Stereological study on the effect of vitamin C in preventing the adverse effects of bisphenol A on rat ovary

Background: Bisphenol A [BPA], an environmental pollutant, can generate free radicals which damages the reproductive system. Vitamin C is an antioxidant which may prevent the adverse effects of free radicals

Objective: The aim was to investigate the effect of vitamin C on the ovary tissue in rats treated with BPA

Materials and Methods: In this experimental study, 24 female Wistar rats [200+/-20 gr] were randomly divided into 4 groups [n=6]: control, BPA [60 microg/Kg/day], vitamin C [150 mg/Kg/day] and BPA + vitamin C and orally treated for 20 days. The left ovaries were taken out, fixed for tissue processing and studied using stereological methods. Data were analyzed with SPSS using one-way ANOVA, and the means were considered significantly different at [p<0.05]

Results: The total volume of ovary and cortex [p<0.01], medulla [p<0.05], the volume of corpus luteum [p<0.001] and the mean number of antral follicles [p<0.001] significantly reduced in BPA group compared with control, while the number of atretic follicles increased [p<0.05]. The volume of oocyte [p<0.01] and its nucleus [p<0.001] in the antral follicles and the thickness of zona pellucida [ZP] in the secondary [p<0.05] and antral [p<0.001] follicles significantly decreased in BPA group compared with controls. The above parameters in the BPA + vitamin C group were compensated to control level

Conclusion: Vitamin C can be used as a potential antioxidant in the case of BPA toxication

[Investigation of the protective effect of epigallocatechingallat on the morphology and viability of the rat bone marrow mesenchymal stem cells following treatment with bisphenol A]

Environmental contaminants such as bisphenol A, in addition to the environmental problems can affect human health. Epigallocatechingallat has been found to have antioxidant and cytoprotective properties in cultured cells but its effect on toxicity induced by bisphenol A has not yet been determined in mesenchymal stem cells. The aim of this investigation was to study the protective role of epigallocatechingallat in rat bone marrow mesenchymal stem cells following treatment with bisphenol A[an oxidative stress inducer]. Material and In this experimental study, rat bone marrow mesenchymal stem cells were extracted using flashing-out method and cultured in DMEM containing 15% FBS and 100U/ml Pen/Strep. At the end of the third passage, cells were divided into 4 groups: control, bisphenol A, bisphenol A + epigallocatechingallat and epigallocatechingallat. The groups were treated for 12, 24, 36, and 48 hrs. After treatment with bisphenol A and epigallocatechingallat, viability, morphology, rate of DNA damage and calcium content of the cells were evaluated. Data were analyzed by one way ANOVA. p<0.05 was considered significant. Bisphenol A caused a significant reduction in the viability and calcium content of the cells. In addition, morphological changes such as nuclear breakage and chromatin condensation, as well as cytoplasm shrinkage, were observed in the group treated with bisphenol A. We found alterations in these parameters in the group of bisphenol A+ epigallocatechingallat which were similar to those observed in the control ones. Epigallocatechingallat can produce a protective role against the toxic effects of bisphenol A in the mesenchymal stem cells

Caspase-mediated apoptosis in sensory neurons of cultured dorsal root ganglia in adult mouse

Sensory neurons in dorsal root ganglia [DRG] undergo apoptosis after peripheral nerve injury. The aim of this study was to investigate sensory neuron death and the mechanism involved in the death of these neurons in cultured DRG. In this experimental study, L5 DRG from adult mouse were dissected and incubated in culture medium for 24, 48, 72 and 96 hours. Freshly dissected and cultured DRG were then fixed and sectioned using a cryostat. Morphological and biochemical features of apoptosis were investigated using fluorescent staining [Propidium iodide and Hoechst 33342] and the terminal Deoxynucleotide transferase dUTP nick end labeling [TUNEL] method respectively. To study the role of caspases, general caspase inhibitor [Z-VAD.fmk, 100 microM] and immunohistochemistry for activated caspase-3 were used. After 24, 48, 72 and 96 hours in culture, sensory neurons not only displayed morphological features of apoptosis but also they appeared TUNEL positive. The application of Z-VAD.fmk inhibited apoptosis in these neurons over the same time period. In addition, intense activated caspase-3 immunoreactivity was found both in the cytoplasm and the nuclei of these neurons after 24 and 48 hours. Results of the present study show caspase-dependent apoptosis in the sensory neurons of cultured DRG from adult mouse

protective role of vitamin E on the testicular tissue in rats exposed to sodium arsenite during the prenatal stage till sex maturity: a stereological analysis

