Characterization of the Sperm Proteome and Reproductive Outcomes with in Vitro, Fertilization after a Reduction in Male Ejaculatory Abstinence Period.

Semen samples from men after a short ejaculatory abstinence show improved sperm quality and result in increased pregnancy rates, but the underlying mechanisms remain unclear. Herein, we report that ejaculates from short (1-3 h) compared with long (3-7 days) periods of abstinence showed increases in motile sperm count, sperm vitality, normal sperm morphology, acrosome reaction capacity, total antioxidant capacity, sperm mitochondrial membrane potential, high DNA stainability, and a decrease in the sperm DNA fragmentation index (p, < 0.05). Sperm proteomic analysis showed 322 differentially expressed proteins (minimal fold change of ±1.5 or greater and p, < 0.05), with 224 upregulated and 98 downregulated. These differentially expressed proteins are profoundly involved in specific cellular processes, such as motility and capacitation, oxidative stress, and metabolism. Interestingly, protein trimethyllysine modification was increased, and butyryllysine, propionyllysine, and malonyllysine modifications were decreased in ejaculates from a short versus, long abstinence (p, < 0.05). Finally, the rates of implantation, clinical pregnancy, and live births from in vitro, fertilization treatments were significantly increased in semen samples after a short abstinence. Our study provides preliminary mechanistic insights into improved sperm quality and pregnancy outcomes associated with spermatozoa retrieved after a short ejaculatory abstinence.

Regulation of blood-testis barrier assembly in vivo by germ cells.

The assembly of the blood-testis barrier (BTB) during postnatal development is crucial to support meiosis. However, the role of germ cells in BTB assembly remains unclear. Herein, KitW/KitWV mice were used as a study model. These mice were infertile, failing to establish a functional BTB to support meiosis due to c-Kit mutation. Transplantation of undifferentiated spermatogonia derived from normal mice into the testis of KitW/KitWV mice triggered functional BTB assembly, displaying cyclic remodeling during the epithelial cycle. Also, transplanted germ cells were capable of inducing Leydig cell testosterone production, which could enhance the expression of integral membrane protein claudin 3 in Sertoli cells. Early spermatocytes were shown to play a vital role in directing BTB assembly by expressing claudin 3, which likely created a transient adhesion structure to mediate BTB and cytoskeleton assembly in adjacent Sertoli cells. In summary, the positive modulation of germ cells on somatic cell function provides useful information regarding somatic-germ cell interactions.-Li, X.-Y., Zhang, Y., Wang, X.-X., Jin, C., Wang, Y.-Q., Sun, T.-C., Li, J., Tang, J.-X., Batool, A., Deng, S.-L., Chen, S.-R., Cheng, C. Y., Liu, Y.-X. Regulation of blood-testis barrier assembly in vivo by germ cells.

CRISPR/Cas9-mediated genome editing induces gene knockdown by altering the pre-mRNA splicing in mice.

BACKGROUND: Clustered Regularly Interspaced Short Palindromic Repeats/CRISPR associated protein 9 (CRISPR/Cas9) has been wildly used to generate gene knockout models through inducing indels causing frame-shift. However, there are few studies concerning the post-transcript effects caused by CRISPR-mediated genome editing. RESULTS: In the present study, we showed that gene knockdown model also could be generated using CRISPR-mediated gene editing by disrupting the boundary of exon and intron in mice (C57BL/6 J). CRISPR induced indel at the boundary of exon and intron (5' splice site) caused alternative splicing and produced multiple different mRNAs, most of these mRNAs introduced premature termination codon causing down expression of the gene. CONCLUSIONS: These results showed that alternative splicing mutants were able to generate through CRISPR-mediated genome editing by deleting the boundary of exon and intron causing disruption of 5' splice site. Although alternative splicing was an unexpected outcome, this finding could be developed as a technology to generate gene knockdown models or to investigate pre-mRNA splicing.

Autophagy is activated in the ovarian tissue of polycystic ovary syndrome.

