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Objective:To study the effectiveness of noninvasive embryo chromosome screening (NICS) based on blastocyst culture medium and cystic fluid in preimplantation genetic detection (PGT) of embryos in different age groups.Methods:A retrospective analysis of 62 couples who underwent PGT assisted pregnancy in Shenzhen Hospital of Peking University from January 2019 to June 2021. A total of 310 blastocysts were biopsied. At the same time, D3-6 blastocyst culture medium and blastocyst cavity fluid were collected for NICS. According to the age of the patients, they were divided into three groups: <35 years old group, 35≤age<40 years old group and ≥40 years old group. The results of NICS were compared with those of embryonic trophoblast (TE) biopsy in each group, and the consistency, sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) were calculated. Then the consistency, sensitivity, specificity, positive predictive value and negative predictive value of NICS and TE among the three age groups were statistically analyzed.Results:There was no statistically significant difference in the consistency rate of NICS and TE among the three age groups ( P>0.05), but there was an upward trend in the elderly group (35≤age<40 years and ≥40 years). There was no statistically significant difference in specificity, sensitivity and PPV among the three age groups ( P>0.05). There was significant difference in NPV between the ≥40 years group and the other two groups ( P<0.05). Conclusions:There was no statistical difference in the effectiveness of NICS among different age groups, However, there was an increasing trend in people ≥35 years of age.
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Objective To evaluate the application value of the blastocysts derived from non-pronucleus (0PN) zygotes by the good quality blastocyst formation rate and the clinical outcomes of frozen-thawed blastocyst transfers. Methods The good quality blastocyst formation rate derived from 0PN zygotes was compared with that derived from2 pronucleus(2PN)zygotes in in vitro fertilization(IVF)or intracytoplasmic sperm injection (ICSI) cycles from January 2015 to December 2016. In addition, the clinical pregnancy, embryo implantation and live birth rates of frozen-thawed blastocyst transfers with blastocysts derived from 0PN and 2PN zygotes were analyzed on corresponding dates. Results (1)In IVF cycles, the high quality blastocysts formation rate of 2PN embryos was significantly higher than that of 0PN (46.64% versus 42.42%, P<0.01). In ICSI cycles, the high quality blastocysts formation rate of 2PN embryos was markedly higher than that of 0PN(41.96% versus 21.73%, P<0.01).(2)In frozen-thawed embryo transfer cycles for IVF, the clinical pregnancy, implantation and live birth rates of D5 0PN blastocysts were significantly higher than those of D6 2PN(52.64% versus 46.78%, 49.91% versus 41.20%, 46.54% versus 39.56%, all P<0.05), however, the abortion and newborn abnormal rates of D5 0PN blastocysts were lower than those of D6 2PN blastocysts(17.37% versus 23.36%, 1.31% versus 4.21%, both P<0.05); the clinical pregnancy, implantation and livebirth rates of D5 2PN blastocysts were significantly higher than those of D5 0PN(59.73% versus 52.64%, 55.95% versus 49.91%, 53.03% versus 46.54%, all P<0.05), but newborn abnormal rate was a little higher than that of D5 0PN(3.90% versus 1.31%, P<0.05);the clinical pregnancy, implantation and live birth rates of D5 2PN blastocysts were significantly higher than those of D6 2PN(59.73% versus 46.78%, 55.95% versus 41.20%, 53.03% versus 39.56%, all P<0.05), and the abortion rate of D5 2PN blastocysts was lower than that of D6 2PN blastocysts(18.23% versus 23.36%, P<0.05). Conclusions Although the blastocysts derived from 0PN could be transffered, the blastocysts derived from 2PN zygotes are preferred in all cycles. In IVF cycles, the good quality blastocysts derived from 2PN or 0PN zygotes will be transferred.
