The effect of biological mechanisms of melatonin on the proliferation of spermatogonial stem cells: a systematic review.

In the last decade, melatonin has gained recognition as a potent scavenger and an effective antioxidant capable of neutralizing free radicals, including reactive oxygen species. Additionally, it exhibits anti-apoptotic properties. In this review, we will examine a compilation of articles that explore the cellular signaling function of melatonin on spermatogonial stem cells (SSCs) and adjacent cells such as Sertoli and Leydig cells. These cells play a crucial role in the proliferation of SSCs both in vitro and in vivo. In this review, we analyze the function of melatonin in the proliferation of SSCs from other aspects. For this purpose, we examine the articles based on the presence of melatonin on SSCs in four groups: As a supplement in SSCs medium culture, SSCs three-dimensional culture system, SSCs freezing medium, and as a therapeutic factor in vivo. Mechanisms of growth and proliferation of SSCs were considered. The purpose of this study is to investigate the potential effects of melatonin as a powerful antioxidant or growth stimulant for SSCs, both in vivo and in vitro.

Peripheral blood mononuclear cells expression of miR-200c, miR-125b, miR-27b, miR-203, and miR-155 in patients with significant or insignificant coronary artery stenosis.

Coronary artery disease (CAD) is one of the principal causes of death worldwide. Among several predisposing factors, inflammation and inflammatory genes play a significant role in disease pathogenesis. Inflammatory microRNAs, small noncoding RNAs involved in regulating inflammation, are promising candidates for understanding pathogenesis of CAD and developing diagnostic biomarkers. The aim of the study was to evaluate the alteration of miR-200c, miR-125b, miR-27b, miR-203 and, miR-155 in patients suffering from coronary artery stenosis and insignificant coronary artery stenosis compared to healthy subjects. In this study we compared expressions of five inflammatory miRNAs in peripheral blood mononuclear cells (PBMCs) of 72 patients suffering significant coronary artery stenosis (CAD), 74 individuals without coronary artery disease and 30 individuals with insignificant coronary artery stenosis (ICAD). After blood collection, PBMCs were isolated and RNA was extracted. Gene expression levels were assessed by SYBR green based real-time PCR. Statistical analysis was performed using R program. Expression levels of miR-200c, miR-203, and miR-155 were lower in subjects with ICAD than that in CAD patients and subjects of the control group. MiR-125b was downregulated in CAD and ICAD groups compared to the control group. PBMC miR-27b was upregulated in the CAD group as compared to the ICAD and control groups. Receiver operating characteristic curve analysis verified potential of three miRNAs in separating subjects with ICAD from CAD patients and healthy individuals. In conclusion, this original investigation suggested that altered expression of these five miRNAs may serve as a novel diagnostic biomarker discriminating clinical presentations of coronary artery diseases.

Assessment of developmental rate of mouse embryos yielded from in vitro fertilization of the oocyte with treatment of melatonin and vitamin C simultaneously.

BACKGROUND: In recent decades, in vitro fertilization (IVF) has been widely used as a method of assisted reproductive technology (ART) to improve fertility in individuals. To be more successful in this laboratory method, we used the presence of two common types of antioxidants (melatonin and vitamin C) simultaneously and exclusively in IVF medium. METHODS: The cumulus-oocyte complexes (COCs) were obtained from Gonadotropin-releasing hormone (GnRH) and Human Chorionic Gonadotropin (HMG) -stimulated mice. Subsequently, metaphase II (MII) oocytes were fertilized in vitro. In the experiment, the IVF medium was randomly divided into two equal groups: The control group did not receive any antioxidants. In the treatment group, 100 µM melatonin and 5 mM vitamin C were added to the IVF medium. Finally, oocytes and putative embryos transferred into developmental medium and cultured 120 h after IVF to the blastocyst stage. After and before IVF, oocytes and putative embryos were stained with dichlorodihydrofluorescein diacetate (DCFDA) and the H2O2 level was measured with an inverted fluorescence microscope using ImageJ software. At the end of the fifth day after IVF, the expression of Bax and B cell lymphoma 2 (Bcl2) was evaluated using real-time PCR. RESULTS: The levels of reactive oxygen species (ROS) in oocytes and putative embryos observed in the treatment group demonstrated a significant reduce compared to the control group (p ≤ 0.01. (.Furthermore, the number of embryos in the blastocycte stage(P < 0.05), the expression level of the Bcl2 (P < 0.05) gene, the Bax unlike gene, significantly increased compared with the control group. CONCLUSION: We conclude that the presence of melatonin and vitamin C antioxidants simultaneously and exclusively in the IVF medium leads to a reduction in ROS and ,as a result, improves the growth of the embryo up to the blastocyst stage.