The importance of autophagy in polycystic ovary syndrome (PCOS)-related metabolic disorders is increasingly being recognized, but few studies have investigated the role of autophagy in PCOS. Here, transmission electron microscopy demonstrated that autophagy was enhanced in the ovarian tissue from both humans and rats with PCOS. Consistent with this, ovarian granulosa cells from PCOS rats showed increases in the autophagy marker protein light chain 3B (LC3B), whereas levels of the autophagy substrate SQSTM1/p62 were decreased. In addition, the ratio of LC3-II/LC3-I was markedly elevated in human PCOS ovarian tissue compared with normal ovarian tissue. Real-time PCR arrays indicated that 7 and 34 autophagy-related genes were down- and up-regulated in human PCOS , Signal-Net, and regression analysis suggested that there are a wide range of interactions among these 41 genes, and a potential network based on EGFR, ERBB2, FOXO1, MAPK1, NFKB1, IGF1,TP53 and MAPK9 may be responsible for autophagy activation in PCOS. Systematic functional analysis of 41 differential autophagy-related genes indicated that these genes are highly involved in specific cellular processes such as response to stress and stimulus, and are linked to four significant pathways, including the insulin, ERBB, mTOR signaling pathways and protein processing in the endoplasmic reticulum. This study provides evidence for a potential role of autophagy disorders in PCOS in which autophagy may be an important molecular event in the pathogenesis of PCOS.

Hyperhomocysteinemia in polycystic ovary syndrome: decreased betaine-homocysteine methyltransferase and cystathionine ß-synthase-mediated homocysteine metabolism.

RESEARCH QUESTION: What are the metabolic characteristics of homocysteine in polycystic ovary syndrome (PCOS)? DESIGN: Homocysteine concentrations were determined in serum samples from non-obese and obese control subjects and PCOS patients. Homocysteine metabolism was studied in a rat model of PCOS established using dehydroepiandrosterone (DHEA) or DHEA in combination with a high-fat diet (HFD). RESULTS: It was shown that (i) serum homocysteine concentrations were greater in PCOS patients than in control subjects in the obese group (P < 0.05) and serum homocysteine concentrations were significantly higher in the obese group than in the non-obese group, regardless of PCOS status (both P < 0.05); (ii) serum homocysteine concentrations were significantly increased in DHEA + HFD-induced rats compared with controls (P < 0.05); (iii) when compared with the control group, mRNA concentrations of homocysteine metabolic enzymes Bhmt and Cbs were significantly reduced in the liver tissues of DHEA + HFD-induced rats (both P < 0.0001); (iv) when compared with the control group, there was a significant decrease in the methylation concentrations of the Cbs (P < 0.05) and Bhmt (P < 0.05 and P < 0.0001) promoter in the DHEA + HFD group. The methylation patterns, together with previous data, indicate that hypomethylated promoter-mediated transcriptional activation of Bhmt and Cbs might be a defence mechanism against PCOS-related hyperhomocysteinemia. CONCLUSIONS: These findings indicate that decreased liver Bhmt and Cbs-mediated homocysteine metabolism might have a role in hyperhomocysteinemia in PCOS and provides further evidence for a potential role of decreased liver function in PCOS.

Assessment of the development of assisted reproductive technology in Liaoning province of China, from 2012 to 2016.

BACKGROUND: The development of assisted reproduction techniques (ART) has resulted in rapid advances in the treatment of infertility. However, a systematic assessment of ART and its processes and outcomes in China has never been carried out. The goal of this study was to assess the features of ART status from 2012 to 2016 in clinics and in vitro fertilization (IVF) laboratories in Liaoning, the largest IVF province in the northeast of China. METHODS: Data from Jan 1, 2012 to Dec 31, 2016 was retrieved from the assisted reproductive certificate registry of Liaoning province. We extracted data from: i) fresh and thawed cycles; ii) donor sperm and donor egg cycles; iii) intrauterine insemination with husband semen and donor semen (AIH and AID). RESULTS: We showed that: (i) there has been a significant increase in the number of IVF fresh and thawed cycles, and the proportion of cases of primary infertility and secondary infertility has decreased and increased, respectively; (ii) standard long GnRH agonist was the major ovarian stimulation protocol. During the observation period, increasing trends in the use of GnRH antagonists, mild stimulation, and natural cycles were observed; (iii) significant differences in the number of cycles, number of retrieved oocytes, fertilization rates, implantation rates, and sex ratio were noticed between conventional IVF and intracytoplasmic sperm injection; (iv) significant differences in age at treatment for infertility, number of cycles, and ectopic pregnancy rates were noticed between donor sperm cycles and donor egg cycles; (v) significant differences in number of thawed cycles, number of thawed embryos, embryo recovery rates, implantation rates, and clinical pregnancy rates were noticed between day 3 and day 5 embryos; (vi) significant differences in age at treatment for infertility, number of cycles, clinical pregnancy rates, ectopic pregnancy rates, and live birth ratio were noticed between AIH and AID. CONCLUSIONS: ART in Liaoning province has undergone substantial development from 2012 to 2016 in clinics and IVF laboratories. This presentation of detailed ART data will provide researchers, policy makers, and potential ART users a rich source of information about IVF characteristics in the northeast of China.

