prospective randomized comparison of early embryo cleavage kinetics between two media culture systems

Objective: To investigate whether early embryo cleavage kinetics were affected by type of culture media

Methods: In this prospective sibling-split study, 620 oocytes from 37 patients were randomly allocated into two groups: Cook group and Vitrolife group. Oocytes/embryos in Cook group, would be cultured with Cook sequential culture medium, while oocytes/embryos in Vitrolife group, would be cultured with Vitrolife sequential culture medium. Time-lapse imaging technology was used to calculate exact timing of early embryo cleavage events which included time to 2PN breakdown, cleavage to 2-, 3-, 4-, 5- cell and the time duration in the 2-,3-cell stage. Then these timing of early embryo cleavage events were compared between Cook group and Vitrolife group. Moreover, fertilization rate, cleavage rate, high quality embryo rate, usable blastocyst rate, pregnancy rate and implantation rate of these two groups were also analyzed

Results: The results showed there were no differences in all timing of early embryo cleavage events between the two groups. In addition, the two groups were similar in fertilization rate [Cook 71.0% vs. Vitrolife 71.3%, P>0.05], cleavage rate [Cook 98.1% vs. Vitrolife 98.2%, P>0.05], high quality embryo rate [Cook 52.1% vs. Vitrolife 52.7%, P>0.05], usable blastocyst rate [Cook 29.7% vs. Vitrolife 28.0%, P>0.05], pregnancy rate [Cook 46.7% VS. Vitrolife 50.0%, P>0.05] and implantation rate [Cook 30.3% VS. Vitrolife 29.0%, P>0.05]

Conclusions: Morphokinetics used for embryo selection are not affected by the two different culture media

Incidence and management of monozygotic twin conceived by assisted reproductive techniques / 中华妇产科杂志

Objective To analysis the incidence and management of monozygotic twin (MZT) conceived by assisted reproductive techniques (ART). Methods A retrospective analysis of clinical pregnancies and MZT that resulted from ART was performed in Reproductive Medical Center,the First Affiliated Hospital,Wenzhou Medical University between January 2011 and January 2014. Results A total of 5 908 pregnancies were diagnosed: 2 012 twins, 157 high-order multiple pregnancy (HOMP), including 4 quadruplets. Overall, 51 MZT pregnancies were identified of them including 32 cases HOMP and 19 cases MZT. The incidence of MZT resulting from cleavage-stage embryo transfer was similar to blastocyst transfer (P=0.960). The percent of MZT resulting from in vitro fertilization [0.93% (28/3 022)], frozen-thawed embryo transfer [0.87% (13/1 502)] and intracytoplamic sperm injection [0.72% (10/1 384)] did not show statistical significance (P=0.794). The expectantly managed MZT was associated with a significantly greater likelihood of miscarriage [6/19 vs 5.11% (101/1 976)], and low birth weight infant [73.91% (17/23) vs 42.89% (1 453/3 388), P<0.01], when compared with dizygotic twin (DZT) did not undergo selective embryo reduction(SER). In monozygotic (MZ)-triplets with SER to 2 fetuses or to 1 fetus, there was no cases of preterm birth or low birth weight infant observed in MZ-triplets with SER to 1 fetus;when compared with MZ-triplets with SER to 2 fetuses, the low birth weight infant [56.00% (14/25), P=0.021] has statistical significance. The likelihood of the survival of two babies was lower in MZ-triplets with SER to 2 fetuses when compared with non-MZ triplets with SER to 2 fetuses [42.86%(9/21)vs 75.21%(91/121), P=0.003]. Conclusions The incidence of MZT pregnancies following ART is high. It plays a significant role in the occurrence of HOMP. MZT pregnancies are at an increased risk of adverse outcomes , it should transform to a single embryo thansfer (SET) program to reduce them incideuce. Reduction of MZT contained in multiple pregnancies appears to be a safe option.

