Oviductal Epithelial Cells Research Articles

Pgp3 monoclonal antibody inhibits the pathogenicity of Chlamydia muridarum to the genital tract of mice.

Chlamydia trachomatis (Ct) is the leading cause of tubal inflammation in women, with a high tendency for persistent asymptomatic infections. Antibiotics are currently the primary treatment for Ct infections of the reproductive tract. However, mounting evidence indicates an increasing incidence of persistent infections and recurrence due to antibiotic treatment failure, highlighting the urgent need for novel therapeutic approaches. In this study, a monoclonal antibody against plasmid-encoded protein Pgp3 was prepared using hybridoma technology and its effects on the pathogenicity of Ct were investigated both in vitro and in vivo. Infectivity of Chlamydia muridarum (Cm) elementary bodies (EBs) increased after incubation with His-Pgp3. When Pgp3mAb-pretreated Ct EBs or Cm-infected cell lysates were used to inoculate HeLa cells, a significantly reduced number of inclusions was observed compared with untreated controls. Cm-infected HeLa cells began to secrete Pgp3 after 6h. Infection with Cm progeny was significantly inhibited by the addition of Pgp3mAb co-cultured during the first developmental cycle of Cm. Immunofluorescence assays revealed that Pgp3mAb could not enter the host cells. His-Pgp3 stimulated the secretion of IL-6 and IL-8 in human fallopian tube epithelial cells, while Pgp3mAb inhibited this pro-inflammatory effect of His-Pgp3. Cm-infected mice subcutaneously injected with Pgp3mAb demonstrated reduced shedding of live organisms in the lower genital tract, shorter infection cycles, reduced hydrosalpinx, and a reduced inflammatory response. Pgp3 enhanced Cm infectivity in host cells. In vitro, Pgp3mAb inhibited Cm infection by binding to secreted Pgp3 and membrane-bound Pgp3, with a more pronounced effect on secreted Pgp3. Furthermore, Pgp3mAb inhibited the pro-inflammatory effects of Pgp3, thereby attenuating the inflammatory response. Subcutaneous administration of Pgp3mAb effectively reduced Cm-induced pathogenicity in the murine reproductive tract.

Pyrroloquinoline Quinone Improved Boar Sperm Quality via Maintaining Mitochondrial Function During Cryopreservation.

Due to oxidative damage and mitochondrial dysfunction, boar semen cryopreservation remains a significant challenge. This study investigates the effects of pyrroloquinoline quinone (PQQ), a mitochondrial-targeted antioxidant, on the post-thaw boar sperm quality during cryopreservation. Boar semen was diluted in a freezing extender containing different concentrations of PQQ (0, 10, 100, 1000, 10,000 nM). After freezing-thawing, the sperm motility, viability, acrosome integrity, mitochondrial activity, adenosine triphosphate (ATP) levels, DNA integrity, malondialdehyde (MDA) levels, reactive oxygen species (ROS) levels, superoxide dismutase (SOD) activity, mitochondrial transcription proteins levels, and fertilization capacity were assessed. The results show that 1000 nM PQQ supplementation to the freezing extender significantly enhanced post-thaw sperm motility, viability, and acrosome integrity compared to the control (p < 0.05). Additionally, 1000 nM PQQ increased mitochondrial membrane potential (MMP) and ATP levels, while decreasing MDA and mitochondrial ROS levels, and reducing DNA damage (p < 0.05). Furthermore, the levels of mitochondrial-encoded proteins were significantly elevated in the 1000 nM PQQ group compared to the control (p < 0.05). Interestingly, sperm in the 1000 nM PQQ group showed a higher binding rate to oviductal epithelial cells and the zona pellucida (ZP), indicating higher fertilization potential. These findings suggest that the use of mitochondria-target antioxidant, PQQ, can improve post-thaw boar sperm quality and fertilization via its capacity to reduce oxidative stress and protect mitochondrial function.

