Boron enhances early embryonic gene expressions and improves fetal development of rats.

Boron is present as several different components in nature. Besides its industrial use, it is an essential element and is playing a very important role in the metabolism. In this study, it was aimed to determine the in vivo effects of boron on mRNA expression of HEX, NANOG, and OCT-3/4 genes in embryo and histological changes during fetal development. Therefore, totally 60 female rats were allocated into 5 equal groups. Experimental groups are as the followings; positive control (fed with standart rat diet), negative control (fed with boron free diet), low boron group (fed with boron free diet and given 0.04 µg boron/ml via gastric gavage), marginal boron group (fed with boron free diet and given 0.3 µg boron/ml via gastric gavage) and normal boron group (fed with boron free diet and given 2 µg boron/ml via gastric gavage). Experimental period was performed for 14 days. Embryos were collected after 4 days of mating and the expression and protein levels of early embryonic genes namely HEX, NANOG, and OCT-3/4 were determined by using Real-Time PCR. Also, 10-20 day embryo and fetus development were histologically determined. According to the results, mRNA expression and protein levels of early embryonic genes were increased in boron groups while decreased in boron deficient group. Histopathologically, tissue and organ developments were definitely observed in the boron groups. In conclusion, mRNA expression levels of early embryonic genes decreased in boron deficient group and boron has an important role for fetal development.

Nonylphenol in Human Breast Milk in Relation to Sociodemographic Variables, Diet, Obstetrics Histories and Lifestyle Habits in a Turkish Population.

BACKGROUND: Human breast milk is the most important food for infants and one of the main roads to exposure to toxic substances. In this study, nonylphenol (NP) levels in human milk samples collected from Turkish mothers (n=100) were determined, and the factors including mothers' demographics, eating habits, obstetric histories, and usage of cleaning and cosmetic products were examined. METHODS: Participants were mothers of randomly selected infants and toddlers from the Primary Health Care Center Number-8 in Afyonkarahisar City in western Turkey. The concentrations of NP in milk samples were measured using high-performance liquid chromatography (HPLC). RESULTS: All the analyzed samples showed the occurrence of NP at levels up to 47 ng/ml. The mean± SE and the median NP concentrations were 10.1±0.98 ng/ml and 8.46 ng/ml or ppb, respectively. A negative correlation with infant age was observed. There was a significant correlation between fresh fish consumption and the level of NP in the breast milk of mothers. No significant association between body mass index (BMI) and the NP level in human milk of mothers was observed. The mothers who were using excessive cleaning products in comparison to those using less had significantly higher NP in their breast milk. CONCLUSION: This study provides the first report about NP levels in a population and characterizes individual variation, thus giving a measure of exposed infants through breastfeeding in Turkey.

Molecular characterization of Fasciola samples, collected from different host species coming from the endemic area of Turkey.

Economical animal breeding programs are important for achieving maximum gain, and any factors resulting in economical loss should be minimized or eliminated. An organism of concern is the liver fluke, Fasciola hepatica, which causes decreased yield and even death in sheep and dairy cattle. In an effort to eliminate or minimize the detrimental effect of this parasite in animals, it is important to understand the genetic diversity within the liver flukes and the relationship between this parasite and the host in the particular geographic area. The aim of this study was to explore genetic diversity by analyzing the mitochondrial ND1 and cyt b genes and Ribosomal ITS1-2 regions. With these analyses, the individual differences, the host animal differences and combined effects of these factors on genetic relationships have been determined.

Electrochemical monitoring of the interaction between 4-nonylphenol and DNA by graphite and carbon nanotube modified graphite electrodes.

The interaction of 4-nonylphenol (NP) with deoxyribonucleic acid (DNA) was explored electrochemically by using differential pulse voltammetry (DPV) in combination with unmodified and modified pencil graphite electrodes (PGE) with single walled carbon nanotubes (SWCNT). The differentiation of the two oxidation signals coming from NP and DNA base, guanine was studied before and after the interaction process. In addition, the effect of NP concentration was investigated in order to determine the optimum experimental conditions. The detection limit and the reproducibility were determined by using CNT-modified electrodes.

Effect of chilling on the motility and acrosomal integrity of rat sperm in the presence of various extenders.

