Comparison of the Cost and Effect of Combined Conditioned Medium and Conventional Medium for Fallopian Tube Organoid Cultures.

Fallopian tube epithelial cells (FTEC) are thought to be the cell of origin of high-grade serous ovarian carcinoma. FTEC organoids can be used as research models for the disease. Nevertheless, culturing organoids requires a medium supplemented with several expensive growth factors. We proposed that a combined conditioned medium based on the composition of the fallopian tubes, including epithelial, stromal, and endothelial cells could enhance FTEC organoid formation. We derived two primary culture cell lines from the fimbria portion of the fallopian tubes. The organoids were split into conventional or combined medium groups based on what medium they were grown in and compared. The number and size of the organoids were evaluated. Quantitative polymerase chain reaction (qPCR) and immunohistochemistry (IHC) were used to evaluate gene and protein expression (PAX8, FOXJ1, beta-catenin, and stemness genes). Enzyme-linked immunosorbent assay was used to measure Wnt3a and RSPO1 in both mediums. DKK1 and LiCl were added to the mediums to evaluate their influence on beta-catenin signaling. The growth factor in the combined medium was evaluated by the growth factor array. We found that the conventional medium was better for organoids regarding proliferation (number and size). In addition, WNT3A and RSPO1 concentrations were too low in the combined medium and needed to be added making the cost equivalent to the conventional medium. However, the organoid formation rate was 100% in both groups. Furthermore, the combined medium group had higher PAX8 and stemness gene expression (OLFM4, SSEA4, LGR5, B3GALT5) when compared with the conventional medium group. Wnt signaling was evident in the organoids grown in the conventional medium but not in the combined medium. PLGF, IGFBP6, VEGF, bFGF, and SCFR were found to be enriched in the combined medium. In conclusion, the combined medium could successfully culture organoids and enhance PAX8 and stemness gene expression. However, the conventional medium was a better medium for organoid proliferation. The expense of both mediums was comparable. The benefit of using a combined medium requires further exploration.

Traditional and contemporary views on the functional morphology of the fallopian tubes and their importance for gynecological practice.

The uterine tube, belonging to the female internal reproductive organs, is the only tubular organ in the human body that has, under physiological conditions, a transport function occurring in two opposite directions. It transports the picked-up oocyte released during ovulation and early embryo towards the uterine cavity. At the same time, it can transport spermatozoa towards the abdominal opening of the fallopian tube. Moreover, the uterine tube has many other vital functions as sperm selection (one of the crucial factors preventing polyspermy) and the production of tubal fluid. This unique secretion is essential not only for the process of fertilization but also for sperm activation and the nourishment of the early embryo during its transport into the uterine cavity. The first part of our review is focused on the historical introduction to the topic in which the reader will become familiar with the views and understanding of these peculiar organs by famous anatomists of the 16th and 17th centuries, namely Gabriele Falloppio and Renier de Graaf. The following section will cover the overview of the latest anatomical, embryological, and histological knowledge, which are also crucial for a better understanding of pathological processes affecting the fallopian tube, such as tubal infertility or tubal pregnancy. Interestingly, recent years have been very fruitful regarding uterine tube morphology, e. g. the discovery of an unique mechanism of lymphatic flow within the uterine tube mucosa, the first description of immunologically-active intraepithelial suppressor T-lymphocytes, or the observation of pacemaker cell population - telocytes - in the muscle layer.

The fertility sparing therapy in ectopic pregnancy.

OBJECTIVE: A review of current knowledge on the possibilities of fertility sparing therapy in case of ectopic pregnancy. METHODS AND RESULTS: Ectopic pregnancy is defined as implantation of an embryo outside the endometrial cavity, most often in the fallopian tube. This dia-gnosis is very common among young women. Ectopic pregnancies can be treated using the following three approaches, which can be combined: expectantly, pharmacologically or surgically. Fertility-sparing salpingostomy may be performed during surgical treatment. Medical (pharmacological) treatment consists in the application of methotrexate with a success rate of 75-96%, depending on the initial level of the free beta subunit of human chorionic gonadotropin (b-hCG). This is a safe treatment with minimal side effects. There is no standardization of the blood b-hCG level limits or of the size of the ectopic pregnancy mass for choosing expectant, surgical or medical treatment. A considerable increase in the rate of Cesarean sections over the last decades has led to an increase in the occurrence of the implantation of the gestational sac in the hysterotomy scar. There are several options to address this dia-gnosis, but none is clearly preferred. This issue is also discussed in the article. CONCLUSION: The goal of ectopic pregnancy treatment is to choose a safe and effective therapy with a low incidence of side effects and maintaining the maximum fertility of women. Properly set indication criteria are most important when choosing the right option.

