Granulosa cells from immature follicles exhibit restricted glycolysis and reduced energy production: a dominant problem in polycystic ovary syndrome.

PURPOSE: We hypothesized that immature oocytes are associated with impaired energy production in surrounding granulosa cells (GCs) in polycystic ovary syndrome (PCOS). Thus, this study investigated mitochondrial function, determined expression of glycolytic regulatory enzymes, and measured ATP levels in GCs of PCOS patients. METHODS: GCs were isolated from forty-five PCOS patients and 45 control women. Intracellular concentration of reactive oxygen species (ROS), mitochondrial membrane potential (Δψm), the rate of glycolysis, total antioxidant capacity (TAC), activities of catalase (CAT) and superoxide dismutase (SOD), and ATP level were measured in GCs. The gene expression and protein levels of glycolytic enzymes (hexokinase, muscular phosphofructokinase, platelet derived phosphofructokinase, and muscular pyruvate kinase) were determined. Association of GC energy level with oocyte maturation was further validated by measuring glycolysis rate and ATP level in GCs isolated from mature and immature follicles from new set of fifteen PCOS patients and 15 controls. RESULTS: PCOS patients showed higher ROS level, decreased TAC, reduced CAT and SOD activities, and lower Δψm together with reduced expression of key glycolytic enzymes. ATP concentration and biochemical pregnancy were lower in PCOS compared with control group. ATP levels were found to be significantly correlated with ROS and Δψm (r = - 0.624 and r = 0.487, respectively). GCs isolated from immature follicles had significantly lower ATP levels and rate of glycolysis compared with the GCs separated from mature follicles in both PCOS patients and control. CONCLUSION: Declined energy due to the mitochondrial dysfunction and restrained glycolysis in GCs is associated with the immature oocytes and lower biochemical pregnancy in PCOS.

Effects of progesterone nanoparticles on the sperm capacitation and acrosome reaction in asthenozoospermia men.

Low motility is one of the causes of male infertility. In this study, the effects of progesterone solid lipid nanoparticles (SLNs) on sperm capacitation, acrosome reaction, oxidative stress and expression of SPACA1 and MAPK way genes were investigated. Progesterone SLNs were synthesized using the solvent emulsification evaporation method. Twenty asthenozoospermia samples were selected, and sperm and acrosome membrane integrity, acrosome reaction, sperm motility, viability, total antioxidant capacity (TAC), total oxidative status tests and PKA, PTK, P38MAPK and SPACA1 gene expressions were assessed. The synthesized nanoparticles were prepared with the size (187.6 nm), PDI (0.184), EE (85.82%), LP (3.43%) and ZP (-23.5mV). Progesterone SLNs increased sperm and acrosome membrane integrity and TAC (p < .05). Also, the expression of P38MAPK, PKA, PTK, and SPACA1 genes in this group showed a significant increase (p < .001). Progesterone SLNs increased acrosome reaction, sperm capacitation and TAC. Also, it increased the expression of PTK PKA, SPACA1 and P38MAPK genes.

Administration of Selenium Nanoparticles Reverses Streptozotocin-Induced Neurotoxicity in the male rats.

Alzheimer's disease is the most common neurodegenerative disease associated with deposition of amyloid-beta and the increased oxidative stress. High free radical scavenging ability of selenium nanoparticles (SeNPs) has been acknowledged, so in the present study, the effects of treatment with SeNPs on Streptozotocin (STZ)-induced neurotoxicity were evaluated in the male rats. Learning and memory impairment was induced by intraventricular injection of STZ. Following induction of memory impairment, the rats received 0.4 mg/kg of SeNPs daily for one month. Memory function, antioxidant capacity, and deposition of Amyloid ß (Aß) were assessed using the shuttle box task, biochemical methods, and Congo red staining. Injection of STZ caused memory impairment, a decrease in the level of total thiol group (TTG), and an increase in the malondialdehyde (MDA) content and deposition of Aß. Administration of SeNPs reversed the neurotoxicity induced by STZ. It seems that SeNPs likely had neuroprotective effects on the animal model of Alzheimer's disease through increasing antioxidants҆ capacity.

