Cervical Secretion Methylation Is Associated with the Pregnancy Outcome of Frozen-Thawed Embryo Transfer.

The causes of implantation failure remain a black box in reproductive medicine. The exact mechanism behind the regulation of endometrial receptivity is still unknown. Epigenetic modifications influence gene expression patterns and may alter the receptivity of human endometrium. Cervical secretions contain endometrial genetic material, which can be used as an indicator of the endometrial condition. This study evaluates the association between the cervical secretion gene methylation profile and pregnancy outcome in a frozen-thawed embryonic transfer (FET) cycle. Cervical secretions were collected from women who entered the FET cycle with a blastocyst transfer (36 pregnant and 36 non-pregnant women). The DNA methylation profiles of six candidate genes selected from the literature review were measured by quantitative methylation-specific PCR (qMSP). Bioinformatic analysis of six selected candidate genes showed significant differences in DNA methylation between receptive and pre-receptive endometrium. All candidate genes showed different degrees of correlation with the pregnancy outcomes in the logistic regression model. A machine learning approach showed that the combination of candidate genes' DNA methylation profiles could differentiate pregnant from non-pregnant samples with an accuracy as high as 86.67% and an AUC of 0.81. This study demonstrated the association between cervical secretion methylation profiles and pregnancy outcomes in an FET cycle and provides a basis for potential clinical application as a non-invasive method for implantation prediction.

Epigenomic Analysis Reveals the KCNK9 Potassium Channel as a Potential Therapeutic Target for Adenomyosis.

Adenomyosis is linked to dysmenorrhea and infertility. The pathogenesis of adenomyosis remains unclear, and little is known of the genetic and epigenetic changes in the eutopic endometrium in adenomyosis, which may predispose patients to the invasion and migration of endometrial tissues into the myometrium. Transcriptome studies have identified genes related to various cell behaviors but no targets for therapeutic intervention. The epigenetics of the eutopic endometrium in adenomyosis have rarely been investigated. Endometrial tissue was obtained from premenopausal women with (n = 32) or without adenomyosis (n = 17) who underwent hysterectomy aged 34-57 years at a tertiary hospital. The methylome and transcriptome were assessed by using a Methylation 450 K BeadChip array and Affymetrix expression microarray. Protein expression was examined by immunohistochemistry. Differential methylation analysis revealed 53 lowly methylated genes and 176 highly methylated genes with consistent gene expression in adenomyosis, including three genes encoding potassium ion channels. High expression of KCNK9 in the eutopic and ectopic endometria in patients with adenomyosis but not in normal controls was observed. Hormone-free, antibody-based KCNK9 targeting is a potential therapeutic strategy for adenomyosis-related dysmenorrhea, menorrhagia, and infertility.

Epigenetic loss of heparan sulfate 3-O-sulfation sensitizes ovarian carcinoma to oncogenic signals and predicts prognosis.

Precision medicine requires markers for therapeutic guidance. The purpose of this study was to determine whether epithelial ovarian cancer (EOC) epigenetics can lead to the identification of biomarkers for precision medicine. Through integrative methylomics, we discovered and validated the epigenetic signature of NEFH and HS3ST2 as an independent prognostic factor for type II EOC in our dataset (n = 84), and two independent methylomics datasets (total n = 467). Integrated transcriptomics dataset (n = 1147) and tissue microarrays (n = 54) of HS3ST2 also related to high-methylation statuses and the EOC prognosis. Mechanistic explorations of HS3ST2 have assessed responses to oncogenic stimulations such as IL-6, EGF, and FGF2 in cancer cells. The combination of HS3ST2 and various oncogenic ligands also confers the worse outcome. 3-O-sulfation of heparan sulfate by HS3ST2 makes ovarian cancer cells intrinsically sensitive to oncogenic signals, which sheds new light on the application of HS3ST2 as a companion diagnostic for targeted therapy using kinase inhibitors or therapeutic antibodies.

Transverse modes of a laser using volume Bragg grating as the cavity mirror.

A simulation and experiment were performed to demonstrate that a laser using volume Bragg grating as one of the cavity mirrors can achieve lasing even if the laser cavity length exceeds the traditional stable cavity condition. The laser transverse mode changes from a Gaussian beam into a ring-shaped mode as the laser cavity length increases from stable to unstable cavity conditions. At the same time, the effective modal reflectivity is reduced as the cavity length increases.

[A case study on duty of care in professional nursing].