Relapse of immune thrombocytopenia after receiving AstraZeneca coronavirus disease-2019 vaccine: A case report.

Immune thrombocytopenic purpura (ITP) is an autoimmune disease characterized by a low platelets count. In this paper, we present a case of ITP relapse in a 31-year-old Iranian woman as a potential complication of the AstraZeneca vaccine.

Epi-miRNAs: Regulators of the Histone Modification Machinery in Human Cancer.

Cancer is a leading cause of death and disability worldwide. Epigenetic deregulation is one of the most critical mechanisms in carcinogenesis and can be classified into effects on DNA methylation and histone modification. MicroRNAs are small noncoding RNAs involved in fine-tuning their target genes after transcription. Various microRNAs control the expression of histone modifiers and are involved in a variety of cancers. Therefore, overexpression or downregulation of microRNAs can alter cell fate and cause malignancies. In this review, we discuss the role of microRNAs in regulating the histone modification machinery in various cancers, with a focus on the histone-modifying enzymes such as acetylases, deacetylases, methyltransferases, demethylases, kinases, phosphatases, desumoylases, ubiquitinases, and deubiquitinases. Understanding of microRNA-related aberrations underlying histone modifiers in pathogenesis of different cancers can help identify novel therapeutic targets or early detection approaches that allow better management of patients or monitoring of treatment response.

Orbital Lateral Cantal Distance Can Predict the Cephalometric Characteristic of Mandible in Iranian Population.

ABSTRACT: Orbitofacial anthropometrics have become an important tool used in reconstructive surgery. The authors attempt to evaluate the relation between orbital lateral canthal distance and the cephalometric characteristic of mandible in Iranian population.In a cross-sectional study, anthropometric parameters of face in 200 subjects (100 males and 100 females) with mean age of 34.39 ±â€Š18.83 were evaluated by three-dimensional computed tomography imaging.In this study, there was not a significant difference in the age of sex groups (P = 0.183). Also, there was no significant difference in the left and right mandible angle in different sex groups (P = 0.25, P  =  0.124, respectively). There were significant differences in the anterior mandible distance, inferior mandible angle distance (P = 0.0001) and lateral cantus distance of sex groups (P = 0.0001). There was a significant correlation between lateral contuse distance and left mandible angle (r = 0.226, P = 0.001), right mandible angle (r = 0.283, P = 0.00), mandible angle (r = -0.266, P = 0.00), anterior mandible angle distance (r = 0.655, P = 0.00), and inferior mandible angle distance (r = 0.582, P = 0.00).Here, we conclude that orbital lateral canthal distance can predict the cephalometric characteristic of mandible in Iranian population.

The Therapeutic Potential of Conditioned Medium from Human Breast Milk Stem Cells in Treating Spinal Cord Injury.