Studies were conducted to determine the effect of chilling on rat sperm and optimal components (extenders) to avoid chilling-induced injury. In the first experiment, the effects of chilling (at 4, 10, or 22 degrees C) on the motility and acrosomal integrity of epididymal sperm from 2 strains of rats (Sprague-Dawley and Fischer 344, F344) were compared. In the second experiment, the motility of epididymal Sprague-Dawley rat sperm after exposure to extenders (HEPES-buffered Tyrode lactate, skim milk, lactose monohydrate, Tris-citrate, and TEST) and cooling and warming was determined. We tested the effects of supplementing base extender solutions with 20% lactose-egg yolk (LEY) alone or in combination with a commercial SDS-based paste (0.5%, v/v) in preventing chilling injury. The motility after each treatment was determined after both cooling and warming. In the third experiment, the motility of Sprague-Dawley rat sperm were compared after supplementing the base extenders with either 0.4 M permeating cryoprotective agent (CPA; glycerol, ethylene glycol, propylene glycol, or DMSO) or 0.1 M nonpermeating CPA (raffinose and sucrose) after cooling and warming. The results showed that chilling significantly reduced the motility-but not acrosomal integrity-of Sprague-Dawley and F344 sperm. Neither motility nor acrosomal integrity differed between Sprague-Dawley and F344 strains. The addition of LEY into each extender significantly prevented motility loss after chilling. These results will be useful during the preparation of optimal extenders and development of successful cryopreservation protocol for rat sperm.

Effects of nonylphenol on motility and subcellular elements of epididymal rat sperm.

Nonylphenol (NP) is an important environmental toxicant and potential endocrine disrupting chemical. The objective of these studies was to determine the effects of NP on epididymal rat sperm in vitro. Epididymal sperm samples from Sprague-Dawley rats were incubated in 1, 10, 100, 250, and 500 microg/ml NP for 1, 2, 3, or 4h. Computer-assisted sperm analysis was used to determine motility. Epifluorescent microscopy was used to determine acrosomal status and flow cytometry was used to determine mitochondrial membrane potential (MMP) and chromatin integrity. Exposure of epididymal rat sperm to 250 or 500 microg/ml NP was highly detrimental to motility (P<0.05), with complete loss of motility observed after exposure to 500 microg/ml NP (P<0.05). The acrosomal integrity of sperm was significantly reduced with the lowest concentration (1 microg/ml) of NP, and higher concentrations resulted in a dose-dependent induction of the acrosomal reaction (P<0.05). Similarly, the percentage of sperm with high MMP declined dramatically after exposure to 100, 250, and 500 microg/ml NP (P<0.05). Duration of NP exposure did not have any effect on motility or MMP and NP did not appear to have detrimental effects on chromatin integrity (P>0.05). These results indicate that major mechanism of action of NP on rat sperm is by adversely affecting their acrosomal integrity. However, NP-induced impaired sperm motility, decreased mitochondrial membrane potential also likely to play an important role in destruction of sperm function.

Motility and acrosomal integrity comparisons between electro-ejaculated and epididymal ram sperm after exposure to a range of anisosmotic solutions, cryoprotective agents and low temperatures.

Effective ram sperm cryopreservation protocols, which would yield acceptable lambing rates following artificial insemination (AI), are currently lacking. The objectives of the current studies were to compare the effects of various anisosmotic conditions, cryoprotective agents (CPAs) and chilling on the motility and acrosomal integrity of electro-ejaculated and epididymal ram sperm. Three experiments were conducted. In experiment 1, ejaculated and epididymal ram sperm were exposed to 75, 150, 225, 600, 900 and 1200 milliosmolal (mOsm)/kg sucrose solutions, held for 5 min and then returned to isosmotic condition. Motility characteristics of sperm during exposure to each anisosmotic solutions and after returning to isosmotic conditions were determined. In experiment 2, ejaculated and epididymal ram sperm were exposed to 1M glycerol (Gly), dimethyl sulfoxide (DMSO), ethylene glycol (EG) and propylene glycol (PG) for 5 min and then returned to isosmotic conditions. Motility characteristics of sperm samples during exposure to each CPA solution and after returning to isosmotic conditions were determined. In experiment 3, effects of various temperatures on motility characteristics of ejaculated and epididymal ram sperm were determined after exposing them to three different sub-physiologic temperatures (4, 10 and 22 degrees C) for 30 min and subsequently returning them to 37 degrees C. The motility of ejaculated ram sperm was significantly more affected from anisosmotic stress than was epididymal ram sperm (P<0.05). While anisosmotic stress had no effects on acrosomal integrity of epididymal ram sperm, there was a significant reduction in acrosomal integrity for ejaculated ram sperm after the addition and removal of a 75 mOsm sucrose solution. The abrupt addition and removal of 1M Gly, DMSO, EG or PG had no effect on the motility and acrosomal integrity of epididymal ram sperm (P>0.05). However, there was a slight decrease in acrosomal integrity for ejaculated ram sperm after exposure to 1M Gly, DMSO or EG (P>0.05). Both epididymal and ejaculated ram sperm exhibited temperature-dependent loss of motility and acrosomal integrity (P<0.05). However, ejaculated ram sperm was more sensitive to chilling stress than epididymal sperm (P<0.05). In conclusion, the current data suggest that while epididymal ram sperm is extremely resilient to various cryobiologically relevant stress conditions, ejaculated ram sperm demonstrate greater sensitivity to such stressors. These findings should be taken into account when developing cryopreservation protocols for ejaculated and epididymal ram sperm.