Interstitial Ectopic Pregnancy-Case Reports and Medical Management.

The term intramural (interstitial) ectopic pregnancy refers to a pregnancy developing outside the uterine cavity, with a gestational sac implanted into the interstitial part of the Fallopian tube, surrounded by a layer of the myometrium. The prevalence rate of interstitial pregnancy (IP) is 2-4% of all ectopic pregnancies. Surgery is the primary treatment for interstitial ectopic pregnancy; the pharmacological management of ectopic pregnancy, including IP, in asymptomatic patients includes systemic administration of methotrexate. In this report, we present two cases of this rare pregnancy type, reviewing our management technique and treatment ways presented in the literature. In our patients, the management was initially conservative and included methotrexate, administered as intravenous bolus injection, regular beta-human chorionic gonadotropins (ß-HCG) level measurements in peripheral blood, and monitoring of the patient's general condition. Due to signs of intra-abdominal bleeding in patient A and inadequate ß-HCG level reduction in patient B, both patients eventually underwent laparoscopic cornual resection. Pregnancy, implanted into the interstitial part of the Fallopian tube and surrounded by myometrial tissue with myometrial invasion of the trophoblast, poses a serious diagnostic challenge to modern gynecology due to particularly low sensitivity and specificity of symptoms, and may require both pharmacological and surgical treatment.

Efficacy and Safety of Platinum-based Chemotherapy With Bevacizumab Followed by Bevacizumab Maintenance for Recurrent Ovarian, Fallopian Tube, and Primary Peritoneal Cancer During PARP Inhibitor Therapy: A Multicenter Retrospective Study.

BACKGROUND/AIM: In recent years, the usefulness of poly ADP-ribose polymerase (PARP) inhibitors as subsequent maintenance therapy with poly ADP-ribose polymerase (PARP) inhibitors has been reported. However, it has been reported shown that platinum-based chemotherapy has a low response rate and short progression-free survival for recurrent platinum-sensitive ovarian cancer during treatment with PARP inhibitor therapy. This retrospective study evaluated platinum-based chemotherapy with bevacizumab (BEV) followed by BEV maintenance in these recurrent patients. PATIENTS AND METHODS: Efficacy and safety were evaluated in 23 patients with ovarian, fallopian tube, or primary peritoneal cancer diagnosed with platinum-sensitive recurrence during PARP inhibitor treatment (administered from April 2019 to December 2022). Platinum-based chemotherapy included either paclitaxel with carboplatin, paclitaxel with cisplatin, docetaxel with carboplatin, or doxorubicin with carboplatin. BEV was administered in combination with any of these chemotherapies agents. Chemotherapy was administered for 6 cycles and BEV was administered up to 21 cycles. RESULTS: The median numbers of cycles of platinum-based chemotherapy and BEV administration were 6 and 8, respectively. Complete response was observed in four patients (17.4%), partial response in 15 (65.2%), stable disease in two (8.7%), and progressive disease in two (8.7%). Objective response and disease control rates were 82.6% and 91.3%, respectively. Grade 3 or higher hematological toxicity occurred in 8 patients, with leukopenia, neutropenia in 14, anemia in 5, and thrombocytopenia in 4. On the other hand, non-hematological toxicities included hypertension in three patients, proteinuria in two, constipation in one, and carboplatin hypersensitivity in four. Only one patient discontinued chemotherapy due to an adverse event of proteinuria. No treatment-related deaths occurred. CONCLUSION: Platinum-based chemotherapy with BEV followed by BEV maintenance for platinum-sensitive recurrence during PARP inhibitor treatment was shown to be efficacious and safe. This combination should be further evaluated in larger randomized clinical trials.

