Putrescine alleviates the oxidative damage of cumulus-oocyte complex via improving fatty acid oxidation.

BACKGROUND: Fatty acid oxidation of cumulus-oocyte complex (COC) provides sufficient energy for oocyte maturation. But, the relationship between fatty acid oxidation and oxidative stress in aging follicles, as well as the effect of putrescine, is still unclear. METHODS: The porcine COCs were randomly divided into four groups and cultured in in vitro maturation (IVM) medium with or without 1 mmol/L putrescine, with 50 µmol/L hydrogen peroxide (H2O2) or with 50 µmol/L H2O2 plus 1 mmol/L putrescine. Oocyte maturation was assessed by the first polar body extrusion. The expressions of genes involved in fatty acid oxidation were detected, and the mitochondrial function was analyzed by themembrane potential. RESULTS: The maturation rate of oocyte was significantly lower in the H2O2 group when compared with the control group (P＜0.001), and putrescine significantly increased this rate in the H2O2 plus putrescine group when compared with the H2O2 group (P＜0.001). The expressions of LKB1, STRAD, ACC2, AMPKα1 and AMPKα2 mRNAs in cumulus cells (CCs) were significantly downregulated by H2O2 treatment, and partly rescued by putrescine addition (P＜0.05-0.001). However, the changes of LKB1, STRAD, ACC2, AMPKα1 and AMPKα2 mRNAs in oocytes were inapparent. The mitochondrial membrane potential of CCs in the H2O2 group was significantly lower than that in the control group, while putrescine addition significantly increased the mitochondrial membrane potential (P＜0.001). CONCLUSION: The decrease of oocyte maturation due to oxidative stress is related with the decreased fatty acid oxidation, and putrescine may alleviate the COCs damage via improving fatty acid oxidation.

Continued spring phenological advance under global warming hiatus over the Pan-Third Pole.

The global surface temperature has witnessed a warming hiatus in the first decade of this century, but how this slowing down of warming will impact spring phenology over Pan-Third Pole remains unclear. Here, we combined multiple satellite-derived vegetation indices with eddy covariance datasets to evaluate the spatiotemporal changes in spring phenological changes over the Pan-Third Pole. We found that the spring phenology over Pan-Third Pole continues to advance at the rate of 4.8 days decade-1 during the warming hiatus period, which is contrasted to a non-significant change over the northern hemisphere. Such a significant and continued advance in spring phenology was mainly attributed to an increase in preseason minimum temperature and water availability. Moreover, there is an overall increasing importance of precipitation on changes in spring phenology during the last four decades. We further demonstrated that this increasingly negative correlation was also found across more than two-thirds of the dryland region, tentatively suggesting that spring phenological changes might shift from temperature to precipitation-controlled over the Pan-Third Pole in a warmer world.

Effects of putrescine on the quality and epigenetic modification of mouse oocytes during in vitro maturation.

CONTEXT: Low ovarian putrescine levels and decreased peak values following luteinising hormone peaks are related to poor oocyte quantity and quality in ageing women. AIMS: To investigate the effects of putrescine supplementation in in vitro maturation (IVM) medium on oocyte quality and epigenetic modification. METHODS: Germinal vesicle oocytes retrieved from the ovaries of 8-week-old and 9-month-old mice were divided into four groups (the young, young+difluoromethylornithine (DFMO), ageing and ageing+putrescine groups) and cultured in IVM medium with or without 1mM putrescine or DFMO for 16h. The first polar body extrusion (PBE), cleavage and embryonic development were evaluated. Spindles, chromosomes, mitochondria and reactive oxygen species (ROS) were measured. The expression levels of SIRT1, H3K9ac, H3K9me2, H3K9me3, and 5mC levels were evaluated. Sirt1 and imprinted genes were detected. RESULTS: The PBE was higher in the ageing+putrescine group than in the ageing group. Putrescine increased the total and inner cell mass cell numbers of blastocysts in ageing oocytes. Putrescine decreased aberrant spindles and chromosome aneuploidy, increased the mitochondrial membrane potential and decreased ROS levels. Putrescine increased SIRT1 expression and attenuated the upregulation of H3K9ac levels in ageing oocytes. Putrescine did not affect 5mC, H3K9me2 or H3K9me3 levels or imprinted gene expression. CONCLUSIONS: Putrescine supplementation during IVM improved the maturation and quality of ageing oocytes and promoted embryonic development by decreasing ROS generation, maintaining mitochondrial and spindle function and correcting aberrant epigenetic modification. IMPLICATIONS: Putrescine shows application potential for human-assisted reproduction, especially for IVM of oocytes from ageing women.

