RESUMO
Few studies have examined the correlation between sperm miRNA levels and clinical outcomes of intracytoplasmic sperm injection (ICSI). In this study, we aimed to assess the correlation of sperm miR-34b, miR-34c, miR-122, and miR-429 levels with ICSI outcomes in men with teratozoospermia and asthenozoospermia. TaqMan microRNA quantitative polymerase chain reaction was used to evaluate the relative expression of miRNAs in sperm. The relative miRNA levels quantified using a comparative method found that the four miRNAs were not associated with fertilization rate and early embryo development. However, revels of miR-34b and miR-34c in teratozoospermia sperm of the live birth group were significantly higher than those in the non-live birth group. Receiver operating characteristic curve analysis revealed that the optimal cut-off delta cycle threshold values of miR-34b and miR-34c were 8.630 and 7.883, respectively. Statistical analysis found that the levels of miR-34b and the miR-34c in teratozoospermic and asthenozoospermic sperm above the thresholds were not associated with the fertilization rate and the high-quality embryo rate above 50%; however, they were more likely to exhibit higher implantation, pregnancy, and live birth rates. miR-34b and miR-34c were significantly associated with ICSI clinical outcomes in male factor infertility, especially teratozoospermia. Further validation is required before it becomes a clinically valid reference indicator.
Assuntos
Astenozoospermia , Infertilidade Masculina , MicroRNAs , Teratozoospermia , Gravidez , Feminino , Masculino , Humanos , Injeções de Esperma Intracitoplásmicas/métodos , Teratozoospermia/metabolismo , Sêmen/metabolismo , Infertilidade Masculina/genética , Infertilidade Masculina/terapia , Infertilidade Masculina/metabolismo , Espermatozoides/metabolismo , Astenozoospermia/genética , Astenozoospermia/terapia , Astenozoospermia/metabolismo , MicroRNAs/genética , MicroRNAs/metabolismo , Ácidos Polimetacrílicos , Estudos Retrospectivos , Taxa de GravidezRESUMO
Lysozyme (LYZ) c-like proteins are primarily present in the testis and epididymis of male reproductive tissues. Here, we report a novel member of the c-type LYZ family, the seminal vesicle-secreted LYZ c-like protein (SVLLP). Three forms of SVLLP were purified from mouse seminal vesicle secretions and characterized as glycoproteins with the same protein core but different N-linked glycans. SVLLP is structurally similar to c-type LYZ proteins. Only one of the 20 invariant residues was altered in the consensus sequence of c-type LYZs; however, the changed residue (N53S) is one of two essential catalytic residues. LYZ activity assays demonstrated that the three glycoforms of SVLLP lacked enzyme activity. SVLLP is primarily expressed in seminal vesicles. Immunohistochemistry revealed that it occurs in the luminal fluid and mucosal epithelium of the seminal vesicles. Testosterone is not the primary regulator for its expression in the seminal vesicle. SVLLP binds to sperm and suppresses bovine serum albumin-induced sperm capacitation, inhibits the acrosome reaction, and blocks sperm-oocyte interactions in vitro, suggesting that SVLLP is a sperm capacitation inhibitor.
Assuntos
Glândulas Seminais/metabolismo , Capacitação Espermática/fisiologia , Espermatozoides/metabolismo , Reação Acrossômica/efeitos dos fármacos , Animais , Transporte Biológico/efeitos dos fármacos , Western Blotting , AMP Cíclico/metabolismo , Imuno-Histoquímica , Masculino , Camundongos , Muramidase/efeitos dos fármacos , Muramidase/metabolismo , Glândulas Seminais/efeitos dos fármacos , Capacitação Espermática/efeitos dos fármacos , Espermatozoides/efeitos dos fármacos , Testosterona/farmacologiaRESUMO
BACKGROUND: Diminished ovarian reserve (DOR) remains one of the greatest obstacles affecting the chance of a successful live birth after fertility treatment. The present study was set to investigate whether using a "dual trigger" consisted of human chorionic gonadotropin (hCG) plus gonadotropin releasing hormone agonist (GnRH-a) for final oocyte maturation could improve the IVF cycle outcomes for patients with diminished ovarian reserve. METHODS: A total of 427 completed GnRH-antagonist downregulated IVF cycles with fresh embryo transfer (ET) were included in this retrospective analysis. DOR was defined as antral follicle count ≤5 and serum anti-Müllerian hormone level ≤ 1.1 ng/mL. The control group (n = 130) used a 6500 IU of recombinant hCG for trigger, and the study group (n = 297) used 0.2 mg of triptorelin plus 6500 IU of recombinant hCG for trigger. RESULTS: The dual-trigger group had significantly higher oocyte fertilization rate (73.1% vs. 58.6%), clinical pregnancy rate (33.0% vs. 20.7%) and live birth rate (26.9% vs. 14.5%) when compared to the hCG trigger group. In addition, the abortion rate (17.4% vs. 37.0%) and embryo transfer cancellation rate (6.1% vs. 15.4%) were both significantly lower in the dual trigger group. The primary outcome measure was the live birth rate per oocyte retrieval cycle. Secondary outcome measures were embryo transfer cancellation rate, clinical pregnancy rate, implantation rate, chemical pregnancy rate and abortion rate per oocyte retrieval cycle. CONCLUSIONS: Dual triggering the final oocyte maturation with GnRH-a and standard dose of hCG can significantly improve the live birth rate, clinical pregnancy rate, and fertilization rate in women with diminished ovarian reserve undergoing GnRH antagonist down-regulated IVF-ICSI cycles.