Abnormal expressions of ADAMTS-1, ADAMTS-9 and progesterone receptors are associated with lower oocyte maturation in women with polycystic ovary syndrome.

PURPOSE: ADAMTS-1 and 9 play a crucial role in the ovulation and their altered levels may play a role in the pathogenesis of polycystic ovary syndrome (PCOS). The aim of this study was to assess ADAMTS-1 and 9 expression and their correlation with the oocyte quality and maturity in the cumulus cells (CCs) of PCOS patients and normovulatory women during an IVF procedure. METHODS: Expression of ADAMTS-1 and 9 and progesterone receptors (PRs) in the CCs containing MII and germinal vesicle (GV) oocytes of 37 PCOS patients and 37 women with normal ovulatory function who underwent IVF treatment was evaluated using qRT-PCR. Moreover, correlation between ADAMTS-1 and 9 expression and oocyte quality were also investigated. RESULTS: mRNA expression levels of ADAMTS-1 and ADAMTS-9 were significantly reduced in the women with PCOS compared to the normovulatory women. ADAMTS-1 and ADAMTS-9 mRNA expression levels in the CCs showed a considerable correlation. Lower expression levels of ADAMTS-1 and ADAMTS-9 in PCOS patients were strongly correlated with diminished oocyte maturation. There was a remarkable association between ADAMTS-1 and ADAMTS-9 mRNA expression levels and oocyte quality. PRs (PRA and PRB) were dramatically decreased in PCOS patients when compared with the control group. CONCLUSIONS: The results of the present study indicated that ADAMTS-1 and ADAMTS-9 as well as PRs are downregulated in the human CCs in PCOS patients, which could be associated with impaired oocyte maturation and may result in a lower oocyte recovery and oocyte maturity rates, as well as lower fertilization rate.

Sperm matrix metalloproteinase-2 activity increased in pregnant couples treated with intrauterine insemination: a prospective case control study.

Matrix metalloproteinase-2 (MMP2) and matrix metalloproteinase-9 (MMP9) have an important role in the reproductive system and in the fertilisation process. The aim of this study was to investigate the MMP2 and MMP9 activity in semen and their association with the pregnancy rate, semen parameters and seminal plasma oxidative stress parameters in couples who were treated with intrauterine insemination (IUI). The semen specimens were obtained from 60 men who attended with their spouse for the IUI in the infertility unit. A controlled ovarian stimulation was performed with clomiphene citrate in IUI cycles. Women with positive pregnancies were recorded (n = 29). The results showed the activity of sperm MMP2 and seminal plasma MMP9 was significantly higher in the pregnant group, compared to the non-pregnant group (p < .05). There was a correlation between the sperm MMP2 activity and the total thiol group (TTG) (r = 0.276, p < .05) and the total antioxidant capacity (TAC) of seminal plasma (r = 0.304, p < .05). The sperm MMP9 showed a positive correlation with the seminal plasma TAC (r = 0.330, p < .05) and an inverse correlation with the lipid peroxidation (LP) of seminal plasma (r = -304, p< 0.05). In addition, the seminal plasma MMP2 activity was correlated to sperm viability (r = 0.266, p< .05) and the TTG of seminal plasma (r = 0.298, p < .05). The MMP2 activity in the sperm may be an important factor for determining the pregnancy rate after IUI. Impact statement What is already known on this subject? Previous studies have reported that the fusion between the sperm and zona pellucida required the activity of matrix metalloproteinase 2 (MMP2), whereas the inhibition of MMP2 can significantly decrease the in vitro fertilisation (IVF) rate. What do the results of this study add? This study has identified that the sperm MMP2 activity was significantly higher in the pregnant couples in comparison with the non-pregnant couples, who treated with intrauterine insemination (IUI). The findings showed there was a correlation between sperm MMP2 activity and the total thiol group (TTG) and the total antioxidant capacity (TAC) of the seminal plasma. What are the implications of these findings for clinical practice and/or further research? MMP2 activity in the sperm could influence the IUI outcome and it is an important factor for IUI success.

Effects of CeO2 nanoparticles on the HO-1, NQO1, and GCLC expression in the testes of diabetic rats.