Epigenetic effect of putrescine supplementation during in vitro maturation of oocytes on offspring in mice.

PURPOSE: To investigate the epigenetic safety of putrescine supplementation during in vitro maturation (IVM) to offspring. METHODS: Germinal vesicle oocytes retrieved from 12-week-old mice were randomly divided into two groups and cultured in IVM medium with or without 1 mmol/L putrescine for 16 h. Then, in vitro fertilization and embryo transplantation were conducted to produce the F1 offspring. The F1 mated with ordinary mice and bred the F2 offspring. The DNA methylation patterns in the brain and heart of F1 were investigated by reduced representation bisulfite sequencing. Imprinted gene expression levels of F1 oocytes were tested. The global methylation of F2 was examined by dot blot. RESULTS: The weight, organ coefficient, and histology were normal in the F1 and F2 offspring from the putrescine-treated oocytes. An overall methylation level of 31.23 to 32.53% was observed for all CpG sites in the brain and heart of the two groups. The DNA methylation patterns of the brain and heart in F1 were not altered in general, with subtle differences. The expression levels of imprinted genes including H19, Snrpn, Peg3, Igf2, and Igf2r did not statistically change. The global 5mC level of F2 was consistent with the control group. CONCLUSION: Putrescine supplementation during IVM did not directly affect the development, health, and reproduction, and did not affect the genome and global epigenetics of mouse offspring derived from those oocytes. The transient putrescine treatment for improving oocyte maturation shows its long-term safety of genome and epigenetics in the offspring of mice.

Sorbitol accumulation decreases oocyte quality in aged mice by altering the intracellular redox balance.

Sorbitol is a product of glucose metabolism through the polyol pathway. Many studies have demonstrated that excessive sorbitol can disrupt the intracellular redox balance. However, we still know very little about the impact of excessive intracellular sorbitol on oocyte quality, oocyte maturation, and embryo developmental potential. This study explored whether intracellular sorbitol accumulates in the oocytes of aged mice during in vitro maturation (IVM) and what roles sorbitol plays in oocyte development and maturation. Our results showed that sorbitol levels were significantly higher in in vitro-matured oocytes from aged mice than in oocytes from young mice (14.08 ± 3.78 vs. 0.23 ± 0.04 ng/oocyte). The expression of aldose reductase (AR) mRNA was significantly higher in the in vitro-cultured oocytes from 9-month-old mice than prior to culture. To decrease the excessive intracellular sorbitol in oocytes from aged mice, sorbinil, a specific inhibitor of aldose reductase, was supplemented in IVM medium, and the sorbitol level was significantly decreased (14.08 ± 3.78 vs. 0.48 ± 0.19 ng/oocyte). Our results indicated that the percentage of oocytes with first polar body extrusion (PBE) was significantly higher in the sorbinil group than in the aged group (82.4% ± 7.2% vs. 66.1% ± 6.9%), and the content of sorbitol was drastically increased in the aged group. The ROS fluorescence intensity in the sorbinil group was drastically lower than that in the aged group, while the GSH fluorescence intensity was significantly higher. Interestingly, SOD1 was upregulated in the sorbinil group. The present study suggests that excessive sorbitol accumulation is induced during IVM in aged mouse oocytes, which negatively influences oocyte quality by altering the intracellular redox balance. Inhibition of sorbitol accumulation may be a potential method to improve the nuclear maturation of aged oocytes.