Assuntos
Gonadotropina Coriônica/uso terapêutico , Hormônio Liberador de Gonadotropina/uso terapêutico , Reserva Ovariana , Indução da Ovulação/métodos , Adulto , Coeficiente de Natalidade , Implantação do Embrião , Feminino , Humanos , Recuperação de Oócitos , Gravidez , Taxa de Gravidez , Estudos Retrospectivos , Injeções de Esperma Intracitoplásmicas/métodosRESUMO
BACKGROUND: Cystatin C (CST3), a cysteine protease inhibitor in seminal plasma, is expressed in animal uteri. However, its expression in the human female reproductive tract and its effect on human sperm capacitation are unclear. METHODS: The cellular localization of CST3 was observed using immunohistochemistry. The binding of CST3 to sperm was examined using immunocytochemistry. Sperm motility parameters were analyzed using computer-assisted sperm analysis. Sperm capacitation was evaluated by analyzing cholesterol content, protein tyrosine phosphorylation levels, and the acrosome reaction. RESULTS: Immunohistochemical staining demonstrated that CST3 is prominently expressed in the female reproductive tract, including the epithelial lining and cervix and endometrium fluids, particularly at times near ovulation. It can bind to human sperm on the post-acrosomal head region and the mid and principal piece of the tail. CST3 enhances sperm motility and inhibits the signal initiating sperm capacitation, i.e., efflux of cholesterol from the sperm plasma membrane and a late sperm capacitation event, i.e., the increase in the sperm protein tyrosine phosphorylation. The suppressive trend on sperm acrosome reaction further supports CST3's ability to inhibit sperm capacitation. CONCLUSIONS: These findings suggest that cervical CST3 may prevent precocious capacitation and acrosome reaction, thus preserving sperm fertilizing ability before it reaches the fallopian tube. Additionally, CST3 may help sperm enter the upper reproductive tract by enhancing sperm motility.
Assuntos
Cistatina C/fisiologia , Capacitação Espermática/fisiologia , Reação Acrossômica , Colo do Útero/metabolismo , Cistatina C/metabolismo , Endométrio/metabolismo , Feminino , Humanos , Imuno-Histoquímica , Masculino , Fosforilação , Interações Espermatozoide-Óvulo , Útero/metabolismoRESUMO
SERPINE2 (serpin peptidase inhibitor, clade E, member 2), predominantly expressed in the seminal vesicle, can inhibit murine sperm capacitation, suggesting its role as a sperm decapacitation factor (DF). A characteristic of DF is its ability to reverse the capacitation process. Here, we investigated whether SERPINE2 can reversibly modulate sperm capacitation. Immunocytochemical staining revealed that SERPINE2 was bound onto both capacitated and uncapacitated sperm. It reversed the increase in BSA-induced sperm protein tyrosine phosphorylation levels. The effective dose and incubation time were found to be >0.1 mg/mL and >60 min, respectively. Calcium ion levels in the capacitated sperm were reduced to a level similar to that in uncapacitated sperm after 90 min of incubation with SERPINE2. In addition, the acrosome reaction of capacitated sperm was inhibited after 90 min of incubation with SERPINE2. Oviductal sperm was readily induced to undergo the acrosome reaction using the A23187 ionophore; however, the acrosome reaction was significantly reduced after incubation with SERPINE2 for 60 and 120 min. These findings suggested that SERPINE2 prevented as well as reversed sperm capacitation in vitro. It also prevented the acrosome reaction in in vivo-capacitated sperm isolated from the oviduct. Thus, SERPINE2 could reversibly modulate murine sperm capacitation.
Assuntos
Reação Acrossômica , Acrossomo/efeitos dos fármacos , Serpina E2/farmacologia , Acrossomo/metabolismo , Animais , Cálcio/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos ICR , Serpina E2/metabolismoRESUMO
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.