Role of WNT signaling in epididymal sperm maturation.

PURPOSE: Spermatozoa maturation, a process required for spermatozoa to acquire progressive motility and the ability to fertilize ova, primarily occurs in the caput and corpus of the epididymis. Despite considerable efforts, the factor(s) promoting epididymal sperm maturation remains unclear. Recently, WNT signaling has been implicated in epididymal sperm maturation. METHODS: To further investigate WNT signaling function in epididymal sperm maturation, we generated Wntless conditional knockout mice (Wls cKO), Wls flox/flox ; Lcn5-Cre. RESULTS: In these mice, WNTLESS (WLS), a conserved membrane protein required for all WNT protein secretion, was specifically disrupted in the principal cells of the caput epididymidis. Immunoblot analysis showed that WLS was significantly reduced in the caput epididymidis of Wls cKO mice. In the caput epididymidis of Wls cKO mice, WNT 10A and WNT 2b, which are typically secreted by the principal cells of the caput epididymis, were not secreted. Interestingly, sperm motility analysis showed that the WLS deficiency in the caput epididymidis had no effect on sperm motility. Moreover, fertility tests showed that Wls cKO male mice had normal fertility. CONCLUSION: These results indicate that the disruption of WLS in principal cells of the caput epididymidis inhibits WNT protein secretion but has no effect on sperm motility and male fertility, suggesting that WNT signaling in the caput epididymidis may be dispensable for epididymal sperm maturation in mice.

EZH2 deletion promotes spermatogonial differentiation and apoptosis.

Spermatogenesis is crucial for male fertility and is therefore tightly controlled by a variety of epigenetic regulators. However, the function of enhancer of zeste homolog 2 (EZH2) in spermatogenesis and the molecular mechanisms underlying its activity remain poorly defined. Here, we demonstrate that deleting EZH2 promoted spermatogonial differentiation and apoptosis. EZH2 is expressed in spermatogonia, spermatocytes and round and elongated spermatids from stage 9 to 11 but not in leptotene and zygotene spermatocytes. Knocking down Ezh2 in vitro using a lentivirus impaired self-renewal in spermatogonial stem cells (SSCs), and the conditional knockout of Ezh2 in spermatogonial progenitors promoted precocious spermatogonial differentiation. EZH2 functions to balance self-renewal and differentiation in spermatogonia by suppressing NEUROG3 and KIT via a direct interaction that is independent of its histone methyltransferase activity. Moreover, deleting Ezh2 enhanced the activation of CASP3 in spermatids, resulting in reduced spermatozoa production. Collectively, these data demonstrate that EZH2 plays a nonclassical role in the regulation of spermatogonial differentiation and apoptosis in murine spermatogenesis.

Selective deletion of Smad4 in postnatal germ cells does not affect spermatogenesis or fertility in mice.