Human amnion-derived mesenchymal stem cells improved the reproductive function of age-related diminished ovarian reserve in mice through Ampk/FoxO3a signaling pathway.

BACKGROUND: Age-related diminished ovarian reserve (AR-DOR) reduced the quality of oocytes, resulting in decreased female fertility. Aging is tightly related to abnormal distribution and function of mitochondria, while mitophagy is a major process to maintain normal quality and quantity of mitochondria in cells, especially in oocytes which containing a large number of mitochondria to meet the demand of energy production during oocyte maturation and subsequent embryonic development. Ampk/FoxO3a signaling is crucial in the regulation of mitophagy. It is reported mesenchymal stem cells (MSCs) can improve ovarian function. Here we aim to explore if human amnion-derived mesenchymal stem cells (hAMSCs) are effective in improving ovarian function in AR-DOR mice and whether Ampk/FoxO3a signaling is involved. METHODS: The AR-DOR model mice were established by 32-week-old mice with 3-8 litters, significantly low serum sex hormone levels and follicle counts. The old mice were divided into 5 treatment groups: normal saline (NS, control), 1% human serum albumin (HSA, resolver), low dose (LD, 5.0 × 106cells/kg), middle dose (MD, 7.5 × 106cells/kg), and high dose (HD, 10.0 × 106cells/kg). The prepared hAMSCs were injected through tail vein. Serum sex hormone level, follicle counts, fertilization rate, gestation rate, little size, apoptosis of granulosa and stromal cells, expression level of Sod2, Ampk, and ratio of phosphorylated FoxO3a to total FoxO3a in ovaries were examined. RESULTS: Our results show that after hAMSC transplantation, the ovarian function in AR-DOR mice was significantly improved, meanwhile the apoptosis of granulosa and stromal cells in the ovaries was significantly repressed, the expression level of Ampk and the ratio of phosphorylated FoxO3a to total FoxO3a both were significantly increased, meanwhile increased Sod2 expression was also observed. CONCLUSION: Our results demonstrate hAMSC transplantation via tail-injection can improve ovarian function of AR-DOR mice through Ampk/FoxO3a signaling pathway.

Peroxiredoxin 4 protects against ovarian ageing by ameliorating D-galactose-induced oxidative damage in mice.

Peroxiredoxin 4 (Prdx4), a member of the Prdx family, is a vital ER-resident antioxidant in cells. As revealed in our previous study, Prdx4 expression was detected in ovarian granulosa cells and was closely related to ovarian function. This research aimed to explore the effect and underlying molecular mechanism of the protective role of Prdx4 against D-gal-induced ovarian ageing in mice. The D-gal-induced ovarian ageing model has been extensively used to study the mechanisms of premature ovarian failure (POF). In this study, adult Prdx4-/- and wild-type mice were intraperitoneally injected with D-gal (150 mg/kg/day) daily for 6 weeks. Ovarian function, granulosa cell apoptosis, oxidative damage and ER stress in the ovaries were evaluated in the two groups. Ovarian weight was significantly lower, the HPO axis was more strongly disrupted, and the numbers of atretic follicles and apoptotic granulosa cells were obviously higher in Prdx4-/- mice. In addition, Prdx4-/- mice showed increased expression of oxidative damage-related factors and the ovarian senescence-related protein P16. Moreover, the levels of the proapoptotic factors CHOP and activated caspase-12 protein, which are involved in the ER stress pathway, and the level of the apoptosis-related BAX protein were elevated in the ovaries of Prdx4-/- mice. Thus, D-gal-induced ovarian ageing is accelerated in Prdx4-/- mice due to granulosa cell apoptosis via oxidative damage and ER stress-related pathways, suggesting that Prdx4 is a protective agent against POF.

The paleoclimatic footprint in the soil carbon stock of the Tibetan permafrost region.