Assuntos
Blastocisto/citologia , Sobrevivência Celular/efeitos dos fármacos , Fator 2 de Crescimento de Fibroblastos/farmacologia , Neovascularização Fisiológica/efeitos dos fármacos , Oócitos/citologia , Ovário/citologia , Fator A de Crescimento do Endotélio Vascular/farmacologia , Animais , Blastocisto/efeitos dos fármacos , Criopreservação , Desenvolvimento Embrionário/efeitos dos fármacos , Ensaio de Imunoadsorção Enzimática , Feminino , Fertilização in vitro , Técnicas Imunoenzimáticas , Camundongos , Oócitos/efeitos dos fármacos , Ovário/efeitos dos fármacos , Tela Subcutânea , Transplante AutólogoRESUMO
BACKGROUND: GJA1 and PTX3 were proposed as gene markers for oocyte and embryo developmental competence, while SERPINE2 was reported to be associated with pregnancy outcome. PRSS35, which is exclusively expressed in the ovary, may be correlated with oocyte competence. This study was conducted to evaluate the correlation of cumulus GJA1, PRSS35, PTX3, and SERPINE2 gene expression levels with oocyte maturation, fertilization, and early embryo development. METHODS: In total, 308 cumulus cell samples separated from individual cumulus-oocyte complex were obtained from 40 patients undergoing the intracytoplasmic sperm injection treatment procedure. Gene expression levels (mRNA levels) in cumulus cells were assessed using quantitative real-time polymerase chain reaction. RESULTS: Gene expression levels of GJA1 and SERPINE2 in cumulus cells surrounding mature oocytes were significantly lower than those in cumulus cells enclosing immature oocytes. PRSS35 mRNA levels in cumulus cells of fertilized oocytes were significantly higher than those in cumulus cells of unfertilized oocytes. GJA1 and SERPINE2 seemed to express higher mRNA levels, while PRSS35 showed lower expression in cumulus cells of oocytes that developed into embryos with good morphology; however, the expression levels of all three genes and PTX3 showed no significant differences between embryos with good or poor morphology. CONCLUSIONS: GJA1 and SERPINE2 represent potential gene markers associated with oocyte maturation. PRSS35 may be correlated with oocyte fertilization potential. However, GJA1, PRSS35, PTX3, and SERPINE2 may not be considered as marker genes for predicting embryo morphology.
Assuntos
Proteína C-Reativa/biossíntese , Conexina 43/biossíntese , Células do Cúmulo/metabolismo , Fertilização/fisiologia , Oogênese/fisiologia , Serina Proteases/biossíntese , Serpina E2/biossíntese , Componente Amiloide P Sérico/biossíntese , Biomarcadores/metabolismo , Proteína C-Reativa/genética , Conexina 43/genética , Células do Cúmulo/citologia , Desenvolvimento Embrionário/fisiologia , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Humanos , Gravidez , Serina Proteases/genética , Serpina E2/genética , Componente Amiloide P Sérico/genética , Injeções de Esperma Intracitoplásmicas/métodosRESUMO
PURPOSE: The aim of this study was to evaluate the correlation between embryonic early-cleavage status and the age of patients receiving either a GnRH agonist long protocol or a GnRH antagonist protocol. METHODS: This retrospective study included 534 patients undergoing a fresh cycle of oocyte retrieval and day-3 embryo transfer. Of the 534 patients treated, 331 received a GnRH agonist long stimulation protocol (GnRH agonist group) for ovarian stimulation and 203 patients received a GnRH antagonist protocol (GnRH antagonist group). RESULTS: By logistic regression analysis, the rate of embryonic early-cleavage was significantly decreased with increasing age of women in the agonist (P < 0.001) but not in antagonist groups (P = 0.61). Based on the results of this study, maternal age is a critical factor for embryonic early-cleavage in agonist protocol but not in antagonist protocol. The results also showed that early-cleavage embryos were of better quality and resulted in a higher pregnancy rate than late-cleavage embryos in the GnRH agonist group. However, embryo quality and pregnancy rate was not significantly different between early and late cleavage embryos in the GnRH antagonist group. CONCLUSIONS: We conclude that embryonic early-cleavage status is negatively correlated with aging in women receiving GnRH agonist long down-regulation but not in GnRH antagonist protocols. We also conclude that early cleavage of the zygote is not a reliable predictor for pregnancy potential using the GnRH antagonist protocol.
