Oocyte vitrification technology has made egg-sharing donation easier in China.

When infertile women undergoing IVF or intracytoplasmic sperm injection (ICSI) have more than 20 mature oocytes retrieved, at least 15 oocytes are inseminated by their husband's spermatozoa. The extra oocytes are cryopreserved by vitrification. If the patients became pregnant and have healthy live births, the patients are encouraged to donate their remaining cryopreserved oocytes. Forty-seven egg-sharing donors were recruited after having normal deliveries and they donated their remaining oocytes, totalling 395 cryopreserved oocytes, to 75 recipients. The survival rate of vitrified-warmed oocytes was 83.0%. Following insemination by ICSI, the fertilization and cleavage rates were 83.8% and 89.8%, respectively. Out of 75 recipients, 71 recipients completed the treatment cycles and 30 of them became pregnant with clinical pregnancy and implantation rates of 42.3% and 25.5%, respectively. The birthweight of the new-born infants (22 from singleton and two from one set of twins) were 3344.5 ± 669.1g and 2425.0 ± 742.5 g, respectively. No birth defects were observed for the live births. These results indicate that oocyte vitrification is an effective methodology for an egg-sharing donation programme, with acceptable pregnancy and implantation rates.

[Sperm chromatin structure assay predicts the outcome of intrauterine insemination].

OBJECTIVE: Sperm chromatin structure assay (SCSA), as a clinically practical technique for the analysis of DNA damage, is rarely reported in China. This study focuses on the correlation of DNA damage with the pregnancy rate of intrauterine insemination (IUI). METHODS: We performed semen analysis for 482 couples undergoing IUI, calculated the DNA fragmentation index (DFI) by SCSA, and observed the relationship between DFI and the pregnancy rate of IUI. RESULTS: Clinical pregnancy was achieved in 5 (5.26%) of the 95 cases with DFI > 25%, and in 59 (15.25%) of the 387 cases with DFI < or = 25%. Those with sperm DFI >25% had significantly lower rates of biochemical pregnancy and clinical pregnancy than those with DFI < or = 25% (OR: 0.37, 95% CI: 0.14 - 0.96 and OR: 0.38, 95% CI: 0.16 - 0.97). No significant differences were found in the DFI of 54 cases between the first and the second cycle ([15.05 +/- 7.98]% vs [17.25 +/- 12.18]%, P > 0.05). Sperm DFI was significantly negatively correlated with sperm concentration, sperm motility and total progressively motile sperm count (P < 0.01). CONCLUSION: The pregnancy rate of IUI is significantly lower in couples with DFI >25% than in those with DFI < or = 25%. Sperm DFI obtained from SCSA is partly correlated with sperm concentration and motility, and it is a robust predictor of the IUI outcome.

Effect of estradiol on proliferation and differentiation of side population stem/progenitor cells from murine endometrium.

BACKGROUND: In our previous study, endometrium side population cells (SP cells) were isolated from postpartum murine uterus, and characterized by a heterogeneous population of stem/progenitor cells. In this study, we investigated the effect of estrogen on the proliferation and differentiation of SP cells. METHODS: SP and non-SP cells of postpartum murine endometrium were isolated by DNA dye Hoechst 33342. The expression of estrogen receptor 1 (ESR1) was measured by reverse transcription polymerase chain reaction (RT-PCR), Real-time PCR, Western blot, immunofluorescence and immunohistochemistry. The proliferation and differentiation of SP cells treated with different concentrations [10(-8) M-10(-6) M] of estradiol (E2) and E2+ ICI182780 (Faslodex, inhibitor of ESR1) were measured by 3-(4, 5-dimethylthiazoly1-2)-2,5-diphenyltetrazolium bromide(MTT) and clonogenic assays. RESULTS: (1) SP cells expressed ESR1 at a higher level than non-SP cells. (2) The level of E2 in the serum and the expression of ESR1 in the uterus of postpartum murine changed in the same manner with the ratio of SP cells to total uterus cells at a different postpartum time point. ESR1, as ABCG2 is also predominantly located in the stroma and the glandular epithelium of the uterus. (3) 10(-6) M E2 notably promoted the proliferation of SP cells after treatment for 24 h. This effect could be inhibited by ICI182780. E2 at the concentration of 10(-7) M or 10(-8) M was sent to impair the large cloning efficiency (CE) of SP cells. CONCLUSIONS: The effect of estrogen on the proliferation and differentiation of endometrium SP cells via ESR1 was observed and it was in a concentration dependent fashion. Clearly, more work is needed to understand the in vivo effect of E2 at the physiological concentration on the differentiation of SP cells.

Isolation and characterization of side population cells in the postpartum murine endometrium.

Endometrium is a highly active organ that is periodically remodeled during the life span. Previous studies have indicated the presence of an adult stem or progenitor cell population in this tissue. In this study, side population (SP) cells were isolated from the endometrium of postpartum murine uterus but not from the endometrium of a uterus undergoing a normal estrus cycle. Phenotype analysis showed that SP cells were negative for hematopoietic, endothelial, and mesenchymal stem cell (MSC) markers, but they expressed stem cell antigen 1 (Sca-1) and c-kit at various degrees. Side population cell is a heterogeneous population of endometrial stem/progenitor cells that have colony-forming capacity. They were found to reside in quiescence in the stroma but not in the luminal epithelium. These data suggest that, like other tissues and organs, the murine endometrium also contains SP cells. Their specific role in the regeneration of the endometrium warrants further study.

[Correlation between serum progesterone level at the day with human chorionic gonadotrophin administration and the outcome of pregnancy in in-vitro fertilization].