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Objective To explore the effects of in vitro culture time of frozen cleavage embryo after thawing on clinical pregnancy rate during frozen-thawed embryo transfer (FET) cycles. Methods The clinical data of 692 patients were analyzed; they received FET in the Department of Reproductive Medicine Center, Changhai Hospital, Navy Medical University (Second Military Medical University) from January to December in 2016. According to the days of in vitro culture between thawing and transferring, the patients were divided into three groups: no in vitro culture group, 1-day in vitro culture group and 2-day in vitro culture group. The pregnancy outcomes were compared between the three groups. Results There were no significant differences in age, body mass index, infertile duration, basic follicle-stimulating hormone (FSH) level or endometrial preparation protocol between the three groups (all P0.05). The embryo loss rates of the no in vitro culture group, the 1-day in vitro culture group and the 2-day in vitro culture group were 0.0% (0/706), 13.3% (64/481) and 43.0% (114/265), respectively, and the difference was statistically significant (P0.01). There were no significant differences in number of transfered embryo or endometrial thickness on transplantation day between the three groups (both P0.05). The good-quality embryo transfer rates in the 2-day in vitro culture group and 1-day in vitro culture group were significantly lower than that in the no in vitro culture group (both P0.01), and the clinical pregnancy rate in the 2-day in vitro culture group was significantly higher than that in the no in vitro culture group (P0.01). Conclusion In vitro culture for 1-2 days after thawing frozen cleavage embryos may increase the embryo loss rate, but it can improve the clinical pregnancy rate by screening the embryos with developmental potential.
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Objective To evaluate the efficiency of the application of array comparative genomic hybridization (array-CGH) in preimplantation genetic diagnosis or screening (PGD/PGS), and compare the clinical outcomes of different stage embryo biopsy. Methods The outcomes of 381 PGD/PGS cycles referred in the First Affiliated Hospital of Nanjing Medical University from July 2011 to August 2015 were retrospectively analyzed. There were 320 PGD cycles with 156 cleavage-stage-biopsy cycles and 164 trophectoderm-biopsy cycles, 61 PGS cycles with 23 cleavage-stage-biopsy cycles and 38 trophectoderm-biopsy cycles.Chromosomal analysis was performed by array-CGH technology combined with whole genome amplification.Single embryo transfer was performed in all transfer cycles.Live birth rate was calculated as the main clinical outcomes. Results The embryo diagnosis rate of PGD/PGS by array-CGH were 96.9%-99.1%. In PGD biopsy cycles, the live birth rate per embryo transfer cycle and live birth rate per embryo biopsy cycle were 50.0%(58/116) and 37.2%(58/156) in cleavage-stage-biopsy group, 67.5%(85/126) and 51.8%(85/164) in trophectoderm-biopsy group (both P<0.01). In PGS biopsy cycles, the live birth rate per embryo transfer cycle and live birth rate per embryo biopsy cycle were the same as 34.8%(8/23) in cleavage-stage-biopsy group, the same as 42.1%(16/38) in trophectoderm-biopsy group (both P>0.05). Conclusions High diagnosis rate and idea live birth rate are achieved in PGD/PGS cycles based on array-CGH technology.The live birth rate of trophectoderm-biopsy group is significantly higher than that of cleavage-stage-biopsy group in PGD cycles;the efficiency of trophectoderm-biopsy is better.
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Objective To explore the influence of modified super-long hormone replacement protocol on pregnancy outcome of previous embryo implantation failure patients who underwent frozen-thawed embryo transfer (FET). Methods A total of 669 women who underwent FET with a failed history of in vitro fertilization/intracytoplasmic sperm injection-embryo transfer (IVF/ICSI-ET) were enrolled in Changhai Hospital of Second Military Medical University from Jan. 2015 to Jan. 2017. Based on different endometrial preparation protocols for FET, the patients were assigned to receive either modified super-long hormone replacement protocol (Modified super-long group, n=184) or conventional hormone replacement protocol (Conventional group, n=485).The pregnancy outcomes were retrospectively analyzed in the two groups. Results Patients in the two groups had similar baseline characteristics, including age, body mass index, duration of infertility, number of transferred embryo, endometrial thickness on transformation day and transplantation day (P>0.05). The good-quality embryo transfer rate in the Modified super-long group was significantly lower than that in the Conventional group (50.9% vs 64.8%, P0.05); however, in the blastocyst transfer cycles, although the good-quality embryo transfer rate had no significant difference between the two groups (45.5% vs 52.7%, P>0.05), the embryo implantation rate and clinical pregnancy rate in the Modified super-long group were significantly higher than those in the Conventional group (39.6% vs 27.2% and 56.9% vs 40.2%, respectively; both P<0.05). Conclusion The modified super-long hormone replacement protocol can improve the embryo implantation rate and clinical pregnancy rate of the patients undergoing FET with a history of embryo implantation failure, and is worthy of clinical popularization.