STUDY DESIGN: Experimental animal study. PURPOSE: This study investigated the therapeutic effects of human breast milk stem cell (BMSC)-conditioned medium (BMSC-CM) in a model of spinal cord injury (SCI) in male Sprague-Dawley rats. OVERVIEW OF LITERATURE: SCI is one of the leading causes of disability in addition to sensory and motor impairment. So far, there have been no successful treatments for SCI. Given the positive outcomes associated with using stem cells and their derivatives as a treatment for various diseases, there is a growing interest in using them as an SCI treatment. Recent research has demonstrated that CM from stem cells has therapeutic advantages. METHODS: Human BMSCs were isolated and characterized, and CM was subsequently collected. Animals received an intrathecal administration of BMSC-CM after SCI. The activity of caspase-3 was measured to assess apoptosis, and levels of tumor necrosis factor-α and interleukin-1ß were measured to assess inflammation. Also, sensory and locomotor performances were assessed after SCI and BMSC-CM administration. RESULTS: Administration of CM from BMSC reduced apoptosis and inflammation at the site of injury in a rat model of SCI (p<0.05). Motor, sensory, locomotor, and sensorimotor performances were significantly improved in rats that received BMSC-CM after SCI. CONCLUSIONS: Intrathecal administration of BMSC-CM improved recovery in a rat model of SCI.

Human Menstrual Blood Stem Cell-Derived Granulosa Cells Participate in Ovarian Follicle Formation in a Rat Model of Premature Ovarian Failure In Vivo.

We recently reported the application of human menstrual blood stem cells' (HuMenSCs) transplantation as a treatment modality in a rat model of premature ovarian failure (POF). We continued to investigate further in this respect. Female rats were injected intraperitoneally with 36 mg/kg busulfan. HuMenSCs were obtained, grown, and analyzed for immunophenotypic features at passage three. The cells were labeled with CM-Dil and infused into the rats. There were four groups: normal, negative control, treatment, and Sham. One month after treatment, the ovaries were collected and weighed. Histological sections were prepared from the ovary and HuMenSCs were tracking. Subsequently, we examined the changes of expression of Bax and B cell lymphoma 2 (Bcl2) genes by real-time polymerase chain reaction assay. One month after HuMenSCs transplantation, these cells were located in the ovarian interstitium and granulosa cells (GCs). The number of TUNEL-positive cells significantly decreased in the treatment group. Also the expression level of Bax genes, unlike Bcl2 gene, significantly decreased compared with negative and sham groups. In our study, HuMenSCs were tracked in ovarian tissues within 2 months after transplantation, and they differentiated into GCs. Therefore, the use of these cells can be a practical and low-cost method for the treatment of POF patients.

Proliferation and differentiation of mouse spermatogonial stem cells on a three-dimensional surface composed of PCL/gel nanofibers / Proliferación y diferenciación de células madre espermatogónicas de ratón en una superficie tridimensional compuesta de nanofibras PCL / gel

Spermatogonial stem cells (SSCs) have self-renewal and differentiation capacity essential for sperm production throughout the male reproductive life. The electrospun polycaprolactone/gelatin (PCL/Gel) nanofibrous scaffold mimics important features of the extracellular matrix (ECM), which can provide a promising technique for the proliferation and differentiation of SSCs in vitro. The goal of the present study was to investigate the effects of PCL/Gel nanofibrous scaffold on the propagation and differentiation of neonate mouse SSCs (mSSCs). mSSCs were enzymatically isolated, and the cells were purified by differential plating method and seeded on scaffold. After 2 weeks, viability, colony number and diameter, and expression of specific spermatogonial cell genes were investigated. After mSSCs propagation, the cells were cultivated in a differentiation medium on the scaffold for another 2 weeks, and differentiating cells were analyzed by real-time PCR. The number of mSSC colony (P<0.01) and expression levels of specific spermatogonial genes Plzf and Inga6 (P<0.01) and also differentiation genes c-Kit, Tp1 and Ptm1 (P<0.05) were higher in scaffold group compared with control during the culture period. We conclude that mSSCs can be expanded and can differentiate toward spermatid cells on PCL/Gel nanofibrous scaffold with improved developmental parameters.

