Ovarian tumor diagnosis using deep convolutional neural networks and a denoising convolutional autoencoder.

Discrimination of ovarian tumors is necessary for proper treatment. In this study, we developed a convolutional neural network model with a convolutional autoencoder (CNN-CAE) to classify ovarian tumors. A total of 1613 ultrasound images of ovaries with known pathological diagnoses were pre-processed and augmented for deep learning analysis. We designed a CNN-CAE model that removes the unnecessary information (e.g., calipers and annotations) from ultrasound images and classifies ovaries into five classes. We used fivefold cross-validation to evaluate the performance of the CNN-CAE model in terms of accuracy, sensitivity, specificity, and the area under the receiver operating characteristic curve (AUC). Gradient-weighted class activation mapping (Grad-CAM) was applied to visualize and verify the CNN-CAE model results qualitatively. In classifying normal versus ovarian tumors, the CNN-CAE model showed 97.2% accuracy, 97.2% sensitivity, and 0.9936 AUC with DenseNet121 CNN architecture. In distinguishing malignant ovarian tumors, the CNN-CAE model showed 90.12% accuracy, 86.67% sensitivity, and 0.9406 AUC with DenseNet161 CNN architecture. Grad-CAM showed that the CNN-CAE model recognizes valid texture and morphology features from the ultrasound images and classifies ovarian tumors from these features. CNN-CAE is a feasible diagnostic tool that is capable of robustly classifying ovarian tumors by eliminating marks on ultrasound images. CNN-CAE demonstrates an important application value in clinical conditions.

Intra and extra pelvic multidisciplinary surgical approach of retroperitoneal sarcoma: Case series report.

BACKGROUND: Retroperitoneal sarcoma (RPS) is a rare malignancy arising from mesenchymal cells that most commonly presents as an abdominal mass and is associated with poor prognosis. Although several studies have assessed the survival benefits of wide excision, few have reported detailed methods for achieving wide excision in patients with RPS. AIM: To describe our experience with multidisciplinary surgical resection of RPS using intra- and extra-pelvic approaches. METHODS: Multidisciplinary surgery is an anatomical approach that combines intra- and extra-peritoneal access within the same surgery to achieve complete RPS removal. This retrospective review of the records of patients who underwent multidisciplinary surgery for RPS analyzed surgical and survival outcomes. RESULTS: Eight patients underwent 10 intra- and extra-pelvic surgical resections, and their median mass size was 12.75 cm (range, 6-45.5 cm). Using an intrapelvic approach, laparoscopy-assisted surgery was performed in four cases and laparotomy surgery in six. Using an extrapelvic approach, ilioinguinal and posterior approaches were used in four cases each, and the prone position and midline skin incision were shared in one. All patients' RPS masses were removed completely, and four achieved R0 resection through intra- and extra-pelvic surgery. The median estimated blood loss was 2000 mL (range, 300-20000 mL) and the median hospitalization was 12.6 d (range, 9-69 d). Reoperation was needed in two patients (one for wound necrosis and the other for bowel perforation and wound necrosis). The median overall survival rate and median progression-free survival were 64.6 and 13.7 mo, respectively. CONCLUSION: RPS is therapeutically challenging because of its location and high risk of recurrence. Therefore, intra- and extra-pelvic surgical approaches can improve the macroscopic security of the surgical margin.

Plasma Protein Biomarkers Associated with Higher Ovarian Cancer Risk in BRCA1/2 Carriers.

Ovarian cancer (OC) is the most lethal gynecologic malignancy and in-time diagnosis is limited because of the absence of effective biomarkers. Germline BRCA1/2 genetic alterations are risk factors for hereditary OC; risk-reducing salpingo-oophorectomy (RRSO) is pursued for disease prevention. However, not all healthy carriers develop the disease. Therefore, identifying predictive markers in the BRCA1/2 carrier population could help improve the identification of candidates for preventive RRSO. In this study, plasma samples from 20 OC patients (10 patients with BRCA1/2 wild type (wt) and 10 with the BRCA1/2 variant (var)) and 20 normal subjects (10 subjects with BRCA1/2wt and 10 with BRCA1/2var) were analyzed for potential biomarkers of hereditary OC. We applied a bottom-up proteomics approach, using nano-flow LC-MS to analyze depleted plasma proteome quantitatively, and potential plasma protein markers specific to the BRCA1/2 variant were identified from a comparative statistical analysis of the four groups. We obtained 1505 protein candidates from the 40 subjects, and SPARC and THBS1 were verified by enzyme-linked immunosorbent assay. Plasma SPARC and THBS1 concentrations in healthy BRCA1/2 carriers were found to be lower than in OC patients with BRCA1/2var. If plasma SPARC concentrations increase over 337.35 ng/mL or plasma THBS1 concentrations increase over 65.28 µg/mL in a healthy BRCA1/2 carrier, oophorectomy may be suggested.