Ovulation sources ROS to confer mutagenic activities on the TP53 gene in the fallopian tube epithelium

IntroductionEpidemiological studies have implicated ovulation as a risk factor for ovarian high-grade serous carcinoma (HGSC) at the initiation stage. Precancerous lesions of HGSC commonly exhibit TP53 mutations attributed to DNA deamination and are frequently localized in the fallopian tube epithelium (FTE), a site regularly exposed to ovulatory follicular fluid (FF). This study aimed to assess the mutagenic potential of FF and investigate the expression levels and functional role of activation-induced cytidine deaminase (AID) following ovulation, along with the resulting TP53 DNA deamination. MethodsThe mutagenic activity of FF toward premalignant and malignant FTE cells was determined using the hypoxanthine phosphoribosyl transferase (HPRT) mutation assay with or without AID knockdown. The sequential activation of AID, including expressional induction, nuclear localization, DNA binding, and deamination, was determined. AID inducers in FF were identified, and the times of action and signaling pathways were determined. ResultsFF induced AID activation and de novo FTE cell mutagenesis in two waves of activity in accordance with post-ovulation FF exposure. The ERK-mediated early activity started at 2 min and peaked at 45 min, and the NF-κB-mediated late activity started at 6 h and peaked at 8.5 h after exposure. ROS, TNF-α, and estradiol, which are abundant in FF, all induced the two activities, while all activities were abolished by antioxidant cotreatment. AID physically bound to and biochemically deaminated the TP53 gene, regardless of known mutational hotspots. It did not act on other prevalent tumor-suppressor genes of HGSC. ConclusionThis study revealed the ROS-dependent AID-mediated mutagenic activity of the ovulatory FF. The results filled up the missing link between ovulation and the initial TP53 mutation and invited a strategy of antioxidation in prevention of HGSC.

Epidermal growth factor receptor ligands enriched in follicular fluid exosomes promote oncogenesis of fallopian tube epithelial cells

BackgroundIncessant ovulation is the main etiologic factor of ovarian high-grade serous carcinomas (HGSC), which mostly originate from the fallopian tube epithelium (FTE). Receptor tyrosine kinase (RTK) ligands essential for follicle development and ovulation wound repair were abundant in the follicular fluid (FF) and promoted the transformation of FTE cells. This study determined whether RTK ligands are present in FF exosomes and whether epidermal growth factor receptor (EGFR) signaling is essential for oncogenic activity.MethodsThe FF of women undergoing in vitro fertilization was fractionated based on the richness of exosomes and tested for transformation toward FTE cells under different RTK inhibitors. EGFR ligands in FF exosomes were identified, and downstream signaling proteins in FTE cells were characterized.ResultsThe transforming activity of FF was almost exclusively enriched in exosomes, which possess a high capacity to induce anchorage-independent growth, clonogenicity, migration, invasion, and proliferation of FTE cells. EGFR inhibition abolished most of these activities. FF and FF exosome exposure markedly increased EGFR phosphorylation and the downstream signal proteins, including AKT, MAPK, and FAK. Multiple EGF family growth factors, such as amphiregulin, epiregulin, betacellulin, and transforming growth factor-alpha, were identified in FF exosomes.ConclusionsOur results demonstrate that FF exosomes serve as carriers of EGFR ligands as well as ligands of other RTKs that mediate the transformation of FTE cells and underscore the need to further explore the content and roles of FF exosomes in HGSC development.

90 Effect of sediment from spontaneously immortalized oviduct epithelial cells on the development and fertility of bovine OPU-IVF embryos

90 Effect of sediment from spontaneously immortalized oviduct epithelial cells on the development and fertility of bovine OPU-IVF embryos

The presence of sexed sperm in bovine oviduct epithelial cells alters the protein profile related to stress and immune response

The presence of sexed sperm in bovine oviduct epithelial cells alters the protein profile related to stress and immune response

A simple, economical, and high-yield method for polyethylene glycol-based extraction of follicular and serum-derived extracellular vesicles.

The optimization of polyethylene glycol (PEG)-based extracellular vesicles (EVs) extraction from human follicular fluid (FF) and serum was investigated, and their functional analysis was confirmed. The PEG-based EV results were compared to the ExoQuick (ExoQ)-based EV. FF-EVs and serum-EVs were extracted by using different concentrations of PEG (8000). Nanoparticle tracking analysis was used to count the particles, and electron microscopy of EVs was performed for visualization. Exosomes were confirmed by the western blot analysis with exosome-specific markers. RNA and microRNA were extracted from exosomes and quantitative polymerase chain reaction analysis was performed. Fallopian tube epithelial (FTE) cells were used for the EV uptake experiment and an anchorage-independent growth test to confirm that extracted EVs harbor transformation activity. The PEG 8% enriched method produced the highest yield and the lowest carry-over protein. Salt containing PEG 8% produced a higher yield than nonsalted PEG 8%. Overnight enrichment increased four times and 18 times for PEG 8% and ExoQ-based EV extraction from FF. For serum EV, the same overnight enrichment moderately increased yield for both PEG 8% and ExoQ methods. Less carry-over protein resulted in more EV-promoted transformation activity. This study overcomes the time-consuming, expensive, laborious, and complicated machine-dependent EV extraction methods. The study highlights that longer incubation time is needed for EV extraction from FF. PEG 8000-based EV extraction provided a higher yield and less carry-over protein than ExoQ-based EV extraction.