CeO2 nanoparticles (CNPs) as effective ROS scavengers exhibit potent antioxidant activity. In this study the effect of CNPs investigated was on HO-1, NQO1, and GCLC expression in the streptozotocin (STZ)-induced diabetic rats. Twenty-four male Wistar rats were divided into 4 groups: controls did not receive any treatment; diabetic rats received STZ (60 mg/kg daily); CNPs group received CNPs 30 mg/kg daily for 2 weeks; and rats in STZ + CNPs group received CNPs 30 mg/kg daily for 2 weeks following STZ injection. Oxidative stress was evaluated by measurement of total antioxidant capacity (TAC) and total oxidative status (TOS levels). HO-1, NQO1, and GCLC expression was measured using quantitative real-time PCR. Following STZ injection, significant lower levels of TAC and higher levels of TOS were observed. CNPs could alleviate deleterious effects of diabetes through the enhancement of TAC levels and a significant decline in TOS levels. HO-1, NQO1, and GCLC expression in the diabetic rats were lower than controls. HO-1, NQO1, and GCLC was upregulated in the diabetic rats treated with CNPs. There were significant correlations between NQO1 and GCLC, NQO1 and HO-1, and between HO-1 and GCLC expression. Moreover, Nrf2 was associated with NQO1, GCLC, and HO-1 expression. CNPs as Nrf2 upregulator confer protection against oxidative stress in the testes of STZ-induced diabetic rats by upregulating HO-1, GCLC, and NQO1 cytoprotective genes.

All-optical wireless wavelength multiplexing and demultiplexing using resonant cavity.

The potential capability of wireless wavelength multiplexing and demultiplexing can enable the next development of smaller photonic counterparts for network architectures. This paper numerically represents a new design of a wireless transmission in C-band infrared wavelengths within two identical resonant cavities between photonic chips. This system consists of an H1 rod-type two-dimensional photonic crystal (PhC) microcavity, which can be operated as both a transmitter and a receiver without interfering with the signal in each PhC waveguide. By using the point-to-point oscillatory light-field exchange between resonant cavities, two independent photonic circuits are linked with each other. The obtained results show that the multi-resonance wavelengths in one chip can be transferred to another chip located far away by ten times the highest resonance wavelength. Such a device can be useful for integrated optical circuit interconnect and small-scale sensors between photonic chips.

Sublethal concentration of H2O2 enhances the protective effect of mesenchymal stem cells in rat model of spinal cord injury.

OBJECTIVE: To investigate the effect of H2O2 on the migration and antioxidant defense of mesenchymal stem cells (MSCs) and the neurotrophic effects of H2O2-treated MSCs on spinal cord injury (SCI). RESULTS: Sublethal concentrations of H2O2 decreased cell migration and expression of CXCR4 and CCR2 as well as Nrf2 expression in MSCs. In the second phase, transplantation of treated and untreated MSCs to SCI caused minor changes in locomotor dysfunction. There was a significantly difference between cell-treated and spinal cord injury groups in expression of BDNF (brain-derived neurotrophic factor). Transplantation of H2O2-treated cells caused an increase in BDNF expression compared to non-treated cells. CONCLUSION: Transplantation of H2O2-treated stem cells may have protective effects against SCI through by increasing neurotrophic factors.

Amelioration of diabetes-induced testicular and sperm damage in rats by cerium oxide nanoparticle treatment.

Cerium oxide nanoparticles (CNPs) as an antioxidant have been used frequently to attenuate hyperglycaemia oxidative damage in different organs. We investigated the impact CNPs on the qualitative and quantitative sperm parameters, spermatogenesis and NFE2-related factor 2 (Nrf2) expression as a major contributor of antioxidant defence in the male diabetic rats. Twenty-four male rats were divided into four groups. Controls received only mouse food and water. Second group were treated with CNPs (30 mg kg-1  day-1 ) for 2 weeks. Rats in third group received streptozotocin (STZ) (60 mg/kg). In fourth group, animals became diabetic and received CNPs (30 mg kg-1  day-1 ) for 2 weeks. The results showed a significant abnormality in the sperm parameters and histopathological patterns of testes in the diabetic group compared to the control group and CNPs treatment significantly improved all testicular parameters. Following CNPs administration, sperm DNA fragmentation significantly reduced in the STZ-treated rats. Moreover, after CNPs intake in the STZ-treated rats, Nfr2 expression levels increased significantly. Overall, CNPs administration on the diabetic rates can attenuate detrimental effects of diabetes on the sperm potential fertility, sperm parameters, DNA integrity and Nrf2 expression levels. This study gives a future prospect to determine the role of CNPs in the context of diabetes.