Plasma-derived exosomal miR-4732-5p is a promising noninvasive diagnostic biomarker for epithelial ovarian cancer.

BACKGROUND: Exosomal miRNAs regulate gene expression and play important roles in several diseases. We used exosomal miRNA profiling to investigate diagnostic biomarkers of epithelial ovarian cancer (EOC). METHODS: In total, 55 individuals were enrolled, comprising healthy (n = 21) and EOC subjects (n = 34). Small mRNA (smRNA) sequencing and real-time PCR (RT-PCR) were performed to identify potential biomarkers. Receiver operating characteristic (ROC) curves were conducted to determine biomarker sensitivity and specificity. RESULTS: Using smRNA sequencing, we identified seven up-regulated (miR-4732-5p, miR-877-5p, miR-574-3p, let-7a-5p, let-7b-5p, let-7c-5p, and let-7f-5p) and two down-regulated miRNAs (miR-1273f and miR-342-3p) in EOC patients when compared with healthy subjects. Of these, miR-4732-5p and miR-1273f were the most up-regulated and down-regulated respectively, therefore they were selected for RT-PCR analysis. Plasma derived exosomal miR-4732-5p had an area under the ROC curve of 0.889, with 85.7% sensitivity and 82.4% specificity in distinguishing EOC patients from healthy subjects (p<0.0001) and could be a potential biomarker for monitoring the EOC progression from early stage to late stage (p = 0.018). CONCLUSIONS: Plasma derived exosomal miR-4732-5p may be a promising candidate biomarker for diagnosing EOC.

Peripheral blood BRCA1 methylation profiling to predict familial ovarian cancer.

OBJECTIVE: Familial cancer appears at a young age and its incidence is increasing. About 12% of familial ovarian cancer cases are associated with BRCA1/2 mutations (BRCAm). In this study, we investigated BRCA1 methylation may predict ovarian cancer in those with a family history of cancer (FHC) but without BRCA1/2 mutations (BRCAwt). METHODS: Using peripheral blood DNA from 55 subjects without a history of cancer [cancer(-)] and 52 ovarian cancer patients, we examined BRCA1 promoter methylation through bisulfite sequencing of the promoter and expressed the results as the cumulative methylation index. Then, we evaluated the BRCA1 promoter methylation according to BRCA1/2 germline mutations. RESULTS: BRCA1 methylation was more prevalent in the BRCAm cancer(-) group than in the BRCAwt cancer(-) group and ovarian cancer patients (p=0.031 and p=0.019, respectively). In the BRCAwt cancer(-) group, BRCA1 methylation was more prevalent in those with an FHC than in those without one and in the BRCAm cancer(-) group with an FHC (p=0.001 and p<0.001, respectively). CONCLUSION: Our data suggest a predictive role of BRCA1 methylation profile for ovarian cancer in those without a history of cancer but with an FHC. BRCA1 methylation has important implications for diagnostic and predictive testing of those with BRCAwt cancer(-) status with FHC.

A Simple Gasless Direct Suturing Technique to Achieve Ovarian Hemostasis During Transvaginal Natural Orifice Transluminal Endoscopic Surgery Ovarian Cystectomy.

