Erratum: miR-424 suppresses proliferation and promotes apoptosis of human ovarian granulosa cells by targeting Apelin and APJ expression.

[This corrects the article on p. 3660 in vol. 12, PMID: 32774725.].

miR-424 suppresses proliferation and promotes apoptosis of human ovarian granulosa cells by targeting Apelin and APJ expression.

OBJECTIVE: Polycystic ovary syndrome (PCOS) is associated with alteration of Apelin signaling in ovarian granulosa cells (GCs). However, the molecular mechanisms regulating Apelin expression remain poorly understood. This study aims to investigate the role of miR-424 in modulating Apelin expression and GC functions. METHODS: miRNA expression in GCs was altered by transfection with specific miR-424 mimics and inhibitors. Apelin level was determined by ELISA. miR-424 and mRNA expression were analyzed by quantitative RT-PCR. Protein abundance was measured by western blotting. Genomic sequence targeted by miR-424 was validated by dual-luciferase reporter assay. Apelin gene was overexpressed by transfection of LV-003 vector carrying its cDNA. GC proliferation was analyzed by MTS method, and its cell cycle progression and apoptosis were measured by flow cytometry. RESULTS: Apelin concentration was increased in serum and follicular fluid from PCOS patients, accompanied by upregulated APJ (Apelin receptor) expression and suppressed miR-424 expression in GCs. miR-424 mimics suppressed Apelin and APJ expression in KGN cells by targeting 3' UTR of Apelin and APJ, whereas miR-424 inhibitors had the opposite effects. miR-424 inhibited KGN cell proliferation and cell cycle progression by down-regulating Cyclin-D/E expression. Moreover, miR-424 promoted KGN cell apoptosis by increasing truncated Caspase-3 level. The regulation of KGN cell proliferation and apoptosis by miR-424 was mediated by directly suppressing Apelin gene expression, instead of inhibiting Apelin peptide activity. CONCLUSION: miR-424 suppresses proliferation and promotes apoptosis of human ovarian granulosa cells by directly targeting and inhibiting Apelin and APJ expression.

Seminal plasma cell-free mitochondrial DNA copy number is associated with human semen quality.

OBJECTIVE: Previous studies have shown that mitochondrial DNA (mtDNA) copy number in human sperm is associated with semen quality. This study examines whether there is association between seminal cell-free mtDNA copy number and human semen parameters. STUDY DESIGN: Seminal cell-free mtDNA copy number was measured in 205 men. Semen reactive oxygen species (ROS) level was also measured. Sperm quality was assessed using World Health Organization criteria, while sperm motility parameters were determined by computer-aided sperm analysis. RESULTS: Comparing with normozoospermia, patients with asthenozoospermia and oligoasthenozoospermia had a significant decrease in cell-free mtDNA copy number and an increase in ROS level in seminal plasma (P < 0.05 for all). Seminal cell-free mtDNA copy number positively correlated with sperm concentration, motility, morphology and motion characteristics (r for 0.247 to 0.673, P < 0.05 for all). Semen ROS level negatively correlated with sperm concentration, motility, and motion characteristics (r for -0.261 to -0.676, P < 0.05 for all). There is a negative relationship between seminal cell-free mtDNA copy number and ROS level (r= -0.573, P < 0.05). CONCLUSION: The results suggested that seminal cell-free mtDNA copy number is associated with semen parameters, which may serve as a novel diagnostic marker of semen quality.

Effects of dehydroepiandrosterone supplementation on mice with diminished ovarian reserve.

Dehydroepiandrosterone (DHEA) has been used to improve the pregnancy rate in women with diminished ovarian reserve(DOR) during in vitro fertilization. We aimed to validate the effects of DHEA and identify the possible mechanisms. We constructed a mice model with DOR and analyzed the hormone parameters and follicle counts. In vivo experiment, FSH and LH concentrations in the serum were significantly elevated in the DOR group. However, the FSH and LH concentrations were partially reversed in the DOR + DHEA group. The E2, AMH and INHB were down-regulated in the DOR group and reversed in the DOR + DHEA group. Our study supported evidences that DHEA might modulate the hormone receptors in the ovary and hormone secretions to the peripheral circulation to regulate the ovary reserve functions.

Live birth sex ratio after in vitro fertilization and embryo transfer in China--an analysis of 121,247 babies from 18 centers.