SMAD4 is the central component of canonical signaling in the transforming growth factor beta (TGFß) superfamily. Loss of Smad4 in Sertoli cells affects the expansion of the fetal testis cords, whereas selective deletion of Smad4 in Leydig cells alone does not appreciably alter fetal or adult testis development. Loss of Smad4 in Sertoli and Leydig cells, on the other hand, leads to testicular dysgenesis, and tumor formation in mice. Within the murine testes, Smad4 is also expressed in germ cells of the seminiferous tubules. We therefore, crossed Ngn3-Cre or Stra8-Cre transgenic mice with Smad4-flox mice to generate conditional knockout animals in which Smad4 was specifically deleted in postnatal germ cells to further uncover cell type-specific requirement of Smad4. Unexpectedly, these germ-cell-knockout mice were fertile and did not exhibit any detectable abnormalities in spermatogenesis, indicating that Smad4 is not required for the production of sperm; instead, these data indicate a cell type-specific requirement of Smad4 primarily during testis development. Mol. Reprod. Dev. 83: 615-623, 2016. © 2016 Wiley Periodicals, Inc.

Melatonin promotes development of haploid germ cells from early developing spermatogenic cells of Suffolk sheep under in vitro condition.

Promotion of spermatogonial stem cell (SSC) differentiation into functional sperms under in vitro conditions is a great challenge for reproductive physiologists. In this study, we observed that melatonin (10(-7) M) supplementation significantly enhanced the cultured SSCs differentiation into haploid germ cells. This was confirmed by the expression of sperm special protein, acrosin. The rate of SSCs differentiation into sperm with melatonin supplementation was 11.85 ± 0.93% which was twofold higher than that in the control. The level of testosterone, the transcriptions of luteinizing hormone receptor (LHR), and the steroidogenic acute regulatory protein (StAR) were upregulated with melatonin treatment. At the early stage of SSCs culture, melatonin suppressed the level of cAMP, while at the later stage, it promoted cAMP production. The similar pattern was observed in testosterone content. Expressions for marker genes of meiosis anaphase, Dnmt3a, and Bcl-2 were upregulated by melatonin. In contrast, Bax expression was downregulated. Importantly, the in vitro-generated sperms were functional and they were capable to fertilize oocytes. These fertilized oocytes have successfully developed to the blastula stage.

[Comparison of clinical outcome of mild-stimulation and conventional ovarian stimulation in in vitro fertilization-embryo transfer (IVF-ET)].

OBJECTIVE: To investigate the practical application value of the mild-stimulation method used in in vitro fertilization and embryo transfer (IVF-ET) treatment for IVF patients with normal ovarian function. METHODS: The patients (ages from 26 to 38 years) with normal ovarian function, who were treated with IVF-ET in Affiliated Reproductive Center of China Medical University from January 2004 to July 2012 were selected as our samples. We compared the clinical results of treatment between the mild-stimulation (104 cycles in experimental group) and the conventional stimulation (198 cycles in control group). RESULTS: The number of retrieved oocytes (3.1±1.6 vs. 10.2±5.7, P<0.05) and clinical pregnancy rate [34% (33/97) vs. 49.7% (93/187), P<0.05] of the mild-stimulation group (104 cycles) were lower than those of the control group (198 cycles), however, the duration of stimulation (4.7±2.1 vs. 9.16±1.87) and total doses of 75 U gonadotropins used (5.7±3.7 vs. 30.9±10.3) were also lower in the mild-stimulation group than in the control group obviously (P<0.05). CONCLUSION: For the normal ovarian function patients, the mild-stimulation method has clear clinical effects, but not as good as the conventional-stimulation one.

[Evaluation for the clinical application of all embryos cryopreservation: a multi-centre study in northern area of China].

OBJECTIVE: To investigate the clinical outcome of patients who underwent cryopreservation of all embryos. METHODS: A retrospective analysis was made in the clinical data of patients with cryopreservation of all embryos between April 2011 and September 2011 in four hospitals of North China. The patients were divided into five groups according to the reasons of cryopreservation of all embryos: ovarian hyperstimulation syndrome (OHSS) group, serum progesterone elevation group, endometrial group, hydrosalpinx group and others. The clinical pregnancy rate per transfer, implantation rate and cumulative clinical pregnancy rate were analyzed. RESULTS: The clinical pregnancy rate, implantation rate and cumulative clinical pregnancy rate of the OHSS group were 55.4%, 34.8% and 73.7%, respectively. The rates of the serum progesterone elevation group were 25.5%, 11.2% and 43.1%, respectively. The rates of the endometrial group were 54.8%, 34.4% and 61.5%, respectively. The rates of the hydrosalpinx group were 60%, 30% and 60%, respectively. The rates of the other factors group were 36.0%, 24.5% and 44.0%, respectively. CONCLUSION: The strategy of cryopreservation of all embryos could improve the clinical outcomes of patients with severe OHSS. It still needs a large multi-centre, randomized trial to evaluate its effectiveness and side effects, although it has the positive clinical application for other reasons of cryopreservation of all embryos.

