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Background and Objective: Degeneration of neurons in the central nervous system occurs during aging. Transplantation of neural stem cells (NSCs) can be preventing the degeneration of neurons. In addition to neuronal replacement, with the production of neurotrophic factors, increased survival and proliferation of endogenous cells. This study was done to compare the cell proliferation, neurotrophic factors expression and features of NSCs harvested from different areas of the central nervous system in vitro. Materials and Methods: In this laboratory study NSCs have been harvested from subgranular zone (SGZ), subventricular zone (SVZ) and central canal of spinal cord from adult Wistar rats with mechanical, enzymatical digestion and subsequently was cultured in α-MEM medium supplemented with serum as monolayer or adherent conditions and passaged for 13 times. Immunocytochemistry was used to determine expression of the nestin and GFAP markers. Semi-quantitative RT–PCR was used to confirm genes expression (NGF, CNTF, NT3, NT4/5, GDNF and BDNF). Results: Morphological features of stem cells extracted from different regions of the central nervous system were similar in the culture. Doubling time NSCs in the SVZ (37.45 hr) is shorter than in the SGZ (44.04 hr) and central canal of spinal cord (57.22 hr). The culture conditions as well as monolayer neural stem cells are capable of producing neurospheres. Also, nestin and GFAP markers, expressed by NSCs. Neurotrophic gene expression pattern profiles were similar to each other in stem cells extracted from the SGZ, SVZ and central canal of spinal cord. Conclusion: Neurotrophic gene expression in stem cells extracted from different regions of the central nervous system were similar, but proliferation capacity was higher in NSCs, which have been harvested from the SVZ.

Background and Objective: Epidermis is the outer layer of skin, regenerating continuously. Epidermal stem cells play important roles in tissue regeneration, scar regeneration and neoplasm formation.This study was displayed for the isolation and culture of interfollicular epidermal stem cells from newborn mouse skin without feeder layer. Materials and Methods: This experimental study was displayed on 0-3 old-day newborn NMRI mouse skin 60-70 gr weight. The epidermal keratinocytes were separated mechanically and enzymatically from 0-3 old day newborn mice skin (NMRI strain) and seeded on fibronectin-collagen culture substrates. Putative epidermal stem cells were selected by rapid adherence for 10 minutes on this composite matrix of type 1 collagen and fibronectin and the unattached cells were discarded and attached cells were cultured in essential minimal eagle medium (EMEM) (ca+2-free culture medium containing 0.05 mM Ca+2, 9% FBS, 50% conditioned medium, EGF (epidermal growth factor) and Cholera Toxin. The immunocytochemistry of β1-integrin analysis used to indicate their stemness nature. Results: The results indicated that rapid adherence yields 50% purity. By using this method, the stem cells have been subcultured continuously without any change in the cell properties. The isolated interfollicular epidermal stem cells, expressed epidermal stem cells special marker (β1-integrin) in high levels, which indicates stem cell nature. Conclusion: This new method yields pure viable epidermal stem cells that can be used in regenerative medicine and cell therapy.

Background and Objective: Research have been focused on the applying the chemical inducer for trans-differentiation the adult BMSCs into neural cell. So that, at the first should investigate the toxcity effect of the chemical inducer on the induced cells. Plasticity and easy accessibility of bone marrow mesenchymal stem cells is a unique charactristic for treatment of neural disorderies. This study was desgined to determine the inductive effect of Deprenyl and Dimethyl sulfoxide on proliferation and survival of the mesenchymal stem cells. Materials and Methods: In this experimental study, BMSCs isolated from the adult rat bone marrow and cultured in αMEM containing 10% FBS. Cell identity for surface antigens was performed in third passage by immunocytochemistry and multipotancy capacity of BMSCs was done by BMSC differentiation into adipocytes and osteocytes. The cells were exposed to chemical agents (a: the αMEM medium supplemented with 2% DMSO, b: the αMEM medium supplemented with 10-8M Deprenyl) for 24 houres and then transferred to αMEM containing 10% FBS cell survival and proliferation was evaluated after the 24, 48, 72 and 96 houres by MTT [3-(4-5-Dimethylthiazolyl-2-y1)-2,5-diphenyltetrazolium bromid] test. Data were analyzed using SPSS-16, One-Way ANOVA and Tukey tests. Results: In addition to expression the surface antigens and adipogenic and osteogenic differentiation by BMSCs, MTT test results showed that proliferation and survival of induced-deprenyl and DMSO cells within 48, 72 and 96 hours after the induction was increased significantly than negative control group. Conclusion: Deprenyl increases survival and cell proliferation compared to Dimethyl Sulfoxide. It can be used as cell inducer.