Assuntos
Blastômeros/citologia , Fase de Clivagem do Zigoto/citologia , Embrião de Mamíferos/citologia , Hormônio Liberador de Gonadotropina/agonistas , Hormônio Liberador de Gonadotropina/antagonistas & inibidores , Oócitos/citologia , Adulto , Blastômeros/efeitos dos fármacos , Fase de Clivagem do Zigoto/efeitos dos fármacos , Implantação do Embrião , Transferência Embrionária , Embrião de Mamíferos/efeitos dos fármacos , Feminino , Fármacos para a Fertilidade Feminina/uso terapêutico , Fertilização in vitro/métodos , Humanos , Infertilidade Feminina/tratamento farmacológico , Oócitos/efeitos dos fármacos , Indução da Ovulação , Gravidez , Taxa de Gravidez , Estudos Retrospectivos , Resultado do TratamentoRESUMO
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 N(6) -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 sperm's 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.
Assuntos
Proteínas Secretadas Inibidoras de Proteinases/metabolismo , Capacitação Espermática/efeitos dos fármacos , Espermatozoides/metabolismo , 1-Metil-3-Isobutilxantina/farmacologia , Reação Acrossômica/efeitos dos fármacos , Animais , Bucladesina/farmacologia , Cálcio/metabolismo , Colesterol/metabolismo , AMP Cíclico/metabolismo , Feminino , Masculino , Camundongos , Camundongos Endogâmicos ICR , Oviductos/metabolismo , Fosforilação , Proteínas Secretadas Inibidoras de Proteinases/farmacologia , Soroalbumina Bovina/farmacologia , Espermatozoides/efeitos dos fármacos , beta-Ciclodextrinas/farmacologiaRESUMO
BACKGROUND: Vitrified M-II oocyte accumulation for later simultaneous insemination has been used for managing POR. Our study aimed to determine whether vitrified oocyte accumulation strategy improves live birth rate (LBR) for managing diminished ovarian reserve (DOR). METHODS: A retrospective study included 440 women with DOR fulfilling Poseidon classification groups 3 and 4, defined as the presence of serum anti-Müllerian hormone (AMH) hormone level < 1.2 ng/ml or antral follicle count (AFC) < 5, from January 1, 2014, to December 31, 2019, in a single department. Patients underwent accumulation of vitrified oocytes (DOR-Accu) and embryo transfer (ET) or controlled ovarian stimulation (COS) using fresh oocytes (DOR-fresh) and ET. Primary outcomes were LBR per ET and cumulative LBR (CLBR) per intention to treat (ITT). Secondary outcomes were clinical pregnancy rate (CPR) and miscarriage rate (MR). RESULTS: Two hundred eleven patients underwent simultaneous insemination of vitrified oocyte accumulation and ET in the DOR-Accu group (maternal age: 39.29 ± 4.23 y, AMH: 0.54 ± 0.35 ng/ml), and 229 patients underwent COS and ET in the DOR-fresh group (maternal age: 38.07 ± 3.77 y, AMH: 0.72 ± 0.32 ng/ml). CPR in the DOR-Accu group was similar in the DOR-fresh group (27.5% vs. 31.0%, p = 0.418). However, MR was statistically higher (41.4% vs. 14.1%, p = 0.001), while LBR per ET was statistically lower (15.2% vs. 26.2%, p < 0.001) in the DOR-Accu group. There is no difference in CLBR per ITT between groups (20.4% vs. 27.5%, p = 0.081). The secondary analysis categorized clinical outcomes into four groups regarding patients' age. CPR, LBR per ET, and CLBR did not improve in the DOR-Accu group. In the group of 31 patients, accumulated vitrified metaphase II (M-II) oocytes reached a total number of ≥ 15, and CPR improved among the DOR-Accu group (48.4% vs. 31.0%, p = 0.054); however, higher MR (40.0% vs. 14.1%, p = 0.03) resulted in similar LBR per ET (29.0% vs. 26.2%, p = 0.738). CONCLUSIONS: Vitrified oocyte accumulation for managing DOR did not improve LBR. Higher MR resulted in lower LBR in the DOR-Accu group. Therefore, the vitrified oocyte accumulation strategy for managing DOR is not clinically practical. TRIAL REGISTRATION: The study protocol was retrospectively registered and was approved by Institutional Review Board of Mackay Memorial Hospital (21MMHIS219e) on August 26, 2021.