Guinea pig spermatozoa adhesion to an immobilized fibronectin matrix alters their physiology and increases their survival†.

Isthmus is the region of the oviduct considered a reservoir for spermatozoa, where they are retained and released synchronously with ovulation. Integrins mediate this interaction, and it is suggested that they regulate the viability and capacitation of spermatozoa. Spermatozoa retained in the oviductal epithelial cells show specific characteristics: normal morphology, intact acrosome and plasma membrane, no DNA fragmentation, and low levels of intracellular Ca2+, and protein phosphorylation at Tyr. This work aimed to define spermatozoa's ability to adhere to an immobilized fibronectin matrix and its effects on their viability and capacitation. We found that guinea pig spermatozoa showed a high affinity for adhering to an immobilized fibronectin matrix but not to those made up of type 1 collagen or laminin. This interaction was mediated by integrins that recognize the RGD domain. Spermatozoa adhered to an immobilized fibronectin matrix were maintained in a state of low capacitation: low levels of intracellular concentration of Ca2+, protein phosphorylation in Tyr, and F-actin. Also, spermatozoa kept their plasma membrane and acrosome intact, flagellum beating and showed low activation of caspases 3/7. The spermatozoa adhered to the immobilized fibronectin matrix, gradually detached, forming rosettes and did not undergo a spontaneous acrosomal reaction but were capable of experiencing a progesterone-induced acrosomal reaction. In conclusion, the adhesion of spermatozoa to an immobilized fibronectin matrix alters the physiology of the spermatozoa, keeping them in a steady state of capacitation, increasing their viability in a similar way to what was reported for spermatozoa adhered to oviductal epithelial cells.

AMH regulates a mosaic population of AMHR2-positive cells in the ovarian surface epithelium

The function and homeostasis of the mammalian ovary depend on complex paracrine interactions between multiple cell types. Using primary mouse tissues and isolated cells, we showed in vitro that ovarian follicles secrete a factor(s) that suppresses growth of ovarian epithelial cells in culture. Most of the growth suppressive activity was accounted for by Anti-Mullerian Hormone/Mullerian Inhibitory Substance (AMH/MIS) secreted by granulosa cells of the follicles, as determined by immune depletion experiments. Additionally, conditioned medium from granulosa cells from wild type control, but not AMH knockout, suppressed epithelial cell growth. Tracing of the AMH regulated cells using AMHR2 (AMH receptor 2)-Cre:ROSA26 mutant mice indicated the presence of populations of AMHR2-positive epithelial cells on the ovarian surface and oviduct epithelia. Cells isolated from the mutant mice indicated that a subpopulation of cells marked by AMHR2-Cre:ROSA26 accounted for most cell growth and expansion in ovarian surface epithelial cells, and the AMHR2 lineage derived cells were regulated by AMH in vitro; whereas, fewer AMHR2-Cre:ROSA26 marked cells accounted for oviduct epithelial cell outgrowth. The results reveal a paracrine pathway in maintaining follicle-epithelial homeostasis in ovary and support a subpopulation of AMHR2 lineage marked epithelial cells as ovarian epithelial stem/progenitor cells with higher proliferative potential regulatable by follicle secreted AMH.