[Pregnancy outcomes of in vitro fertilization and embryo transfer in infertile women with polycystic ovarian syndrome].

OBJECTIVE: To investigate the pregnancy outcomes of in vitro fertilization (IVF) and embryo transfer (ET) in infertile women with polycystic ovarian syndrome (PCOS). METHODS: From August 2005 to June 2011, 200 IVF-ET cycles performed in women with polycystic ovarian syndrome in Shengjing Hospital and Shenyang 204 Hospital were enrolled in this retrospective study, matched with 400 IVF-ET cycles in infertile women with fallopian tube factors as control group. The incidence of abortion, gestational diabetes mellitus (GDM), hypertensive disorders in pregnancy, preterm birth, small for gestational age infant (SGA), large for gestational age infant (LGA), neonatal asphyxia, neonatal death and deformity was compared between two groups. RESULTS: The incidence of spontaneous abortion was 26.0% (52/200) in PCOS group, which was significantly higher than 10.2% (41/400) in control group (P < 0.05). The incidence of GDM, hypertensive disorders in pregnancy, preterm birth, cesarean section in PCOS group was 23.6% (35/148), 16.2% (24/148), 17.6% (26/148), 83.1% (123/148), which were significantly higher than 4.2% (15/359), 6.1% (22/359), 7.8% (28/359), 73.8% (265/359) in control group (P < 0.05). The incidence of SGA, LGA, neonatal asphyxia, neonatal death and deformity did not show remarkable difference between two groups, which were 2.7% (4/148), 4.7% (7/148), 5.4% (8/148), 0 in PCOS group and 1.4% (5/359), 2.2% (8/359), 2.8% (10/359), 0 in control group (P > 0.05). CONCLUSION: IVF-ET is an effective treatment for infertile women with PCOS, however, the incidence of spontaneous abortion, GDM, hypertensive disorders in pregnancy, preterm birth, cesarean section in PCOS patients was increased.

[Outcomes of IVF-ET and percentage of morphologically normal sperm before and after swim-up treatment].

OBJECTIVE: To study the relationship between sperm morphology and the outcomes of in vitro fertilization-embryo transfer (IVF-ET) before and after swim-up treatment of sperm on the day of oocyte retrieval. METHODS: This study included 94 couples to be treated by IVF-ET for tubal factor infertility. Sperm samples were collected on the day of oocyte retrieval and sperm morphology evaluated according to the Kruger criteria before and after swim-up treatment. Based on the results of morphological evaluation, the sperm samples were divided into groups A1 (morphologically normal sperm > or = 10% after swim-up treatment), A2 (morphologically normal sperm < 10% after swim-up treatment), B1 (morphologically normal sperm > or = 10% before swim-up treatment) and B2 (morphologically normal sperm < 10% before swim-up treatment). The outcomes of IVF-ET treatment were compared between groups A1 and A2 as well as between B1 and B2. RESULTS: After swim-up treatment, the rates of fertilization, cleavage, good quality embryo, clinical pregnancy and embryo implantation of group A1 were (72.72 +/- 3.35)%, (95.64 +/- 2.04)%, (24.39 +/- 4.57)%, 50.00% and 23.87%, respectively, while those of group A2 were (70.27 +/- 8.82)%, (94.82 +/- 4.94)%, (13.45 +/- 7.39)%, 9.52% and 6.25%, respectively, the latter three indexes remarkably higher in A1 than in A2 (P < 0.05), but the differences in the former two not statistically significant (P > 0.05). Before swim-up treatment, the above five indexes were (72.90 +/- 4.23)%, (95.20 +/- 2.61)%, (23.35 +/- 5.19)%, 39.58% and 18.35% in group B1, as compared with (71.33 +/- 5.10)%, (95.71 +/- 2.88)%, (20.18 +/- 6.15)%, 41.86% and 21.28% in group B2, with no statistically significant differences between the two groups (P > 0.05). CONCLUSION: The percentage of morphologically normal sperm after swim-up treatment on the day of oocyte retrieval may be a valuable predictor of the outcomes of IVF-ET.