Assuntos
Aborto Espontâneo , Doenças Ovarianas , Reserva Ovariana , Feminino , Gravidez , Humanos , Estudos Retrospectivos , Coeficiente de Natalidade , Oócitos , Hormônio AntimüllerianoRESUMO
We examined whether there is a correlation among early embryo cleavage, speed of cleavage, and implantation potential for in-vitro fertilization (IVF) treatment and intracytoplasmic sperm injection (ICSI). This retrospective study examined 112 cycles of IVF and 82 cycles of ICSI in patients less than 40 years of age. Early cleavage was defined as embryonic mitosis occurring 25-27 h after insemination. These day-3 embryos were then grouped according to cleavage speed (rapid, normal, and slow) and morphological quality (good or poor). A larger proportion of early-cleavage embryos developed normally compared to non-early-cleavage embryos (IVF: 69.1 % vs. 47.1 %, respectively; ICSI: 63.0 % vs. 45.6 %, respectively). The early-cleavage embryos also produced more good quality embryos than the non-early-cleavage embryos (IVF: 80.2 % vs. 56.4 %, respectively; ICSI: 73.4 % vs. 59.4 %). The implantation rate was significantly higher with early-cleavage embryos in both IVF (42.9 % vs. 19.7 %) and ICSI (48.1 % vs. 24 %). These results indicate that early-cleavage embryos have a higher rate of normal development and develop into better quality embryos on day 3, resulting in more and higher quality embryos to choose from for day-3 embryo transfer. Thus, early cleavage may be a useful criterion when selecting embryos for IVF or ICSI.
Assuntos
Fase de Clivagem do Zigoto/fisiologia , Implantação do Embrião , Fertilização in vitro/métodos , Injeções de Esperma Intracitoplásmicas/métodos , Adulto , Transferência Embrionária/métodos , Embrião de Mamíferos/fisiologia , Desenvolvimento Embrionário , Feminino , Humanos , Masculino , Mitose , Gravidez , Estudos Retrospectivos , Fatores de Tempo , Resultado do TratamentoRESUMO
OBJECTIVE: To compare the clinical outcomes between fresh and vitrified-thawed day 5 blastocyst transfers. DESIGN: Retrospective case control study. SETTING: Tertiary referral center. PATIENT(S): Patients 38 years of age or less who underwent IVF/ICSI cycles with fresh or frozen-thawed blastocysts transferred from June 1, 2009 to November 30, 2011 INTERVENTION(S): Vitrification and thawing of day 5 blastocysts using the Cryotop method. (Kitazato BioPharma Co., Ltd., Fuji city, Shizuoka, Japan) MAIN OUTCOME MEASURE(S): Clinical pregnancy rate, implantation rate, ongoing pregnancy rate, and multiple pregnancy rates. RESULTS: Of the 118 cycles in the fresh transfer group, 234 blastocysts were transferred. The clinical pregnancy rate was 66.1 % and implantation rate was 50.9 %. The ongoing pregnancy rate was 56.8 % and the rates for singleton and twin pregnancies were 53.7 % and 44.8 %. Of the 59 cycles in the vitrified-thawed group, 111 blastocysts were transferred. The clinical pregnancy rate was 59.3 % and implantation rate was 43.2 %. The ongoing pregnancy rate was 47.5 % and the rates for singleton and twin pregnancies were 60.7 % and 39.3 %. The clinical pregnancy rate, implantation rate and ongoing pregnancy rate did not differ significantly between the two groups. CONCLUSIONS: The implantation rates were not significantly different between the fresh and the vitrified-thawed groups. Thus, single embryo transfer may be considered in fresh cycles to decrease multiple pregnancy rates. The surplus embryos should be vitrified for the frozen embryo transfer to improve the cumulative pregnancy rate.
Assuntos
Blastocisto , Transferência Embrionária/métodos , Vitrificação , Adulto , Blastocisto/citologia , Blastocisto/fisiologia , Estudos de Casos e Controles , Criopreservação/métodos , Implantação do Embrião , Feminino , Fertilização in vitro , Humanos , Gravidez , Taxa de Gravidez , Estudos Retrospectivos , Transferência de Embrião Único , Injeções de Esperma IntracitoplásmicasRESUMO
SERPINE2, one of the potent serine protease inhibitors that modulates the activity of plasminogen activator and thrombin, is implicated in many biological processes. In the present study, we purified SERPINE2 from mouse seminal vesicle secretion (SVS), using liquid chromatography and identified it by liquid chromatography/tandem mass spectrometry, and it showed potent inhibitory activity against the urokinase-type plasminogen activator. SERPINE2 was expressed predominantly in seminal vesicles among murine male reproductive tissues. It was immunolocalized to the SVS and mucosal epithelium of the seminal vesicle, epididymis, coagulating gland, and vas deferens. In the testes, SERPINE2 was immunostained in spermatogonia, spermatocytes, spermatids, Leydig cells, and spermatozoa. SERPINE2 was also detected on the acrosomal cap of testicular and epididymal sperm and was suggested to be an intrinsic sperm surface protein. The purified SERPINE2 protein could bind to epididymal sperm. A prominent amount of SERPINE2 was detected on ejaculated and oviductal spermatozoa. Nevertheless, SERPINE2 was detected predominantly on uncapacitated sperm, indicating that SERPINE2 is lost before initiation of the capacitation process. Moreover, SERPINE2 could inhibit in vitro bovine serum albumin-induced sperm capacitation and prevent sperm binding to the egg, thus blocking fertilization. It acts through preventing cholesterol efflux, one of the initiation events of capacitation, from the sperm. These findings suggest that the SERPINE2 protein may play a role as a sperm decapacitation factor.