Maternal stress and the early embryonic microenvironment: investigating long-term cortisol effects on bovine oviductal epithelial cells using air–liquid interface culture

The oviduct epithelium is the initial maternal contact site for embryos after fertilization, offering the microenvironment before implantation. This early gestation period is particularly sensitive to stress, which can cause reduced fertility and reproductive disorders in mammals. Nevertheless, the local impact of elevated stress hormones on the oviduct epithelium has received limited attention to date, except for a few reports on polyovulatory species like mice and pigs. In this study, we focused on the effects of chronic maternal stress on cattle, given its association with infertility issues in this monoovulatory species. Bovine oviduct epithelial cells (BOEC) differentiated at the air–liquid interface (ALI) were stimulated with 250 nmol/L cortisol for 1 or 3 weeks. Subsequently, they were assessed for morphology, bioelectrical properties, and gene expression related to oviduct function, glucocorticoid pathway, cortisol metabolism, inflammation, and apoptosis. Results revealed adverse effects of cortisol on epithelium structure, featured by deciliation, vacuole formation, and multilayering. Additionally, cortisol exposure led to an increase in transepithelial potential difference, downregulated mRNA expression of the major glucocorticoid receptor (NR3C1), upregulated the expression of cortisol-responsive genes (FKBP5, TSC22D3), and significant downregulation of oviductal glycoprotein 1 (OVGP1) and steroid receptors PGR and ESR1. The systematic comparison to a similar experiment previously performed by us in porcine oviduct epithelial cells, indicated that bovine cultures were more susceptible to elevated cortisol levels than porcine. The distinct responses between both species are likely linked to their divergence in the cortisol-induced expression changes of HSD11B2, an enzyme controlling the cellular capacity to metabolise cortisol. These findings provide insights into the species-specific reactions and reproductive consequences triggered by maternal stress.

CCL2 may contribute to the damage of fallopian tube epithelium in women with tubal endometriosis

Research QuestionHow does tubal endometriosis (TEM) impact the morphology and ultrastructure of the fallopian tube epithelium? What are the underlying pathophysiological mechanisms of TEM? DesignEpithelial samples of the fallopian tubes from patients with (TEM group) and without (non-TEM group) TEM were collected. The morphological characteristics, ultrastructure, ciliated cell percentage, and ciliary beat frequency (CBF) were assessed via hematoxylin-eosin staining, electron microscopy, and high-speed video microscopy. mRNA microarray analysis was conducted to identify differentially expressed genes (DEGs) in the fallopian tube epithelium, which were further validated using qPCR, immunohistochemistry, and Western blotting. The effects of recombinant CCL2 protein on primary human fallopian tube epithelial cells (FTEC) were examined in vitro. ResultsPatients with TEM exhibited significant abnormalities in the morphology and ultrastructure of the fallopian tube epithelium with reduced percentage of ciliated cells and CBF, compared to the non-TEM patients. Among the 765 DEGs identified in the fallopian tube epithelium, 512 genes were upregulated and 253 genes were downregulated in the TEM group. qPCR, immunohistochemistry, and Western blotting validation confirmed a significant upregulation of CCL2 expression in the TEM group. In vitro experiments revealed that recombinant CCL2 protein significantly reduced ciliated cell differentiation, length of cilia, and CBF in FTEC, suggesting a role for CCL2 in the development of the TEM-related pathological phenotype. ConclusionsThese findings indicate that CCL2 may contribute to the pathological phenotype associated with TEM and could be a crucial signaling molecule in the damage of fallopian tube epithelium in patients with TEM.

Vitamin D Significantly Inhibits Carcinogenesis in the Mogp-TAg Mouse Model of Fallopian Tube Ovarian Cancer.

Epidemiological and observational studies suggest that vitamin D has potential for the chemoprevention of ovarian cancer. The anticancer effect of vitamin D in the fallopian tube epithelium (FTE), which is now thought to harbor the precursor cells for high grade ovarian cancer, is not known. The purpose of this study was to investigate whether vitamin D can inhibit carcinogenesis in the mogp-TAg fallopian tube (FT) ovarian cancer mouse model and examine underlying mechanisms. To test this hypothesis, 3 groups of 40 5-week-old female mogp-TAg mice were divided equally into two cohorts of 20 mice, treated with either vehicle (vitamin D solvent) or the active 1,25(OH)2D3 analogue EB1089, delivered via mini-pump or IP injection or cholecalciferol delivered in the feed. The FTs were characterized histologically and pathologically after 3 and 7 weeks of treatment. The effect of vitamin D on cultured human FTE cells was also examined. After 3 weeks, vitamin D, delivered as either cholecalciferol or EB1089 significantly inhibited FT carcinogenesis. After 7 weeks, cholecalciferol significantly reduced p53 signatures, serous tubal epithelial carcinoma, FT cancer, and plasma CA125 while increasing apoptosis in the FTE. EB1089 had no significant effect on FT carcinogenesis at 7 weeks. Cholecalciferol significantly reduced proliferation and increased apoptosis in vitro in p53-altered FTE cells. In conclusion, vitamin D inhibited FT carcinogenesis by clearing cells with p53 alterations. These data suggest that vitamin D has merit for the chemoprevention of fallopian tube/ovarian cancer. The optimal chemopreventive effect may be dependent on the route of vitamin D administration.