Background and Objective: Neurotrophic factors are diffusible polypeptides that have critical roles in survival, proliferation and differentiation of stem cells. This study was done to assess the role of neurotrophic factors (CNTF‎, BDNF, ‎GDN‎F, ‎NT-‎3‎) expression and proliferation rate of neural stem cells (NSCs) in coculture with mesenchymal stem cells (MSCs). Materials and Methods: In this experimental study, NSCs and MSCs were isolated from adult Wistar rat. Initially, NSCs was harvested from temporal lobe after mechanical digestion by a sterile flamed Pasteur pipette and enzymatic digestion with trypsin and Dnase. The cell suspension was cultivated in a flask with DMEM/F12 medium supplemented with 10% FBS 100U/ml Penicillin and 100 mg/ml Streptomycin. To obtain MSCs, bone marrow of femur and tibia bones were flashed out and cultured. MSCs and NSCs‎ cocultured by transwell ‎system in DMEM/F12 medium supplemented with 10% FBS 100U/ml Penicillin and 100 mg/ml Streptomycin. Haemocytometer, immunocytochemistry and RT-PCR methods were performed to identify and evaluate cell proliferation, purity levels and neurotrophic factors expression. Results: There ‎is‎ no differences in NTFs profile of ‎neurotrophic‎ factors expression between ‎coculture ‎group‎ ‎and‎ control ‎NSCs, but interactions between MSCs and NSCs significantly promoted NSCs proliferation (P

Intrduction: Phoenix dactylifera pollen (DPP) is one of the traditional herbal medicines used in developing countries for male infertility treatment. The goal of this study was to assess in vitro effects of aqueous extract of DPP on the efficiency of in vitro colony formation in neonate mouse spermatogonial stem cells (SSCs). Materials and Methods: Testicular cell suspension of SSCs and sertoli cells were isolated from neonatal 6 day-old mice during 2 steps enzymatic digestion and cultured in a DMEM and FCS 4% in the absence or presence of different doses of aqueous extract of DPP (0.06, 0.25 and 0.62 mg/ml). The number of colonies was determined at the end of the day 9 of culture in order to evaluate the rate of SSCs expansion. Data were analyzed using ANOVA and Tukey posttest. Results: The results showed that the spermatogonial cells significantly increased in vitro spermatogonial cell cluster formation at the presence of 0.62 mg/ml of aqueous extract of DPP in comparison with the 0.06 mg/ml concentration group (P⩽ 0.05). Conclusion: Our results indicated that 0.62 mg/ml of aqueous extract of DPP can be used for enrichment of SSCs and treatment of male infertility caused by oligospermia. However, further studies are needed to determine its beneficial effects on man. [ABSTRACT FROM AUTHOR]

Oxidative stress hurts the survival of transplanted mesenchymal stem cells (MSCs). Lipopolysaccharide (LPS) preconditioning inhibits apoptotic death in MSCs. Also, Lovastatin's protective effect was reported on MSCs. Here, we investigated the potential of LPS and Lovastatin combination therapy on the survival and proliferation of MSCs. MSCs harvested from adult rats (240-260 g) femur and tibia bone marrow. Third passage MSCs were divided into 6 groups control group, LPS, LPS + Lovastatin (10 and 15 µM), and Lovastatin (10 and 15 µM). Cell survival and proliferation were assessed using an MTT assay 24 h after LPS, Lovastatin, or LPS + Lovastatin treatment. Also, Malondialdehyde (MDA) as a lipid peroxidation marker and antioxidant enzymes such as Glutathione peroxidase (GPX) and Superoxide dismutase (SOD) activity levels evaluated. Finally, the expression level of tumor protein P53 (P53) and octamer-binding transcription factor 4 (OCT4) genes were measured by qRT-PCR test. Lovastatin 10 μM potentiated proliferation and survival of MSCs. It can increase the activity of GPX and SOD. 10 µM Lovastatin could not affect MDA amounts but decreased the expression levels of P53 and Oct4 significantly. Nevertheless, treatment with LPS reduced the survival and proliferation of MSCs, along with a significant reduction in GPX activity. LPS + Lovastatin could increase SOD activity, however, GPX enzyme activity and MSCs proliferation did not change so, and it was not effective. We propose Lovastatin at the dose of 10 µM as a suitable combination agent to increase the survival and proliferation of MSCs in oxidative stress conditions., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Ethanol can have harmful effects on the development of the embryos. The aim of this study was to evaluate the effect of vitamin E, as an antioxidant, on changes in liver tissue damaged by ethanol in rats. Rats were divided into 11 groups, control, naive, sunflower oil (oil), ethanol, vitamin E (100, 200, and 400 mg/kg), ethanol + vitamin E (100, 200, and 400 mg/kg), and oily ethanol. In the experimental groups, rats received ethanol (v/v 40%) and vitamin E (100, 200, and 400 mg/kg) orally once a day from gestational day 0 to 28 days after delivery. Then, we evaluated the weight of rats and their offspring, the number of offspring, and the level of malondialdehyde (MDA), as an index of lipid peroxidation, superoxide dismutase (SOD), and glutathione peroxidase (GPX), as antioxidant enzymes, in the liver tissue of the offspring. Vitamin E significantly increases in weight of pregnant mothers and their offspring on the 21st day of pregnancy. The level of MDA in the groups receiving vitamin E was significantly reduced compared to the ethanol group. The activity of GPx and SOD antioxidants enzymes was significantly increased in the offspring. Vitamin E could reduce ethanol-induced liver damage in male offspring by reducing oxidative stress., (© 2024. Akadémiai Kiadó Zrt.)