Prediction of clinical pregnancy in vitrified-warmed single blastocyst transfer cycles by pre-freeze morphology

Background: The selection of blastocyst warmed for transfer is based on pre-freeze morphology in vitrified-warmed single blastocyst transfer cycles. But, it is controversial which parameter of blastocyst morphology most closely related to the clinical outcomes

Objective: To estimate the effect of blastocoele expansion, trophectoderm [TE] morphology grade, and inner cell mass [ICM] morphology grade on clinical pregnancy in vitrified-warmed single blastocyst transfers

Materials and Methods: There were 172 vitrified-warmed single blastocyst transfer cycles during the year 2012 included in this analysis. Comparison of clinical results between pregnancy and no pregnancy group based on patient and blastocyst morphology characteristics was done. Then stepwise logistic regression analysis was used to select the best morphological predictor for clinical pregnancy. Last, comparison of patient characteristics and clinical outcomes separated by the best independent morphological predictor was done

Results: Comparison of clinical results between pregnancy and no pregnancy group and logistic regression showed the clinical pregnancy rate was affected by ICM. Comparison of patient characteristics separated by ICM grade, ICM grade A cycles got higher clinical pregnancy rate than ICM grade B cycles [54.3% vs. 35.0% respectively, p=0.037]

Conclusion: Blastocyst with good ICM morphology could increase clinical pregnancy rate in vitrified-warmed single blastocyst transfer cycles

Comparision of in vitro maturation applied in PCOS and non-PCOS patients undergo stimulated and unstimulated protocols / 中华妇产科杂志

Objective To compare the laboratory and clinical results between unstimulated in vitro maturation (IVM) and IVM converted from in vitro fertilization (IVF) in polycystic ovarian syndrome (PCOS) and non-PCOS patients.Methods We divided 591 IVM cycles in the First Affiliated Hospital of Wenzhou Medical Univesity from Jan.2008 to Dec.2013 into 4 groups:group A1B1,PCOS patients underwent unstimulated IVM protocol,240 cycles; group A1B2,PCOS patients underwent IVM converted from conventional stimulated IVF protocol,153 cycles; group A2B1,non-PCOS patients underwent unstimutlated IVM protocol,103 cycles; group A2B2,non-PCOS patient underwent IVM converted from conventional stimulated IVF protocol,95 cycles.Multiple linear regression method and binary logistic regression method were used to assess the influence of PCOS and protocols for IVM on laboratory and clinical outcomes.Results The mean number of oocytes retrieved was positively related with PCOS [partial regression coefficient (B)=3.37,P＜0.01].The maturation rate of oocytes was positively related with hCG-prime prior to oocyte aspiration (B=0.05,P=0.010).High-quality embryo rate was positively related with PCOS and IVM converted from IVF (B=0.08,P=0.010; B=0.09,P=0.001),as well as implantation rate related with them (B=0.07,P=0.010; B=0.10,P＜0.01).PCOS and IVM converted from IVF improved hCG positive (hCG＞10 U/L) rate (OR=1.636,95％CI:1.113-2.204,P＜0.05; OR=1.861,95％CI:1.307-2.649,P＜0.05) and the clinical pregnancy rate (OR=1.507,95％CI:1.041-2.240,P＜0.05; OR=1.881,95％CI:1.312-2.696,P＜0.05).IVM converted from IVF protocol decreased the spontaneous abortion rate (OR=0.490,95％CI:0.245-0.978,P＜0.05).Multiple gestation rate and ectopic pregnancy rate were not affected by PCOS condition and protocol used (P＞0.05).Conclusions PCOS and IVM converted from IVF protocol improved the high-quality embryo rate,implantation rate,hCG positive rate and clinical pregnancy rate.IVM converted from IVF protocol reduced the spontaneous abortion rate.PCOS patients may be more suitable for the IVM treatment.No matter PCOS or non-PCOS patients,IVM converted from IVF protocol had better pregnancy outcome than that of unstimulated cycle.

Mutation analysis of the mtND4 gene associated with asthenospermia patients / 中华泌尿外科杂志

Objective To investigate the relationship between mtND4 point mutation in sperms and asthenospermia. Methods Fifty-six asthenospermia cases and 44 control cases were collected using the WHO criterion for defining asthenospermia, the regions of mtND4 gene were amplified by using PCR of 3 pairs primers. Consequently, the point mutation, missense mutation and multiple single nucleotide polymorphisms (SNP) were analyzed by employing sequencing technology and bioinformatics tools. Results Six mutations never before identified were found. The frequency of single point mutation T10873C and T11944C in the control group were significantly higher than those in the asthenospermia group (P＜0.05). Eight cases involved T10873C or T11944C among the 10 cases in control groups with missense mutations were found. But, there were only 2 cases with such mutation in the 10 asthenospermia cases with missense mutations (P＜0.05). The previous 20 cases of missense mutations can be described as either multiple SNP group (with T10873C or T11944C) or nonmultiple SNP group. The percentage of a range and a plus b range of multiple SNP group of sperm was significantly higher than the non-multiple SNP group(P＜0.05). Conclusions mtND4 gene mutation, especially the missense mutation may induce loss of sperm motility. The mutations of T10873C and T11944C may be useful for sperm motility or counteract the influence for the sperm motility caused by these harmful mutations.