Las células madre espermatogónicas (CME) tienen capacidad de auto renovación y diferenciación esenciales para la producción de esperma a lo largo de la vida reproductiva masculina. El «scaffold¼ nanofibroso de policaprolactona / gelatina (PCL / Gel) electrohilado imita características importantes de la matriz extracelular (MEC), que puede proporcionar una técnica prometedora para la proliferación y diferenciación de CME in vitro. El objetivo del presente estudio fue investigar los efectos del «scaffold¼ nanofibroso PCL / Gel en la propagación y diferenciación de CME de ratones neonatos (mSSC). Los mSSC se aislaron enzimáticamente y las células se purificaron mediante un método de siembra diferencial y se sembraron en un «scaffold¼. Después de 2 semanas, se investigaron la viabilidad, el número y el diámetro de las colonias y la expresión de genes específicos de células espermatogónicas. Después de la propagación de mSSC, las células se cultivaron en un medio de diferenciación en el «scaffold¼ durante otras 2 semanas, y las células se analizaron mediante PCR en tiempo real. El número de colonias mSSC (P <0,01) y los niveles de expresión de los genes espermatogónicos específicos Plzf e Inga6 (P <0,01) y también los genes de diferenciación c-Kit, Tp1 y Ptm1 (P <0,05) fueron mayores en el grupo de «scaffold¼ en comparación con el control durante el período de cultivo. Concluimos que los mSSC pueden expandirse y diferenciarse en células espermátidas en un «scaffold¼ de nanofibras PCL / Gel con parámetros de desarrollo mejorados.

The effects of human menstrual blood stem cells-derived granulosa cells on ovarian follicle formation in a rat model of premature ovarian failure.

Many studies have reported that human endometrial mesenchymal stem cells (HuMenSCs) are capable of repairing damaged tissues. The aim of the present study was to investigate the effects of HuMenSCs transplantation as a treatment modality in premature ovarian failure (POF) associated with chemotherapy-induced ovarian damage. HuMenSCs were isolated from menstrual blood samples of five women. After the in vitro culture of HuMenSCs, purity of the cells was assessed by cytometry using CD44, CD90, CD34, and CD45 FITC conjugate antibody. Twenty-four female Wistar rats were randomly divided into four groups: negative control, positive control, sham, and treatment groups. The rat models of POF used in our study were established by injecting busulfan intraperitoneally into the rats during the first estrus cycle. HuMenSCs were transplanted by injection via the tail vein into the POF-induced rats. Four weeks after POF induction, ovaries were collected and the levels of Amh, Fst, and Fshr expression in the granulosa cell (GC) layer, as well as plasma estradiol (E2) and progesterone (P4) levels were evaluated. Moreover, migration and localization of DiI-labeled HuMenSCs were detected, and the labeled cells were found to be localized in GCs layer of immature follicles. In addition to DiI-labelled HuMenSCs tracking, increased levels of expression of Amh and Fshr and Fst, and the high plasma levels of E2 and P4 confirmed that HuMenSC transplantation had a significant effect on follicle formation and ovulation in the treatment group compared with the negative control (POF) group.

Colonization of Mouse Spermatogonial Cells in Modified Soft Agar Culture System Utilizing Nanofibrous Scaffold: A New Approach.

