Effects of melatonin and human follicular fluid supplementation of in vitro maturation medium on mouse vitrified germinal vesicle oocytes: A laboratory study

Background: Vitrification as the most efficient method of cryopreservation, enables successful storage of oocytes for couples who undergo specific procedures including surgery and chemotherapy. However, the efficacy of in vitro maturation (IVM) methods with vitrified germinal vesicle (GV) oocytes could be improved. Objective: As melatonin and follicular fluid (FF) might enhance IVM conditions, we used these supplements to assess the maturation rate of vitrified GV oocytes and their artificial fertilization rate. Materials and Methods: Four hundred mouse GV oocytes were harvested, vitrified, and assigned into control (C-Vit-GV) and treatment groups of melatonin (M-Vit-GV), human follicular fluid (HFF-Vit-GV), and a combination (M + HFF-Vit-GV). A non-vitrified group of GV oocytes (non-Vit-GV) and a group of in vivo matured metaphase II (VivoMII) oocytes served as control groups to evaluate the vitrification and IVM conditions, respectively. Maturation of GV oocytes to MII and further development to two-cell-stage embryos were determined in the different groups. Results: Development to two-cell embryos was comparable between the Vivo-MII and non-Vit-GV groups. IVM and in vitro fertilization (IVF) results in the non-Vit-GV group were also comparable with the C-Vit-GV oocytes. In addition, the IVM and IVF outcomes were similar across the different treatment groups including the M-Vit-GV, HFF-Vit-GV, M + HFF-Vit-GV, and C-Vit-GV oocytes. Conclusion: Employing an appropriate technique of vitrification followed by suitable IVM conditions can lead to reasonable IVF outcomes which may not benefit from extra supplementations. However, whether utilizing other supplementation formulas could improve the outcome requires further investigation. Key words: Vitrification, Germinal vesicle, In vitro oocyte maturation, Melatonin, Follicular fluid

Effect of embryonic fibroblast cell co-culture on development of mouse embryos following exposure to visible light

Purpose To determine the effects of visible light on development of mouse embryos and the potential of fibroblast cells to overcome deleterious effects of visible light on mouse preimplantation stage embryos. Methods Two-cell mouse embryos were randomly allocated to un-exposed group (control) and exposed group receiving 1600 lx visible light for various time lengths. Both exposed and un-exposed embryos were co-cultured with either Mouse Embryonic Fibroblast (MEF) or Human Embryonic Fibroblast (HEF). Developmental rate of embryos at day 3 (morula), 4 (expanded blastocyst) and 5 (hatching or hatched blastocyst) was evaluated. Results Exposure of embryos to visible light for 30 min decreased developmental rate significantly (P<0.01). Developmental rate of exposed embryos co-cultured with MEF (58%; p<0.05 both at day 4 and 5) and HEF (67%; P<0.01 both at day 4 and 5) was higher than control. Conclusions Visible light adversely affects embryo development in a time-dependent manner. Feeder cells may enhance embryo development particularly when suboptimal conditions are involved

Antioxidants rescue stressed embryos at a rate comparable with co-culturing of embryos with human umbilical cord mesenchymal cells

Purpose During laboratory manipulations, oocytes and embryos are inevitably exposed to suboptimal conditions that interfere with the normal development of embryos. Materials and methods In this study, we examined the effects of antioxidants, feeder cells and a conditioned medium on embryo development and cleavage rate following exposure of the embryos to suboptimal conditions. We exposed mouse two-cell embryos to visible light and divided them into four groups: control (E-ctr), co-culture (Co-c), conditioned medium (Cndm) and antioxidant-plus medium (Aopm). We used human umbilical cord matrixderived mesenchymal cells for co-culture. A group of embryos was not exposed to visible light and served as the non-exposed control (NE-ctr) group. Results The developmental rate was higher in NE-ctr embryos than in the E-ctr group. Exposed embryos in the various groups showed a comparable developmental rate at different stages. Blastomere number significantly increased (P<0.05) in the Co-c and Aopm groups compared with the E-ctr and Cndm groups. No significant difference was observed between the Co-c and Aopm groups. Conclusions Our data indicate that in suboptimal conditions, antioxidants could improve the embryo cleavage rate in the same way as feeder cells. Antioxidants probably improve embryo quality through their ability to scavenge reactive oxygen species

Suspension Culture Alters Insulin Secretion in Induced Human Umbilical Cord Matrix-Derived Mesenchymal Cells