Vitamin E is an effective antioxidant, protecting cells against oxidative stress. In this investigation the protective effect of vitamin E on the testis during development and spermatogenesis in rats exposed to sodium arsenite was evaluated. Pregnant Wistar rats were divided into 4 groups [n=8] control, sodium arsenite [8 mg/kg/day], sodium arsenite+vitamin E [100 mg/kg/day] and vitamin E. Treatment was carried out from day seven of pregnancy till 90 days. Finally the right testis was stereologically studied. The obtained data was analyzed using one way ANOVA and Tukey's test and the means difference was considered significant at p<0.05. The weight and volume of testis, volume of seminiferous tubules and its diameter, volume of interstitial tissue, height of germinal epithelium and the total number of types A and B spermatogonia, spermatocyte, spermatid and sertoli cells reduced significantly in sodium arsenite group compared to the control. Co-administration of vitamin E and sodium arsenite compensated the adverse effects of sodium arsenite on the above parameters. We concluded co-treatment of rats with sodium arsenite and vitamin E could prevent the adverse effects of sodium arsenite exposure on the testicular tissue during the prenatal stage till sex maturity

Effects of L-carnitine and L-acetyl-carnitine on testicular sperm motility and chromatin quality

Sperm cells extracted from testes [TESE] have poor chromatin quality and motility. Various substances are used in the laboratory to increase sperm motility and improve the ART outcomes; however, there are few research which considered improving both sperm motility and chromatin quality. The aim of this investigation was to evaluate the improvement of the testicular sperm motility and chromatin quality exposed to L-carnitine [LC] and L-acetyl-carnitine [LAC], which are normally concentrated in testis and epididymis, compared with Pentoxifylline [PF], which used for sperm motility enhancement in IVF procedures. TESE samples from 30 male mice divided into four parts. The sperm samples were added to Ham' F10 [control] or the media contained 1.76mM of LC, LAC or PF], then, the samples were kept in the room temperature for 30, 90 and 180 min. At each time step, sperm motility and chromatin quality were assessed. Chromatin quality was evaluated by chromomycin A3 and aniline blue. Statistical analysis was performed using one way analysis of variance [ANOVA]. A p-value less than 0.05 were accepted as a statistically significant difference. The results showed LC, LAC and PF significantly increased the sperm motility. However, sperm chromatin quality only improved significantly by administration of LC and LAC. Administration of LC and LAC to the testicular sperm samples can lead to improve both sperm motility and chromatin quality. It may be because they can mimic in vivo sperm condition during late spermatogenesis

[Effect of the polyamine component, TDMTN, on apoptosis of motor neurons in adult mouse spinal cord slices]

The aim of this study was to investigate the preventive effect of the polyamine component N, N, N, N-tetrakis[2-aminoethyl]2,2-dimethylpropane-1,3-diamine [tdmtn] on apoptotic motor neurons in adult mouse spinal cord slices. Thoracic region of adult mouse spinal cords was sliced by a tissue chopper into 400 micro m slices and cultured in medium in the presence or absence of tdmtn for 6 hours. Morphological a features of apoptosis were evaluated using fluorescent staining with propidium iodide and Hoechst 33342. The appearance of nucleosomal DNA fragmentation was studied using agarose gel electrophoresis. Our results were analyzed using the one-way ANOVA and Tukey's tests. After 6 hours in culture, motor neurons displayed morphological sings of apoptosis including cell shrinkage, as well as nuclear and chromatin condensation. DNA extracted from slices cultured for 24 hours revealed nucleosomal DNA fragmentation on agarose gel electrophoresis. Tdmtn reduced the occurrence of apoptosis in motor neurons and significantly [p<0.01] increased the viability of these neurons after 6 hours of culturing. Tdmtn, as a polyamine component, could probably prevent the occurrence of motor neuron apoptosis through calcium chelation