BACKGROUND: Nurses are expected to discharge their duty of care effectively and professionally to prevent medical negligence. Only three articles have previously focused on medical negligence. Duty of care and medical negligence in nursing are topics that have been neglected in Taiwan. PURPOSE: (1) Classify the duty of care of professional nurses; (2) Investigate the facts and disputes in the current case; (3) Clarify the legal issues involved with regard to duty-of-care violations in the current case; (4) Explore the causal relationships in a legal context between nurses' duty-of-care violations and patient harm / injury. METHODS: Literature analysis and a case study are used to analyze Supreme Court Verdict No.5550 (2010). RESULTS: Duty of care for nursing professionals may be classified into seven broad categories. Each category has its distinct correlatives. In nursing practice, every nursing behavior has a corresponding duty. In this case, the case study nurse did not discharge her obstetric professional duty and failed to inform the doctor in a timely manner. Negligence resulted in prenatal death and the case study nurse was found guilty. CONCLUSIONS: In order to prevent committing a crime, nurses should gain a better understanding of their duty of care and adequately discharge these duties in daily practice.

Accuracy of digital impressions for three-unit and four-unit implant supported fixed dental prostheses using a novel device.

Background/ purpose: Accuracy of digital implant impressions was considered questionable due to the lack of anatomical reference points between implants and the similarities in scan body morphology, which lead to the purpose of this research is to propose a simple and convenient technique to improve the accuracy of scanning. Materials and methods: Four implant analogues (teeth: 15, 17, 24, and 27) were inserted into a stone model of a partially edentulous maxilla; two implants were inserted on each side, creating a three-unit span and a four-unit span. The model was scanned using a 3Shape E4 dental laboratory scanner for reference and a TRIOS 3 intraoral scanner for testing. Each side was scanned 10 times, both with and without the novel device attached to the scan bodies. The trueness and precision of interimplant distances (linear deviations), and interimplant angulations (angle deviations) between the scan bodies were determined using software. A Mann-Whitney U test was used to determine statistical differences between subgroups. Results: Significant differences were discovered in the trueness of angular deviations (-0.20° ± 0.15° vs. -0.01° ± 0.11º) and precision of linear deviations (11.14 ± 6.35 vs. 3.10 ± 2.14 µm) for the four-unit groups. Conclusion: The novel device significantly improved scanning accuracy for a four-unit groups (approximately 22.93 mm) compared to three-unit groups.

Intraperitoneal splenosis mimics peritoneal carcinomatosis of leiomyosarcoma and ovarian cancer.

OBJECTIVE: Leiomyosarcoma and ovarian cancer are often diagnosed late due to the absence of initial symptoms. Patients seek help when abdominal distension occurs; this is associated with pelvic tumor and carcinomatosis. Initial imaging often reveals pelvic tumors with diffuse abdominal nodules; however, this imaging could be misleading, such as in the cases of splenosis. CASE REPORT: A female presented with vaginal bleeding at our outpatient department. Serum CA125 level was elevated. Abdominal and pelvic CT showed multiple uterine masses and left adnexal cysts with peritoneal nodules. Leiomyosarcoma or ovarian cancer with carcinomatosis was suspected. Exploratory laparotomy was performed. Multiple purple spots spreading over peritoneal cavity were noted during the surgery. Pathological examination revealed adenomyosis with multiple uterine myomas and left ovarian endometrioma. Splenic tissues peritoneal implants were observed. CONCLUSION: In patients with a history of spleen rupture or splenectomy, splenosis should be considered in the differential diagnosis, especially in young patients.

Transition from multiport to single-site surgery: A single institution experience in robotic supracervical hysterectomy for benign gynecological diseases.

OBJECTIVE: To share our experience of transition from multiport to single-site robotic surgery for benign gynecological conditions as well as to assess the selection criteria of candidates for robotic single-site supracervical hysterectomy (RSSH). MATERIALS AND METHODS: A retrospective review was conducted on patients undergoing robotic supracervical hysterectomy by a single surgeon in a single institute between June 2014 and December 2017. Patients who underwent additional procedures along with supracervical hysterectomy and who had unexpectant corpus malignancy proved pathologically were excluded from comparisons between patients undergoing RSSH and robotic multiport supracervical hysterectomy (RMSH). RESULTS: Between June 2014 and December 2017, we accomplished 26 RSSH and 57 RMSH. There were no conversions, intraoperative complications, and readmissions within 30 days after surgery. In the RSSH group, the mean uterine weight was 264.6 ± 140.9 g with mean docking time of 15.8 ± 5.5 min, mean console time of 61.1 ± 35.6 min and mean operative time of 140.3 ± 34.4 min. In comparison to the RMSH group, the percentage of overweight/obese patients was lower (p = 0.018) and the uterine size was smaller (p < 0.001) with adenomyosis diagnosed more frequently (p = 0.002) in the RSSH group. While the operative time in the RSSH group was significantly shorter (p = 0.002), the RSSH group took longer time in docking (p < 0.001) and comparable time in console (p = 0.254). In view of chronological change, docking time and console time in the RMSH group remained steady, whereas steep decreases were observed in the RSSH group. The intraoperative blood loss and hemoglobin drop were comparable. The length of hospital stay was significantly shorter in the RSSH group (p = 0.005). CONCLUSION: Transition from multiport to single-site surgery can be smooth for a surgical team experienced in the conventional multiport robotic system. RSSH is safe and feasible in properly selected patients.