Assuntos
Genitália Masculina/metabolismo , Serpina E2/metabolismo , Serpina E2/fisiologia , Capacitação Espermática , Sequência de Aminoácidos , Animais , Desdiferenciação Celular/efeitos dos fármacos , Desdiferenciação Celular/fisiologia , Ejaculação , Genitália Masculina/citologia , Masculino , Camundongos , Camundongos Endogâmicos ICR , Dados de Sequência Molecular , Ligação Proteica , Reprodução/fisiologia , Sêmen/citologia , Sêmen/metabolismo , Inibidores de Serina Proteinase/metabolismo , Inibidores de Serina Proteinase/farmacologia , Inibidores de Serina Proteinase/fisiologia , Serpina E2/isolamento & purificação , Serpina E2/farmacologia , Capacitação Espermática/efeitos dos fármacos , Capacitação Espermática/fisiologia , Distribuição TecidualRESUMO
BACKGROUND: SERPINE2, also known as protease nexin-1, belongs to the serine protease inhibitor (SERPIN) superfamily. It is one of the potent SERPINs that modulates the activity of plasminogen activators (PAs). PAs and their SERPIN inhibitors, such as SERPINB2 and SERPINE1, were expressed in the human endometrium and were implicated in implantation. However, expression data about SERPINE2 in the human endometrium is still unknown. Thus, we conducted an investigation to reveal the spatiotemporal and cellular expression of SERPINE2 in the human uterus during the menstrual cycle. METHODS: Seven patients who underwent a hysterectomy and samples of 120 archived patients' endometrial curettage or parts of the uterus that were formalin-fixed and embedded in paraffin. Western blotting was performed to evaluate the specificity and sensitivity of the antibody. Immunohistochemistry was conducted to localize the SERPINE2 expression site. Quantitative analysis was conducted to evaluate expression levels of SERPINE2 in various sub-phases of the menstrual cycle. RESULTS: The SERPINE2 protein was primarily detected in the uterine fluid during the mid- and late-secretory phases of the menstrual cycle. It was predominantly expressed in the luminal and glandular epithelium, less in the myometrium, and only dispersedly in certain stromal cells throughout the menstrual cycle. A quantitative analysis of expression levels of SERPINE2 in the glandular epithelium revealed that it was highly expressed in the endometrium during the secretory phase compared to the proliferative phase. CONCLUSIONS: The SERPINE2 protein is highly expressed in the endometrium during the secretory phase, indicating that it may participate in tissue remodeling involved in implantation.
Assuntos
Fase Luteal/metabolismo , Serpina E2/biossíntese , Líquidos Corporais/química , Endométrio/metabolismo , Feminino , Humanos , Útero/químicaRESUMO
BACKGROUND: Serum 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. METHODS: A 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. RESULTS: Mean 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. CONCLUSIONS: Both 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.
Assuntos
Hormônio Antimülleriano/sangue , Endometriose/sangue , Endometriose/cirurgia , Cistos Ovarianos/cirurgia , Adulto , Feminino , Humanos , Laparoscopia , Pessoa de Meia-Idade , Ovário/fisiologia , Ovário/cirurgia , Estudos RetrospectivosRESUMO
Reactive oxygen species may cause oxidative damage in the placenta, yet some mechanisms must exist to reduce or prevent such damage. We investigated whether oxidative injury to placental endothelial cells is inhibited by activation of antioxidant enzymes by paracrine factors secreted by human placental multipotent mesenchymal stromal cells (hPMSC). hPMSC-conditioned medium and umbilical endothelial cells were assayed for cytokines and cytokine receptor expression by immunoassay and real-time PCR. Endothelial cell survival was evaluated by MTS [3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt] assay and caspase 3 activity assay. tert-Butyl hydroperoxide was used to induce oxidative injury in endothelial cells, with fluorescent microscopy and flow cytometry used to detect intracellular peroxides and cell apoptosis. Western blot, real-time PCR, STAT3 DNA-binding activity assay, and STAT3 siRNA were used to assess endothelial cell antioxidant enzymes. hPMSC-conditioned medium supported endothelial cell survival and reduced endothelial cell intracellular peroxides and apoptosis. hPMSCs expressed the transcripts of the interleukin (IL) 6 cytokine family, including IL6 and leukemia-inhibitory factor. hPMSC-conditioned medium activated STAT3 expression in endothelial cells, which was inhibited by neutralizing antibody to interleukin 6 signal transducer (IL6ST) but not to IL6 or leukemia-inhibitory factor. STAT3 siRNA or manganese superoxide dismutase (SOD2) siRNA transfected into endothelial cells inhibited the antiapoptotic effect of conditioned medium. SOD2 was significantly upregulated in endothelial cells by conditioned medium via STAT3 activation that, in turn, was inhibited by IL6ST-neutralizing antibody or STAT3 siRNA. Paracrine factors secreted by hPMSCs support endothelial cell survival. STAT3 activation and SOD2 production protect against oxidative stress-induced endothelial cell damage.