Background: Transvaginal natural orifice transluminal endoscopic surgery (vNOTES) was introduced in 2012, but the technique is not yet widely used for ovarian cystectomy. We aim to introduce a new gasless ovarian hemostatic suturing technique for ovarian cystectomy using vNOTES. Methods: We conducted a prospective study using a novel technique for vNOTES ovarian cystectomy. Our vNOTES port (a wound retractor and a disposable glove) was inserted transvaginally through a posterior colpotomy. After ovarian cystectomy, removal of the glove created a gasless state. Hemostatic suturing of the ovary was performed through a vaginal speculum inserted through the wound retractor, under direct observation. Results: Twenty ovarian cystectomies were performed through vNOTES at our institution between June 2019 and February 2020. The mean patient age was 34.2 years (range, 24-51 years). Four patients (20%) underwent bilateral cystectomy and 16 patients (80%) underwent unilateral cystectomy. The mean operative time was 58.7 minutes (bilateral, 57.5 minutes; unilateral, 58.9 minutes), and the mean ovarian hemostatic suturing time was 4.3 minutes (bilateral, 5 minutes; unilateral, 4.1 minutes). Ten patients (50%) received additional medication for pain control within 30 minutes of surgery. All patients were discharged within 24 hours, and 11 were discharged within 12 hours. Conclusion: The gasless hemostatic suturing technique for vNOTES, using a speculum to observe the suturing process, is easy to perform and allows for rapid ovarian hemostasis.

Incidence and clinical course of septic shock in neutropenic patients during chemotherapy for gynecological cancers.

OBJECTIVE: To identify the incidence and clinical course of septic shock combined with neutropenia during chemotherapy in gynecological cancer patients. METHODS: We retrospectively reviewed the medical records of all gynecological cancer patients who received intravenous chemotherapy between March 2009 and March 2018. Patients diagnosed with neutropenic septic shock (NSS) during the course of chemotherapy were identified. We calculated the overall incidence and mortality rate of NSS, and analyzed risk factors and clinical course. RESULTS: A total of 1,009 patients received 10,239 cycles of chemotherapy during the study period. Among these, 30 (3.0%) patients had 32 NSS events, of which 12 (1.2%) died. With respect to patient age during the first course of chemotherapy, the incidence of NSS after the age of 50 was significantly higher than that in patients under 50 (3.9% vs. 1.4%, p=0.034). As the number of chemotherapy courses increased, the incidence of NSS increased, and linear-by-linear association analysis showed a positive correlation (p=0.004). NSS events occurred on average 7.8 days after the last cycle of chemotherapy, and the median duration of vasopressor administration was 23.3 hours. The median age (64.0 vs. 56.5, p=0.017) and peak heart rate (149.5 min-1 vs. 123.5 min-1, p=0.015) were significantly higher in the group of patients who subsequently died of NSS than in those who survived. CONCLUSION: The overall incidence of NSS in gynecological cancer patients receiving chemotherapy was 3.0%, which is higher than previously estimated. Peak heart rate during NSS events may be an indicator for predicting survival.

Clinical factors that affect diagnostic discrepancy between colposcopically directed biopsies and loop electrosurgical excision procedure conization of the uterine cervix.

OBJECTIVE: This study aimed to determine the factors affecting pathologic discrepancy and final diagnosis between colposcopic biopsy and pathology by loop electrosurgical excision procedure (LEEP). METHODS: Between 2004 and 2016, 1,200 patients who underwent LEEP were enrolled for this study. 667 underwent cervical cytology, human papillomavirus (HPV) test, colposcopic biopsy, and LEEP. We analyzed patient's age, menopausal status, number of delivery, abortion times, cervical cytology, number of punch biopsies, HPV type, LEEP, and interval between colposcopic biopsy and LEEP. RESULTS: Logistic regression analysis of the final diagnosis showed that age 30-39 years and other high HPV group types were associated with cancer diagnosis, whereas atypical squamous cells cannot exclude high-grade squamous intraepithelial lesion (ASC-H), high-grade squamous intraepithelial lesion (HSIL), and HPV type 16 affected the diagnosis of cervical intraepithelial neoplasia (CIN) 2. The overall concordance rate of histopathology between punch biopsy and LEEP was 43.3%. The rates of detecting a more severe lesion by LEEP than those by biopsy were 23.1%. The rates of a less severe lesion detected by LEEP than those by biopsy were 33.6%. Factors related with biopsy underestimation were as follows: <1 vaginal delivery, HSIL, number of punch biopsies and HPV type. Punch biopsy number is a unique factor of biopsy overestimation. CONCLUSION: Patients with ASC-H, HSIL, and HPV type 16 may undergo conization immediately without colposcopic biopsy. We suggest that colposcopically directed 3 to 5 punch biopsies may be used to determine the need for conization.

Amelioration of Cognitive Dysfunction in APP/PS1 Double Transgenic Mice by Long-Term Treatment of 4-O-Methylhonokiol.