BACKGROUND: Spermatogonial stem cells (SSCs) are considered in fertility management approaches of prepubertal boys facing cancer therapies. However, in vitro propagation has become an important issue due to a small number of SSCs in testicular tissue. The present study aimed to investigate a modified soft agar culture system by using a nanofibrous scaffold as a new approach to mimic in vivo conditions of SSCs development. MATERIALS AND METHODS: The SSCs were isolated from neonate mouse testes, cultured on polycaprolactone scaffold, and covered by a layer of soft agar for 2 weeks. Then, the number and diameter of colonies formed in experimental groups were measured and spermatogonial markers (i.e., Plzf, Gfrα1, Id4, and c-Kit) in SSCs colonies were evaluated by a real-time polymerase chain reaction and immunostaining. RESULTS: Our results indicated that the colonization rate of SSCs was significantly higher in the present modified soft agar culture system (P<0.05). Only Plzf indicated a significant increased at the levels (P<0.05), the gene expression levels of Id4, Plzf, and Gfrα1 were higher in the present culture system. In addition, the expression of the c-Kit gene as a differentiating spermatogonia marker was higher in presence of scaffold and soft agar compared with the amount of other experimental groups (P<0.05). CONCLUSION: The culture system by using nanofibrous scaffold and soft agar as a new culture method suggests the potential of this approach in SSCs enrichment and differentiation strategies for male infertility treatments, as well as in vitro spermatogenesis.

Efficiency of colony formation and differentiation of human spermatogenic cells in two different culture systems.

Optimization of in vitro culture system for the expansion and the maturation of male germ cells to post meiotic stages is a valuable tool for studies exploring spermatogenesis regulation and the management of male infertility. Several studies have reported promising results of mouse spermatogonial stem cells culture in three-dimensional (3D) culture systems and a subsequent production of sperm. In the present study, we investigated the capacity of a three-dimensional soft agar culture system (SACS) supplemented with Knockout Serum Replacement (KSR) in colony formation and inducing human germ cells to reach post-meiotic stages. Testicular cells from testes of brain -dead donors were first cultured for three weeks in proliferation medium. The cells were subsequently cultured in the upper layer of the SACS (3D group) in a medium supplemented with KSR and hormones, and the results were compared with that of a two-dimensional (2D) culture system. We found that the number and diameter of colonies and the levels of expression of Scp3 and Integrin α6 in the 3D culture group were significantly higher than in the 2D group. Our findings indicate that SACS can reconstruct a microenvironment capable of regulating both proliferation and differentiation of cell colonies.

The effects of melatonin on colonization of neonate spermatogonial mouse stem cells in a three-dimensional soft agar culture system.

BACKGROUND: Melatonin is a pleiotropic hormone with powerful antioxidant activity both in vivo and in vitro. The present study aimed to investigate the effects of melatonin on the proliferation efficiency of neonatal mouse spermatogonial stem cells (SSCs) using a three-dimensional soft agar culture system (SACS) which has the capacity to induce development of SSCs similar to in vivo conditions. METHODS: SSCs were isolated from testes of neonate mice and their purities were assessed by flow cytometry using PLZF antibody. Isolated testicular cells were cultured in the upper layer of the SACS in αMEM medium in the absence or presence of melatonin extract for 4 weeks. RESULTS: The identity of colonies was confirmed by alkaline phosphatase staining and immunocytochemistry using PLZF and α6 integrin antibodies. The number and diameter of colonies of SSCs in the upper layer were evaluated at days 14 and 28 of culture. The number and diameter of colonies of SSCs were significantly higher in the melatonin group compared with the control group. The levels of expression of ID-4 and Plzf, unlike c-kit, were significantly higher in the melatonin group than in the control group. CONCLUSIONS: Results of the present study show that supplementation of the culture medium (SACS) with 100 µM melatonin significantly decreased reactive oxygen species (ROS) production in the treated group compared with the control group, and increased SSC proliferation.

In vitro effects of melatonin on colonization of neonate mouse spermatogonial stem cells.