Expression patterns of folate metabolism-related enzymes in the bovine oviduct: estrous cycle-dependent modulation and responsiveness to folic acid

Folate metabolism is required for important biochemical processes that regulate cell functioning, but its role in female reproductive physiology in cattle during peri- and post-conceptional periods has not been thoroughly explored. Previous studies have shown the presence of folate in bovine oviductal fluid, as well as finely regulated gene expression of folate receptors and transporters in bovine oviduct epithelial cells (BOECs). Additionally, extracellular folic acid (FA) affects the transcriptional levels of genes important for the functioning of BOECs. However, it remains unknown whether the anatomical and cyclic features inherent to the oviduct affect regulation of folate metabolism. The present study aimed to characterize the gene expression pattern of folate cycle enzymes in BOECs from different anatomical regions during the estrous cycle and to determine the transcriptional response of these genes to increasing concentrations of exogenous FA. A first PCR screening showed the presence of transcripts encoding dihydrofolate reductase (DHFR), methylenetetrahydrofolate reductase (MTHFR), and methionine synthase (MTR) in bovine reproductive tissues (ovary, oviduct and uterus), with expression levels in oviductal tissues comparable to, or even higher than, those detected in ovarian and uterine tissues. Moreover, expression analysis through RT-qPCR in BOECs from the ampulla and isthmus during different stages of the estrous cycle demonstrated that folate metabolism-related enzymes exhibited cycle-dependent variations. In both anatomical regions, DHFR was upregulated during the preovulatory stage, while MTHFR and MTR exhibited increased expression levels during the postovulatory stage. Under in vitro culture conditions, ampullary and isthmic cells were cultured in the presence of 10, 50, and 100 μM FA for 24 h. Under these conditions, isthmus epithelial cells exhibited a unique transcriptional response to exogenous FA, showing a pronounced increase in MTR expression levels. Our results suggest that the expression of folate metabolism-related genes in BOECs is differentially regulated during the estrous cycle and may respond to exogenous levels of folate. This offers a new perspective on the transcriptional regulation of genes associated with the folate cycle in oviductal cells and provides groundwork for future studies on their functional and epigenetic implications within the oviductal microenvironment.

Extracellular vesicles affecting embryo development in vitro: a potential culture medium supplement.

Extracellular vesicles (EVs) are nanometer-sized lipid bilayer vesicles released by cells, playing a crucial role in mediating cellular communication. This review evaluates the effect of EVs on early embryonic development in vitro by systematically searching the literature across three databases, Embase, PubMed, and Scopus, from inception (Embase, 1947; PubMed, 1996; and Scopus, 2004) to 30 June 2024. A total of 28 studies were considered relevant and included in this review. The EVs included in these investigations have been recovered from a range of sources, including oviduct fluid, follicular fluid, uterine fluid, seminal plasma, embryos, oviduct epithelial cells, endometrial epithelial cells, amniotic cells, and endometrial-derived mesenchymal stem cells collected primarily from mice, rabbits, cattle and pigs. This diversity in EV sources highlights the broad interest and potential applications of EVs in embryo culture systems. These studies have demonstrated that supplementation with EVs derived from physiologically normal biofluids and cells to the embryo culture medium system has positive effects on embryonic development. Conversely, EVs derived from cells under pathological conditions have shown a negative impact. This finding underscores the importance of the source and condition of EVs used in culture media. Further, the addition of EVs as a culture medium supplement holds significant therapeutic potential for optimizing in vitro embryo culture systems. In conclusion, this evaluation offers a thorough assessment of the available data on the role of EVs in embryo culture media and highlights the potential and challenges of using EVs in vitro embryo production.