Treadmill exercise attenuates 3,4-methylenedioxymethamphetamine-induced memory impairment through a decrease apoptosis in male rat hippocampus.

3,4-methylenedioxymethamphetamine (MDMA) leads to apoptosis in the hippocampus with consequent induction of learning and memory impairment. In this study, we have investigated the effects of treadmill exercise on memory in relation to apoptosis and oxidative stress in the hippocampi of MDMA-treated rats. Male Wistar rats received multiple intraperitoneal (IP) injections of MDMA (10 mg/kg) and exercised for one month on a treadmill (simultaneously or asynchronously with MDMA). We assessed memory function with the Morris water maze (MWM) test. Lipid peroxidation (LPO) and expression of caspase 3, Bax, and Bcl-2 were examined by the thiobarbituric acid assay (TBA) and western blot, respectively. Our results showed that asynchronous treadmill exercise could significantly improve MDMA-induced memory impairment in the MWM test. Caspase 3 expression decreased in the exercise group compared to the MDMA group. Although MDMA treatment caused an increase in the Bax/Bcl-2 ratio, the treadmill exercise reduced this ratio. Simultaneous exercise caused a reduction in lipid peroxidation in the hippocampus. This data suggests that treadmill exercise can be a useful strategy for treating memory impairment in persons with neurodegenerative disease and stimulant drug users. © 2017 Wiley Periodicals, Inc.

Treadmill exercise alters ecstasy- induced long- term potentiation disruption in the hippocampus of male rats.

3, 4-methylenedioxymethamphetamine (MDMA) or ecstasy is a derivative of amphetamine that leads to long term potentiation (LTP) disruption in the hippocampal dentate gyrus (DG). Exercise has been accepted as a treatment for the improvement of neurodegenerative disease. Herein, the effects of exercise on the MDMA- induced neurotoxicity were assessed. Male Wistar rats received intraperitoneal injection of MDMA (10 mg/kg) and exercised for one month on a treadmill (Simultaneously or asynchronously with MDMA). LTP and expression of BDNF were assessed using electrophysiology and western blotting methods, respectively. MDMA attenuated the field excitatory post-synaptic potential (fEPSP) slope in comparison with the control group, whereas treadmill exercise increased this parameter when compared to MDMA group. Furthermore, BDNF expression significantly decreased in MDMA group and treadmill exercise could increase that. In conclusion, results of this study suggest that synchronous exercise is able to improve MDMA-induced LTP changes through increase of BDNF expression in the hippocampus of rats.

Erratum to: Treadmill exercise alters ecstasy- induced long- term potentiation disruption in the hippocampus of male rats.

In the original publication of the article, author name Masoumeh Asadbegi was incorrectly written as Masoumeh Asadbeigi. The authors regret the oversight.

Downregulation of adiponectin system in granulosa cells and low levels of HMW adiponectin in PCOS.