A novel technique for large-fragment knock-in animal production without ex vivo handling of zygotes.

CRISPR/Cas-based genome editing has dramatically improved genetic modification technology. In situ electroporation called genome editing via oviductal nucleic acid delivery (GONAD), which eliminates the need for ex vivo embryo handling, is technically the simplest method for gene transfer and can be performed in laboratories without developmental engineering expertise including micromanipulation techniques. However, the use of this method remains challenging in the case of large-fragment knock-in, such as gene expression cassettes. Adeno-associated viruses (AAV) act as donor DNA for homologous recombination in infected cells, including rodent embryos. In this study, we demonstrated simultaneous electroporation of AAV donors and CRISPR/Cas9 components into embryos to create knock-in animals, and successfully generated knock-in rats carrying a gene cassette with a length of 3.0 kb using a small number of animals and in situ electroporation. These findings indicate that this technique is an efficient high-throughput strategy for producing genetically modified rodents and may be applicable to other animal species.

CytoSaLPs score: a promising new tool for the detection and screening of extrauterine high grade serous carcinoma.

BACKGROUND: A recent theory supports that high-grade serous epithelial ovarian cancer derives from the fimbrial end of the fallopian tube and during the last decade, a few studies have examined the fallopian tube cytology. Our study aims to determine the cytomorphological characteristics of both benign and non-benign salpingeal samples, in order to establish fallopian cytology as a valuable diagnostic test for women with high risk for development of ovarian/fallopian/peritoneal cancer. METHODS: Our study included patients undergoing salpingoophorectomy or total hysterectomy and salpingoophorectomy for any gynecological pathology. Using a soft brush, fallopian tube smears from the fimbrial end were collected ex vivo. The Cytologists of our Institution described the morphological characteristics of the fallopian cells by adopting a proposed Table, which had a calibration system ranging from 3 to 29. This Table is referred to as the CytoSaLPs Score. Our study compared the two diagnostic cytological methods, the one of the conventional cytology and the other using the CytoSaLPs Score, having as gold standard the tubal's pathological findings. RESULTS: A total of 230 tubal specimens from 144 patients were included in this study. The Score's mean for the benign and non-benign arm was 12.8 and 18.7 respectively. The cut-off point for both arms was 16.5. The CytoSaLPs Score tool showed significantly higher specificity (87.50% vs. 75.96, p-value < 0.001) and positive predictive value PPV (40.91% vs. 26.47%, p-value < 0.001) compared to conventional cytology. Regarding the accuracy, the Score's superiority is highlighted (86.96% vs. 76.52%, p-value < 0.001). CONCLUTIONS: The evaluation of tubal cytology using the CytoSaLPs Score could be used as a reliable diagnostic method. Further evaluation with larger studies is warranted.

[Morphological precursors of high-grade serous ovarian cancer]. / Morfologicheskie predshestvenniki seroznogo raka yaichnikov vysokoi stepeni zlokachestvennosti.

At the beginning of this century, there was a paradigm shift in understanding the histogenesis of high-grade serous carcinomas. The theory of the origin of these tumors from the ovarian surface epithelium was replaced by the concept of their origin from the secretory epithelium of the fallopian tubes. In recent years, researchers have put forward the hypothesis of the "escape" of the precursor of high-grade serous carcinomas. It allows looking at the carcinogenesis of these neoplasms as a natural history of tumor transformation of the serous epithelium without reference to a specific localization.

High visceral fat-to-muscle ratio is an independent factor that predicts worse overall survival in patients with primary epithelial ovarian, fallopian tube, and peritoneal cancer.