Objective: Neural stem cells (NSCs) are suitable therapeutic candidates. Here, we compare the proliferation rate, differentiation potential, and expression levels of specific markers in two groups of cultured NSCs derived from rat subgranular (SGZ) and subventricular (SVZ) zones., Materials and Methods: In this experimental study, NSCs isolated from SGZ and SVZ were cultured in α-minimal essential medium (α-MEM) supplemented with 1% penicillin/streptomycin, 10% fetal bovine serum (FBS), 20 ng/ml basic fibroblast growth factor (bFGF), 20 ng/ml epidermal growth factor (EGF), and B27 supplement. Glial fibrillary acidic protein ( Gfap ), p75 neurotrophin receptor ( Ngfr ), tyrosine kinase receptor A ( TrkA ), beta-tubulin III ( βTIII ), and Nestin gene levels were compared via reverse transcription polymerase chain reaction (RT-PCR) in these NSCs. Nestin and Gfap protein levels were compared by immunoassay. Subsequently, both populations were induced with 10-8 M selegiline for 48 hours, followed by immunohistochemical analysis of tyrosine hydroxylase (TH) levels. One-way ANOVA and Tukey's post-test were used with a significance level of P<0.05., Results: Both groups were successfully expanded in vitro and expressed the neurotrophin receptor genes. The SGZNSCs had a significantly higher proliferation rate and significantly higher numbers of Nestin and Gfap-positive cells. Although the majority of selegiline-induced NSCs were TH-positive, we observed more TH-positive cells in SGZ-derived NSCs and these SGZ-NSCs displayed a shorter differentiation time., Conclusion: SGZ-derived NSCs appear to be a more appropriate candidate for therapeutic purposes based on proliferation rate, neurosphere size, and Gfap and Nestin expression levels, as well as differentiation time and TH expression level after dopaminergic induction.

Objective: Parkinson's disease (PD) is a neurodegenerative disorder described by the dynamic decline of dopaminergic neurons in the substantia nigra pars compacta (SNpc). Stem cell transplantation is a new therapeutic strategy in the treatment of PD. The objective of the study was to assess the impact of intravenous infusion of adipose-derived mesenchymal stem cells (AD-MSCs) on memory disorder in Parkinsonian rats., Materials and Methods: In this experimental study, male Wistar rats were randomly divided to four groups containing sham, cell treatment, control, and lesion. The cell treatment group received intravenous injection of AD-MSCs 12 days after PD induction by bilateral injection of 6-hydroxydopamine. Four weeks after lesion formation, spatial memory was examined using the Morris water maze (MWM) assessment. The rats' brains were removed and assessed by bromodeoxyuridine (BrdU), tyrosine hydroxylase (TH), and glial fibrillary acidic protein (Gfap) immunostaining., Results: Statistical analyses revealed a significant addition and reduction in time spent and escape latency in the target quadrant, respectively, in the cell group as compared to the lesion group. Also, BrdU-labeled cells were present in the substantia nigra (SN). The density of TH-positive cells was significantly increased in the AD-MSCs transplantation group as compared to the lesion group, and the density of astrocytes significantly diminished in the AD-MSCs transplantation group as compared to the lesion group., Conclusion: It appears that AD-MSCs treatment for Parkinson's could decrease the density of astrocytes and promote the density of TH-positive neurons. It appears that AD-MSCs could improve spatial memory impairment in PD.

Human adipose stem cells (hASCs) were introduced as appropriate candidate due to advantages like ease of isolation, in vitro expansion and lack of immune response. Deprenyl (Dep) was used to induce bone marrow stem cells into neuron-like cells. We investigated the Dep effect on neurotrophin genes expression in hASCs and their differentiation into neuron-like cells. The cells were isolated from small pieces of abdominal adipose tissue and subjected to flow cytometry to confirm purification. The osteogenic and adipogenic differentiation were identified. The proliferation rate and neurotrophin genes expression of treated cells were evaluated by MTT, TH immunostaining and RT-PCR. hASCs had positive response to CD44, CD73, CD90, CD105 markers and negative response to CD34 and CD45 markers and differentiated into adipocytes and osteocytes. Exposure to 10 -7 M of Dep for 24 hours caused a significant increase of viable cells and BDNF, NTF-3 genes expression as compared to cultured cells in serum free medium and had no effect on the expression of NGF and GDNF genes. Based on our results, Dep is able to induce BDNF, NTF-3 and NTF-4 genes expression and neroun-like morphology in hASCs.