We have recently reported that antioxidant supplements enhance the efficacy of cryopreserved spermatogonial stem cells. Melatonin is considered a free radical scavenger which has direct and indirect antioxidant effects in in vitro and in vivo microenvironments. Due to the anti-apoptotic properties of melatonin, researchers have proposed that melatonin may improve the efficiency of spermatogonial stem cell (SSC) transplantation. However, the appropriate methodology which facilitates SSC proliferation remains to be determined. Identification of a proper propagation system is essential for the future application of SSCs in the field of infertility. The aim of the present study was to investigate the effects of melatonin on the colonization of SSCs. SSCs were isolated from the testes of three to six day old mice, and their purities were assessed by cytometry using Plzf antibody. Isolated testicular cells were cultured in the absence or presence of melatonin extract for two weeks. Suppression of differentiation and maintenance of spermatogonial stem cells was confirmed by alkaline phosphatase staining and immunocytochemistry using Plzf antibody. The number and diameter of the colonies of SSCs were assessed during the 7th and 14th days of culture, and the expression of Id4, Plzf, and C-kit were evaluated using real-time PCR at the end of the culture period. The survival rate of the cultured cells in the presence of melatonin was significantly higher than the control group. The number and diameter of colonies also increased in the cells treated with melatonin. The results of our study suggest that culture of SSCs with 100 µM melatonin supplementation can increase SSCs proliferation significantly.

Comparison of Human Amniotic, Chorionic, and Umbilical Cord Multipotent Mesenchymal Stem Cells Regarding Their Capacity for Differentiation Toward Female Germ Cells.

Placenta harbors a plentiful source of various cells with stem cells or stem-like cell properties, which can be used in therapeutic procedures and research. Mesenchymal stem cells (MSCs) have attracted much attention due to their specific differentiation potential and tolerogenic properties. MSCs have been isolated from different parts of placenta; however, in this study, we isolated MSCs from amnion and chorion membrane, as well as umbilical cord (Wharton's jelly [WJ]) and compared their capacity regarding differentiation toward female germ cells under influence of 10 ng/mL BMP4. All placenta samples were collected from delivering mothers by normal cesarean section and cells were isolated by different methods. Results showed that all isolated cells were mostly positive for the MSC markers CD73, CD166, and CD105, and minimally reacted with CD34 and CD45 (hematopoietic markers). After differentiation induction using third passage cultured cells, immunocytochemistry staining showed that cells were positive for germline cell-related genes Ssea4, Oct4, and Ddx4, and oocyte-related gene Gdf9. RT-qPCR results indicated that human chorion MSCs (hCMSCs) had a greater potential to be differentiated into female germline cells. Moreover, the results of this study indicate that human umbilical cord MSCs originated from either male or female umbilical cord have the same differentiation potential into female germline cells. We recommend that for presumptive application of MSCs for infertility treatment and research, hUMSCs are best candidates due to their higher differentiation potential, ease of proliferation and expansion, and low immunogenicity.

The effect of aminoguanidine on sperm motility and mitochondrial membrane potential in varicocelized rats.

OBJECTIVES: Increased levels of nitric oxide (NO) in the testicular veins of people suffering from varicocele have already been reported. However, the role of NO-synthase (NOS) isozymes and their inhibitors have not been extensively studied. We aimed to evaluate the inhibitory effects of aminoguanidine (AG), on sperm motility, vitality, and mitochondrial membrane potential (MMP) in varicocelized rats. MATERIALS AND METHODS: Twenty fore male Wister rats were divided into control, sham, varicocele, and treatment groups. Varicocele and treatment groups underwent partial ligation of left renal vein. Rats in the sham group underwent the same procedures as the varicocele group with the exception of vein ligation. 10 weeks after varicocele induction, sperm parameters were evaluated in all groups. The treatment group received 50 mg/kg AG injection daily for 10 weeks after which they were sacrificed prior to assessment of the parameters. Sperm viability and MMP were assessed by flow cytometry using propidium iodide (PI) and rhodamine 123 (Rh123), respectively. RESULTS: The results of this study show a decrease in sperm viability, motility and MMP in the varicocele group compared with the other groups. After AG injection, we observed that all the parameters were significantly enhanced in the treatment group compared with the other groups. Rh123 staining revealed a positive relation between MMP and sperm motility, whereas PI staining showed a positive relation between sperm motility and viability. CONCLUSION: The findings of our study show that AG improves sperm motility and MMP, and thus, might be useful in the management of varicocele-related infertility.