Various physical stress factors on rat sperm motility, integrity of acrosome, and plasma membrane.

The objective of this study was to determine the effects of various physical interventions such as centrifugation regimes, Percoll gradient separation, and repeated pipetting on various viability parameters of epididymal sperm of Fischer 344 (F-344) and Sprague-Dawley (SD) rat strains. Three experiments were conducted. In experiment 1, sperm motility and acrosomal and membrane integrity were compared after exposing sperm samples to 200, 400, 600, and 800 x g centrifugal forces for 5, 10, or 15 minutes. In experiment 2, sperm motility and acrosomal and membrane integrity were compared after passing them through a Percoll separation using centrifugal forces of 600, 800, 1000, and 1200 x g for either 15 or 30 minutes. In experiment 3, the effect of repeated pipetting (2, 4, 6, 8, and 10 times) on motility and membrane integrity of rat sperm was compared with that on mouse, ram, bull, and boar sperm. The results revealed that both F-344 and SD rat sperm motility and membrane integrity were significantly affected by centrifugation (P < .05). The acrosomal integrity of SD rat sperm was affected after using 800 x g centrifugation force for 10 or 15 minutes (P < .05), whereas F-344 rat sperm acrosomal integrity was not affected by any centrifugation regimes (P > .05). Sperm from SD rats also had higher motility and membrane integrity loss than did sperm from F-344 rats after centrifugation and pipetting (P < .05). Percoll gradient separation did not cause significant motility loss or acrosomal damage to either F-344 or SD sperm (P > .05). Repeated pipetting had a dramatic adverse effect on both rat and mouse sperm motility (P < .05) as compared with sperm from bull, boar, and ram, which were not affected at all (P > .05). These data suggest that rat sperm have unique properties that need to be considered during centrifugation, Percoll gradient separation, and pipetting procedures.

Histological evaluation of gonadal differentiation in fathead minnows (Pimephales promelas).

The timing of sex determination and the pattern of sex differentiation have not been studied in fathead minnow even though this species of fish are commonly used as a research model for toxicological studies. In this study, the developmental histology of gonadal development was investigated. Fish were cultured in the laboratory conditions and spawning obtained at a photoperiod of 16 h-light and 8 h-dark. Samples were collected from day 7 fish post-spawning (day 7 fps) to day 150 fps and their gonads were processed for histological examination. Developmental histology was assessed by using a light microscopy. The results showed that ovarian differentiation normally occurs at around day 13 fps, while testicular differentiation normally occurs at around day 22 fps.

The effects of vitamin A, pentoxyfylline and methylprednisolone on experimentally induced amyloid arthropathy in brown layer chicks.

The effects of vitamin A, pentoxyfylline and methylprednisolone on experimentally induced amyloid arthropathy were investigated. In this study, 175 1-day-old brown layer chicks were used. Throughout the study Group II (vitamin A) received high doses of vitamin A (75,000 IU/kg), whereas Group I (negative control), Group III (positive control), Group IV (pentoxyfylline) and Group V (methylprednisolone) received normal levels of vitamin A in the diet. At the fifth week, the experimental Groups II, III, IV and V were injected with Freund's adjuvant intra-articularly to induce amyloid arthropathy. Group IV received pentoxyfylline and Group V received methylprednisolone (10 mg/kg, intramuscularly) once. Joint and blood samples were examined 13 weeks after the injections. The values in Groups I, II, III, IV and V, respectively, were as follows: amyloid arthropathy formation (%), 0, 100, 87, 76, 66; serum amyloid A (ng/ml), 166+/-17, 607+/-40, 423+/-39, 342+/-27, 293+/-22; serum retinol (microg/dl): 59.75+/-3.8, 42.72+/-3, 59.24+/-3.6, 102+/-9.1, 101.3+/-12.3; heterophil/lymphocyte ratio: 0.504, 0.75, 0.75, 0.087, 0.44. In conclusion, it was observed that vitamin A enhanced the development of amyloid arthropathy and there were positive associations between amyloidosis, increased levels of serum amyloid A and increased numbers of tissue infiltrating macrophages. Methylprednisolone had a more successful inhibitory effect on amyloid arthropathy than pentoxyfylline.