In order to study the impact of procedures of IVF/ICSI technology on sex ratio in China, we conducted this multi-center retrospective study including 121,247 babies born to 93,895 women in China. There were 62,700 male babies and 58,477 female babies, making the sex ratio being 51.8% (Male: Female  = 107:100). In univariate logistic regression analysis, sex ratio was imbalance toward females of 50.3% when ICSI was preformed compared to 47.7% when IVF was used (P<0.01). The sex ratio in IVF/ICSI babies was significantly higher toward males in transfers of blastocyst (54.9%) and thawed embryo (52.4%) when compared with transfers of cleavage stage embryo (51.4%) and fresh embryo (51.5%), respectively. Multiple delivery was not associated with sex ratio. However, in multivariable logistic regression analysis after controlling for related factors, only ICSI (adjusted OR =  .90, 95%CI: 0.88-0.93; P<0.01) and blastocyst transfer (adjusted OR = 1.14, 95% CI: 1.09-1.20; P<0.01) were associated with sex ratio in IVF/ICSI babies. In conclusion, the live birth sex ratio in IVF/ICSI babies was influenced by the use of ICSI, which may decrease the percentage of male offspring, or the use of blastocyst transfer, which may increase the percentage of male offspring.

Hepatitis B virus s protein enhances sperm apoptosis and reduces sperm fertilizing capacity in vitro.

OBJECTIVE: Studying the impact of Hepatitis B virus S protein (HBs) on early apoptotic events in human spermatozoa and sperm fertilizing capacity. METHODOLOGY/PRINCIPAL FINDINGS: Spermatozoa were exposed to HBs (0, 25, 50, 100 µg/ml) for 3 h, and then fluo-4 AM calcium assay, Calcein/Co(2+) assay, protein extraction and ELISA, ADP/ATP ratio assay, sperm motility and hyperactivation and sperm-zona pellucida (ZP) binding and ZP-induced acrosome reaction (ZPIAR) tests were performed. The results showed that in the spermatozoa, with increasing concentration of HBs, (1) average cytosolic free Ca(2+) concentration ([Ca(2+)]i) rose; (2) fluorescence intensity of Cal-AM declined; (3) average levels of cytochrome c decreased in mitochondrial fraction and increased in cytosolic fraction; (4) ADP/ATP ratios rose; (5) average rates of total motility and mean hyperactivation declined; (6) average rate of ZPIAR declined. In the above groups the effects of HBs exhibited dose dependency. However, there was no significant difference in the number of sperms bound to ZP between the control and all test groups. CONCLUSION: HBs could induce early events in the apoptotic cascade in human spermatozoa, such as elevation of [Ca(2+)]i, opening of mitochondrial permeability transition pore (MPTP), release of cytochrome c (cyt c) and increase of ADP/ATP ratio, but exerted a negative impact on sperm fertilizing capacity.

[Usefulness of serum inhibin B measurement for evaluation of ovarian reserve in controlled ovarian hyperstimulation].

OBJECTIVE: To assess usefulness of serum inhibin B (INHB) measurement for evaluation of ovarian function. METHODS: Serum INHB level on day 3 of menstrual cycle was determined by enzyme labeled immunosorbent assay (ELISA) in 96 cases of in vitro fertilization and embryo transfer (IVF-ET). The patients were classified into 3 groups including (1) poor response (n = 6), normal response (n = 72), and over response (n = 18) according to their response to ovary stimulation. In addition, serum follicle-stimulating hormone (FSH), luteinizing hormone (LH) and estradiol (E(2)) levels were also determined by chemiluminescent microparticle immunoassay in these patients. Evaluation of the INHB measurement and related factors was performed by statistical analysis. RESULTS: Serum INHB levels in poor, normal and over-response groups were (28 +/- 20), (85 +/- 42), (92 +/- 34) pg/ml; FSH levels were (11.9 +/- 5.3), (7.5 +/- 2.6), (7.2 +/- 1.7) U/L; E(2) levels on day 3 of human chorionic gonadotropin (hCG) administration were (2558 +/- 2108), (9366 +/- 4472), (18 392 +/- 9655) pmol/L; numbers of retrieved oocytes per cycle were (0.6 +/- 0.4), (8.7 +/- 3.6), (14.3 +/- 2.9); top grade embryos were (0.4 +/- 0.3), (3.8 +/- 1.9), (4.6 +/- 1.7); pregnancy rates were 16.7%, 36.1%, 61.1%, respectively. INHB level was negatively correlated to FSH (r = -0.222, P < 0.05) and FSH/LH (r = -0.371, P < 0.05); while positively correlated to E(2) on the day of hCG administration (r = 0.336, P < 0.05), number of retrieved oocytes (r = 0.404, P < 0.05), number of quality embryos (r = 0.323, P < 0.05) and pregnancy rate (r = 0.246, P < 0.05), respectively. CONCLUSIONS: INHB test may reflect the ovarian reserve which is of clinic importance in the guidance of controlled ovarian hyperstimulation.