Extremely fast-charging lithium ion battery enabled by dual-gradient structure design.

Extremely fast-charging lithium-ion batteries are highly desirable to shorten the recharging time for electric vehicles, but it is hampered by the poor rate capability of graphite anodes. Here, we present a previously unreported particle size and electrode porosity dual-gradient structure design in the graphite anode for achieving extremely fast-charging lithium ion battery under strict electrode conditions. We develop a polymer binder-free slurry route to construct this previously unreported type particle size-porosity dual-gradient structure in the practical graphite anode showing the extremely fast-charging capability with 60% of recharge in 10 min. On the basis of dual-gradient graphite anode, we demonstrate extremely fast-charging lithium ion battery realizing 60% recharge in 6 min and high volumetric energy density of 701 Wh liter-1 at the high charging rate of 6 C.

Superior Fast-Charging Lithium-Ion Batteries Enabled by the High-Speed Solid-State Lithium Transport of an Intermetallic Cu6 Sn5 Network.

Superior fast charging is a desirable capability of lithium-ion batteries, which can make electric vehicles a strong competition to traditional fuel vehicles. However, the slow transport of solvated lithium ions in liquid electrolytes is a limiting factor. Here, a Lix Cu6 Sn5 intermetallic network is reported to address this issue. Based on electrochemical analysis and X-ray photoelectron spectroscopy mapping, it is demonstrated that the reported intermetallic network can form a high-speed solid-state lithium transport matrix throughout the electrode, which largely reduces the lithium-ion-concentration polarization effect in the graphite anode. Employing this design, superior fast-charging graphite/lithium cobalt oxide full cells are fabricated and tested under strict electrode conditions. At the charging rate of 6 C, the fabricated full cells show a capacity of 145 mAh g-1 with an extraordinary capacity retention of 96.6%. In addition, the full cell also exhibits good electrochemical stability at a high charging rate of 2 C over 100 cycles (96.0% of capacity retention) in comparison to traditional graphite-anode-based cells (86.1% of capacity retention). This work presents a new strategy for fast-charging lithium-ion batteries on the basis of high-speed solid-state lithium transport in intermetallic alloy hosts.

Embryo selection through non-invasive preimplantation genetic testing with cell-free DNA in spent culture media: a protocol for a multicentre, double-blind, randomised controlled trial.

INTRODUCTION: Morphological evaluation is used to select embryos for in vitro fertilisation. However, it does not fully reflect the implantation potential. Preimplantation genetic testing for aneuploidies (PGT-A) can detect embryonic aneuploidy, but biopsy procedure is invasive. Currently, a non-invasive PGT (ni-PGT) approach using spent medium is being evaluated. However, the clinical benefit of ni-PGT has not been clearly demonstrated. A multicentre randomised trial is needed to verify whether ni-PGT can be an new effective tool for evaluating embryos. METHODS AND ANALYSIS: Overall, 1148 couples aged 35~42 (women) receiving in vitro fertilization-intracytoplasmic sperm injection are planned to be enrolled. Couples will be digitally randomised to (1) ni-PGT and (2) conventional morphology groups at a 1:1 treatment ratio. The primary outcome will be the ongoing pregnancy rate related to the first transfer cycle within 6 months after oocyte retrieval. ETHICS AND DISSEMINATION: The study protocol is approved by the Ethics Committee of Peking University Third Hospital and the participating hospitals. The results will be disseminated through international conferences and scientific journals. TRIAL REGISTRATION NUMBER: NCT04339166.

Association between prenatal air pollution exposure and risk of hypospadias in offspring: a systematic review and meta-analysis of observational studies.