Several antiepileptic drugs (AEDs) are administrated during pregnancy according to recent therapeutic protocols. Ten percent of pregnant women with epilepsy give birth to offspring with malformations and teratogenic defects. Since the mechanism of action of AEDs is not yet completely understood, therefore, it could be hypothesized that they may cause cyto- or genotoxicity in embryonic fetus cells. To investigate this hypothesis, the genotoxicity and cell survival of AEDs treated human embryonic stem cells (hESCs) were investigated by single-cell gel electrophoresis (Comet assay) and MTS assay, respectively. HESCs (Royan H6 cell line) were treated in-vitro with high therapeutic doses of Carbamazepine, Gabapentin, Lamotrigine, Levetiracetam or Topiramate as monotherapy or combination therapy of each drug with Folic acid. After hESCs pluripotency confirmation, the effect of AEDs on cellular DNA damage of hESCs was investigated. levetiracetam and topiramate were found to damage the DNA significantly compared to untreated cells. The amount of DNA damage produced by carbamazepine and lamotrigine was similar while for gabapentin, the amount of DNA migration was very low and produced less DNA damage than the others. A considerable reduction in DNA damages occurred in genotoxicity in the presence of Folic acid in comparison to AEDs monotherapies. According to our results, all mentioned AEDs caused DNA damage, while Levetiracetam and topiramate caused more extensive DNA damages than the others. Noticeably, the addition of Folic acid to the treated cells decreased the DNA damages considerably., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Objectives: This study aimed to determine whether exposure to pulsed electromagnetic field (PEMF) can impair behavioral failure as induced by PTSD, and also its possible effects on hippocampal neurogenesis. PEMF was used as a non-invasive therapeutic tool in psychiatry., Materials and Methods: Male rats were divided into Control-Sham exposed, Control-PEMF, PTSD-Sham exposed, and PTSD-PEMF groups. PTSD rats were conducted by the single prolonged stress procedures and then conditioned by the contextual fear conditioning apparatus. Control rats were only conditioned. Experimental rats were submitted to daily PEMF (7 mT, 30 Hz for 16 min/day, 14 days). Sham-exposed groups were submitted to the turned off PEMF apparatus. Fear extinction, sensitized fear and anxiety, cell density in the hippocampus, and proliferation and survival rate of BrdU-labeled cells were evaluated., Results: Freezing of PTSD-PEMF rats was significantly lower than PTSD-Sham exposed. In the PTSD-PEMF, center and total crossing in open field, also the percentage of open arms entry and time in the elevated plus maze, significantly increased as compared with PTSD-Sham exposed ( P <0.001). Numbers of CA1, CA3, and DG cells in PTSD-PEMF and Control-Sham exposed groups were significantly more than PTSD-Sham exposed ( P <0.001). There were more BrdU-positive cells in the DG of the PTSD-PEMF as compared with the PTSD-Sham exposed. Qualitative observations showed an increased number of surviving BrdU-positive cells in the PTSD-PEMF as compared with PTSD-Sham exposed., Conclusion: Using 14-day PEM attenuates the PTSD-induced failure of conditioned fear extinction and exaggerated sensitized fear, and this might be related to the neuroprotective effects of magnetic fields on the hippocampus., Competing Interests: The authors declare that they have no conflicts of interests.

The transcriptional factor OCT4 regulates pluripotency of stem cells and has an important role during oocyte growth. Whereas, its role has remained ambiguous in ovarian tissue during reproductive cycle. Therefore, this study was aimed to investigate the expression patterns of OCT4 in mouse ovaries during the normal estrous cycle. Adult National Medical Research Institute mice were classified as proestrous, estrous, metestrous and diestrous on the basis of vaginal smear cytology. Their ovaries were removed and the protein and gene expression levels of OCT4 were assessed using immunohistochemical staining and real-time quantitative reverse-transcription PCR, respectively. Immunohistochemical staining revealed the expression of OCT4 in the cytoplasm of corpus luteum cells. In the follicles, OCT4 was expressed in the cytoplasm of granulosa cells. Furthermore, the gene expression levels of OCT4 was significantly higher in the proestrous phase than in the other phases of the estrous cycle ( p < 0.05). The results indicated that OCT4 gene expression levels are affected by the cyclic pattern of the estrous cycle.

Background: The uterus is a dynamic tissue responding to hormonal changes during reproductive cycles. As such, uterine stem cells have been studied in recent years. Transcription factors oct4 and sox2 are critical for effective maintenance of pluripotent cell identity., Objective: The present research evaluated the mRNA expression of oct4 and sox2 in the uterine tissues of ovariectomized mice treated with steroid hormones., Materials and Methods: In this experimental study, adult virgin female mice were ovariectomized and treated with estradiol 17β (E2), progesterone (P4), and a combination of E2 and P4 (E2 & P4) for 5 days. Uterine tissues were removed, and immunofluorescent (IF) staining and quantitative real-time PCR of oct4 and sox2 markers were performed., Results: IF showed oct4 and sox2 expression in the uterine endometrium and myometrium among all groups. The mRNA expression of oct4 (p=0.022) and sox2 (p=0.042) in the E2-treated group significantly were decreased compared to that in the control group. By contrast, the mRNA expression of oct4 and sox2 in the P4 (p=0.641 and 0.489 respectively) and E2 & P4-treated groups (p=0.267 and 0.264 respectively) did not show significant differences compared to the control group., Conclusion: The results indicate ovarian steroid hormones change the expression of oct4 and sox2 in the mice uterine tissues, which suggest the involvement of steroid hormonal regulation in uterine stem cells.

Ethnopharmacological Relevance: Date palm (Phoenix dactylifera L.) pollen (DPP) is widely used as a folk remedy for male infertility treatment, and has well known medicinal effects., Aim of the Study: This study aimed to determine the in vitro effects of DPP on the efficiency of neonate mouse spermatogonial stem cells (SSCs) proliferation., Material and Methods: Sertoli and SSCs were isolated from 6 to 10-days-old mouse testes, and their identity was confirmed using immunocytochemistry against cytokeratin for sertoli cells and PLZF, Oct-4 and CDH-1 for SSCs. Isolated testicular cells were cultured in the absence or presence of 0.06, 0.25 and 0.62mg/ml concentrations of DPP aqueous extract for 2 weeks. The number and diameter of SSC colonies were assessed during third, 7th, 9th and 14th day of culture, and the expression of the Mvh, GFRα-1 and Oct-4 was evaluated using quantitative PCR at the end of the culture period. The significance of the data was analyzed using ANOVA and paired samples t-test and Tukey and Bonferroni test as post hoc tests at the level of p≤0.05., Results: Pattern assay of colony formation showed that SSCs numbers increased in the present of 0.62mg/ml concentration of DPP extract with higher slop relative to other groups (P <0.05). Colony diameters had no significant difference between groups in 3th, 7th, 9th and 14th days after culture. The Mvh and Oct-4 genes expression had no significant difference between groups, while GFRα1 expression was increased significantly in cells treated with 0.06mg/ml concentration relative to other groups (P<0.05)., Conclusion: It seems that co-culture of SSCs with sertoli sells in the presence of low doses of DPP can increase SSCs proliferation and keep their stemness state, while higher concentrations can differentiate the treated cells., (Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.)

