Embryonic stem cell conditioned medium supports in vitro maturation of mouse oocytes

Background: This study aimed to investigate the maturation and fertilization rates of immature mouse oocytes using Embryonic Stem Cell Conditioned Medium [ESCM]

Methods: Germinal Vesicle [GV] stage oocytes were observed in 120 NMRI mice, aged 4-6 weeks. GV oocytes with or without cumulus cells were subjected to IVM in either ESCM, Embryonic Stem Cell Growth Medium [ESGM], or alpha-minimum essential medium [alpha-MEM]. After recording the Metaphase II [MII] oocyte maturation rate, the oocytes were fertilized in vitro. The fertilization success rate was recorded after 24 hr. The embryos were maintained in potassium Simplex Optimization Medium [KSOM] for 96 hr and allowed to grow until the blastocyst stage. After recording developmental competence, they were transferred into the uteri of pseudopregnant mice and their birth rates were recorded

Results: No significant difference existed between the maturation rates in alpha-MEM [68.18%] and ESCM [64.67%; p>0.05], whereas this rate was significantly higher for both alpha-MEM and ESCM compared to ESGM [32.22%; p<0.05]. A significant difference in IVF success rate existed for oocytes grown in alpha-MEM [69.44%], ESCM [61.53%], and ESGM [0%]. A significantly higher developmental competence was observed at the blastocyst stage for oocytes grown in alpha-MEM [51.2%] compared to ESCM [35%; p<0.05]. 17 days after embryo transfer into the uteri of pseudopregnant mice, there was a nonsignficant [p>0.05], similar birth rate between alpha-MEM and ESCM [47 vs. 40%]

Conclusion: ESCM is an effective medium for preantral follicle growth, oocyte maturation, and subsequent embryo development

Stromal vascular fraction combined with silicone rubber chamber improves sciatic nerve regeneration in diabetes / 中华创伤杂志(英文版)

<p><b>PURPOSE</b>To study the effects of transplantation of characterized uncultured stromal vascular fraction (SVF) on sciatic nerve regeneration.</p><p><b>METHODS</b>A 10-mm sciatic nerve defect was bridged using a silicone conduit filled with SVF. In control group, silicone conduit was filled with phosphate-buffered saline alone. In sham-operated group, the sciatic nerve was only exposed and manipulated. The regenerated nerve fibers were studied 8 and 12 weeks after surgery.</p><p><b>RESULTS</b>Behavioral and functional studies confirmed faster recovery of regenerated axons in SVF transplanted animals than in control group (p<0.05). Gastrocnemius muscle mass in SVF transplanted animal was found to be significantly more than that in control group. Morphometric indices of the regenerated fibers showed the number and diameter of the myelinated fibers to be significantly higher in SVF transplanted animals than in control group. In immunohistochemistry, the location of reactions to S- 100 in SVF transplanted animals was clearly more positive than that in control group.</p><p><b>CONCLUSION</b>SVF transplantation combined with silicone conduit could be considered as a readily accessible source of stromal cells that improves functional recovery of sciatic nerve. It may have clinical implications for the surgical management of acute diabetic patients after facial nerve transection.</p>

Effect of leptin on in vitro nuclear maturation and apoptosis of buffalo [bubalus bubalis] oocyte

Leptin, as a 16 kDa adipokine, is a pleiotropic cytokine-like hormone that primarily secreted from adipose tissue. It also involves in the regulation of energy homeostasis, neuroendocrine function, immunity, lipid and glucose homeostasis, fatty acid oxidation, angiogenesis, puberty and reproduction. The aim of this study was to investigate the effects of in vitro addition of leptin to in vitro maturation [IVM] medium on buffalo oocyte maturation and apoptosis. In this experimental study, Ovaries from apparently normal reproductive organs of slaughtered adult buffaloes [Bubalus bubalis] with unknown breeding history were collected from Urmia Abattoir, Urmia, Iran, and were transported immediately to the laboratory in a thermos flask containing sterile normal saline with added antibiotics. Oocytes were aspirated from 2-8 mm visible follicles of the ovaries using an 18-G needle attached to a 10 ml syringe. IVM medium included tissue culture medium-199 [TCM-199], 10% fetal bovine serum [FBS], 22 microg/ml sodium pyruvate, 0.5 IU/ml ovine follicle-stimulating hormone [oFSH], 0.5 IU/ml ovine luteinizing hormone [oLH], 1 microg/ml oestradiol, 50 microg/ml gentamycin, and leptin [0 [control], 10, 50, and 100 ng/ml]. The good quality buffalo oocytes [batches of 10 oocytes] were placed in a culture plate containing six 50 microl droplets of maturation medium, covered with sterilized mineral oil, and then incubated at 38.5?C with 5% CO2 in air for 24 hours. The maturation of oocytes was evaluated under a stereomicroscope by detecting the first polar body extrusion of oocytes. FITC-Annexin V - propidium iodide [PI] staining method was used to detect oocyte apoptosis. From a total of 115 collected ovaries, 1100 oocytes were recovered among which 283 oocyte were suitable for IVM. In the groups of leptin treated with 0 [control], 10, 50 and 100 ng/ml, the percentage of oocytes maturation was 74.65, 83.81, 77.85, and 75.40%, while the percentage of oocytes apoptosis was 9.83, 9.54, 9.93, and 10.42%, respectively. Our results showed that addition of 10 ng/ml leptin to buffalo IVM medium increased oocyte maturation, significantly, as compared with that in control group. However, addition of leptin to IVM medium had no significant influence on buffalo oocyte apoptosis. Our findings suggested that addition of 10 ng/ml leptin to IVM medium of buffalo oocyte can improve oocyte nuclear maturation. Furthermore, we showed that there is no relation between in vitro addition of leptin to buffalo oocyte IVM medium and oocyte apoptosis