Objective: Worldwide, diabetes mellitus (DM) is an ever-increasing metabolic disorder. A promising approach to the treatment of DM is the implantation of insulin producing cells (IPC) that have been derived from various stem cells. Culture conditions play a pivotal role in the quality and quantity of the differentiated cells. In this experimental study, we have applied various culture conditions to differentiate human umbilical cord matrix-derived mesenchymal cells (hUCMs) into IPCs and measured insulin production. Materials and Methods: In this experimental study, we exposed hUCMs cells to pancreatic medium and differentiated them into IPCs in monolayer and suspension cultures. Pancreatic medium consisted of serum-free Dulbecco’s modified eagle’s medium Nutrient mixture F12 (DMEM/F12) medium with 17.5 mM glucose supplemented by 10 mM nicotinamide, 10 nM exendin-4, 10 nM pentagastrin, 100 pM hepatocyte growth factor, and B-27 serum-free supplement. After differentiation, insulin content was analyzed by gene expression, immunocytochemistry (IHC) and the chemiluminesence immunoassay (CLIA). Results: Reverse transcription-polymerase chain reaction (RT-PCR) showed efficient expressions of NKX2.2, PDX1 and INSULIN genes in both groups. IHC analysis showed higher expression of insulin protein in the hanging drop group, and CLIA revealed a significant higher insulin production in hanging drops compared with the monolayer group following the glucose challenge test. Conclusion: We showed by this novel, simple technique that the suspension culture played an important role in differentiation of hUCMs into IPC. This culture was more efficient than the conventional culture method commonly used in IPC differentiation and cultivation

The role of stem cells in tumor targeting and growth suppression of gliomas

Glioma remains the most challenging solid organ tumor to treat successfully. Based on the capacity of stem cells to migrate extensively and target invading glioma cells, the transplantation of stem cells as a cell-based delivery system may provide additional tools for the treatment of gliomas. In addition to the use of modified stem cells for the delivery of therapeutic agents, unmodified stem cells have been shown to have growth-suppressing effects on tumors in vitro and in vivo. This review outlines the probable factors involved in tumor tropism and tumor growth suppression, with a specific focus on the use of unmodified stem cells in the treatment of gliomas. Based on these and further future data, clinical trials may be justified

The role of co-culture systems on developmental competence of preimplantation mouse embryos against pH fluctuations

Purpose To determine the effect of pH fluctuations of culture media, and the role of co-culture systems on embryo development. Methods Mouse embryos were incubated in phosphate buffered solutions (PBSs) with different pH for various lengths of time. After 3 h incubation of embryos at various pH, the embryos were transferred into four media with human (HEF) and mouse (MEF) embryonic fibroblast cells, and without feeder cells; HTF and MEM-α. Developmental rate at day three (morula), four (expanded blastocyst) and five (hatching or hatched blastocyst) was evaluated. Results Developmental rate at day three, four and five decreased when the incubation time at pH 6.2 and 8 increased to 3 h and more. In addition, significantly less embryos incubated at pH 6.2 and 8 developed to hatching and hatched blastocysts compared with pH 7.35. Embryos incubated at pH 6.2, co-cultured with MEF or HEF showed a significant improvement (P<0.05) at day three in HEF compared to HTF, and at day five in MEF compared to HTF. At pH 8, a significant improvement (P<0.05) was observed at day five in HEF and MEF compared to MEM-α. Conclusions Mouse 2-cell embryos could tolerate minor pH fluctuations, but that major pH changes affect subsequent development. Besides, feeder cells could improve embryo development, especially when embryos are prone to rise or fall in pH

Cationic b-Cyclodextrin–Chitosan Conjugates as Potential Carrier for pmCherry-C1 Gene Delivery

In this study cationic b-cyclodextrin-chitosanmediated nanoparticles were used to transfer pmCherry-C1 into glioblastoma cells and their transfection efficiency were compared to lipofectamine and electroporation. Physicochemical characteristics of nanoparticles were evaluated by photon correlation spectroscopy and scanning electron microscopy. Electrophoretic nuclease resistance and stability assays were used to check the protection of DNA from nucleases digestion. mCherry reporter construct was used for visualization, followed by quantitation of cell survival and gene expression by fluorescence-activated cell sorting analysis and fluorescence microscopy. Particle size was approximately 200 nm and did not change at 4 �C even after 12 weeks. Importantly, the positively charged complexes interacted with DNA could serve as an efficient DNA delivery systems. Most of the gene was associated with the nanoparticles and was efficiently protected from DNAse I digestion. More than 80 % of transfected cells expressed mCherry efficiently

Do Insulin Replacement and Omega3 Protect the Male Reproductive Function of the Streptozotocin-Induced Diabetic Mice?