It was assumed that uterine stem cells are responsible for the unique regenerative capacity of uterine. Therefore, the aim of the present study was to investigate the expression of the pluripotent stem cell markers in the mice uterine tissue during different stages of estrous cycles. Twelve virgin female NMRI mice (6 to 8 weeks old) were considered at proestrus, estrus, metestrus and diestrus according to the cell types observed in the vaginal smear and underwent hysterectomy operation. Quantitative real-time polymerase chain reaction (PCR) and immunohistochemical staining for pluripotent stem cell markers (SOX2, OCT4, KLF4, and NANOG) were performed. Immunofluorescence staining revealed that expression and localization of the pluripotency markers SOX2, OCT4, KLF4, and NANOG at the protein level were not different throughout estrous cycle. Also, mRNA of pluripotency markers was detected in all tested samples. However, there were no significant differences in their genes expression at each stage and during the estrous cycle. Different hormonal profile during the estrous cycle could not affect expression of pluripotent stem cell markers in uterine tissue.

Background: One of the most major obstacles of ovarian tissue vitrification is suboptimal developmental competence of follicles. Matrix metalloproteinases 2 (MMP-2) and 9 (MMP-9) and their tissue inhibitors TIMP-1 and TIMP-2 are involved in the remodeling of the extracellular matrix in the ovaries., Objective: This study aimed to evaluate the expression of MMP-2, MMP-9, TIMP-1, and TIMP-2 genes in the preantral follicles derived from vitrified mouse ovaries., Materials and Methods: In this experimental study, the gene expression of MMP-2, MMP-9, TIMP-1, and TIMP-2 in the isolated preantral follicles derived from fresh and vitrified ovaries of 14-16 days old female mice through real time qRT-PCR was evaluated. Developmental parameters, including survival rate, growth, antrum formation and metaphase II oocytes were also analyzed., Results: The developmental parameters of fresh preantral follicles were significantly higher than vitrified preantral follicles. The TIMP-1 and MMP-9 expression levels showed no differences between fresh and vitrified preantral follicles (p=0.22, p=0.11 respectively). By contrast, TIMP-2 expression significantly decreased (p=0.00) and MMP-2 expression increased significantly (p=0.00) in vitrified preantral follicles compared with to fresh ones., Conclusion: Changes in expression of MMP-2 and TIMP-2 after ovarian tissues vitrification is partially correlated with decrease in follicle development., Competing Interests: There is no conflict of interest in this research.

There is a fast growing tendency in the use of herbal remedies in developing countries. One of the traditional medicines used for male infertility treatment is date palm (Phoenix dactylifera) pollen (DPP). Isolated spermatogonial stem cells and sertoli cells using enzymatic digestion were grown in Dulbecco's modified Eagle's medium supplemented with 4% foetal bovine serum in the absence or presence of 0.06, 0.25 and 0.62 mg/mL concentrations of aqueous extract of DPP for 2 weeks. The assessment of mean number of the whole cells and the living cells showed that there were no significant differences between the mean viability percentage and proliferation rate between control and experimental groups (P>0.05). As there are no cytotoxicity effects of DPP in our cultural system, this system can be utilised for the enrichment or differentiation of these cells in clinical applications, cell replacement therapy, tissue regeneration and tissue engineering applications.

Background: Cryopreservation of pre-antral follicles is a hopeful technique to preserve female fertility. The aim of the present study was to evaluate reactive oxygen species (ROS) and total antioxidant capacity (TAC) levels of mouse vitrified pre-antral follicles in the presence of alpha lipoic acid (ALA)., Methods: Isolated pre-antral follicles (140-150 µm in diameter) were divided into vitrified-warmed and fresh groups. Each group was subjected to in vitro maturation with or without ALA for 12 days, followed by adding human chronic gonadotropin to induce ovulation. In vitro fertilization was performed to evaluate their developmental competence. In parallel, the amount of ROS and TAC were assessed after 0, 24, 48, 72, and 96 h of culture by 2',7'-dichlorofluorescin assay and ferric reducing/antioxidant power assay, respectively., Results: The respective rates of survival, antrum formation, and metaphase II oocytes were significantly higher in ALA-supplemented groups compared to the groups not treated with ALA. TAC and ROS levels were significantly decreased and increased, respectively during the culture period up to 96 h in the absence of ALA in both vitrified and non-vitrified samples. However, with pretreatment of ALA, TAC levels were increased significantly and remained constant up to 96 h in vitrified-warmed pre-antral follicles, while ROS levels completely returned to the level of starting point after 96 h of culture in the presence of ALA., Conclusion: Pretreatment of ALA positively influences development of pre-antral follicles in vitrified and non-vitrified samples through increasing follicular TAC level and decreasing ROS levels.