PURPOSE: The purpose of the study was to investigate changes in adiponectin system expression in granulosa cells (GCs) and high molecular weight adiponectin levels in serum and follicular fluid (FF) of 40 women with polycystic ovary syndrome (PCOS) compared to those in 40 women with normal ovary function. METHODS: Adiponectin (Adipo), adiponectin receptor 1 (AdipoR1), and adiponectin receptor 2 (AdipoR2) messenger RNA (mRNA) expression levels were measured using quantitative real-time polymerase chain reaction (qRT-PCR). High molecular weight (HMW) adiponectin protein concentration was evaluated by ELISA method. Data were analyzed using Student's t test and one-way ANOVA in SPSS 21 software. At oocyte retrieval, FF was aspirated and GCs were obtained from a pooled collection of FF per each patient. RESULTS: PCR results showed expression of adiponectin, AdipoR1, AdipoR2, follicle-stimulating hormone receptor (FSHR), and luteinizing hormone receptor (LHR) in GCs. After controlling body mass index (BMI) values, qRT-PCR demonstrated a decreased expression of adiponectin system in GCs of PCOS patients compared to those in controls (p = 0.001). There was a strong positive correlation among AdipoR1 and AdipoR2 expression and also among FSH and LH receptor expression. (Both r = 0.8, p = 0.001). There were low levels of high molecular weight adiponectin in the serum of PCOS patients with controlled ovarian hyperstimulation (30.19 ± 4.3 ng/ml) compared to the controls (48.47 ± 5.9 ng/ml) and in the FF of PCOS patients with controlled ovarian hyperstimulation (7.86 ± 1.44 ng/ml) compared to the controls (14.22 ± 2.01 ng/ml; p = 0.02). CONCLUSIONS: Lower expression of adiponectin and its receptors in GCs might be an important manifestation in gonadotropin-stimulated PCOS patients which could influence the physiologic adiponectin roles such as interaction with insulin and LH in induction of GC gene expression.

PROTECTIVE EFFECTS OF GLUTATHIONE SUPPLEMENTATION AGAINST OXIDATIVE STRESS DURING CRYOPRESERVATION OF HUMAN SPERMATOZOA.

BACKGROUND: Freeze damage is one of the most important factors which impair the membrane and DNA integrity of sperm cells. OBJECTIVE: The present study aims to investigate glutathione (GSH) supplementation on human spermatozoa cryopreservation. We determined sperm motility and viability, sperm lipid peroxidation, DNA damage, and the amount of hydrogen peroxide (H(2)O(2)) and superoxide (O(2)(-)). MATERIALS AND METHODS: Twenty pooled semen samples were freeze with 5mM GSH for 10 minute (test) and without GSH as control and stored in liquid nitrogen. RESULTS: After thawing, cryovials supplemented with 5mM GSH led to higher sperm viability compared with control samples (p < 0.05). Furthermore, the addition of 5mM GSH decreased sperm lipid peroxidation, DNA fragmentation, and H(2)O(2) and O(2)(-) content compared with controls (p < 0.05). CONCLUSION: GSH can be a good free radical scavenger in the freezing media and can support function of sperm cell after a cycle of freezing and thawing.

Estrogen and progesterone receptor subtype expression in granulosa cells from women with polycystic ovary syndrome.

We evaluated gene expression of estrogen and progesterone nuclear receptors in granulosa cells (GCs) of polycystic ovary syndrome (PCOS) women compared to women with normal cycling ovaries (control group) to achieve a better understanding of ovarian steroid status in patients with PCOS. In this prospective study, 40 patients with PCOS and 40 women with normal ovulatory function who underwent in vitro fertilization (IVF) for treatment of tubal and/or male infertility were recruited. Follicular fluid was collected from patients and GCs were isolated from follicular fluid and then were purified with Micro Beads conjugated to monoclonal anti-human CD45 antibodies. RNA was extracted and reverse transcription was performed. Gene expression of estrogen and progesterone receptors was determined by quantitative real time PCR (qRT-PCR). Estrogen receptor ß (ERß) expression was significantly higher than ERα expression in both groups (p < 0.002). ERα and ERß mRNA expression in PCOS was significantly lower than control group (p < 0.002). The expression levels of PRA and PRB in PCOS was significantly lower than control group (p < 0.002). In conclusion, a significant reduction of these genes in GCs from follicles of women with PCOS could be considered as a sign for maturation defect or follicular arrest in GCs.

Comparing Viability of Periodontal Ligament Stem Cells Isolated From Erupted and Impacted Tooth Root.