BACKGROUND: The intra-abdominal cavity, surrounded by adipocytes, is the main metastatic site of epithelial ovarian, fallopian tube, and peritoneal cancer. Epidemiological and molecular studies have demonstrated a link between adipose tissue and ovarian cancer. However, the clinical significance of fatty tissue has not been elucidated. Thus, we investigated the clinical significance of body composition in patients with epithelial ovarian, fallopian tube, and peritoneal cancer. METHODS: Fat and skeletal muscle areas were measured using software based on pretreatment computed tomography scans at the third lumbar vertebra. Fat-to-muscle ratios were calculated using the total (visceral and subcutaneous) fat area or visceral fat area. High fat-to-muscle ratios were defined by values greater than the mean. Sarcopenia was defined as a skeletal muscle index < 38.7 cm2/m2. The clinicopathological parameters and survival of 153 patients were analyzed. RESULTS: High visceral fat-to-muscle ratios and sarcopenia at the time of diagnosis were observed in 43.8% and 33.3% of the patients, respectively. Multivariate analysis showed that high visceral fat-to-muscle ratio (p = 0.014), advanced Federation of Gynecology and Obstetrics stage (p = 0.008), and chemoresistance (p = 0.027) were independent factors for worse overall survival. Patients with high visceral fat-to-muscle ratios were older, had higher body mass indexes, and were more likely to have diabetes/hypertension, serous cancer subtypes, and implementation of neoadjuvant chemotherapy than those with low visceral fat-to-muscle ratios. The platelet count was significantly higher in the high visceral fat-to-muscle ratio group than in the low visceral fat-to-muscle ratio group (p = 0.011). CONCLUSIONS: Pretreatment visceral fat area could be an independent predictive factor of overall survival in patients with epithelial ovarian, fallopian tube, and peritoneal cancer and may be significantly associated with thrombocytosis.

LINE-1 ORF1p as a candidate biomarker in high grade serous ovarian carcinoma.

Long interspersed element 1 (LINE-1) open reading frame 1 protein (ORF1p) expression is a common feature of many cancer types, including high-grade serous ovarian carcinoma (HGSOC). Here, we report that ORF1p is not only expressed but also released by ovarian cancer and primary tumor cells. Immuno-multiple reaction monitoring-mass spectrometry assays showed that released ORF1p is confidently detectable in conditioned media, ascites, and patients' plasma, implicating ORF1p as a potential biomarker. Interestingly, ORF1p expression is detectable in fallopian tube (FT) epithelial precursors of HGSOC but not in benign FT, suggesting that ORF1p expression in an early event in HGSOC development. Finally, treatment of FT cells with DNA methyltransferase inhibitors led to robust expression and release of ORF1p, validating the regulatory role of DNA methylation in LINE-1 repression in non-tumorigenic tissue.

When a balloon catheter or tenaculum is required for cervical traction during hysterosalpingography.

The aim of this study was to define the actual rate of the traction needed and the balloon catheter or tenaculum requirement for hysterosalpingography (HSG) examinations, and to investigate the correlation between pain scores with the type of traction, operator, parity or the type of infertility. 788 patients undergoing HSG participated in the trial. The HSG examinations were completed in 58% of the patients (458) without any traction. Traction was needed in 42% of patients, those with the balloon catheter in 26.9%, and those with the tenaculum only at 15%. Patients with balloon catheter traction had similar pain scores to those using tenaculum traction. The pain scores changed according to the operator. HSG examinations should be performed step-by-step and the need for traction evaluated during the procedure.IMPACT STATEMENTWhat is already known on this subject? HSG is a technique to evaluate fallopian tube patency and other potential intrauterine pathology in infertile women.What do the results of this study add? Traction was not needed in more than half of the HSG examinations. The pain due to the balloon catheter and tenaculum is similar.What the implications are of these findings for clinical practice and/or further research? HSG examinations should be performed step-by-step by checking the need for traction. Traction with the balloon catheter prevents the uterine spasm, infection and bleeding complications during or after the HSG.

Involvement of gap junctions in the increased contraction of bovine oviducts at pre-ovulation observed in vitro.