FT-IR spectroscopic analysis of rainbow trout liver exposed to nonylphenol.

Nonylphenol (NP) is a biodegradation product of nonylphenol ethoxylates (NPEs) belonging to the alkylphenol ethoxylates (APEs) group. APEs are widely used nonionic surfactants in detergents, herbicides, pesticides, paints, and cosmetics. The present work investigates the effects of NP on rainbow trout (Oncorhynchus mykiss) livers at the molecular level using Fourier transform infrared (FT-IR) spectroscopy. The FT-IR spectra revealed dramatic differences between the NP-treated and control tissues, which mainly indicated that the level of triglycerides increased, the lipid order increased, and the protein concentration decreased in the treated samples. Moreover, it was also found that glycogen levels significantly decreased and the relative content of nucleic acids increased in NP-treated fish. The 17beta-Estradiol-treated fish liver spectra were found to be quite similar to those of NP-treated fish. All these results implied that rainbow trout may offer considerable promise to be used as a bioindicator for NP in the future.

The bioaccumulation of nonyphenol and its adverse effect on the liver of rainbow trout (Onchorynchus mykiss).

Alkylphenol polyethoxylates (APEs) are widely used as nonionic surfactants. Nonylphenol (NP), one of the derivatives of APEs, has been found in the aquatic environment in ranges from nanograms per liter to milligrams per liter. In this study, juvenile rainbow trout were exposed to 0 (control), 66, 220, or 660 micro g NP/L for up to 28 days. Fish remained healthy under NP exposures of 0, 66, and 220 micro g/L for the length of the experiment. All fish died after 4 days of exposure to 660 micro g NP/L. Time-dependent NP bioaccumulation was detected in the tissues of fish exposed to 220 micro g NP/L (P<0.05) and histopathological changes were observed in the livers of fish exposed to 220 micro g NP/L. Furthermore, an increase in the activity of glutathione-S-transferase (GST) was found in the liver of fish exposed to 220 micro g NP/L for 1 week (P<0.05). There was an increase in GST activity in the liver of fish exposed to 66 micro g NP/L but it did not occur before 2 weeks of exposure to NP. The GST activity then decreased in a time-dependent manner in treatment groups, and this decrease was lower in the livers of fish treated with 66 and 220 micro g NP/L than in control fish after 3 weeks of exposure (P<0.05). These results indicated that sublethal doses of NP were accumulating in the bodies of the fish and causing histopathological and biochemical changes in the livers of rainbow trout.

Developmental genetics and physiology of sex differentiation in vertabrates.

The involvement of the Y chromosome in sex determination was determined by the development and the application of techniques for karyotyping the mammalian chromosome in 1960s. There were many reports on the particular region of the Y chromosome, such as histocompatibility (H-Y) antigen, bandit krait minor satellite (Bkm) the zinc finger Y gene (ZFY) and the sex-determining region of the Y chromosome (SRY) which were believed to be the testis determining factors. However, converging experimental evidence have indicated that the sex determining region of the Y chromosome (sry) is the testis determining factor (TDF) in mammalian species since sex is determined genetically at the time of fertilization in these species. In non-mammalian vertebrates especially in fishes, amphibians and reptiles, genotypic sex can be overridden by the external application of steroid hormones and temperature. In this review paper, after reviewing the complex literature on the molecular and biochemical mechanisms of sex determination and differentiation in all vertebrates, the potential danger of environmentally induced sex determination will be focused on.

Alkylphenol concentrations in two rivers of Turkey.

Alkylphenol polyethoxylates (APEs) called environmental endocrine disruptors has been shown to accumulate in water around the world. In this study, the pollution level of alkylphenolic compounds was measured and quantified in water, sediment, and the tissues of fishes collected in two rivers, Sakarya and Degirmendere Rivers, Turkey. Butylphenol (BP) were detected in sediment samples at one sampling stations of both rivers with 1.68 and 3.15 µg/g sediment, while nonylphenol (NP) were detected with the amount of 4.46 µg/g sediment in one sampling station in Degirmendere river. Fish samples also showed the presence of alkylphenolic compounds in both rivers. The level of alkylphenol pollution in two rivers of Turkey was determined to be in the range of alkylphenol level reported in Europe but lower than that of in the USA.