HPV-YAP1 oncogenic alliance drives malignant transformation of fallopian tube epithelial cells

High grade serous ovarian carcinoma (HGSOC) is the most common and aggressive ovarian malignancy. Accumulating evidence indicates that HGSOC may originate from human fallopian tube epithelial cells (FTECs), although the exact pathogen(s) and/or molecular mechanism underlying the malignant transformation of FTECs is unclear. Here we show that human papillomavirus (HPV), which could reach FTECs via retrograde menstruation or sperm-carrying, interacts with the yes-associated protein 1 (YAP1) to drive the malignant transformation of FTECs. HPV prevents FTECs from natural replicative and YAP1-induced senescence, thereby promoting YAP1-induced malignant transformation of FTECs. HPV also stimulates proliferation and drives metastasis of YAP1-transformed FTECs. YAP1, in turn, stimulates the expression of the putative HPV receptors and suppresses the innate immune system to facilitate HPV acquisition. These findings provide critical clues for developing new strategies to prevent and treat HGSOC.

Effect of the estrous cycle on zinc transporter expression in bovine cumulus-oocyte complexes and oviduct epithelial cells.

During the luteal and follicular phases of the estrous cycle, cumulus-oocyte complexes (COC) and oviduct epithelial cells (OEC) undergo notable physiological and morphological changes. Maintaining proper zinc (Zn) homeostasis is crucial in both somatic and germinal mammalian cells. This study aimed to assess the impact of the estrous phase (luteal or follicular) on Zn transporter expression in bovine COC and OEC (BOEC). The expression of Zn transporters Slc39a6 (ZIP6), Slc39a8 (ZIP8), Slc39a14 (ZIP14), Slc30a3 (ZnT3), Slc30a7 (ZnT7), and Slc30a9 (ZnT9) was analyzed in COC and BOEC from cows during the luteal or follicular phases. Gene expression of ZIP6, ZIP14, and ZnT9 was quantified in COC and BOEC. The gene expression in the remaining transporters could not be quantified due to low mRNA levels (ZIP8 and ZnT3 in COC and BOEC; ZnT7 in BOEC) or absence of expression (ZnT7 in COC). In COC, the relative expression (RE) of all three transporters was higher in the luteal phase compared to the follicular phase (P ≤ 0.05). In BOEC, the luteal phase increased the RE of ZIP 6 (P ≤ 0.05), decreased the RE of ZnT9 (P ≤ 0.05), and did not modify the RE of ZIP14 (P > 0.05) compared to the follicular phase. In conclusion, the study reveals differences in the gene expression of ZIP6, ZIP14, and ZnT9 according to the estrous cycle phase in ex vivo samples of bovine COC and OEC.

ISGylation enhances dsRNA-induced interferon response and NFκB signaling in fallopian tube epithelial cells

Heritable mutations in BRCA1 associate with increased risk of high-grade serous tubo-ovarian cancer (HGSTOC). Non-genetic risk factors associated with this cancer, which arises from fallopian tube epithelial (FTE) cells, suggests a role for repetitive ovulation wherein FTE cells are exposed to inflammatory signaling molecules within follicular fluid. We previously reported increased NFκB and EGFR signaling in BRCA1-deficient primary FTE cells, with follicular fluid exposure further increasing abundance of interferon-stimulated gene (ISG) transcripts, including the ubiquitin-like protein ISG15 and other ISGylation pathway members. Both NFκB and type I interferon signaling are upregulated by stimulation of cGAS-STING or MDA5 and RIGI pattern recognition receptors (PRRs). Since some PRRs and their signal transduction pathway members are ISGylated, we tested the impact of ISG15 and ISGylation on IRF3 and NFκB signaling through cGAS-STING or RIGI and MDA5 activation. Expression of ISG15 or UBA7, the E1-like ISG15 activating enzyme, in immortalized FTE cells was disrupted by CRISPR gene editing. Activation of IRF3 by RIGI or MDA5 but not cGAS-STING was attenuated by loss of either ISG15 or UBA7 and this was reflected by a similar effect on NFκB activation and downstream targets. Loss of ISGylation decreased levels of both MDA5 and RIGI, with knock-down of RIGI but not MDA5, decreasing IRF3 and NFκB activation in parental cells. These finding indicate that ISGylation enhances the ability of dsRNA to activate cytokine release and pro-inflammatory signaling. Further work to explore ISGylation as a target for prevention of HGSTOC in BRCA1 mutation carriers is warranted.

Unveiling the role of protein kinase A (PKA) activity in bovine oviductal epithelial cells: implications on apoptotic signaling pathways during the estrous cycle.