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Objective To explore the relevant parameters and clinical outcome of frozen-thawed blastocyst transfer compared with fresh blastocyst transfer. Methods A historic cohort study was conducted to analyze the blastocyst implantation rate and clinical pregnancy rate of 61 patients receiving fresh blastocyst transfer and 372 receiving frozen-thawed blastocyst transfer from Mar. 2015 to Dec. 2016 in Changhai Hospital of Second Military Medical University. And then the clinical outcomes of day five (n=308) or day six (n=64) embryo were analyzed in the frozen-thawed blastocyst transfer group. Results There was no significant difference in age or number of the transferred blastocysts between the two groups. Blastocyst implantation rate in frozen-thawed blastocyst transfer group was significantly higher than that in fresh blastocyst transfer group (40.2% vs 29.6%, P=0.036), while no significant difference was found in clinical pregnancy rate between the two groups (57.8% vs 47.5%, P=0.134). Blastocyst implantation rate and clinical pregnancy rate of day five embryo were significantly higher than those of day six (42.3% vs 30.1%, P=0.016, and 60.7% vs 43.8%, P=0.012, respectively). Conclusion The pregnancy outcome of frozen-thawed blastocyst transfer is better compared with fresh blastocyst transfer, especially when transferring on day five.
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Objective To compare the pregnancy outcome of blastocyst which were cryopreserved in different carrier:cryotop and cryoLogic vitrification method (CVM) embryo freezing carriers.Methods Data were collected retrospectively from January 2014 to August 2015.All the thawed-frozen embryo transfer (FET) cycles were the first one after their fresh embryo transfer.According to the freezing carrier,they were divided into two groups:Cryotop group (n =114) and CVM group (n =232).The pregnancy outcome in FET cycles was compared between two groups.Results The clinical pregnancy rate,implantation rate,and the birth rate were significantly higher in group cryotop.The recovery rate,the rate of multiple births,ectopic pregnancy,miscarriage,premature birth rate,birth weight,and birth defect rate were similar in two groups.Conclusions Compared to CVM frozen carrier,the thawed-frozen embryo cyropreserved in Cryotop has higher clinical pregnancy rate,higher implantation rate,and live birth rate.
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Objective To compare the outcome of in vitro fertilization (IVF) between sequential embryo transfer and simple blastocyst transplantation in patients with previous multiple IVF failures. Methods A total of 170 patients with repeated implantation failure were divided into sequential transfer group (n=71) and blastocyst transfer only group (n=99). A retrospective matched case-control analysis was made for the medical files of 71 patients who underwent sequential transfer of D3 embryos and blastocysts. The control group included 99 matched women who underwent embryo transfer on D5/6 only. All of the patients in two groups used the same protocols of emdometrium preparation (natural cycle or hormone-replacement cycle) and ultrasound-guided transplantation. The embryo implantation rate and clinical pregnancy rate were compared and analyzed between two groups. Results Sequential transfer of embryos resulted in a clinical pregnancy rate of 60.6%compared with that of 31.3%following D5/6 embryo transfer, and the implantation rate was 34.8%and 23.8%respectively (P<0.05). Although the total number of transfered embryos were higher in sequential transfer group than that of blastula transfer only group, the number of D5/6 embryo transfered in sequential transfer group were less than blastula transfer only group (P<0.05). And the number of high quality blastula transfered showed no statistical significant difference between two groups. There were 20 cases of twin and 5 cases of triplet pregnancy in sequential transfer group, which were 5 cases and 1 case in blastula transfer only group respectively. While, there was no case of muliple pregnancy beyond triplet in both groups Conclusion Sequential transfer of embryos can be used for women with repeated IVF cycles. The program avoids the possibility of eliminating the transplant, and which is effective in patients with more transplant embryos.