Combined genetic mutations and DNA-methylated genes as biomarkers for endometrial cancer detection from cervical scrapings.

BACKGROUND: Endometrial cancer is a common gynecologic cancer. Noninvasive molecular biomarkers for triage of high-risk patients for invasive procedures are needed. Based on the success of cytological Pap smear screening, cervical scrapings are a good source of DNA for molecular testing. In addition to genetic lesions, DNA methylation is a promising biomarker. We assessed the usefulness of combining genetic and epigenetic biomarkers from cervical scrapings to detect endometrial carcinomas. METHODS: We performed a retrospective case-control study of 96 consecutive cervical scrapings from patients with abnormal uterine bleeding who underwent surgery for diagnostic evaluation. Thirty and 16 cases were diagnosed with type I and type II endometrial cancers, respectively. The remaining non-cancer cases included normal endometrium (n = 12), benign uterine lesions (n = 20), and endometrial hyperplasia (n = 18). Quantitative methylation-specific PCR and mass spectrometry were used for DNA methylation and genetic mutation analysis. Logistic regression was used to evaluate the clinical performance of these candidate biomarkers. RESULTS: We tested the effectiveness of the methylation status of four genes (BHLHE22, CDO1, TBX5, and HAND2) in endometrial cancer detection. The area under the receiver operating characteristic curves ranged from 0.703 to 0.878, and panels of hypermethylated BHLHE22/CDO1/HAND2 (87.0% sensitivity and 86.0% specificity) and BHLHE22/CDO1/TBX5 (89.1% sensitivity and 80.0% specificity) showed significant differences and could distinguish benign from malignant endometrial lesions. The sensitivity and specificity in endometrial cancer detection for BHLHE22/CDO1 were 84.8% and 88.0%, respectively. Both type I and II endometrial carcinomas could be detected using a BHLHE22/CDO1-based methylation profile, suggesting that they may have common epigenomes. Moreover, PTEN and TP53 mutations were found in 63.3% of type I and 93.6% of type II endometrial cancers. Unexpectedly, PTEN and TP53 mutations were commonly found in cervical scrapings of the normal endometrium (25% and 33.3%, respectively) and in cases with benign uterine lesions (10% and 50%, respectively). Finally, combinations of any one mutation of PTEN and TP53 mutations had a sensitivity of 91.3%, but a specificity of only 42.0%. CONCLUSIONS: Adding PTEN/TP53 mutation testing to BHLHE22/CDO1-based methylation testing did not improve the detection of endometrial cancer.

TET1 promotes 5hmC-dependent stemness, and inhibits a 5hmC-independent epithelial-mesenchymal transition, in cervical precancerous lesions.

DNA hypermethylation is a driving force in carcinogenesis. However, the role of active DNA hypomethylation in cancer remains largely unknown. This process, facilitated by ten-eleven translocation methylcytosine dioxygenase 1 (TET1), which oxidizes 5-methylcytosine (5 mC) to 5-hydroxymethylcytosine (5hmC), has never been studied in cervical cancer. Here, we found that TET1 and 5hmC correlative increases from normal cervix to Low-grade squamous intraepithelial lesion (LSIL), maximizing in High-grade squamous intraepithelial lesion (HSIL), and decreasing in invasive cancer. Full-length HPV-immortalized HSIL cells demonstrated higher TET1/5hmC levels, and stemness properties, compared to invasive cancer cells. TET1 silencing promoted the epithelial-mesenchymal transition (EMT), to transform precancerous cells in vivo. TET1 increased 5hmC in the ZEB1 and VIM promoters, surprisingly, silencing both genes. TET1 interaction with the histone modifiers, LSD1 and EZH2, on the ZEB1 promoter, resulted in gene silencing, via loss of histone H3K4 trimethylation, and gain of histone H3K27 trimethylation. Taken together, TET1 promotes stemness properties, and inhibits EMT, in HSIL cells, through 5hmC-dependent and -independent mechanisms.

Paired Box-1 (PAX1) Activates Multiple Phosphatases and Inhibits Kinase Cascades in Cervical Cancer.