PURPOSE: The aim of the study was to compare the viability of periodontal ligament-derived stem/progenitor cells (PDLSCs) from 2 different sources. MATERIALS AND METHODS: Periodontal ligament (PDL) tissue was obtained from 20 surgically extracted human third molars and 20 healthy premolars extracted for orthodontic reasons. Periodontal ligament-derived stem/progenitor cells were isolated from 2 different PDL tissue sources and characterized by colony forming unit assay, cell surface marker characterizations, and their osteogenic differentiation potential. To determine cell viability within 2 groups, the colorimetric 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl tetrazolium bromide (MTT) metabolic activity assay was used. Data were statistically analyzed using independent t-test by SPSS 16 software (SPSS Inc, Chicago, IL). RESULTS: According to the MTT assay, the mean viability rate ± standard deviation of PDLSCs in the impacted third molar sample cells was 0.355 ± 0.411 and for erupted premolar sample cells was 0.331 ± 0.556. Based on One-Sample Kolmogorov-Smirnov test, P value for impacted and erupted teeth was 0.954 and 0.863, respectively. No statistical difference was seen between 2 groups. (P value > 0.05) CONCLUSIONS: Our results demonstrated that if surgical aseptic technique is a method employed to maintain asepsis, PDLSCs obtained from impacted and erupted tooth root would have the same viability rate.

Minocycline effects on memory and learning impairment in the beta-amyloid-induced Alzheimer's disease model in male rats using behavioral, biochemical, and histological methods.

Alzheimer's disease (AD), as an advanced neurodegenerative disease, is characterized by the everlasting impairment of memory, which is determined by hyperphosphorylation of intracellular Tau protein and accumulation of beta-amyloid (Aß) in the extracellular space. Minocycline is an antioxidant with neuroprotective effects that can freely cross the blood-brain barrier (BBB). This study investigated the effect of minocycline on the changes in learning and memory functions, activities of blood serum antioxidant enzymes, neuronal loss, and the number of Aß plaques after AD induced by Aß in male rats. Healthy adult male Wistar rats (200-220g) were divided randomly into 11 groups (n = 10). The rats received minocycline (50 and 100 mg/kg/day; per os (P.O.)) before, after, and before/after AD induction for 30 days. At the end of the treatment course, behavioral performance was measured by standardized behavioral paradigms. Subsequently, brain samples and blood serum were collected for histological and biochemical analysis. The results indicated that Aß injection impaired learning and memory performances in the Morris water maze test, reduced exploratory/locomotor activities in the open field test, and enhanced anxiety-like behavior in the elevated plus maze. The behavioral deficits were accompanied by hippocampal oxidative stress (decreased glutathione (GSH) peroxidase enzyme activity and increased malondialdehyde (MDA) levels in the brain (hippocampus) tissue), increased number of Aß plaques, and neuronal loss in the hippocampus evidenced by Thioflavin S and H&E staining, respectively. Minocycline improved anxiety-like behavior, recovered Aß-induced learning and memory deficits, increased GSH and decreased MDA levels, and prevented neuronal loss and the accumulation of Aß plaques. Our results demonstrated that minocycline has neuroprotective effects and can reduce memory dysfunction, which are due to its antioxidant and anti-apoptotic effects.

The effect of minocycline on beta-amyloid-induced memory and learning deficit in male rats: A behavioral, biochemical, and histological study.

OBJECTIVES: Minocycline hydrochloride is a semi-synthetic, second-generation tetracycline with neuroprotective, neurorestorative, anti-amyloidogenic, anti-inflammatory, antioxidant, and anti-apoptotic properties. The present study was designed to investigate the potential protective effects of minocycline against beta-amyloid (Aß)-induced Alzheimer's disease (AD), recognition memory decline, and the possible involved anti-apoptotic mechanisms. METHODS: The rats were treated with minocycline (50 and 100 mg/kg/day; P.O.) after AD induction for 30 days. Behavioral functions were assessed by employing standard behavioral tests, including novel object recognition (NOR) and passive avoidance learning (PAL) tasks. Then, total antioxidant capacity (TAC) and total oxidant status (TOS) were measured in blood serum using ELISA kits. Apoptosis and the number of Aß plaques were examined by the TUNEL and Congo red staining, respectively. RESULTS: Treatment of Aß rats with minocycline improved memory deficit in the PAL task and a decline in recognition memory in the NOR test. Minocycline at 50 and 100 mg/kg significantly reduced the TOS levels and increased the TAC levels (P < 0.0001). Also, minocycline at 50 and 100 mg/kg reduced the apoptotic index in the hippocampus of Aß rats. After Congo red staining, the minocycline group showed improved cell morphology and markedly fewer Aß plaques. CONCLUSIONS: Minocycline reduced memory and learning deficit in behavioral experiments after Aß injection, which may be due to its anti-inflammatory and anti-apoptotic effects.