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Objective To analyze the expression of the placenta Grb10 from women conceived by transferred thawed blastocyst,and to evaluate the security of blastocysts vitrification.Methods A cross-sectional study was performed in the Department of Obstetrics and Gynecology of Fujian Provincial Maternity and Children's Hospital from January 2012 to May 2014,50 women conceived by transferring thawing blastocyst and 50 natural pregnancy control women were enrolled in this study.The expression of Grb10 protein was detected by immunohistochemistry and Western blot,and the expression of Grb10 mRNA was detected by Realtime PCR method.Results Comparison of two cases of gestational age,gestational age,fetal sex,fetal body weight,body length,head circumference,abdominal circumference,there were no significant differences(P＞0.05),comparison of placental area,placental weight,the difference was statistically significant(P＜0.05).Real-time PCR and Western blot results showed that,there was no significant difference in the expression of Grb10 mRNA and protein between the two groups(P＞0.05).Conclusion Blastocysts vitrification may increase the area and quality of delivery of placenta,however,there was no significant change in the expression of Grb10 in placenta.
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El pargo colorado (Lutjanus colorado) es una especie con un alto valor comercial en el mercado mexicano, con potencial para su cultivo. Hasta la fecha no existen estudios sobre su reproducción, cultivo larvario y engorda en cautiverio. El presente trabajo es el primer reporte sobre la descripción a detalle del desarrollo embrionario de la especie bajo condiciones de cultivo. Los huevos fertilizados viables del pargo colorado son pelágicos, esféricos, transparentes y con una sola gota de aceite. Midieron 0,77±0,09 mm de diámetro y la gota de aceite 0,14±0,01 mm. La primera división ocurrió a las 0,05 horas post fertilización (HPF). La eclosión se llevó a cabo a las 17,22 HPF bajo las condiciones del presente estudio. Las larvas recién eclosionadas midieron 1,8±0,1 mm de longitud total (LT). El desarrollo embrionario de esta especie fue similar a la descrita para especies de la misma familia. Los resultados del presente estudio aportan información básica para iniciar el desarrollo de la biotecnología para la producción de semilla de esta especie a escala comercial.
The Colorado snapper (Lutjanus colorado) is one of the most commercially important fish species in México and it is considered a suitable candidate for culture. Until now, no research has been carried out on its reproduction, larviculture and fattening in captivity. This study is the first description of embryonic development of this species under controlled conditions. Fertilized eggs of Colorado snapper are pelagic, spherical and transparent and contain one drop of oil. Eggs measured 0.77±0.09 mm and the drop of oil 0.14±0.01 mm. First cell division occurred at 0.05 h post-fertilization (HPF), hatching at 17.22 HPF under the above described conditions. Larvae total length (LT) was 1.8±0.1 mm. Embryonic development of this species was similar to other lutjanidae species. These results provide basic information for developing the necessary biotechnology for commercial seed production of the Colorado snapper.