Tibetan permafrost largely formed during the late Pleistocene glacial period and shrank in the Holocene Thermal Maximum period. Quantifying the impacts of paleoclimatic extremes on soil carbon stock can shed light on the vulnerability of permafrost carbon in the future. Here, we synthesize data from 1114 sites across the Tibetan permafrost region to report that paleoclimate is more important than modern climate in shaping current permafrost carbon distribution, and its importance increases with soil depth, mainly through forming the soil's physiochemical properties. We derive a new estimate of modern soil carbon stock to 3 m depth by including the paleoclimate effects, and find that the stock ([Formula: see text] PgC) is triple that predicted by ecosystem models (11.5 ± 4.2 s.e.m PgC), which use pre-industrial climate to initialize the soil carbon pool. The discrepancy highlights the urgent need to incorporate paleoclimate information into model initialization for simulating permafrost soil carbon stocks.

Deciphering impacts of climate extremes on Tibetan grasslands in the last fifteen years.

Climate extremes have emerged as a crucial driver of changes in terrestrial ecosystems. The Tibetan Plateau, facing a rapid climate change, tends to favor climate extremes. But we lack a clear understanding of the impacts of such extremes on alpine grasslands. Here we show that extreme events (drought, extreme wet, extreme cold and extreme hot) occurred at a frequency of 0.67-4 months decade-1 during 2001-2015, with extreme precipitation predominantly occurring in June-to-August and extreme temperatures in May. Drought and extreme wet cause opposite and asymmetric effects on grassland growth, with drought-induced reductions greater than increases due to extreme wet. Grassland responses to extreme temperatures, which predominantly occur in May, show a dipole-like spatial pattern, with extreme hot (cold) events enhanced (reduced) growth in the eastern plateau but slightly reduced (enhanced) growth in the western plateau. These opposite responses to extreme temperatures over the eastern plateau are explained by the possibility that the occurrence of extreme cold slows the preseason temperature accumulation, delaying the triggering of spring phenology, while extreme hot hastens the accumulation. In the western plateau, in contrast, positive responses to extreme cold are induced by accompanying high precipitation. Furthermore, high extremeness of climate events generally led to a much lower extremeness in growth response, implying that the Tibetan grasslands have a relatively high resistance to climate extremes. The ecosystem models tested could not accurately simulate grassland responses to drought and extreme temperatures, and require re-parameterization before trust can be placed in their output for this region.

Putrescine delays postovulatory aging of mouse oocytes by upregulating PDK4 expression and improving mitochondrial activity.

If fertilization does not occur for a prolonged period in vivo or in vitro, the postovulatory oocytes will deteriorate, which called the postovulatory aging. This process disrupts the developmental competence. In the present study, we showed that the reactive oxygen species (ROS) was accumulated in oocytes during the postovulatory aging. ROS inhibited Sirt1 expression, and then increased oxidative stress by downregulating the intracellular Sirt1-FOXO3a-SOD2 axis. Moreover, the inhibited Sirt1 expression was related to the decreased mitochondrial function and the lowered level of autophagy. The mitochondrial-related apoptosis was increased by inhibiting the AKT and ERK1/2 pathways, due to the accumulation of ROS in the postovulatory oocytes. The mitochondrial pyruvate dehydrogenase kinase-4 (PDK4) can reduce ROS by inhibiting the tricarboxylic acid (TAC) cycle. We found that PDK4 was significantly decreased in the postovulatory aging oocytes. Putrescine, one of the abundant biogenic amines, ameliorated the effects of ROS and therefore improved the quality of the postovulatory aging oocytes by increasing the expression of PDK4. When PDK4 was downregulated using siRNAs, the effects of putrescine were significantly receded. We concluded that putrescine delayed the aging process of postovulatory oocytes by upregulating PDK4 expression and improving mitochondrial activity.

Curcumin exerts a protective effect against premature ovarian failure in mice.