Assuntos
Células Endoteliais/metabolismo , Células-Tronco Mesenquimais/metabolismo , Células-Tronco Multipotentes/metabolismo , Placenta/citologia , Fator de Transcrição STAT3/fisiologia , Superóxido Dismutase/metabolismo , Apoptose/efeitos dos fármacos , Apoptose/fisiologia , Sobrevivência Celular/efeitos dos fármacos , Meios de Cultivo Condicionados/farmacologia , Células Endoteliais/efeitos dos fármacos , Feminino , Humanos , Interleucina-6/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Estresse Oxidativo/fisiologia , Comunicação Parácrina/fisiologia , Placenta/metabolismo , Gravidez , Fator de Transcrição STAT3/efeitos dos fármacos , Superóxido Dismutase/efeitos dos fármacosRESUMO
OBJECTIVE: Intrauterine adhesion after hysteroscopic myomectomy contributes to infertility, recurrent miscarriages, menstrual irregularities, and hinders pregnancy outcomes. The aim of this study was to apply the indwelling Malecot catheter in prevention of intrauterine adhesion after hysteroscopic myomectomy and to further evaluate the effectiveness of this approach with reported live birth rates in infertile patients who underwent subsequent infertility treatment. MATERIALS AND METHODS: Seventeen patients with FIGO Classification System PALM-COIEN Type 0 or 1 submucous myoma that received hysteroscopic myomectomy were recruited in this retrospective analysis. Post-operative insertion of the Malecot catheter via the aid of the uterine sound was performed and the catheter was left in place for seven days. RESULTS: The mean duration of TTP (time to pregnancy) was 15.6 months after hysteroscopy. Within three years after the operation, 10 out of 17 infertility patients achieved ongoing pregnancy over 12 weeks. Ongoing pregnancy rate was 58.8% (10/17). Eight patients achieved live birth (seven singletons, one twin pregnancy) with mean gestational age of 38 weeks. Live birth rate was 47.1% (8/17). CONCLUSION: The Malecot catheter is an inexpensive, easy-to-operate, and effective physical barrier method for preventing IUA in infertile patients undergoing hysteroscopic myomectomy with high live birth rate and no obvious visible post-operative adhesions.
Assuntos
Catéteres , Histeroscopia/instrumentação , Complicações Pós-Operatórias/prevenção & controle , Complicações na Gravidez/prevenção & controle , Doenças Uterinas/prevenção & controle , Miomectomia Uterina/instrumentação , Adulto , Coeficiente de Natalidade , Feminino , Humanos , Histeroscopia/efeitos adversos , Histeroscopia/métodos , Infertilidade Feminina/cirurgia , Nascido Vivo , Complicações Pós-Operatórias/etiologia , Gravidez , Complicações na Gravidez/etiologia , Taxa de Gravidez , Estudos Retrospectivos , Aderências Teciduais/etiologia , Aderências Teciduais/prevenção & controle , Doenças Uterinas/etiologia , Miomectomia Uterina/efeitos adversos , Miomectomia Uterina/métodosRESUMO
BACKGROUND: To test if early-cleavage was a strong predictor of pregnancy in patients receiving either a GnRH agonist long protocol or a GnRH antagonist protocol for in-vitro fertilization treatment (IVF) and intracytoplasmic sperm injection (ICSI). METHODS: This retrospective study included 534 patients undergoing a fresh cycle of oocyte retrieval and the day-3 embryo transfer (from 22 to 46 years old). Of the 534 patients treated, 331 received a GnRH agonist long stimulation protocol (GnRH agonist group) for ovarian stimulation and 203 patients received a GnRH antagonist protocol (GnRH antagonist group). In each group, patients who had at least one early-cleavage embryo transferred were designated as the 'early-cleavage' subgroup. Patients who had no early-cleavage embryos transferred were designated as the 'late-cleavage' subgroup. RESULTS: The early cleavage rate was significantly lower in the GnRH antagonist group compared with that in the GnRH agonist group (IVF cycles: 34% versus 20%; ICSI cycles: 50% versus 37.8%, respectively, P < 0.0001). In the GnRH agonist group, the pregnancy rates were significantly higher in the early-cleavage subgroup than those in the late-cleavage subgroup (53.7% vs 33.9%, P < 0.0001). In the GnRH antagonist group, the pregnancy rates were not significantly different between the early-cleavage and late-cleavage subgroups (45.9% vs 43.8%, P > 0.05). CONCLUSION: Early cleavage of zygote is not a reliable predictor for embryo implantation potential in using the GnRH antagonist protocol. Furthermore, the implantation rates between the GnRH agonist and GnRH antagonist groups were comparable.
