Sheng-Hsiang Li

The impact of endometrioma and laparoscopic cystectomy on serum anti-Müllerian hormone levels

BackgroundSerum anti-Müllerian hormone (AMH) had been proposed as a marker of ovarian reserve. The aim of this study was to evaluate the impact of endometrioma and laparoscopic cystectomy on ovarian reserve as measured by serum AMH levels.MethodsA total of 1,642 patients were recruited in this retrospective analysis. Control group (group 1) included 1,323 infertility patients without endometrioma. Endometrioma group (group 2) included 141 patients with ovarian endometrioma. Previous cystectomy group (group 3) included 147 patients who underwent unilateral or bilateral laparoscopic cystectomy due to ovarian endometrioma more than 6 months before enrollment. Current cystectomy group (group 4) included 31 patients who underwent cystectomy during study period. Serum anti-müllerian hormone (AMH) levels were measured upon enrollment with all patients. For patients in group 4, AMH levels were measured before and 3 months after cystectomy.ResultsMean AMH level of patients in control group was significantly higher than that of endometrioma group or previous cystectomy group in each age subgroup, while the mean serum AMH level of the endometrioma group was also significantly higher than that of the previous cystectomy group in each age subgroup. The mean AMH level was significantly lower in patients with previous bilateral cystectomy compared to that of patients with unilateral cystectomy. The mean serum AMH level was also significantly lower in patients with bilateral endometrioma compared to that of patients with unilateral endometrioma. In group 4, mean AMH level significantly decreased from 3.95 +/- 0.42 preoperation to 2.01 +/- 0.21 ng/ml at 3-month postoperation.ConclusionsBoth ovarian endometrioma and cystectomy are associated with a significant reduction on ovarian reserve. Bilateral endometrioma exerts a more profound negative impact on ovarian reserve than unilateral endometrioma, regardless of either conservative or surgical intervention.

Dual trigger with combination of gonadotropin-releasing hormone agonist and human chorionic gonadotropin significantly improves the live-birth rate for normal responders in GnRH-antagonist cycles

OBJECTIVE To investigate whether dual triggering of final oocyte maturation with a combination of gonadotropin-releasing hormone (GnRH) agonist and human chorionic gonadotropin (hCG) can improve the live-birth rate for normal responders in GnRH-antagonist in vitro fertilization/intracytoplasmic sperm injection (IVF-ICSI) cycles. DESIGN Retrospective cohort study. SETTING Infertility unit of a university-affiliated medical center. PATIENT(S) Normal responders to controlled ovarian hyperstimulation who were undergoing IVF-ICSI with a GnRH antagonist protocol. INTERVENTION(S) Standard dosage of hCG trigger (6,500 IU of recombinant hCG) versus dual trigger (0.2 mg of triptorelin and 6,500 IU of recombinant hCG). MAIN OUTCOME MEASURE(S) Live-birth, clinical pregnancy, and implantation rates per cycle. RESULT(S) A total of 376 patients with 378 completed cycles with embryo transfer were enrolled (hCG trigger/control group: n = 187; dual trigger/study group: n = 191). The dual trigger group demonstrated statistically significantly higher implantation (29.6% vs. 18.4%), clinical pregnancy (50.7% vs. 40.1%), and live-birth (41.3% vs. 30.4%) rates as compared with the hCG trigger group. There was no statistically significant difference in terms of patient demographics, cycle parameters, or embryo quality. CONCLUSION(S) Dual trigger of final oocyte maturation with a GnRH-agonist and a standard dosage of hCG in normal responders statistically significantly improves implantation, clinical pregnancy, and live-birth rates in GnRH-antagonist IVF cycles.

Abnormally low expression of connexin 37 and connexin 43 in subcutaneously transplanted cryopreserved mouse ovarian tissue

PurposeTo analyze the gap junction proteins connexin 37 (Cx37) and connexin 43 (Cx43) after subcutaneous transplantation of cryopreserved mouse ovarian tissue.MethodsExpression of gap junction genes was assessed by immunohistochemistry and real-time polymerase chain reaction (PCR) in transplanted cryopreserved ovarian tissue compared with that of normal ovarian tissue. Apoptosis of ovarian cells was evaluated by using the terminal deoxynucleotidyl transferase-mediated biotinylated deoxyuridine triphosphates nick end-labeling method.ResultsAfter subcutaneous transplantation, Cx37 and Cx43 mRNA and protein expression were significantly lower in cryopreserved than in normal ovarian tissue. Apoptosis was increased in granulosa cells from antral follicles of the cryopreserved tissue.ConclusionAfter cryopreservation and subcutaneous transplantation of ovarian tissue, proteins forming gap junctions between oocytes and granulosa cells are under-expressed compared with normal controls.