DNA methylation alteration, such as global hypomethylation and localized hypermethylation, within the promoters of tumor suppressor genes, is an important risk factor in cervical cancer. The potential use of DNA methylation detection, in cervical cancer screening or triage of mildly abnormal cytology, has recently been demonstrated. In particular, PAX1 DNA methylation testing was approved as an adjunct to cytology, in Taiwan, and is now undergoing registration trials in China. However, the function of PAX1 in cancer biology remains largely unknown. Here, we show that PAX1 inhibits malignant phenotypes upon oncogenic stress. Specifically, PAX1 expression inhibited the phosphorylation of multiple kinases, after challenges with oncogenic growth factors such as EGF and IL-6. Analogously, PAX1 activated a panel of phosphatases, including DUSP1, 5, and 6, and inhibited EGF/MAPK signaling. PAX1 also interacted with SET1B, increasing histone H3K4 methylation and DNA demethylation of numerous phosphatase-encoding genes. Furthermore, hypermethylated PAX1 associated with poor prognosis in cervical cancer. Taken together, this study reveals, for the first time, the functional relevance of PAX1 in cancer biology, and further supports the prospect of targeting multifold oncogenic kinase cascades, which jointly contribute to multiresistance, via epigenetic reactivation of PAX1.

Genotype-specific methylation of HPV in cervical intraepithelial neoplasia.

OBJECTIVE: Hypermethylation of human papillomavirus (HPV) and host genes has been reported in cervical cancer. However, the degree of methylation of different HPV types relative to the severity of the cervical lesions remains controversial. Studies of the degree of methylation associated with the host gene and the HPV genome to the severity of cervical lesions are rare. We examined the association of methylation status between host genes and late gene 1 (L1) regions of HPV16, 18, 52, and 58 in cervical brushings. METHODS: Cervical brushings from 147 HPV-infected patients were obtained. The samples comprised normal (n=28), cervical intraepithelial neoplasia (CIN) 1 (n=45), CIN2 (n=13), and CIN3/carcinoma in situ (n=61). The methylation status of HPV and host genes was measured using bisulfite pyrosequencing and quantitative methylation-specific polymerase chain reaction (PCR). RESULTS: The degree of methylation of L1 in HPV16, 18, and 52 was associated with the severity of the cervical lesion. In HPV52, C-phosphate-G (CpG) sites 6368m, 6405m, and 6443m showed significantly higher methylation in lesions ≥CIN3 (p=0.005, 0.003, and 0.026, respectively). Methylation of most HPV types except HPV52 (r<-0.1) was positively correlated with the degree of methylation of host genes including PAX1 and SOX1 (0.4≤r≤0.7). Combining HPV methylation with PAX1 methylation improved the clustering for ≥CIN2. CONCLUSION: Our study showed that the degree of L1 methylation of HPV16, 18, and 52 but not 58 is associated with the severity of cervical lesions. The association between HPV methylation and host gene methylation suggests different responses of host cellular epigenetic machinery to different HPV genotypes.

Outcomes of anti-Müllerian hormone-tailored ovarian stimulation protocols in in vitro fertilization/intracytoplasmic sperm injection cycles in women of advanced age.

OBJECTIVE: We aimed to compare the outcomes of in vitro fertilization/intracytoplasmic sperm injection (IVF/ICSI) treatments in women of advanced age (>40 years) using anti-Müllerian hormone (AMH)-tailored ovarian stimulation protocols versus conventional protocols based on antral follicle count (AFC). MATERIALS AND METHODS: We retrospectively reviewed 210 women who underwent IVF/ICSI cycles: 116 women underwent stimulation protocols that were tailored to their AMH levels, whereas 94 women received treatment using conventional stimulation protocols based on AFC as the ovarian reserve marker. RESULTS: The following parameters were significantly higher in the AMH-tailored group than in the conventional group: initial and total doses (IU) of recombinant follicle-stimulating hormone (rFSH) used for stimulation (514.2 ± 137.9 vs. 452.3 ± 135.3, p = 0.001; 4713.8 ± 1618.8 vs. 4047.2 ± 1366.0, p = 0.007, respectively), ovum pick-up rate (OPU; 88.8% vs. 75.5%, p = 0.016), serum estradiol (E2) level on the day of human chorionic gonadotropin (hCG) administration (1818.5 ± 1422.4 vs. 1394.0 ± 929.0 pg/mL, p = 0.028), number of oocytes retrieved (7.4 ± 5.1 vs. 5.5 ± 3.4, p = 0.007), number of embryos per case (4.2 ± 3.2 vs. 3.3 ± 2.5, p = 0.048), clinical pregnancy rates (22.4% vs. 8.5%, p = 0.008), implantation rates (13.1% vs. 3.9%, p = 0.001), and live birth rates per cycle (15.5% vs. 6.4%, p = 0.049). CONCLUSION: Individualized controlled ovarian stimulation (COS) protocols tailored to patients' AMH levels may improve the pregnancy rate, implantation rate, and live birth rate in women of advanced age undergoing IVF/ICSI compared with those receiving conventional stimulation protocols.