Assuntos
Fase de Clivagem do Zigoto/citologia , Implantação do Embrião/fisiologia , Hormônio Liberador de Gonadotropina/agonistas , Indução da Ovulação , Taxa de Gravidez , Adulto , Blastômeros/citologia , Protocolos Clínicos , Feminino , Fármacos para a Fertilidade Feminina/administração & dosagem , Fármacos para a Fertilidade Feminina/uso terapêutico , Hormônio Liberador de Gonadotropina/antagonistas & inibidores , Humanos , Infertilidade Feminina/terapia , Pessoa de Meia-Idade , Gravidez , Estudos Retrospectivos , Resultado do TratamentoRESUMO
We report a secreted serine protease inhibitor Kazal-type-like (SPINKL) protein. The SPINKL protein was purified from mouse seminal vesicle secretions through a series of steps, including ion-exchange chromatography on a diethylaminoethyl-Sephacel column, gel filtration on a Sephadex G-75 column, and ion-exchange HPLC on a Q strong anion exchange column. Further analysis identified several SPINKL proteins with various N-linked carbohydrates. The SPINKL protein has six conserved cysteine residues that are nearly identical to those of members of the SPINK protein family. It was noted that the SPINKL protein showed no inhibitory activities against common serine proteases such as trypsin, chymotrypsin, subtilisin, or elastase. Spinkl mRNA and SPINKL proteins were found to be primarily expressed in seminal vesicles. Immunohistochemistry revealed that the SPINKL protein occurred in the luminal fluid and mucosal epithelium of the seminal vesicles and was regulated by testosterone. The SPINKL protein was able to bind onto sperm and enhance sperm motility. Also, it was able to suppress BSA-stimulated sperm capacitation and block sperm-oocyte interactions in vitro, suggesting that SPINKL may be a decapacitation factor.
Assuntos
Proteínas Secretadas Inibidoras de Proteinases/isolamento & purificação , Glândulas Seminais/química , Inibidores de Serina Proteinase/farmacologia , Capacitação Espermática/efeitos dos fármacos , Espermatozoides/efeitos dos fármacos , Reação Acrossômica/efeitos dos fármacos , Sequência de Aminoácidos , Animais , Western Blotting/métodos , Líquidos Corporais/química , Cromatografia Líquida de Alta Pressão , Eletroforese em Gel de Poliacrilamida , Feminino , Fertilidade/efeitos dos fármacos , Masculino , Camundongos , Dados de Sequência Molecular , Proteínas Secretadas Inibidoras de Proteinases/genética , Proteínas Secretadas Inibidoras de Proteinases/farmacologia , RNA Mensageiro/análise , Alinhamento de Sequência , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por MatrizRESUMO
OBJECTIVE: It is known that embryos with faster growing potential, especially in blastocyst development, correlate with the increased euploid rate. Our study investigated the preimplantation genetic screening cycle to analyze the correlation between early blastulation (EB) on day 4 embryo and the euploid rate. MATERIALS AND METHODS: This is a retrospective study examining 273 biopsied blastocysts after preimplantation genetic screening obtained from 54 patients from March 2013 to March 2017. Of the 273 biopsied embryos, 81 had early blastulation on day 4 and were classified as the EB (+) group, while the other 192 had no early blastulation and were classified as the EB (-) group. Euploid rates were compared between the two groups. A total of 34 single euploid embryos were transferred, with 14 from the EB (+) group and 20 from the EB (-) group. Clinical pregnancy was compared between the groups. RESULTS: There is a statistically significant increase in the euploid rate in the EB (+) group (49.4% vs. 34.4%, p = 0.02). The clinical pregnancy rate was also increased in the single euploid embryo transfer group with early blastulation, but did not reach statistical significance (71.4% vs. 50.0%, p = 0.211). CONCLUSIONS: Early blastulation of day 4 embryo correlates significantly with the euploid rate. Early blastulation of day 4 embryo may serve as a potential aid for embryo selection for transfer in preimplantation genetic screening cycles.