Background: Cryopreservation of ovarian tissues and pre-antral follicles is a promising prospect for preservation of women fertility., Objective: The aim of this study was to evaluate the in vitro developmental competence of mouse vitrified pre-antral follicles in comparison to isolated pre-antral follicles derived from vitrified ovaries in the presence of alpha lipoic acid (ALA)., Materials and Methods: Pre-antral follicles derived from fresh, vitrified-warmed ovarian tissues and vitrified-warmed pre-antral follicles were cultured individually with or without ALA, followed by adding hCG to induce ovulation. The follicle growth, oocyte maturation, and embryo development were assessed., Results: The diameter and development of follicles, oocyte maturation and embryo development rates were significantly higher in ALA supplemented groups compared to the respective ALA-free conditions groups. Aforementioned parameters were significantly higher in vitrified-warmed follicles in comparison to follicles derived from vitrified-warmed ovaries., Conclusion: These findings support a superior performance of pre-antral follicles when vitrified rather than when isolated from vitrified ovaries with regard to increasing the rates of developmental parameters. Moreover, ALA improves the in vitro maturation of pre-antral follicles in vitrified and non-vitrified samples. This article extracted from M.Sc. thesis. (Sahar Hatami).

Objective(s): Stem cell therapy is believed to be as a promising treatment strategy for tissue repair and regeneration. The plasticity specification of the adult stem cells, such as MSCs, has enabled that these cells to be used in the treatment of a broad spectrum of diseases like liver disorders. In this study, the production of urea and Albumin (Alb), glycogen storage, and expression of some liver genes including α-fetoprotein (AFP), Alb, cytokeratin18 (CK18) and cytokeratin19 (CK19) was compared between mesenchymal stem cells (MSCs) and isolated rat hepatocytes., Materials and Methods: The MSCs were isolated from rat femurs and tibias and cultured in α-MEM, DMEM and RPMI mediums supplemented with serum. Hepatocytes were isolated from Rat livers and cultured in DMEM with serum. The expression of AFP, Alb, CK18, and CK19 genes was evaluated using the reverse transcription-polymerase chain reaction (RT-PCR). Furthermore, the synthesis of albumin and urea of the cells was measured., Results: In vitro conditions, MSCs and hepatocytes exhibited the characteristic functions of the liver such as capacity to synthesize Alb, urea, the storage of glycogen. In this study, the expression of some liver genes such as AFP, Alb, CK18 and CK19 at mRNA levels was also shown., Conclusion: The results showed that MSCs exhibited some liver functions, and may be considered as an alternative source for adult stem cell transplantation in liver repair due to the excellent proliferation and differentiation capacities.

Background: In spite of extensive efforts to improve in vitro oocyte maturation, the obtained results are not very efficient. This study was conducted to assess impacts of cAMP elevating agents and alpha lipoic acid (ALA) on in vitro oocyte maturation and fertilization., Methods: Mouse germinal vesicle (GV) oocytes were categorized into cumulus denuded oocytes (DOs; n=896) and cumulus oocyte complexes (COCs; n=1077) groups. GV oocytes were matured in vitro with or without ALA; (I) without the meiotic inhibitors; (II) supplemented with cilostamide; (III) supplemented with forskolin and (IV) supplemented with Forskolin plus cilostamide. The obtained metaphase II (MII) oocytes were subjected to in vitro fertilization. Independent-samples t-testand ANOVA were used for data analysis. A p-value less than 0.05 was considered to be statistically significant., Results: The COCs maturation, fertilization and two cell embryo rates were higher than those of DOs in the control group, while no significant difference was observed between relevant COCs and DOs when they were cultured with cilostamide meiotic inhibitors in two step manner. Combined treatment of cilostamide and forskolin significantly elevated the developmental rates in both COCs and DOs as compared to other groups. The developmental rates of COCs and DOs in the presence of ALA were similar to their respective groups without ALA., Conclusion: cAMP elevating agents were more effective on DOs than COCs with regard to rates of maturation and fertilization. However, ALA did not affect the developmental rates of both COCs and DOs in in vitro maturation in one or two step manner.

Objective: There is longstanding experimental and clinical evidence that supports the idea that replacement of dopaminergic (DAergic) neurons can ameliorate functional disabilities of Parkinson's disease (PD). The purpose of the present study is to examine the efficacy of transplantation of rat bone marrow stromal cell (BMSCs)-derived tyrosine hydroxylase-positive (TH(+)) cells induced by deprenyl into 6-hydroxydopamine (6-OHDA)-lesioned rat models, using behavioral tests and immunohistochemical evaluations., Materials and Methods: In this experimental study, undifferentiated BrdU-labeled BMSCs were incubated in serum-free medium that contained 10(-8) M deprenyl for 24 hours. Afterwards, BMSCs were cultured for 48 hours in α-minimal essential medium (α-MEM) supplemented with 10% FBS, then differentiated into TH(+) neurons. We randomly divided 24 hemiparkinsonian rats as follows: group 1 (control) received only medium, while groups 2 and 3 were injected with 2×10(5) BMSCs and deprenyl-treated cells in 4 µl medium. Injections were made into the injured strata of the rats. Rotational behavior in response to apomorphine was tested before transplantation and at 2, 4, and 6 weeks post-graft. Animals were then sacrificed, and the brains were extracted for immunohistochemical and electron microscopic studies., Results: Apomorphine-induced rotation analysis indicated that animals with grafted cells in groups 2 and 3 exhibited significantly less rotational behavior than those in the control group at 2, 4, and 6 weeks after transplantation. Immunohistochemical analysis demonstrated that BrdU-labeled cells expressed specific neuronal markers, such as NF 200 and TH, at the implantation site. The presence of TH(+) cells in conjunction with the reduction in rotation might show the capacity of grafted cells to release dopamine. Ultrastructural analysis revealed the presence of immature neurons and astrocyte-like cells at the graft site., Conclusion: TH(+) neurons induced by deprenyl can be considered as a cell source for PD autograft therapy.