This study was designed to investigate the protective effect of curcumin against d-galactose (d-gal)-induced premature ovarian failure (POF) in mice. A mouse POF model was induced by subcutaneous injection of d-gal (200 mg/kg/day) daily for 42 days. Mice in the curcumin group received both d-gal treatment and intraperitoneal injection of curcumin (100 mg/kg/day) for 42 days. Ovarian function, oxidative stress and apoptosis were evaluated. The P, E2 and SOD levels were higher, and the FSH, LH and MDA levels were significantly lower in the curcumin group than those in the d-gal group. The proportion of primordial follicles was also significantly higher in the curcumin group than that in the d-gal group. In addition, curcumin treatment after d-gal administration resulted in significantly lower Sod2, Cat, 8-OhdG, 4-HNE, NTY and senescence-associated protein P16 expression levels, higher Amh expression levels and less apoptosis in granulosa cells than was observed in the d-gal group. Moreover, the p-Akt, Nrf2 and HO-1 protein expression levels were significantly higher and the apoptosis-related cleaved caspase-3 and -9 protein expression levels were markedly lower in the curcumin group than in the d-gal group. In conclusion, curcumin effectively inhibited d-gal-induced oxidative stress, apoptosis and ovarian injury via a mechanism involving the Nrf2/HO-1 and PI3K/Akt signaling pathways, suggesting that curcumin is a potential protective agent against POF.

Analysis of Pigeon (Columba) Ovary Transcriptomes to Identify Genes Involved in Blue Light Regulation.

Monochromatic light is widely applied to promote poultry reproductive performance, yet little is currently known regarding the mechanism by which light wavelengths affect pigeon reproduction. Recently, high-throughput sequencing technologies have been used to provide genomic information for solving this problem. In this study, we employed Illumina Hiseq 2000 to identify differentially expressed genes in ovary tissue from pigeons under blue and white light conditions and de novo transcriptome assembly to construct a comprehensive sequence database containing information on the mechanisms of follicle development. A total of 157,774 unigenes (mean length: 790 bp) were obtained by the Trinity program, and 35.83% of these unigenes were matched to genes in a non-redundant protein database. Gene description, gene ontology, and the clustering of orthologous group terms were performed to annotate the transcriptome assembly. Differentially expressed genes between blue and white light conditions included those related to oocyte maturation, hormone biosynthesis, and circadian rhythm. Furthermore, 17,574 SSRs and 533,887 potential SNPs were identified in this transcriptome assembly. This work is the first transcriptome analysis of the Columba ovary using Illumina technology, and the resulting transcriptome and differentially expressed gene data can facilitate further investigations into the molecular mechanism of the effect of blue light on follicle development and reproduction in pigeons and other bird species.

Genomic expression profiles in cumulus cells derived from germinal vesicle and MII mouse oocytes.

Cumulus cells (CCs) are distinct from other granulosa cells and the mutual communication between CCs and oocytes is essential for the establishment of oocyte competence. In the present study we assessed genomic expression profiles in mouse CCs before and after oocyte maturation in vitro. Microarray analysis revealed significant changes in gene expression in CCs between the germinal vesicle (GV) and metaphase II (MII) stages, with 2615 upregulated and 2808 downregulated genes. Genes related to epidermal growth factor, extracellular matrix (Ptgs2, Ereg, Tnfaip6 and Efemp1), mitochondrial metabolism (Fdx1 and Aifm2), gap junctions and the cell cycle (Gja1, Gja4, Ccnd2, Ccna2 and Ccnb2) were highlighted as being differentially expressed between the two development stages. Real-time polymerase chain reaction confirmed the validity and reproducibility of the results for the selected differentially expressed genes. Similar expression patterns were identified by western blot analysis for some functional proteins, including EFEMP1, FDX1, GJA1 and CCND2, followed by immunofluorescence localisation. These genes may be potential biomarkers for oocyte developmental competence following fertilisation and will be investigated further in future studies.