In brief: Spontaneous contraction of oviductal smooth muscle is essential for gamete transport to the fertilization site in mammals. This study sheds light on the mechanism of elevated contraction amplitude in the bovine oviductal isthmus just before ovulation. Abstract: Rhythmic contraction of the oviducts is essential for transporting gametes and embryos at peri-ovulation; however, its regulatory mechanism during the estrous cycle is unclear. Meanwhile, it is reported that ion currents regulate muscle contraction. Our study aimed to clarify the involvement of ion channels and gap junctions in regulating oviductal motility during the estrous cycle in cattle. The isthmic sections of bovine oviducts collected just after ovulation (0-4 days after ovulation), at the mid-late luteal stage (10-17 days), and at the follicular stage (1-3 days before ovulation) were used in the experiments. The frequency and amplitude of contraction of the oviductal strips in the longitudinal direction were examined using the Magnus system. The frequency was not different among the estrous stages. Conversely, the amplitude was significantly higher at the follicular stage. The blockers of voltage-dependent calcium channels, both IP3 receptor and ryanodine receptors, chloride channel, and gap junction reduced the amplitude. Additionally, mRNA and protein expression of GJA1, a component of the gap junction, in the smooth muscle tissues of the oviductal isthmus were significantly higher in the follicular stage. In addition, estradiol-17ß (E2; 1.0 ng/mL) significantly increased GJA1 mRNA expression in cultured smooth muscle tissues after 24 h and GJA1 protein expression in cultured smooth muscle cells after 48 h. These results suggest that local levels of E2 in the oviductal isthmus ipsilateral to an ovary with a dominant follicle support the increased contraction amplitude of bovine ipsilateral oviducts by elevating the gap junction expression.

Metabolic memory determines oviductal gene expression of underfed ewes during early gestation.

Our aim was to investigate the oviduct environment by studying oviduct gene expression after undernutrition in day-5 pregnant ewes with different initial (i) BCS, and its association with the number of embryos recovered. Thirty-six ewes were divided into 2 groups with different iBCS: iBCS ≥2.75 (n = 19; high, H) and iBCS ≤2.25 (n = 17; low, L), and were randomly assigned to two nutritional treatments for 20 days: 1.5 (control, C) or 0.5 (underfed, U) times the daily maintenance requirements. Thus, the final four groups were: high-iBCS control (HC, n = 9), high-iBCS underfed (HU, n = 10), low-iBCS control (LC, n = 9) and low-iBCS underfed (LU, n = 8). Samples of oviduct were collected and the expression of target genes was quantified using real-time PCR. While high-iBCS control ewes presented more ADIPOR1 mRNA than the high-iBCS underfed group (P < 0.05) and low-iBCS control ewes (P = 0.01), high-iBCS underfed group presented higher ADIPOR2 gene expression than low-iBCS underfed ewes (P < 0.01) evidencing a differential oviductal gene expression for these receptors. In high-iBCS ewes, control animals presented higher IGFBP2 gene expression than underfed ewes (P < 0.05), associated these results with a poor oviductal environment. High-iBCS underfed ewes presented higher IGFBP4 gene expression than high-iBCS control ewes (P < 0.05). Stepwise regression models, using various combinations of data on metabolic and reproductive hormones, and oviduct gene expression as independent variables, identified a set of variables that accounted for 75% of the variation in the number of embryos recovered. In conclusion, the oviductal gene expression depends on body reserves and nutritional treatment, and the effect is gene-specific.

Optical coherence tomography for dynamic investigation of mammalian reproductive processes.

The biological events associated with mammalian reproductive processes are highly dynamic and tightly regulated by molecular, genetic, and biomechanical factors. Implementation of live imaging in reproductive research is vital for the advancement of our understanding of normal reproductive physiology and for improving the management of reproductive disorders. Optical coherence tomography (OCT) is emerging as a promising tool for dynamic volumetric imaging of various reproductive processes in mice and other animal models. In this review, we summarize recent studies employing OCT-based approaches toward the investigation of reproductive processes in both, males and females. We describe how OCT can be applied to study structural features of the male reproductive system and sperm transport through the male reproductive tract. We review OCT applications for in vitro and dynamic in vivo imaging of the female reproductive system, staging and tracking of oocytes and embryos, and investigations of the oocyte/embryo transport through the oviduct. We describe how the functional OCT approach can be applied to the analysis of cilia dynamics within the male and female reproductive systems. We also discuss the areas of research, where OCT could find potential applications to progress our understanding of normal reproductive physiology and reproductive disorders.

Immune disturbances in female genital tuberculosis and latent genital tuberculosis.

