Masahide Shiotani

Fertilization and pregnancy using cryopreserved testicular sperm for intracytoplasmic sperm injection with azoospermia

OBJECTIVES To report fertility for men with obstructive (OA) and nonobstructive (NOA) azoospermia after frozen and thawed spermatozoa recovered from the seminiferous tubules and intracytoplasmic sperm injection (ICSI) and to evaluate the factors other than spermatozoa. DESIGN Retrospective clinical analysis. SETTING Male infertility clinic for testicular sperm extraction and freezing/thawing (cryoTESE) and assisted reproductive technologies clinic for ICSI. PATIENT(S) Seventy-four men with OA and 140 men with NOA undergoing attempted cryoTESE-ICSI. INTERVENTION(S) Seventy-three couples with OA underwent a total of 184 cryoTESE-ICSI cycles, and 46 couples with NOA underwent a total of 75 cryoTESE-ICSI cycles. MAIN OUTCOME MEASURE(S) The numbers of eggs at metaphase II injected, two-pronuclei oocytes, normal cleaved embryos, embryos transferred, transfer cycles, biochemical pregnancies, and clinical pregnancies, as well as the implantation and delivery rates, were examined. RESULT(S) Fertilization rate in NOA was significantly lower than in OA. Neither the pathology, the source, nor the quantity of spermatozoa had any effect on fertilization or pregnancy rates. Maternal age had no effect on fertilization or embryo cleavage, but did dramatically affect the implantation, pregnancy, and delivery rates in NOA and OA. CONCLUSION(S) Good pregnancy rates were achieved without significant differences among the sperm sources. The pregnancy and the delivery rate were dependent strictly on the age of the female partner but not on her ovarian reserve.

Rapidly progressing, late-onset obstructive azoospermia linked to herniorrhaphy with mesh

OBJECTIVE To report a case of a man presenting with rapidly progressing inguinal vasal obstruction 5 years after bilateral herniorrhaphy with polypropylene mesh. DESIGN Case report with a review of the scientific literature. SETTING Male infertility clinic, Kobe University Hospital (Kobe, Japan). PATIENT(S) A 30-year-old patient who had undergone bilateral inguinal herniorrhaphy using polypropylene mesh 5 years previously. INTERVENTION(S) The patient underwent bilateral vasography and left testicular sperm extraction (TESE). MAIN OUTCOME MEASURE(S) Long-term effects of the adult inguinal herniorrhaphy with polypropylene mesh. RESULT(S) Rapid progression of vas deferens obstruction occurred within several months. We performed left TESE for intracytoplasmic sperm injection (ICSI) and retrieved many motile sperm. CONCLUSION(S) Before azoospermia, men who undergo inguinal herniorrhaphy using polypropylene mesh need to rapidly cryopreserve their sperm for future fertility; TESE-ICSI is also a selectable treatment.

Hypocoagulable State of Human Preovulatory Ovarian Follicular Fluid: Role of Sulfated Proteoglycan and Tissue Factor Pathway Inhibitor in the Fluid

Abstract Ovulation accompanied by tissue damage can cause an increase in the level of tissue factor (TF) in the follicular fluid, triggering the extrinsic coagulation pathway. However, follicular fluid must block fibrin formation and maintain fluidity until the release of the oocyte at ovulation. The combination of sulfated proteoglycan, antithrombin, and TF pathway inhibitor (TFPI) appears to play a critical role in the hypocoagulability of human follicular fluid. When compared with plasma, folicular fluid differs markedly in the levels of a number of important coagulation proteins. Principal among these are 15-fold, 13-fold, and 3.7-fold increases in free TFPI, thrombin-antithrombin complex, and TF, respectively. The excessively prolonged activated partial thromboplastin time (APTT) and prothrombin time (PT) of human ovarian follicular fluid appear to be primarily due to high concentrations of sulfated proteoglycans, which accelerate the inactivation of thrombin and the anti-Xa activity of TFPI. Thus, heparitinase treatment shortened the clotting times of follicular fluid and reduced the inhibition of thrombin by the proteoglycan fraction combined with a fraction containing antithrombin. The remaining prolongation of APTT and PT may be caused by high levels of free TFPI in follicular fluid, which were confirmed by Northern blotting analysis, demonstrating TFPI mRNA expression by granulosa cells.

Molecular changes induced by bisphenol-A in rat Sertoli cell culture.