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.

[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.

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.

[Clinical effects of oocyte cryopreservation in assisted reproduction technology].

OBJECTIVE: To investigate the clinical effects of oocyte cryopreservation in assisted reproduction technology (ART). METHODS: 258 patients undergoing retrieval of more than 20 oocytes during in vitro fertilization/intracytoplasmic sperm injection (IVF/ICSI) were divided into 2 groups:Group A, undergoing surplus oocytes cryopreservation (84 cycles) and Group B undergoing embryo cryopreservation (174 cycles) according to the patients' choices. Fertilization rate and clinical pregnancy rate of fresh embryo transfer cycle were compared between these two groups. Twenty-three infertile couples' frozen oocytes were thawed for further ART treatment. Among them, fifteen couples received embryo transfer using their own frozen-thawed oocytes, and other four couples used donated frozen-thawed oocytes. The survival rate, fertilization rate, cleavage rate, implantation rate, and clinical pregnancy rate of these 19 cycles were compared to the outcome of 56 frozen-thawed embryo transfer cycles. RESULTS: The fertilization rate of Group A who underwent IVF was 65.9%, not significantly different from that of Group B who received IVF (66.9%, P > 0.05), and the fertilization rate of Group A who underwent ICSI was 71.6%, not significantly different from that of Group B who received ICSI (64.1%, P > 0.05). The clinical pregnancy rate (per embryo transfer cycle) of Group A who received IVF was 52.9%, not significantly different from that of Group B who received IVF (42.3%, P > 0.05), and the clinical pregnancy rate (per embryo transfer cycle) of Group A who received ICSI was 35.5%, not significantly different from that of Group B who received ICSI (34.4%, P > 0.05). The clinical pregnancy rate of frozen-thawed oocyte group (per embryo transfer cycle) was 47.4% (9/19). Four couples used donated frozen-thawed oocytes, two of them got clinical pregnancy and one of them had term delivery. CONCLUSION: For women who undergo retrieval of more than 20 oocytes in IVF/ICSI, the clinical outcome of fresh embryo transfer cycle, such as fertilization rate and clinical pregnancy rate, are not influenced by oocyte cryopreservation and embryo cryopreservation. There is no significant difference in the clinical pregnancy rate (per embryo transfer cycle) between frozen-thawed oocyte group and frozen-thawed embryo group. Compared with embryo cryopreservation, oocyte cryopreservation has obvious advantages in fertility preservation and oocyte donation.

The clinical significance of sperm DNA damage detection combined with routine semen testing in assisted reproduction.

Traditional routine semen analysis and biochemical assays are insufficient for the determination of fertility status in individual men. In recent years, clinical evidence has shown that damaged human sperm DNA may adversely affect assisted reproductive outcomes and that the spermatozoa of infertile men possess substantially more DNA damage than the spermatozoa of fertile men. In this study, we combined these two methods to test their clinical significance in assisted reproductive techniques (ART). A total of 302 in vitro fertilization (IVF) and 67 intracytoplasmic sperm injection (ICSI) couples were included in the study. The acridine orange technique (AOT) was used to detect the DNA integrity of the sperm. Correlation analysis showed that the DNA fragmentation index (DFI) was negatively related to the no. of good quality embryos and the clinical pregnancy rate in IVF patients. No significant correlations were found between DFI and ICSI patients. We then combined the results of the routine semen testing of IVF patients with their DFI values. Statistical analysis revealed notable differences in the no. of good quality embryos (P=0.015) and the clinical pregnancy rate (P=0.015) between subgroups divided according to DFI value in a group with normal semen test results (Group 1). In a group with abnormal semen test results (Group 2), the fertilization rate (P=0.034) and the pregnancy rate (P=0.018) showed remarkable variation between DFI subgroups. In conclusion, the detection of damaged DNA in spermatozoa needs to be conducted along with standard semen analysis. This might prove to be a promising predictor of ART outcome, particularly in IVF.