Demonstration of a Glycoprotein Derived From the Ceacam10 Gene in Mouse Seminal Vesicle Secretions

Abstract CEACAM10 was purified from mouse seminal vesicle secretions by a series of purification steps that included ion exchange chromatography on a DEAE-Sephacel column and ion exchange high-performance liquid chromatography on a sulfopropyl column. It was shown to be a 36-kDa glycoprotein with an N-linked carbohydrate moiety. The circular dichromoism spectrum of CEACAM10 in 50 mM phosphate buffer at pH 7.4 appeared as one negative band arising from the β form at 217 nm. CEACAM10 was expressed predominantly in seminal vesicles of adult mice. Both CEACAM10 and its mRNA were demonstrated on the luminal epithelium of the mucosal folds in the seminal vesicle. The amount of Ceacam10 mRNA in the seminal vesicle was correlated with the stage of animal maturation. Castration of adult mice resulted in cessation of Ceacam10 expression, while treatment of castrated mice with testosterone propionate in corn oil restored Ceacam10 expression in the seminal vesicle. During the entire course of pregnancy, Ceacam10 might be silent in the embryo. A cytochemical study illustrated the presence of the CEACAM10 binding region on the entire surface of mouse sperm. CEACAM10-sperm binding greatly enhanced sperm motility in vitro.

Involvement of the Serine Protease Inhibitor, SERPINE2, and the Urokinase Plasminogen Activator in Cumulus Expansion and Oocyte Maturation

The serpin peptidase inhibitor, clade E, member 2 (SERPINE2) inhibits urokinase-type plasminogen activator (PLAU) and tissue-type plasminogen activator. Higher SERPINE2 expression levels were detected in cumulus cells of human immature oocytes than in those of mature oocytes. The objective of this study was to evaluate whether high SERPINE2 levels in cumulus cells are associated with oocyte immaturity. Using the mouse cumulus–oocyte complex as an experimental model, the effects of elimination and overexpression of SERPINE2 in cumulus cells on cumulus expansion and oocyte maturation were assayed by in vitro maturation. Serpine2 and PLAU transcripts were the most highly expressed serpins and plasminogen activators, respectively. Their expression was coordinately regulated in cumulus cells during gonadotropin-induced oocyte maturation. Silencing of Serpine2 expression using small interfering RNAs or blockage of SERPINE2 protein using a specific antibody had no effect on oocyte maturation. However, overexpression of Serpine2 or exogenous supplementation with high levels of SERPINE2 impaired cumulus expansion and oocyte maturation, probably by decreasing hyaluronan synthase 2 (Has2) and versican (Vcan) mRNA expression. Amiloride, a specific PLAU inhibitor, also suppressed these processes. PLAU supplementation of the oocyte in vitro maturation medium caused earlier and more extensive expansion of cumulus cells and oocyte maturation that may be mediated by increased Has2 mRNA expression. However, these effects were neutralized by coincubation with SERPINE2 or amiloride and PLAU. In conclusion, SERPINE2 and PLAU are involved in cumulus expansion and oocyte maturation. High SERPINE2 levels impair these processes, probably by decreasing cumulus matrix gene expression as well as reducing cumulus hyaluronan contents and inhibiting PLAU activity. These findings may explain why cumulus cells surrounding immature human oocytes express high SERPINE2 levels.

Mechanisms underlying the inhibition of murine sperm capacitation by the seminal protein, SPINKL.

SPINKL, a serine protease inhibitor kazal‐type‐like protein initially found in mouse seminal vesicle secretions, possesses structurally conserved six‐cysteine residues of the kazal‐type serine protease inhibitor family. However, it has no inhibitory activity against serine proteases. Previously, it was found to have the ability to suppress murine sperm capacitation in vitro. Herein, we investigated the mechanisms underlying the suppressive effect of SPINKL on sperm capacitation. Three in vitro capacitation‐enhancing agents, including bovine serum albumin (BSA), methyl‐beta‐cyclodextrin (MBCD), and dibutyryl cyclic AMP (dbcAMP), coupled with 3‐isobutyl‐1‐methylxanthine (IBMX), were used to evaluate the influence of SPINKL on capacitation signaling. Preincubation of sperm with SPINKL suppressed BSA‐ and MBCD‐induced sperm capacitation by blocking three upstream signals of capacitation that is the cholesterol efflux from sperm plasma membranes, extracellular calcium ion influx into sperm, and increases in intracellular cAMP. Moreover, SPINKL also inhibited downstream signal transduction of capacitation since it suppressed dbcAMP/IBMX and N6‐phenyl cAMP (6‐Phe‐cAMP)‐activated cAMP‐dependent protein kinase‐associated protein tyrosine phosphorylation. Such inhibition is probably mediated by attenuation of SRC tyrosine kinase activity. Furthermore, SPINKL could not reverse capacitation once sperm had been capacitated by capacitation‐enhancing agents or capacitated in vivo in the oviduct. SPINKL bound to sperm existed in the uterus but had disappeared from sperm in the oviduct during the sperms transit through the female reproductive tract. Therefore, SPINKL may serve as an uncapacitation factor in the uterus to prevent sperm from precocious capacitation and the subsequent acrosome reaction and thus preserve the fertilization ability of sperm. J. Cell. Biochem. 114: 888–898, 2013.