Diabetes mellitus (DM), the most common metabolic disease, might affect different organs such as male reproductive system. Experiments have shown that n-3 fatty acids could improve male reproductive function. Present study was performed to examine the effects of omega3 on sperms and testicular parameters in diabetic mice. Adult NMRI male mice were randomly divided into intact and diabetic groups (n=8). Streptozotocin-induced diabetic animals were divided into 4 groups of diabetic-saline (Dia-Sa), diabetic-insulin (Dia-Ins), diabetic-omega3 (Dia-omg3), and diabetic-insulin-omega3 (Dia-Ins-omg3). Following confirmation of diabetes, different treatments including 3 U/100 g insulin subcutaneously and 400 mg/kg omega3 orally were administered, where applicable according to the treatment groups. Thirty-five days later, the sperm number, motility, progression, and normal morphology were determined. Also, testes diameters and structure including germinal epithelium thickness, seminiferous tubule diameters, Leydig cell number, and testosterone level were assessed. Sperm number, viability, fast motility, testes volume, and serum testosterone level decreased insignificantly in the Dia-Sa group compared with the intact animals. Neither insulin replacement nor omega3 administration could significantly improve the outcome. We might conclude that short periods of diabetes could not significantly affect the male reproductive function. In addition, insulin replacement and/or omega-3 supplementation does not have any profound effects on male reproductive system

Report an educational intense decline in medical student with excellent educational background

Educational decline among university students during the first university terms is generally observed as a reason of the big change in lifestyle and not being accustomed to dormitory atmosphere, or it can be as a result of being involved with emotional matters, due to the special age at which they enter university. This research concerns the situation of a medical student ((Mohammad)) who enters Kerman medical university with a perfect educational background and a top rank in entrance exam of university but he encounters a noticeable underachievement in educating as soon as he starts his courses at university. Mohammad who is from a relatively supportive family and has a suitable financial situation in his family, gets involved with an emotional case and its following problems and matters, facing a breakup at this stage , he is suffered with depression and severe educational decline . His family, in order to help him, acts mistakenly and it brings about bigger problems. Therefore, after finishing two university terms with conditional educational situation, he exposes with the warning of being dropped out of university. Noticing the importance of this stage of life among university students, more than the consideration of university teachers, needs the contribution of consulting center of university in an effective way. so that they can identify students with emotional conflicts that leads them toward educational underachievement, and then offering them suitable and helpful consultation aimed to help them pass this serious phase of life and moreover, how to adopt themselves with new situations, surely with having their family support as well. Key Words: Educational Decline, Medical Student, Consultatio

Clinical safety and primary efficacy of bone marrow mesenchymal cell transplantation in subacute spinal cord injured patients

Background: In recent years, some studies were conducted to evaluate the effects of stem cells from different sources on patients with spinal cord injury (SCI). This study was carried out to evaluate the feasibility and therapeutic potential of autologous bone marrow cell (BMC) transplantation in 11 complete spinal cord injured patients at thoracic level. Methods and materials: This nonrandomized clinical trial compared the results of autologous BMC transplantation into cerebrospinal fluid (CSF) via lumbar puncture (LP) in 11 patients having complete SCI, with 20 patients as control group who received conventional treatment without BMC transplantation. The patients underwent preoperative and follow-up neurological assessments using the American Spinal Injury Association (ASIA) impairment scale. Then, the participants were followed for 12–33 months. Results: Eleven patients with the mean age of 33.2 ± 8.9 years and 20 patients with the mean age of 33.5 ± 7.2 years were enrolled in the study and in the control group, respectively. None of the patients in the study and control group experienced any adverse reaction and complications, neither after routine treatment nor after cell transplantation. Five patients out of 11 (45.5%) in the study group and three patients in the control group (15%) showed marked recovery, but the result was statistically borderline (P = 0.095). Conclusion: We conclude that transplantation of autologous BMC via LP is a feasible and safe technique, but at the moment, no clear answer can be given regarding the clinical potential, despite a potential tendency to treat SCI patients, observed through statistics