OBJECTIVE: To investigate the relationship between serum progesterone level at the day with human chorionic gonadotrophin (hCG) administration and pregnant outcome from in in-vitro fertilization-embryo transfer (IVF-ET). METHODS: From Mar. 2002 to Apr. 2007, 786 cycles with serum progesterone measurement on the day of hCG administration for final oocyte maturation in IVF were analyzed retrospectively in Reproductive Medicine Center in First Affiliated Hospital of Nanjing Medical University. All stimulations were down-regulated with gronadotrophin release hormone agonist (GnRH-a) in both long protocols and short protocols before gonadotrophin stimulation. When the thresholds of serum progesterone were set at 5.5, 6.0, 6.5, 7.0, 7.5, 8.0, 8.5 and 9.0 nmol/L, respectively. If the level of progesterone was less than the thresholds, those patients were in lower progesterone group, on the contrary, more than the threshold value, those patients were in higher progesterone group. The laboratory results and the clinical outcomes between all patients at lower and higher progesterone group at different thresholds value were analyzed. RESULTS: The rate of normal fertilization, quality embryos, successful implantation, chemical pregnancy, clinical pregnancy and live birth did not exhibit remarkable difference between patients with higher and lower serum progesterone level at multiple thresholds on the day of hCG administration in the 786 cycles (P > 0.05). However, when the thresholds of serum progesterone were at 8.5 and 9.0 nmol/L, early abortion rates of 27.3% (3/11) and 3/7 in higher progesterone group were significantly higher than 8.8% (26/297) and 8.6% (26/301) in lower progesterone group (P < 0.05). And the total abortion rates of 3/7 in higher progesterone group were significantly higher than 11.0% (34/301) in lower progesterone group when the thresholds of serum progesterone were 9.0 nmol/L (P < 0.05). CONCLUSIONS: This study did not prove the correlationship between progesterone level at the day with hCG administration and the probability of clinical pregnancy or live birth. However, early abortion rates or the total abortion rates were associated with higher progesterone level when the thresholds of serum progesterone were at 8.5 nmol/L or 9.0 nmol/L.

[Clinical value of artificial insemination by donor].

OBJECTIVE: To investigate the clinical value of artificial insemination by donor (AID). METHODS: We retrospectively analyzed 480 cycles of AID among 258 infertile couples, who were divided according to the women's age into a < or = 30 yr group and a > or = 31 yr group. RESULTS: A total of 120 pregnancies were achieved in 480 AID cycles, with a cycle pregnancy rate of 25.00% and a cumulative pregnancy rate of 46.51%. In the natural cycles, the cycle pregnancy rate was 29.65% and the cumulative pregnancy rate was 51.00% in the < or = 30 yr group, significantly higher than 13.33% and 25.00% in the > or = 31 yr group (P < 0.05). In the ovulation induction cycles, no significant differences were found in the cycle and cumulative pregnancy rates between the two groups (24.02 and 48.86% versus 23.81 and 43.48% , P > 0.05). The cycle and cumulative pregnancy rates decreased with the increase of infertility duration and the women's age, but had no significant differences. In the first four cycles, the cycle pregnancy rates were 24.03, 24.94, 24.69 and 25.00% (P > 0.05), and the cumulative pregnancy rates were 24.03, 39.53, 45.74 and 46.51%, with significant differences between the first cycle and the other three (P < 0.01). CONCLUSION: Ovulation induction is superior to natural cycle in AID for older women. IVF/ICSI can be resorted to only after AID has failed three or four times.

Evaluation of x-inactivation status and cytogenetic stability of human dermal fibroblasts after long-term culture.

Human primary fibroblasts are a popular type of somatic cells for the production of induced pluripotent stem (iPS) cells. Here we characterized biological properties of primary fibroblasts in terms of cell-growth rate, cytogenetic stability, and the number of inactive X chromosomes during long-term passaging. We produced eight lines of female human dermal fibroblasts (HDFs) and found normal karyotype and expected pattern of X chromosome inactivation (XCI) at low passages (Passage P1-5). However, four out of the eight HDF lines at high passage numbers (≥ P10) exhibited duplicated hallmarks of inactive X chromosome including two punctuate signals of histone H3 lysine 27 trimethylation (H3K27me3) and X inactive-specific transcript (XIST) RNA signals in approximately 8.5-18.5% of the cells. Our data suggest that the copy number of inactive X chromosomes in a subset of female HDF is increased by a two-fold. Consistently, DNA fluorescent in situ hybridization (FISH) identified 3-4 copies of X chromosomes in one nucleus in this subset of cells with two inactive Xs. We conclude that female HDF cultures exhibit a higher risk of genetic anomalies such as carrying an increased number of X chromosomes including both active and inactive X chromosomes at a high passage (≥ P10).

[Polypronuclear zygotes and clinical pregnancy after IVF].

OBJECTIVE: To study the relationship between the percentage of polypronuclear zygotes and clinical pregnancy following IVF. METHODS: We collected the data of 954 IVF cycles, and according the percentage of polypronuclear zygotes in the IVF cycles, allocated them to Groups A (without polypronuclear zygotes) , B (with < 30% polypronuclear zygotes) and C (with > or = 30% polypronuclear zygotes). Then we analyzed the relationship between the percentage of polypronuclear zygotes and the rate of clinical pregnancy. RESULTS: Compared with Group A, Group C showed a significantly lower rate of clinical pregnancy (43.2% vs 28. 1%, P < 0.05), while Group B exhibited a markedly higher rate (43.2% vs 52.36%, P < 0.05) and obviously decreased polypronuclear zygote formation with the increase of age (35.6% vs 24.1%, P < 0.05). CONCLUSION: The percentage of polypronuclear zygotes in IVF cycles may serve as a prognostic indicator of the clinical outcome.