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Animals , Ovum , Perciformes/embryology , Larva/growth & development , Perciformes/growth & development , Blastula/embryology , Organogenesis , Embryonic Development , Embryo, Nonmammalian , Gastrula/embryologyABSTRACT
Objective To investigate the effects on pregnancy outcome and neonate by artificial shrinkage by microsucting the fluid of expanded blastocysts before vitrification using glass micropipette (GMP).Methods From Jan.2006 to Dec.2009, 342 vitrified-thawed blastocyst cycles from patients that performed in vitro fertilization-embryo transfer (IVF-ET) or intracyteplasmic sperm injection ( ICSI ) were enrolled in this study in Reproductive Medicine center, Maternal and Child Health Hospital of Guangxi Zhuang Autonomous Region.Three hundred and fourteen cycles of expanded blastocysts were artificially shranked by microsucting blastocoelic fluid with a micro-needle before vitrification as artificial shrinkage group, in the mean time, 28 cycles without artificial shrinkage were chosed as control group.The survival rate, implantation rate, clinical pregnancy rate and transfer canceled rate were compared between artificial shrinkage group and control group.Among pregnant women, the miscarriage rate, live birth rate, congenital birth defect rate, neonatal weight and gestational age were compared with those of fresh embryo transfers in 520 cycles.Results The blastocyst survival rate, implantation rate and clinical pregnancy rate were 95.3%(403/423), 38.0% ( 153/403), 44.6% (140/314) in artificial shrinkage group and 64.3 % (27/42),7.4% (2/27), 7.1% (2/28) in control group, respectively, which reached statistical difference (P＜0.05).The transfer canceled rate was 0 in artificial shrinkage group and 25.0% (7/28) in control group, which also reached statistical difference ( P ＜ 0.05 ).Among pregnant patients, the miscarriage rate of 18.2% (10/55), live birth rate of 80.0% (44/55), gestational age of (38.2 ± 1.3) weeks, congenital birth defect rate of 2.1% (1/47), birth weight of newborns of (2989 ±640) gram in artificial shrinkage group were not significantly different with 17.5% (91/520), 74.0% (385/520), (37.9 ±2.3) weeks,1.7% (8/479) and (2856±640) gramin fresh embryo transfer group (P＞0.05).Conclusion Artificial shrinkage by microsucting blastocoelic fluid with a micro-needle before vitrification significantly improved the vitrification effects of expanded blastocyst and no distinct increasing rate of neonates congenital anomality were observed.
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To study the effects of mouse cytomegalovirus (MCMV) on the in vitro maturation, fertilization, cleavage and blastula formation of mouse oocytes, the immature oocytes were infected in vitro by MCMVs of different dosages (100 TCID50, 10 TCID50 and 1 TCID50). The oocytes were then observed for in vitro maturation, fertilization, cleavage and blastula formation and the ultrastructural changes after the culture with the viruses. Our results showed that no significant differences were found in IVM, IVF, cleavage and blastula formation among the groups treated with of virus of various dosages. And ultrastructural abnormality was observed in the oocytes treated by 100 TCID50 of viruses. It is concluded that MCMV did not have any conspicuous effects on IVM, IVF, cleavage and blastula formation of murine immature oocytes.
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Objective To determine whether cleavage developmentally retarded embryos have not cleaved during a 24 hour period could develop into blastocysts and produce hESC cell lines.Methods A total of 120 such embryos were cultured to blastocyst stage by sequential culture.Blastocysts formation rate and quality of blastocyst were detected under microscope.The relation between blastocyst formation rate and blastomere number,the fragment of blastomere and blastomere symmetry were analyzed by stepwise Logistical regression analysis.Inner cell masses(ICMs)were isolated by immunosurgery.Colonies derived from the ICMs were passed every 4-7 days and the derivatives were passaged and identified.Results A total of 22 blastocysts were obtained from 120 embryos.The blastulation rate was 18.7%.Early blatocyst, blastocyst,full blastocyst,expanded blastocyst,hatching blastoeyst and hatched blastocyst accounted for 5.9%,23.5%,35.3%,23.5%,5.9%,and 5.9% respectively.The grade of ICM and trophoblast was mostly scored C or B.Blastocyst formation rate was related to cell number and blastomere symmetry but not fragment.Immunosurgery resulted in the formation of 7 ICMs and 3 primary colonies,which produced 2 cell lines.The cell lines satisfied the criteria that characterize pluripotent hESC cells.Undifferentiated cells were positive for AKP,SSEA-4,TRA-1-60,and TRA-1-81.It could continue to proliferate in vitro and form embryoid bodies when cultured in suspension.It had capability to form teratoma in SCID mice.Both cell lines had normal karyotypes after 45 and 34 passages respectively.Conclusions Our results suggest that a subset of developmentally retarded embryos can form blastocysts and give rise to hESC cell lines.