The complex interactome crucial for successful pregnancy is constituted by the intricate network of endocrine and paracrine signaling pathways, involving gametes, embryos, and the female reproductive tract. Specifically, the oviduct exhibits distinct responses to gametes and early embryos during particular phases of the estrus cycle, a process tightly regulated by reproductive hormones. Moreover, these hormones play a pivotal role in orchestrating cyclical changes within oviductal epithelial cells. To unravel the molecular mechanisms underlying these dynamic changes, our study aimed to investigate the involvement of protein kinase A (PKA) in oviductal epithelial cells throughout the estrus cycle and in advanced pregnancy, extending our studies to oviductal epithelial cell in primary culture. By a combination of 2D-gel electrophoresis, Western blotting, and mass spectrometry, we identified 17 proteins exhibiting differential phosphorylation status mediated by PKA. Among these proteins, we successfully validated the phosphorylation status of heat shock 70kDa protein (HSP70), aconitase 2 (ACO2), and lamin B1 (LMNB1). Our findings unequivocally demonstrate the dynamic regulation of PKA throughout the estrus cycle in oviductal epithelial cells. Also, analysis by bioinformatics tools suggest its pivotal role in mediating cyclical changes possibly through modulation of apoptotic pathways. This research sheds light on the intricate molecular mechanisms underlying reproductive processes, with implications for understanding fertility and reproductive health.

A bovine oviduct microfluidic platform to evaluate the effects of cisplatin and hormones on cilia beating frequencies

Chemotherapeutics such as cisplatin modulate the function of cells in fallopian tubes (oviducts) reducing the formation of sperm reservoirs that promote gamete interaction during the fertilization process. The mechanisms behind these resulting deficiencies must be better understood to improve fertility for cancer patients. A microfluidic platform was developed by immobilizing a transparent polycarbonate membrane between two poly(dimethyl siloxane) molds to make channels for the growth of a functional monolayer of oviduct cells on one side and medium on the other side. Freshly isolated bovine oviduct epithelial cells were cultured on the membrane with an air-liquid interface after 5 days, and the resulting cells had consistent cilia beating frequencies (CBF) for 12 weeks between 4.92 ± 0.59 Hz and 5.76 ± 0.94 Hz. Incubation with 75 pg ml−1 17-beta-estradiol and 100 ng ml−1 progesterone increased the CBF from 4.62 ± 0.16 Hz to 7.61 ± 0.41 Hz. When incubated with 50 µM of cisplatin, the CBF dropped by 40 ± 19 % without hormones and 52 ± 9 % with hormones, with similarly decreased CBFs. Short 2-h incubations of bovine oviduct cells with cisplatin directly reduced the beating of ciliated cells, illustrating that the short-term effects (hours) of chemotherapeutics should be considered in oncofertility.

Investigations into the role of platelet-activating factor in the peri-conception period of the mare.

In many mammals, the lipid platelet-activating factor (PAF) has important functions in female reproduction and fertility. This study shows that PAF is present in the reproductive tissues of mares and is involved in processes related to ovulation and early pregnancy. Platelet-activating factor (PAF) has been implicated in a number of reproductive processes ranging from ovulation to embryo motility but has not been widely explored in the mare. To identify the presence and examine the role of PAF in the equine periconception processes, targeted mass spectrometry coupled with chromatographic separation was performed on equine follicular fluid (FF), and PAF was quantitatively detected. Subsequently, untargeted high-resolution mass spectrometry-based lipidomic analysis was carried out to quantify PAF in different-sized pre-ovulatory follicles, whereby different molecular species of PAF, PAF (14:0) and PAF (16:1), were both seen to be increasing with follicle diameter. These findings suggest that PAF within FF is increasing as preovulatory follicles approach ovulation. Additionally, immunofluorescence staining identified the PAF receptor in the luminal pericellular, apical, and basal aspect of equine oviductal epithelial cells. Lastly, an equine oviductal epithelial organoid model was generated and showed that the addition of PAF significantly increased the ciliary beat frequency (CBF) (Hz), an action consistent with a role for PAF in embryo migration. It is proposed that the local action of PAF on the ciliated cells of the oviduct propels both the oocyte and the conceptus towards the uterus. In the mare, it appears that PAF is a contributor during the periconception period, potentially being a mediator in the mechanisms of ovulation and in the dialogue of very early pregnancy.