Objective: This research study is an attempt to examine whether the administration of ethanol after memory reactivation would modulate subsequent expression of memory in rats. Additionally, we examined whether this administration alters the density of Cornu Ammonis (CA)1 and CA3 pyramidal and dentate gyrus (DG) granule cells., Materials and Methods: In this experimental study, adult male Wistar rats (200-300 g) were trained in a fear conditioning system using two 1 second, 0.6 mA shocks with an interval of 180 seconds. Twenty four hours later rats were returned to the chamber for 120 seconds. Immediately after reactivation they were injected with ethanol (0.5, 1, 1.5 mg/ kg) or saline. 1, 7 and 14 days after reactivation, rats were returned to the context for 5 minutes. Seconds of freezing (absence of all movement except respiration) were scored. In the second experiment (described in the previous paragraph), after test 1, animals were anesthetized with sodium pentobarbital and perfused transcardially with phosphate buffer (10 minutes) and 4% paraformaldehyde (15 minutes). The brains were postfixed in phosphate-buffered 4% paraformaldehyde (24 hours) and 30% sucrose. 10-µm sections were stained with cresyl violet. Data were analyzed by 1-and 2-way ANOVA for repeated measurements by means of SPSS 16.0. Tukey's post hoc test was performed to determine the source of detected significant differences. P <0 .05 were considered significant. Data are presented as mean ± SEM., Results: Findings from the first experiment indicated that ethanol at a dose of 1.5 mg/kg significantly impaired recall of memory only in the first test. The density of CA1 and CA3 pyramidal and DG granule cells in the ethanol group was decreased (p< 0.01) compared with control group respectively 43.7%, 35.8%, and 37.8., Conclusion: The data demonstrate that ethanol exposure impairs post retrieval processes. Moreover, ethanol decreases the density of CA1, CA3 and DG cells. Presumably it would be a correlation between our behavioral and histological results.

Objective: It has been reported that rat bone marrow stromal cells (BMSCs) can be spontaneously differentiated into neural-like cells without any supplemental growth factors and/or chemical treatment after long-term culture.This study aims to determineWhether, growth factors secreted by MSCs could induce self-differentiation into neural-like cells in a long-term culture., Materials and Methods: THIS STUDY CONSISTED OF TWO GROUPS: i. rat BMSCs (passage 5) were cultured in alfa- minimal essential medium (α-MEM) and 10% fetal bovine serum (FBS) without the addition of inducer and exchanging medium for three weeks, as the experimental group and ii.rat BMSCs (passage 5) as the control group. Each group was analysed by reverse transcriptase polymerase chain reaction (RT-PCR) to evaluate the expressions of neurotrophic factors and neural marker genes. Statistical analyses were carried out using one-way analysis of variance (ANOVA) and Tukey's multiple comparison with SPSS software (version 16). P< 0.05 was considered statistically significant., Results: The experimental group (fifth passage of BMSCs) obtained from adult rats spontaneously differentiated into neural precursor cells after long-term culture. Cultured cells expressed tyrosine hydroxylase (TH), Nurr1 and nestin genes. Furthermore, some growing cells in suspension became neurosphere-like. Self-differentiated rat MSCs (SDrMSCs) expressed significantly higher levels of NGF (0.96 ± 0.16), nestin (0.63 ± 0.08), and Nurr1 (0.80 ± 0.10) genes (p<0.05)., Conclusion: In this study, we reported that rMSCs in long-term culture underwent spontaneous transformation to neural precursors without the supplement of growth factors and specific chemicals. Cells expressed neural markers such as: TH, Nurr1, and nestin genes.

