Transcriptomic Analysis Reveals Intrinsic Abnormalities in Endometrial Polyps.

Endometrial polyps (EPs) are benign overgrowths of the endometrial tissue lining the uterus, often causing abnormal bleeding or infertility. This study analyzed gene expression differences between EPs and adjacent endometrial tissue to elucidate intrinsic abnormalities promoting pathological overgrowth. RNA sequencing of 12 pairs of EPs and the surrounding endometrial tissue from infertile women revealed 322 differentially expressed genes. Protein-protein interaction network analysis revealed significant alterations in specific signaling pathways, notably Wnt signaling and vascular smooth muscle regulation, suggesting these pathways play critical roles in the pathophysiology of EPs. Wnt-related genes DKK1 and DKKL1 were upregulated, while GPC3, GREM1, RSPO3, SFRP5, and WNT10B were downregulated. Relevant genes for vascular smooth muscle contraction were nearly all downregulated in EPs, including ACTA2, ACTG2, KCNMB1, KCNMB2, MYL9, PPP1R12B, and TAGLN. Overall, the results indicate fundamental gene expression changes promote EP formation through unrestrained growth signaling and vascular defects. The intrinsic signaling abnormalities likely contribute to clinical symptoms of abnormal uterine bleeding and infertility common in EP patients. This analysis provides molecular insights into abnormal endometrial overgrowth to guide improved diagnostic and therapeutic approaches for this troublesome women's health condition. Confirmation of expanded cohorts and further investigations into implicated regulatory relationships are warranted.

Correlation between Sperm Micro Ribonucleic Acid-34b and -34c Levels and Clinical Outcomes of Intracytoplasmic Sperm Injection in Men with Male Factor Infertility.

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.

Secretory mouse quiescin sulfhydryl oxidase 1 aggregates defected human and mouse spermatozoa in vitro and in vivo.

A flavin-dependent enzyme quiescin Q6 sulfhydryl oxidase 1 (QSOX1) catalyzes the oxidation of thiol groups into disulfide bonds. QSOX1 is prominently expressed in the seminal plasma. However, its role in male reproduction is elusive. Here, we purified the secreted form of QSOX1, i.e., QSOX1c, from mouse seminal vesicle secretions and revealed for the first time its function involved in sperm physiology. Exogenous addition of QSOX1c time-dependently promoted the in vitro aggregation of thiol-rich, oxidative stressed, and apoptotic mouse and human sperm cells. Also, in vivo aggregated sperm cells collected from mouse uterine and human ejaculates also showed high levels of QSOX1c, intracellular reactive oxygen species, annexin V, and free thiols. In summary, our studies demonstrated that QSOX1c could agglutinate spermatozoa susceptible to free radical attack and apoptosis. This characteristic may provide an opportunity to separate defective sperm cells and improve sperm quality before artificial insemination in humans and animals.

A mouse seminal vesicle-secreted lysozyme c-like protein modulates sperm capacitation.

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.

Serine Protease Inhibitor SERPINE2 Reversibly Modulates Murine Sperm Capacitation.

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.

Expression of cystatin C in the female reproductive tract and its effect on human sperm capacitation.

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.

Correlation of cumulus gene expression of GJA1, PRSS35, PTX3, and SERPINE2 with oocyte maturation, fertilization, and embryo development.

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.

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.