Alzheimer's disease (AD) is the most common neurodegenerative disease without known ways to cure. A key neuropathologic manifestation of the disease is extracellular deposition of beta-amyloid peptide (Aß). Specific mechanisms underlying the development of the disease have not yet been fully understood. In this study, we investigated effects of 4-O-methylhonokiol on memory dysfunction in APP/PS1 double transgenic mice. 4-O-methylhonokiol (1 mg/kg for 3 month) significantly reduced deficit in learning and memory of the transgenic mice, as determined by the Morris water maze test and step-through passive avoidance test. Our biochemical analysis suggested that 4-O-methylhonokiol ameliorated Aß accumulation in the cortex and hippocampus via reduction in beta-site APP-cleaving enzyme 1 expression. In addition, 4-O-methylhonokiol attenuated lipid peroxidation and elevated glutathione peroxidase activity in the double transgenic mice brains. Thus, suppressive effects of 4-O-methylhonokiol on Aß generation and oxidative stress in the brains of transgenic mice may be responsible for the enhancement in cognitive function. These results suggest that the natural compound has potential to intervene memory deficit and progressive neurodegeneration in AD patients.

Decreased pain responses of C-C chemokine receptor 5 knockout mice to chemical or inflammatory stimuli.

Chemokines are small chemotactic cytokines that elicit many physiological and pathological effects through binding to their corresponding receptors. Recent studies have suggested that C-C chemokine receptor (CCR) 5 interacts with µ-opioid receptor and modifies a nociceptive reaction. We examined effects of CCR5 deficiency on pain responses by employing CCR5 knockout (KO) mice. We found that pain responses of CCR5 KO mice to chemical or inflammation stimuli were milder than those of CCR5 wild type (WT) mice. However, there was no remarkable change in thermal nociception. To prove the involvement of CCR5 deletion in lowered nociception, we examined pain reactions with CCR5 WT mice following treatment of a CCR5 antagonist (D-Ala(1)-peptide T-NH(2,) DAPTA). Chemical or inflammatory pain behavior was significantly relieved by intracerebroventricular infusion of the inhibitor. When we assessed expression level of µ-opioid receptor (MOR) in the periaqueductal gray where the receptors are critical for analgesic effects, immunoreactivity of MOR was significantly higher in CCR5 KO mice than WT mice without change in phosphorylation level of the receptor. Reduced nociceptive responses in CCR5 KO mice were moderated by administration of naloxone and d-Phe-Cys-Tyr-d-Trp-Arg-Thr-Pen-Thr-NH2 (CTAP), MOR antagonists. Our data indicate that CCR5 deficiency is related to up-regulation of MOR without an increase in the receptor desensitization which might result in increased analgesic effects against chemical or inflammatory stimuli. Alternatively, higher amount of opioid ligands in CCR5 mice might be linked to these results. Therefore, CCR5 appears to be a therapeutic target for treatment of pain related diseases such as inflammatory hyperalgesia.

Redox regulation of lipopolysaccharide-mediated sulfiredoxin induction, which depends on both AP-1 and Nrf2.

Sulfiredoxin (Srx) is an enzyme that catalyzes the reduction of cysteine sulfinic acid of hyperoxidized peroxiredoxins and exerts a protective antioxidant role. Here we investigated the regulatory mechanism of Srx induction by lipopolysaccharide (LPS) in mouse macrophages. LPS up-regulated Srx expression on the transcriptional level. The promoter region of the Srx gene contained putative NF-κB and AP-1 (activator protein-1) sites, and the proximal site of three AP-1 sites was embedded within the antioxidant response element (ARE), a cis-acting element for Nrf2 (nuclear factor erythroid 2-related factor). Mutational analysis of the Srx promoter revealed that Srx induction is dependent on AP-1 sites and ARE but not on NF-κB sites. Consistently, both transcription factors, AP-1 and Nrf2, were required for LPS-mediated Srx induction, as revealed by chromatin immunoprecipitation using antibodies specific for c-Jun and c-Fos and little Srx induction in Nrf2-null bone marrow-derived macrophages. Among mitogen-activated protein kinases that mediate the signal transduction by LPS, JNK played a major role in Srx induction. Moreover, chemical antioxidants, such as N-acetylcysteine and butylated hydroxyanisole, and the NADPH oxidase inhibitor diphenyleneiodonium inhibited Srx induction as well as generation of reactive oxygen species, both of which were also suppressed in Nox2 (NADPH oxidase 2)-deficient bone marrow-derived macrophages. These results suggest that LPS-mediated Srx induction is dependent on both AP-1 and Nrf2, which is regulated by Nox2-derived reactive oxygen species.