First trimester ultrasound estimation of gestational age in pregnancies conceived after in vitro fertilization

OBJECTIVE To evaluate BPD as an alternative to CRL for the estimation of gestation age in late first trimester pregnancies (between 9th and 13th gestational weeks), and to construct a first trimester reference chart of fetal BPD growth. STUDY DESIGN A prospective study that compared the gestational age estimated by BPD and CRL with the IVF gestational age in 167 first trimester pregnancies (127 singletons, 40 twins). RESULTS Both BPD and CRL correlated well with the IVF gestational age (GA) from 9th to 13th gestation weeks. When comparing the difference of the GA (in days) estimated from the two respective ultrasound parameters versus the GA based on IVF (oocyte retrieval day +14 days), BPD estimations had a significantly lower mean difference than CRL (0.013 vs. 0.746; p<0.01), as well as a lower standard deviation (2.414 vs. 3.008; p<0.05). In addition, the 95% limits of agreement between the BPD estimated GA and IVF GA were also smaller than CRL estimated GA versus IVF GA (-4.719 to 4.745 vs. -5.149 to 6.641). CONCLUSION Biparietal diameter shares similar accuracy with crown rump length in late first trimester ultrasound estimation, with additional advantages of lower random measurement errors.

Luteinizing hormone increases platelet-derived growth factor-D gene expression in human granulosa-luteal cells.

We aimed to investigate the effects of LH and FSH on the gene expression of the platelet-derived growth factor (PDGF) gene family in human granulosa-luteal cells. We found that LH significantly increased PDGF-D messenger RNA (mRNA) expression but suppressed PDGF-B and PDGF-C mRNA in human granulosa-luteal cells, and FSH also increases PDGF-D gene expression, but to a lesser extent.

VEGF and FGF2 Improve Revascularization, Survival, and Oocyte Quality of Cryopreserved, Subcutaneously-Transplanted Mouse Ovarian Tissues.

This study was conducted to investigate the effect of the vascular endothelial growth factor (VEGF) and fibroblast growth factor 2 (FGF2) on revascularization, survival, and oocyte quality of cryopreserved, subcutaneously-transplanted mouse ovarian tissue. Autologous subcutaneous transplantation of vitrified-thawed mouse ovarian tissues treated with (experimental group) or without (control group) VEGF and FGF2 was performed. After transplantation to the inguinal region for two or three weeks, graft survival, angiogenesis, follicle development, and oocyte quality were examined after gonadotropin administration. VEGF coupled with FGF2 (VEGF/FGF2) promoted revascularization and significantly increased the survival rate of subcutaneously-transplanted cryopreserved ovarian tissues compared with untreated controls. The two growth factors did not show long-term effects on the ovarian grafts. In contrast to the untreated ovarian grafts, active folliculogenesis was revealed as the number of follicles at various stages and of mature oocytes in antral follicles after gonadotropin administration were remarkably higher in the VEGF/FGF2-treated groups. Although the fertilization rate was similar between the VEGF/FGF2 and control groups, the oocyte quality was much better in the VEGF/FGF2-treated grafts as demonstrated by the higher ratio of blastocyst development. Introducing angiogenic factors, such as VEGF and FGF2, may be a promising strategy to improve revascularization, survival, and oocyte quality of cryopreserved, subcutaneously-transplanted mouse ovarian tissue.

The efficiency of conventional microscopic selection is comparable to the hyaluronic acid binding method in selecting spermatozoa for male infertility patients

OBJECTIVE To evaluate if hyaluronic acid (HA)-bound spermatozoa surpassed conventional microscopy-selected spermatozoa in the status of sperm DNA integrity by acridine orange (AO) fluorescence staining. MATERIALS AND METHODS Spermatozoa obtained from couples with indication for the intracytoplasmic sperm injection (ICSI) procedure due to male infertility (n = 34) and control males with normal sperm parameters (n = 12) were analyzed using AO fluorescence staining after density-gradient centrifugation (DGC), polyvinylpyrrolidone (PVP)-microscopic selection, and HA-binding selection to determine sperm DNA integrity. RESULTS Percentages of DNA intact spermatozoa with green fluorescence were significantly higher in both PVP-microscopic selected spermatozoa (82.1 ± 24.0%) and HA-bound spermatozoa (83.9 ± 21.1%) than in spermatozoa prepared by DGC (66.8 ± 24.0%). However, there was no significant difference between the PVP-sperm and HA-sperm groups. When the percentage of green fluorescent spermatozoa prepared by DGC fell initially below 68%, both PVP-microscopic and HA-binding selection failed to select over 90% spermatozoa with intact DNA for ICSI in the male infertility group. Compared to control males with normal sperm parameters (99.3 ± 1.8%), the proportion of green fluorescence sperm after HA-binding selection from couples with male infertility (83.9 ± 21.1%) did not reach the range of > 99% reported by Yagci et al. CONCLUSION The percentages of DNA intact spermatozoa between the PVP-sperm and HA-sperm groups were not significantly different. In an ICSI procedure, a well-trained embryologist will have the same ability to choose sperm with intact DNA by conventional microscopic selection as with HA-bound spermatozoa selection.