Bisphenol-A (BPA) is an industrial chemical and is known to act as an endocrine disrupter. This study was designed to evaluate how BPA regulates Sertoli cell (SC) signal molecules. Purified rat SCs were cultured and treated with BPA (200 µmol/l) at various time points. Western blot analysis was used to determine the activation of extracellular signal-related kinases 1 and 2 (ERK1/2), c-Jun N-terminal kinase (JNK), p38 mitogen activated protein kinase (MAPK), nuclear factor kappa B (NF-κB), cyclooxygenase-1,2 (COX-1, 2), estrogen receptor-α (ER-α), and androgen receptor (AR). The levels of transferrin (TF), prostaglandin E2 (PGE2), and prostaglandin F2α (PGF2α) in culture medium were quantified by ELISA. Interleukin (IL)-1β and IL-6 mRNAs were measured by quantitative real-time PCR (QRT-PCR). Compared with the control, BPA activated the phosphorylation of ERK1/2 (p-ERK1/2) through 30 min to 6 h. TF was down-regulated at 6 and 24 h. Furthermore, IL-1β was up-regulated at 30 min and IL-6 was up-regulated at 1 and 24 h. ERK activity inhibitor (PD98059, 10 µmol/l) inhibited these molecular changes. These results reveal the possibility that BPA may have adverse effects on spermatogenesis via ERK1/2.

Stimulation of endometrium embryo transfer can improve implantation and pregnancy rates for patients undergoing assisted reproductive technology for the first time with a high-grade blastocyst

OBJECTIVE To evaluate whether stimulation of endometrium embryo transfer (SEET) can improve implantation rate and pregnancy rate (PR) for patients undergoing assisted reproductive technology (ART) for the first time by injecting embryo culture supernatant into the uterus before blastocyst transfer (BT). DESIGN Randomized, controlled trial. SETTING Private in vitro fertilization clinic. PATIENT(S) Forty-eight women in the BT group, 48 women in the stimulation group who had culture medium injected into the uterus before BT, and 48 women in the SEET group. INTERVENTION(S) Injection of embryo culture supernatant and injection of culture medium. MAIN OUTCOME MEASURE(S) Implantation rates and PRs. RESULT(S) Odds ratios of successful implantation rate for stimulation and SEET in patients with high-grade blastocysts, having BT as reference, were 2.58 and 6.46 without adjustment, and 5.91 and 9.20 after adjusting for basal FSH levels and period of infertility. Odds ratios of clinical pregnancies were 2.47 and 4.32 without adjustment, and 4.46 and 5.10 with adjustment, respectively. In groups with low-grade blastocysts, such tendencies were not observed. CONCLUSION(S) The SEET may be an effective method for increasing implantation rate and PR for first-time ART patients who have a high-grade blastocyst.

A healthy birth after intracytoplasmic sperm injection using ejaculated spermatozoa from a patient with Kartagener's syndrome

OBJECTIVE To report a healthy birth that was achieved by intracytoplasmic sperm injection (ICSI) with use of ejaculated spermatozoa from a patient with Kartageners syndrome. DESIGN Case report. SETTING Private infertility clinic. PATIENT(S) Couple with male factor infertility due to Kartageners syndrome. INTERVENTION(S) Intracytoplasmic sperm injection with ejaculated sperm. MAIN OUTCOME MEASURE(S) Semen characteristics, sperm motility, fertilization, pregnancy, and birth after ICSI. RESULT(S) With ejaculated sperm, the fertilization rates were 73% in the first stimulation cycle and 100% in the second cycle. Intracytoplasmic sperm injection was successful. The pregnancy resulted in birth of a single healthy child. CONCLUSION(S) With ejaculated sperm, successful pregnancy after ICSI in couples with Kartageners syndrome is possible. Kartageners syndrome is a heterogeneous group of disorders with similar clinical presentations, and treatment should be individualized depending on sperm motility.

The Fate of Embryos Transferred into the Uterus

PurposeThe purpose of this study is to elucidate the fate of embryos transferred into the uterus.MethodsImplantation rates were compared between synchronous and asynchronous intrauterine embryo transfers in mice. Then transferred embryos were recovered from the uterus and examined morphologically 24 hr after transfer. Moreover, the same transfer-recovery experiments were performed after ligation of the uterus two-thirds from the uterotubal junction immediately before transfer.ResultsThe implantation rate was high (58.4% per embryo, 87.5% per recipient) when blastocysts were transferred into the uteri of recipients of pseudopregnant Day 4, but it was very low (0–2.4% per embryo, 0–8.3% per recipient) when blastocysts were transferred to recipients of pseudopregnant Day 2, or two-cell embryos into recipients of pseudopregnant Days 2 and 4. When transferred embryos were recovered from the uterus 24 hr after transfer, few embryos (1.4%) were recovered in asynchronous transfers, while 66.3% of the embryos were recovered in synchronous transfers. However, from recipients in which the lower portions of uteri were ligated, embryos at more advanced stages were recovered even in asynchronous transfers.ConclusionsThese results suggest that discharge from the uterus is one of the major causes of the low implantation rate for asynchronous transfer.