In vitro production of mouse morphological sperm in artificial testis bioengineered by 3D printing of extracellular matrix.

Since autologous stem cell transplantation is prone to cancer recurrence, in vitro sperm production is regarded a safer approach to fertility preservation. In this study, the spermatogenesis process on testicular tissue extracellular matrix (T-ECM)-derived printing structure was evaluated. Ram testicular tissue was decellularized using a hypertonic solution containing triton and the extracted ECM was used as a bio-ink to print an artificial testis. Following cell adhesion and viability examination, pre-meiotic and post-meiotic cells in the study groups (as testicular suspension and co-culture with Sertoli cells) were confirmed by real-time PCR, flow-cytometry and immunocytochemistry methods. Morphology of differentiated cells was evaluated using transmission electron microscopy (TEM), toluidine blue, Giemsa, and hematoxylin and eosin (H&E) staining. The functionality of Leydig and Sertoli cells was determined by their ability for hormone secretion. The decellularization of testicular tissue fragments was successful and had efficiently removed the cellular debris and preserved the ECM compounds. High cell viability, colonization, and increased expression of pre-meiotic markers in cultured testicular cells on T-ECM-enriched scaffolds confirmed their proliferation. Furthermore, the inoculation of neonatal mouse testicular cells onto T-ECM-enriched scaffolds resulted in the generation of sperm. Morphology evaluation showed that the structure of these cells was quite similar to mature sperm with a specialized tail structure. The hormonal analysis also confirmed production and secretion of testosterone and inhibin B by Leydig and Sertoli cells. T-ECM printed artificial testis is a future milestone that promises for enhancing germ cell maintenance and differentiation, toxicology studies, and fertility restoration to pave the way for new human infertility treatments in the future.

The Association of Mitochondrial Translocator Protein and Voltage-Dependent Anion Channel-1 in Granulosa Cells with Estradiol Levels and Presence of Immature Follicles in Polycystic Ovary Syndrome.

Background: Granulosa cells (GCs) play key roles in oocyte maturation by providing required estradiol (E2). Since the presence of immature oocytes has been reported in cases with polycystic ovary syndrome (PCOS), in this study, the levels of mitochondrial membrane transporter proteins involved in E2 synthesis were determined. E2 concentration and parameters of oxidative status were also measured in follicular fluids of PCOS women. Methods: Forty-three women with PCOS and 43 healthy women who were candidates for IVF procedure due to their husbands' infertility were enrolled in this case-control study. The gene expression and protein levels of mitochondrial translocator protein (TSPO) and voltage-dependent anion channel 1 (VDAC1) were determined in GCs using RT-qPCR and immunocytochemistry assay, respectively. E2 level was measured with electrochemiluminescence, whereas total cholesterol, total antioxidant capacity (TAC), total oxidant status (TOS), and malondialdehyde (MDA) were determined using colorimetric methods in follicular fluids. Data were analyzed using unpaired t-test or Mann-Whitney U test, and Spearman's correlation coefficient. Results: VDAC1 and TSPO were significantly lower in mRNA (p<0.05) and protein levels (p<0.001) of PCOS patients. PCOS patients had lower cholesterol, estradiol, and TAC levels, and higher TOS and MDA contents. E2 level had direct correlation with VDAC1, TSPO, and TAC while it was negatively correlated with TOS, oxidative stress index (OSI), and MDA (p<0.001). Higher E2 levels were associated with higher numbers of high-quality oocytes and conceived embryos (p<0.001). Conclusion: Decreased E2 levels and increased oxidative stress in the follicular fluid may be the cause of immature oocytes in PCOS cases.