BACKGROUND: The findings of associations between prenatal air pollution exposure and hypospadias risk in offspring are inconsistent. No systematic review or meta-analysis has yet summarized the present knowledge on the aforementioned topic. METHODS: Relevant manuscripts were identified by searching PubMed and Web of Science databases through January 31, 2020. Summary odds ratios (ORs) with 95% confidence intervals (CIs) in meta-analyses were estimated based on a random effects model. Publication bias was evaluated by funnel plots, Begg's test, and Egger's test. RESULTS: The search identified 3,032 relevant studies. Sixteen studies cumulatively involving 21,701 hypospadias cases and 1,465,364 participants were included. All of these studies were classified as having a low risk of bias. We classified pollutants as nitrogen oxides, particulate matter (PM), ozone, and other exposures. The exposure window to pollutants varied from three months before conception to seven days after delivery. In the meta-analyses, only PM2.5 exposure in the first trimester was related to increased risk of hypospadias (per 10 µg/m3 OR = 1.34; 95% CI: 1.06-1.68). CONCLUSION: We found evidence for an effect of PM2.5 exposure on hypospadias risk. Improvements in the areas of study design, exposure assessment, and specific exposure window are needed to advance this field.

[Surveying on menstruation status of reproductive age women of Han nationality in Liaoning province in 2008].

OBJECTIVE: To investigate the menstruation status of reproductive age women of Han Nationality in Liaoning province. METHODS: From Apr. 2008 to Dec. 2008, 1611 women at age of 19-45 years from Shenyang, Yingkou, Benxi, Zhangwu were enrolled in this study according to epidemiologic cluster sampling method. The study was performed by questionnaire consisting of age of menarche, regularity of menstruation, menstrual cycle and dysmenorrhea, et al. RESULTS: In the survey of 1611 women, the average menarche age is 14.4 years old, there were significant difference on menarche between (14.2 +/- 1.5) years in women from city and (14.6 +/- 1.5) years in women from country (t = 6.58, P < 0.01). The linear regression statistic method was used to analyze the relationship between age and menarche age, the linear regression equation was gotten as Y = 0.074X + 11.855, which means 1 year increase was associated with decrease by 0.074 years in menarche age approximately. About 86.34% (1391/1611) of women have normal and regular menstrual cycle at range of 21 to 35 days, while 11.05% (178/1611) of women have longer menstrual cycle (> 35 days) and 2.61% (42/1611) of women have shorter menstrual cycle (< 21 days). Of which 65.67% (1058/1611) women have regular menstrual cycle just after menarche, 94.97% (1530/1611) of women would have regular menstruation in 2 years after menarche. The rate of dysmenorrhea was 42.09% (678/1611), of which 13.6% (92/678) women have high severe pelvic pain. CONCLUSION: Our results suggested that the trend went toward younger ages of menarche, which was younger menarche age in women from city than country. About 95% women would have regular menstrual cycle within 2 years after menarche.

[Intracytoplasmic sperm injection for obstructive azoospermia].

OBJECTIVE: To sum up the experience in the treatment of obstructive azoospermia by intracytoplasmic sperm injection (ICSI). METHODS: We retrospectively analyzed 107 cases of obstructive azoospermia treated by ICSI in our center from Jan. 2006 to Dec. 2008, and compared the rates of fertilization, cleavage and pregnancy between the patients with congenital bilateral absence of vas deferens (CBAVD) and those with non-CBAVD. RESULTS: A total of 949 oocytes were injected for the 107 patients undergoing ICSI, of which 678 (71.4%) were fertilized and 605 (89.2%) cleaved, with 44 pregnancies (41.4%). Of the 442 oocytes injected for the 49 patients with CBAVD, 308 (69.6%) were fertilized and 279 (90.6%) cleaved, with 27 pregnancies (55.1%), and of the 507 oocytes injected for the 58 cases induced by inflammation or surgery, 370 (72.9%) were fertilized and 326 (88.1%) cleaved, with 17 pregnancies (29.3%). The rate of pregnancy was significantly higher in the CBAVD than in the non-CBAVD group (P < 0.01), but there were no significant differences in the rates of fertilization and cleavage between the two groups (P > 0.05). CONCLUSION: PESA or TESE combined with ICSI is an effective approach to the treatment of male infertility induced by obstructive azoospermia, which may achieve a higher rate of pregnancy in patients with CBAVD than in those with non-CBAVD. Inflammation or surgery may not only cause the obstruction of the deferent duct, but also affect sperm quality, and consequently reduce the potentiality of embryonic development.