ABSTRACT
OBJECTIVE: Some reports have indicated that conditioned medium from growing mouse embryonic stem cells (ESCs) provides a supportive condition for small follicles growing, oocyte maturation, and following embryo growth. The aim of this study is assessing in vitro maturation (IVM) and consequent in vitro fertilization (IVF) outcome of immature mouse oocytes using human embryonic stem cells conditioned medium (HESCM). MATERIALS AND METHODS: In this experimental study, 240 germinal vesicle (GV) oocytes were took from NMRI female mice, aged 4-6 weeks, 48 hours before injection of 5 IU pregnant mare serum gonadotropin (PMSG). 120 GV oocytes without cumulus cells were cultured in each of the groups. 120 GV were cultured in HESCM as test groups and also 120 GV cultured in human embryonic stem cells medium (HESM) as control groups. After evaluating the metaphase II (MII) oocyte maturation rate at 8, 16 and 24 hours, the MII oocytes subsequently were fertilized in vitro and the two-cell embryo development rate was recorded at days 1, 2, and 3. Statistical analysis was performed by using the generalized estimating equations (GEE) method that calculated their rate ratio. RESULTS: Our data indicated there are significant differences between the maturation rates in HESCM and HESM (P=0.004), also the two-cell embryo development was significant between two culture media (P=0.00). CONCLUSION: Similar to some other studies, the secretome of the HESCM showed a significant impact on the IVM outcomes in mice.
ABSTRACT
BACKGROUND: Testicular cell conditioned medium (TCCM) has been shown to induce female germ cell development In Vitro from embryonic stem cells (ESCs). Testicular cells (TCs) secrete a variety of growth factors such as growth differentiation factor-9 (GDF-9), bone morphogenetic protein 4 (BMP-4), stem cell factor (SCF), leukemia inhibitory factor (LIF), and other, that could improve oocyte maturation. Here we have investigated the effect of human TCCM (hTCCM) on in vitro maturation (IVM) and morphology of mouse oocytes. MATERIALS AND METHODS: In this experimental study, 360 germinal vesicle (GV) oocytes were obtained from NMRI mice, aged 4-6 weeks that had received 5 IU pregnant mare's serum gonadotropin (PMSG) 48 hours before. GV oocytes were subjected to IVM. 120 GV oocytes were cultured in each medium; hTCCM as the test group, DMEM + 20%FBS as the control group and Ham's F10 + HFF medium as the sham group. The rates of the IVM and perivitelline space (PVS) changes were recorded at 8, 16 and 24 hours after culture. The metaphase II (MII) oocytes were subjected for in vitro fertilization (IVF) and the fertilization rate was evaluated after 1, 2, and 3 days. RESULTS: There was a significant difference between the maturation rates in hTCCM (31.67% MII) and the control [0% MII, P<0.05, (7.5% MI, 52.5% deg. and 40%GV)] groups but there was not a significant difference between the maturation rates in hTCCM and the sham group (53.33% MII, P>0.05). IVF success rate for MII oocytes obtained from IVM in the hTCCM group was 28.94% (n=11). Our data showed that hTCCM is an effective medium for GV oocyte growth and maturation compared to the control medium. CONCLUSION: Our findings show that TCCM supports oocyte IVM in mice and affect oocyte morphology.
ABSTRACT
BACKGROUND: To increase the results of infertility treatment, many efforts have been made to improve the treatment methods. As assisted reproductive technology is mainly using cell culture methods, one of the approaches to improve this technology is conditioned medium from different sources. It is desirable to apply in vitro maturation (IVM) and use oocytes from normal cycles instead of stimulating ovulation. OBJECTIVE: To investigate the effect of human cumulus cell condition medium (hCCCM) on the IVM of immature mouse oocytes and morphology. MATERIALS AND METHODS: In this experimental study, 240 germinal vesile oocytes were collected from four-six wk-old mice after 48 hr of 5IU pregnant mare serum gonadotropin (PMSG) injection and cultured in hCCCM (test group, n = 120) and DMEM + 20% FBS (control group, n = 120). The IVM rates and changes in perivitelline space (PVS) and shape were investigated at 8, 16, and 24 hr following the culture. The mature (MII) oocytes were subjected to in vitro fertilization (IVF) and the fertilization rate was assessed in three days. RESULTS: A significant difference was observed between the maturation rates in the hCCCM and control groups (24.16% vs 0%; p = 0.001), as well as morphologic changes between the two groups (p = 0.04, p = 0.05). The development rate for MII oocytes attained from IVM in the hCCCM group was 27.58% (2-cell) and 6.89% (4-cell). Data displayed that hCCCM is an effective medium for oocytes maturation compared to the control medium. CONCLUSION: hCCCM supports oocyte in vitro growth and maturation. Moreover, hCCCM changes the oocyte shape and size of perivitelline space.
ABSTRACT
PURPOSE: Our aim was to evaluate the effect of neat semen vitrification on human sperm vital parameters and DNA integrity in men with normal and abnormal sperm parameters. MATERIALS AND METHODS: Semen samples were 17 normozoospermic samples and 17 specimens with abnormal sperm parameters. Semen analysis was performed according to World Health Organization (WHO) criteria. Then, the smear was provided from each sample and fixed for terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining. Vitrification of neat semen was done by plunging cryoloops directly into liquid nitrogen and preserved for 7 days. The samples were warmed and re-evaluated for sperm parameters as well as DNA integrity. Besides, the correlation between sperm parameters and DNA fragmentation was assessed pre- and post vitrification. RESULTS: Cryopreserved spermatozoa showed significant decrease in sperm motility, viability and normal morphology after thawing in both normal and abnormal semen. Also, the rate of sperm DNA fragmentation was significantly higher after vitrification compared to fresh samples in normal (24.76 ± 5.03 and 16.41 ± 4.53, P = .002) and abnormal (34.29 ± 10.02 and 23.5 ± 8.31, P < .0001), respectively. There was negative correlation between sperm motility and sperm DNA integrity in both groups after vitrification. CONCLUSION: Vitrification of neat ejaculates has negative impact on sperm parameters as well as DNA integrity, particularly among abnormal semen subjects. It is, therefore, recommend to process semen samples and vitrify the sperm pellets.