Association between asthenospermia and mtDNA mutations in ND3 and ND4L genes / 中国病理生理杂志

AIM: To explore the molecular mechanism of asthenospermia(AST) by preliminary screening of nucleotide sequences from the ND3 and ND4L genes of mitochondrial DNA(mtDNA). METHODS: Samples from 50 AST patients and 42 age-matched normal controls were collected according to the WHO criteria. Density gradient centrifugation was applied to separate spermatozoa with different vigor. The ND3 and ND4L genes of mtDNA were amplified and sequenced directly from the extracted genomic DNA from AST patients and normal controls. The sequences were compared with revised Cambridge Reference Sequence(rCRS) to analyze the variants. RESULTS: A total of 22 nucleotide variations were found in ND3 and ND4L genes of mtDNA in asthenospermia group and control group. G10320A, A10398G and T10609C were missense mutations, while A10157G and A10313C were the reported for the first time in this study. Haplotype N in patients with AST(33/50) was higher than that in control group(14/42, P<0.05), and haplotype R9 in patients with AST(15/50) was also higher than that in control group(4/42, P<0.05) through genetic testing of ND3 gene. Rates of sperm progressive motility of haplotype F1, F2 and R9 were significantly lower than those of haplotype M and M rest. Two haplotype differences, haplotype M and N, were found in the same AST patient's spermatozoas which had different vigor. Haplotype M had stronger vigor, while haplotype N had lower vigor. By sequencing ND3 gene of mtDNA from 50 AST patients, we detected G10310A heteroplasmic mutation in 2 specimens of asthenospermia with poor and moderate motility spermatozoa, respectively. No mutation occurred in good motility spermatozoa. CONCLUSION: Haplotype of mitochondrial may have some correlation with sperm motility. The nt10398G-10400T polymorphisms may have benefit for sperm motility, whereas the mutation in nt10310A may impair sperm motility.

Clinical study on in vitro maturation of immature oocytes transferred from conventional in vitro fertilization / 中华妇产科杂志

Objective To investigate clinical effect of in vitro maturation(IVM)of immature oocytes transferred from conventional in vitro fertilization embryo transfer(IVF-ET) cycles.Methods From January 2008 to June 2009.medical documents of 155 infertile patients underwent IVF-ET in the Reproductive Medical Center of Fimt Affiliated Hospital of Wenzhou Medical College were analyzed retrospectively.If more than 20 oocytes were monitored after 5-7 days of ovulation induction or follicular developmental retardation were confirmed after 8-13 days of ovulation induction.according to patients' wish,IVM were transferred in 60 cycles(group A).In the mean time.IVF was continued in 95 cycles (group B).The mean dosage of gonadotropin,the cancel lation rate of cycles,the mean numbers of oocytes retrieved and maturation,the rate of fertilization and excellent quality embryos.pregnancy outcome and the incidence rate of ovarian hyperstimulation syndrome(OHSS)were compared and analyzed.Results The rates of embryo transfer were 92%(55/60)in group A and 63%(60/95)in group B,which showed significant differences (P<0.05).In group A,the mean dosage of the gonadotropin,the mean number of oocytes retrieved,the cleavage rate and OHSS rate were(1030±468)U,10±6,82.2%(231/281)and O,and were(1544±338)U,14±4,94.0%(502/534)and 35%(21/60)in group B,respectively,all data abeve exhibited statistieal difference between two groups(P<0.05).However,the rates of fertilization and excellent quality embryos had no significant differences between two groups(P>0.05).In group A,the rate of clinical pregnancy per transfer was 53%(29/55)and multiple pregnancy was 14%(4/29),and were 47%(28/60) and 32%(9/28)in group B,they all had no significant differences (P>0.05).Conclusion IVM of immature oocytes used in conventional IVF cycles not only obtained a high clinical pregnancy rate,but also reduced gonadotropin using dosage and avoided OHSS completely.