This study is an attempt to examine whether administration of ethanol after memory reactivation will modulate expression of memory in rats or not. We further examined whether this administration alters the number of tunnel positive cells in hippocampus. Adult male Wistar rats were trained in a fear conditioning system using two 1s , 0.6 mA shock with an interval of 180 s. 24 h later the rats were returned to the chamber for reactivation, and then they were injected with ethanol (0.5, 1, 1.5 mg/kg) or saline, ip. Again, one, seven and fourteen days after reactivation, the rats were returned to the context for 5 min. The freezing time (absence of all movements except respiration) was scored in seconds. In the second experiment, after test 1, the animals were anesthetized and a transcardial perfuse with phosphate buffer and paraformaldehyde 4% was conducted. After post-fixation of brains 5-μm sections were stained with cresyl violet. Finally, paraffin-embedded sections of 10 μm were cut out throughout the tissue and each sample was processed with TUNEL. The number of apoptotic cells in a 130 μm-long segment of the hippocampal CA1 and CA3 fields and dentate gyrus was counted. The data demonstrate that ethanol exposure impairs post retrieval processes. Rats receiving ethanol (1.5 mg/kg) showed lower freezing levels during the first test. Moreover, ethanol decreases the density of CA1, CA3 and DG cells and increases the density of apoptotic cells in all regions of hippocampus. Therefore, ethanol exposure impairs reconsolidation of contextual fear conditioning probably via decreasing the density of CA1, CA3 and DG cells., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Objective(s): The purpose of this study was to investigate the role of oxidative stress in Purkinje cell neurotoxicity of ethanol-treated rat., Materials and Methods: Male rat pups 4-day-old was used in this study. Ethanol was administered to rat pups at a dose of 6 g/kg from postnatal days (PDs) 4 to 5. Pups were killed 90 min after the second alcohol treatment on PD 5 by decapitation and the brain was immediately removed. The cerebellum was dissected for analyzing the oxidative stress parameters and histological study. The activities of several antioxidant enzymes including superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx) in vermis of cerebellum were assayed. Thiobarbituric acid reactive substances (TBARS) levels were also measured as a marker of lipid peroxidation., Results: Administration of ethanol significantly increased TBARS levels in the cerebellum compared to control pups (P< 0.01). The treated pups with ethanol exhibited a marked decrease in the GPx activity (P< 0.01) whereas, in spite of decrease in the activities of SOD and CAT, when compared to control, there were not significant differences. The spherical cell bodies of Purkinje cells in control rats are aligned nicely between the granular and molecular layers. In ethanol treated pups, Purkinje cells scattered within the Purkinje cell layer and shrinkage of the cell somata is seen., Conclusion: The results of the present work demonstrated that ethanol exposure during the vulnerable window could increase TBARS levels (lipid peroxidation) and decrease GPx levels in pup's cerebellum. Also, the results confirmed ethanol-induced microencephaly, cerebellar Purkinje cell loss. These findings suggest that Purkinje cell loss is, in part through decrease in the activity of GPx and increase of lipid peroxidation in the rat cerebellum.

During particular periods of central nervous system (CNS) development, exposure to ethanol can decrease regional brain growth and can result in selective loss of neurons. Unfortunately, there are few effective means of attenuating damage in the immature brain. In this study, the possible antioxidant and neuroprotective properties of 17β-estradiol against ethanol-induced neurotoxicity was investigated. 17β-estradiol (600 μg/kg) was injected subcutaneously in postnatal day (PD) 4 and 5, 30 min prior to intraperitoneal injection of ethanol (6g/kg) in rat pups. Ninety minutes after injection of ethanol, the activities of several antioxidant enzymes including superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (Gpx) in vermis of cerebellum were assayed. Thiobarbituric acid reactive substance (TBARS) levels were also measured as a marker of lipid peroxidation. Behavioral studies, including rotarod and locomotor activity tests were performed in PD 21-23 and histological study was performed after completion of behavioral measurements in postnatal day 23. The results of the present work demonstrated that ethanol could induce lipid peroxidation, increase TBARS levels and decrease glutathione peroxidase levels in pup cerebellum. We also observed that ethanol impaired performance on the rotarod and locomotor activities of rat pups. However, treatment with 17β-estradiol significantly attenuated motoric impairment, the lipid peroxidation process and restored the levels of antioxidants. Histological analysis also indicated that ethanol could decrease vermis Purkinje cell count and 17β-estradiol prevented this toxic effect. These results suggest that ethanol may induce lipid peroxidation in the rat pups cerebellum while treatment with 17β-estradiol improves motor deficits by protecting the cerebellum against ethanol toxicity., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Objective: Bone marrow stromal cells (BMSCs) were documented as a feasible candidate for cell therapy. Several protocols with one or more steps, using different chemicals, were used for inducing BMSCs differentiation into neuronal phenotype. Many of these chemicals were reported to be of mutagenic, teratogenic or carcinogenic properties. The purpose of this work was to evaluate the neuronal inductivity of selegiline to BMSCs., Methods: Selegiline was used to induce BMSCs following pre-induction with beta-mercaptoethanol and retinoic acid. The neuronal phenotype was evaluated using antineurofilament 200, neurofilament 68, anti-Map-2 and synapsin I antibodies. In addition, reverse transcription polymerase chain reaction was used for the expression of brain-derived neurotrophic factor (BDNF), nerve growth factor (NGF), neurotrophin 3 (NT-3), neuroligin 1 and post-synaptic density protein 95 (PSD-95)., Results: The result of the work showed that the induced BMSCs were immunoreactive for neurofilaments 200 and 68. Moreover, anti-Map-2 and synapsin I antibodies showed a structure morphologically consistent with synapses. Reverse transcription polymerase chain reaction showed that the induced BMSCs could express BDNF, NGF, NT-3, neuroligin 1 and PSD-95. The time course used was 1, 3, 6, 12, 24, 36 and 48 hours, and the percentages of neurofilament 200 immunoreactive cells were analysed using the logistic growth curve., Discussion: The results showed that selegiline was efficient and a potent inducer for BMSCs into neuronal phenotype, which can be used in the cell therapy in neurodegeneration and traumatic nervous tissue injuries.