BACKGROUND: Female genital tuberculosis (FGTB), an important clinical sub-type of extra-pulmonary tuberculosis (EPTB) is responsible for about 10% cases of infertility in India. Both FGTB and latent genital tuberculosis (LGTB) can cause infertility through blockage of fallopian tubes and through altered uterine endometrial receptivity. AIMS: This review tries to elucidates the role of various immune factors in FGTB and LGTB. CONTENT: Various immune disturbances are observed in FGTB and LGTB like growth factors and cytokines which inhibit implantation and several inflammatory signaling pathways like mitogen activated protein kinase (MAPK), natural killer (NK) cells, nuclear factor kappa-B (NF-KB), tumor necrosis factor (TNF), and toll like receptors (TLR) signaling are dysregulated. These altered immune factors and pathways may be detected in the endometrial biopsies at the early stages of disease before permanent damage. Prompt and adequate treatment with the four anti-tubercular drugs (rifampicin [R], isoniazid [H], pyrazinamide [Z], and ethambutol [E]) can increase pregnancy rates in some of these women. Assisted reproduction especially in-vitro fertilization and embryo transfer may be required for some women. IMPLICATIONS: Inflammatory pathways identified from the gene profiling have enabled development of potential biomarkers for early diagnosis of FGTB. Immunomodulation and novel biotechniques like stem cell transplantation, nanoparticles and host directed therapies are being tried in selected patients of FGTB and LGTB with promising results.

Comparative transcriptomes of nine tissues for the Heilongjiang brown frog (Rana amurensis).

The Heilongjiang brown frog (Rana amurensis) is widely used in traditional Chinese medicine. In particular, the oviduct and skin have been developed into various health products. However, limited numbers of complete genomes of amphibian species have been reported, excluding the Heilongjiang brown frog. Here, the transcriptomes of 45 samples from the liver, spleen, heart, ovaries, thigh muscles, skin, oviduct, stomach and intestine of five Heilongjiang brown frog were reassembled and analyzed. A total of 1,085,532 unigenes with an average length of 676.6 bp and N50 of 722 bp were obtained. Comparative transcriptomics of different tissues detected tissue-specific expression. There were 3248 differentially expressed genes (DEGs) in the ovary, and the number of unique DEGs between the ovary and spleen was the largest. The results of DEGs enrichment showed there were many pathways and items related to protein synthesis and metabolism in the oviduct. The DEGs of the skin were enriched with many bacterial defense items, indicating that there were a large number of antimicrobial peptides in the skin. Thus, these were suitable as biological sources for the development and extraction of antimicrobial peptides. Through the assembly of transcriptome sequencing data and functional annotation of the Heilongjiang brown frog genome, this study provides reference materials for further exploring and utilizing functional gene resources of frogs and lays a foundation for medical research and the development of new products.

Pretreatment Nutritional Status in Combination with Inflammation Affects Chemotherapy Interruption in Women with Ovarian, Fallopian Tube, and Peritoneal Cancer.

BACKGROUND: Discontinuing chemotherapy worsens cancer prognosis. This study aimed to investigate the relationship between nutritional status at the start of chemotherapy and chemotherapy discontinuation in patients with ovarian, fallopian tube, and primary peritoneal cancer. METHODS: This was a retrospective cohort study. One hundred and forty-six patients to whom weekly paclitaxel and carboplatin were administered as postoperative chemotherapy were included. Six courses in 21-day cycles were defined as complete treatment. As nutritional indicators, body mass index, weight change rate, serum albumin, total lymphocyte count, prognostic nutritional index, and C-reactive protein-to-albumin ratio (CAR) were compared between complete and incomplete treatment groups. Patients were divided into two groups according to CAR. The number of chemotherapy cycles was compared between these two groups. A Cox proportional hazard model was used for covariate adjustment. RESULTS: Several indicators differed between complete and incomplete treatment groups, and among the indicators, CAR had the highest discriminatory ability. The number of chemotherapy cycles was shorter in the high CAR group than in the low CAR group. A high CAR was associated with chemotherapy interruption even after adjusting for covariates. CONCLUSION: Based on CAR, nutritional status before chemotherapy is suggested to be associated with the risk of chemotherapy discontinuation.