol A (BPA), has been suggested to affect reproductive health. Animal studies indicate that BPA may play a role in the process of reproduction and impact on maturing oocytes, meiotic cell division or fertilization rate. Nevertheless, data regarding the effects of exposure to BPA on women’s ovarian function are still limited. Therefore, the aim of the current study is to assess the effects of environmental exposure to BPA on ovarian reserve. Methods: The study participants consisted of 511 women in reproductive age (25–39 years) who attended an infertility clinic for diagnosis, due to the couples’ infertility. BPA urinary concentrations were assessed by the validated gas chromatography ion-trap mass spectrometry method. The ovarian reserve was assessed using ovarian reserve parameters: Hormones concentrations: E2 (estradiol), FSH (follicle stimulating hormone), AMH (anti-Müllerian hormone), and AFC (antral follicle count). Results: In the present study, the negative association between BPA urinary concentrations and AMH (p = 0.02) and AFC (p = 0.03) levels was found. Exposure to BPA was not related to other examined parameters of ovarian reserve (FSH, E2). Conclusions: Our results suggest that BPA exposure may affect women ovarian reserve parameters and reduce ovarian reserve. As this is one of the first studies of its kind, the findings need confirmation in a further investigation

The effect of serum dilution on BChE activity.

<p>The activity was estimated based on the mean of hydrolysis rate (OD/min) determined from at least 5 spectrophotometer reads (every 1 minute) to ensure linearity and regularity of slopes. Assays were done in triplicate, error bars represent mean ± standard error (Student’s t-test, p<0.05). <b>A.</b> The effect of BTC concentration on the BChE activity in 100, 400, 4000 -fold diluted serum samples. Inset shows a Lineweaver–Burk replot of the results presented in Fig 2A at [BTCh] = 0.1, 0.5 and 1 mM. <b>B.</b> The effect of 5mM and 2.5 mM BTC concentration on the BChE activity in variously diluted serum samples. <b>C.</b> BChE activity in 100, 400, 2000 -fold diluted eleven randomly selected serum samples from our serum collection samples.</p

Optimization of the BChE assay linearity.

<p>Measurement of TNB ion concentration at 412 nm produced in Ellman's reaction in the presence of 5 mM BTC and 40 (5), 100 (2), 200 (1), 400 (0.5), 600 (0.3), 800 (0.25)-fold diluted serum. Volume (μl) of serum in the reaction mixture (200 μl total volume) is shown above in brackets. Assays were performed in triplicate, standard error for all points is less than 2.5% of the values.</p

Effects of Synergistic Massage and Physical Exercise on the Expression of Angiogenic Markers in Rat Tendons

Physical exercise and massage are regarded as key factors in regulating tendon structure. However, information on the mechanism through which massage influences the structure and biology of a tendon is scarce. In this study, we attempted to define the impact of these two activities on rat tendons by using morphological and molecular techniques, determining the expression of VEGF-A, FGF-2, and CD34 in the tendons of rats subjected to 10 weeks of physical exercise (running) with massage of varied duration. The group of rats that was trained and massaged during the entire study was characterized by the highest expression of these markers, compared to the rats subjected to massage before training and to the control group subjected to physical exercises only. The greatest significant differences, compared to the control, were noted in the expression of all the studied markers at mRNA level, and in the case of VEGF-A, at protein level, in the third and fifth weeks of the experiment. The results of this study could point to the synergistic impact of simultaneous massage and physical exercise on the expression of angiogenesis markers in rat tendons

Characteristics of dibucaine inhibition of “usual” and “atypical” forms of BChE.

<p>DN values were calculated in assays performed at various dibucaine concentration at 25°C using 5mM BTC (A) or PTC (B) as substrate with two types of 400-fold diluted serum samples from heterozygous individuals with usual UU and UA phenotype. Dibucaine Number (DN) = (1 - (butyrylcholinesterase activity in presence of inhibitor/butyrylcholinesterase activity in absence of inhibitor)) x 100. Assays were performed in triplicate, standard error for all points is less than 3% of the values.</p