Effects of vitamin E on ovarian tissue of rats following treatment with p-nonylphenol: a stereological study

Para-Nonylphenol [p-NP] is one of the environmental pollutants which cause reproductive system disorders. The effects of vitamin E on ovary structure during its development in rats treated with p-NP. 32 Wistar female rats after mating were divided into 4 groups; control, vitamin E [100mg/kg/day], p-NP [250mg/kg/day] and p-NP + vitamin E. The rats were treated from the day 7 of pregnancy till 21st day of postnatal through sucking period. After weaning, the female pups were treated by gavages for 120 days. The total volume of ovary, number of follicles, volume of oocyte, follicular cells and their nuclei and the thickness of zona pellucida were estimated stereologically. The results were analyzed using one way ANOVA and p<0.05 was considered significant. The ovary weight, mean total volume of ovary and cortex, number of antral and graafian follicles and body weight were decreased significantly [p<0.05] in the p-NP treated rats compared to control and other groups, while the number of atretic follicles was increased significantly [p<0.05]. A significant reduction [p<0.05] in volume of oocyte, follicular cells and their nuclei in antral and graafian follicles was found in p-NP group. In addition, treatment with only vitamin E showed an improving effect on folliculogenesis due to a highly significant increase [p<0.01] in the number of primordial follicles. Vitamin E could compensate the adverse effects of p-NP on the ovary structure during its development

Bio treatment protects rat marrow-derived mesenchymal stem cell culture against the TNF-a induced apoptosis

This study is an attempt to examine the anti apoptotic effects of BIO on rat MSC culture. Rat marrow primary cell culture was established and exposure groups were defined; cultures with 0.01, 0.1, 1 micro M BIO. Cells cultured without BIO treatment were used as controls. During culture expantion, the average doubling time, as an index of the rate of cell growth, were determined and compared. To examine whether or not BIO is able to protect MSCs against apoptosis, the passaged-3 cells from each group were induced to undergo apoptosis with the addition of TNF-alpha [Tumor necrotic factor-alpha]. Three days after, the cultures were quantified in terms of the percentages of apoptotic cells using either the Tunnel or Annexin V staining method. Marrow cells cultivated with 0.1 and 1 micro M BIO appeared to expand at a significantly more rapid rate than the 0.01 micro M BIO and the control cultures [p < 0.05]. Tunnel staining indicated that in 1 micro M BIO-treated groups, there were lower percentages of apoptotic nuclei than in groups with other concentrations of BIO [p < 0.05]. The BIO protective effect appeared to be dose-dependent in that the cultures with high BIO content possessed less apoptotic nuclei. The results obtained by Annexin staining were in agreement with the results of Tunnel staining. The Annexin method additionally takes into account the early apoptotic cells which are not detectable by the Tunnel method. Taken together, it seems that cultivation with BIO could both increase the growth rate of marrow cells and protect MSCs against induced apoptosis

Effect of staurosporine on neural differentiation of CD133[+] umbilical cord blood cells

CD133[+] umbilical cord blood cells were identified as a hematopoietic stem cell which has the capacity for extensive self-renewal and differentiation. The aim of this study was to identify the effect of staurosporine [STS], a well-known protein kinase inhibitor on differentiation of CD133[+] cells into neural cells. CD133[+] cells were enriched by immunomagnetic beads from human mononuclear cells of umbilical cord blood and the purity of higher than 94% was achieved by flowcytometry. Induction of differentiation was performed by addition of STS [12.5, 25, and 50 nM]. The differentiated cells were evaluated by immunofluorescence and RT-PCR for neuron-specific proteins and transcripts. STS-treated CD133[+] cells expressed mRNA transcripts for neuron-specific neurofilament protein [NFM], and several basic helix-loop-helix [bHLH] transcription factors important for early neurogenesis, including Otx2, Wnt1, and Hash1. The structural proteins characteristics of neurons including beta-tubulinlll and Microtubule-Associated Protein-2 [MAP-2], were shown by immunocytochemistry. STS-treated CD133[+] cells also expressed the astrocyte-specific marker, glial fibrillary acidic protein [GFAP] by immunofluorescence. The human cord blood-derived CD133[+] hematopoietic stem cells could differentiate into neural cell types of neuron-like cells and astrocytes by STS treatment