Successful pregnancy outcome in a case of heterotopic intrauterine and cervical pregnancy and a literature review

It is difficult to treat cervical pregnancy without harming the intrauterine pregnancy. We report the case of a patient who was 37 years old and had an abdominal myomectomy. She had become pregnant by in vitro fertilization and egg transfer, and was transferred because of massive bleeding at 6 weeks of gestation. Afterwards, Escherichia coli infection was detected and intravenous antibiotics were administered. Careful intracervical mass reduction and curettage were performed under abdominal ultrasound guidance. The postoperative course was uneventful and she gave birth by cesarean section at 38 weeks. Sixteen cases exist in the literature. Treatment of cervical pregnancy was performed by KCl injection in six cases, hyperosmotic glucose injection in one and complete cervical evacuation in nine. The nine patients that had cervical evacuation delivered at term. In seven cervical mass preservation cases, one patient with twin pregnancy delivered at 34 weeks, and six with singletons delivered at 31–36 weeks. The best therapy is complete evacuation in early gestation in order to avoid infection, bleeding and premature birth. The authors emphasize abdominal ultrasound usage through the bladder window with 150–200 mL of water (or urine) for careful cervical mass reduction and curettage without harming the intrauterine pregnancy.

Grade and looseness of the inner cell mass may lead to the development of monochorionic diamniotic twins.

OBJECTIVE To examine the relationship between the inner cell mass (ICM) grade and its morphological configuration on the occurrence of monochorionic diamniotic (M-D) twinning. DESIGN Retrospective embryo cohort study. SETTING Private IVF clinic. PATIENT(S) Evaluation of frozen-thawed single blastocyst transfers with hormone replacement treatment in 8,435. This cohort included 71 blastocysts and their ICMs observed by time-lapse photography. INTERVENTION(S) Any changes in configuration of the ICMs observed by time-lapse photography were analyzed retrospectively. MAIN OUTCOME MEASURE(S) The amount of loosening of blastomeres within the ICM was evaluated by time-lapse observations. The number of cells that were involved in the loosening process was also assessed. Both of these parameters were correlated with the type of monozygotic twinning that eventuated. RESULT(S) The M-D twinning incidence resulting from blastocysts with a high grade ICM (grade A) were transferred was 0.38% (3/796), whereas it was significantly higher, 1.38% (34/2,463), when blastocysts with a poorer (B and C) grade ICM were transferred. Among 71 transferred frozen-thawed blastocysts that were studied with time-lapse photography, there were two dichorionic diamniotic and one M-D twins. Careful observations of the embryo that resulted in the one M-D case, revealed that the ICM acquired a looser appearance due to decompaction of at least eight cells. This type of decompaction was not observed in the ICMs of other transferred blastocysts. CONCLUSION(S) The occurrence of M-D twinning may be avoided by excluding blastocysts that contain decompacting ICMs.

Isoflavones regulate secretion of leukemia inhibitory factor and transforming growth factor β and expression of glycodelin in human endometrial epithelial cells

Isoflavones have attracted much attention due to their association with health benefits; however, comprehensive understanding of the beneficial impacts of isoflavones on uterine biology at the molecular level remains unexplored. In the present study, our data showed that isoflavones aglycones AglyMax, genistein, and equol, but not daidzein, within the range of plasma concentration, displayed bioavailability in regulating the secretion of leukemia inhibitory factor (LIF) and transforming growth factor beta (TGF-beta) in Ishikawa cells, which was blocked by an estrogen receptor antagonist ICI 182 780, mitogen-activated protein kinase kinase (MEK)1/2 inhibitor PD98059, and p38 mitogen-activated protein kinase inhibitor SB203580. We also found that AglyMax and genistein increased in cyclic AMP release and the expression of glycodelin protein in Ishikawa cells assayed using western blot and immunochemical staining. The MEK1/2 inhibitor PD98059 and the protein kinase A inhibitor H89, but not SB203580, attenuated this glycoprotein expression. Moreover, isoflavone aglycones AglyMax stimulated LIF, and TGF-beta secretion, and glycodelin expression in separate primary endometrial epithelial cells in the follicular phase or luteal phase from healthy subject donors. Overall, our findings suggest that isoflavones may alter the uterine expression of estrogen-responsive genes.