Thioredoxin-related protein 14, a new member of the thioredoxin family with disulfide reductase activity: implication in the redox regulation of TNF-alpha signaling.

Thioredoxin-related protein 14 (TRP14) is a novel 14-kDa disulfide reductase with two active site Cys residues in its WCPDC motif, which is comparable to the WCGPC motif of thioredoxin (Trx). Although the active site cysteine of TRP14 is sufficiently nucleophilic, its redox potential is similar to that of Trx1, and it receives the electrons from Trx reductase 1 (TrxR1) as does Trx1. TRP14 does not target the same substrate as Trx1, suggesting that TRP14 and Trx1 might act on distinct substrate proteins. Comparison of the crystal structures of TRP14 and Trx1 reveals distinct surface structures in the vicinity of their active sites. Both TRP14 and Trx1 inhibit the pathways of nuclear factor-kappaB (NF-kappaB), mitogen-activated protein kinases, and apoptosis in cells stimulated with tumor necrosis factor-alpha (TNF-alpha), but they appear to do so by acting on target proteins, some of which do not overlap. TRP14 inhibits the TNF-alpha-induced NF-kappaB activation to a greater extent than Trx1. The dynein light chain LC8 was identified as a new target of disulfide reductase activity of TRP14, and LC8 was shown to bind IkappaBalpha in a redox-dependent manner, thereby preventing its phosphorylation by IkappaB kinase. These findings elucidate the molecular mechanism by which NF-kappaB activation is regulated through TRP14.

Dynein light chain LC8 negatively regulates NF-kappaB through the redox-dependent interaction with IkappaBalpha.

Redox regulation of nuclear factor kappaB (NF-kappaB) has been described, but the molecular mechanism underlying such regulation has remained unclear. We recently showed that a novel disulfide reductase, TRP14, inhibits tumor necrosis factor alpha (TNFalpha)-induced NF-kappaB activation, and we identified the dynein light chain LC8, which interacts with the NF-kappaB inhibitor IkappaBalpha, as a potential substrate of TRP14. We now show the molecular mechanism by which NF-kappaB activation is redox-dependently regulated through LC8. LC8 inhibited TNFalpha-induced NF-kappaB activation in HeLa cells by interacting with IkappaBalpha and thereby preventing its phosphorylation by IkappaB kinase (IKK), without affecting the activity of IKK itself. TNFalpha induced the production of reactive oxygen species, which oxidized LC8 to a homodimer linked by the reversible formation of a disulfide bond between the Cys(2) residues of each subunit and thereby resulted in its dissociation from IkappaBalpha. Butylated hydroxyanisol, an antioxidant, and diphenyleneiodonium, an inhibitor of NADPH oxidase, attenuated the phosphorylation and degradation of IkappaBalpha by TNFalpha stimulation. In addition LC8 inhibited NF-kappaB activation by other stimuli including interleukin-1beta and lipopolysaccharide, both of which generated reactive oxygen species. Furthermore, TRP14 catalyzed reduction of oxidized LC8. Together, our results indicate that LC8 binds IkappaBalpha in a redox-dependent manner and thereby prevents its phosphorylation by IKK. TRP14 contributes to this inhibitory activity by maintaining LC8 in a reduced state.

Peroxisome proliferator activated receptor gamma agonists suppress TNFalpha-induced ICAM-1 expression by endothelial cells in a manner potentially dependent on inhibition of reactive oxygen species.

In this study, we investigated the anti-inflammatory effect of various peroxisome proliferator activated receptor gamma (PPARgamma) agonists (15-deoxy-Delta12,14-prostaglandin J(2), troglitazone, rosiglitazone, ciglitazone) on human aortic endothelial cells. Pretreatment with PPARgamma agonists abrogated tumor necrosis factor alpha (TNFalpha)-induced expression of intercellular adhesion molecule-1 (ICAM-1) and subsequent monocytic adhesion by endothelial cells. Because reactive oxygen species (ROS) have been reported to play important roles in pro-inflammatory signal transduction, the involvement of ROS was investigated as a potential mechanism of anti-inflammatory effect of PPARgamma ligands. Consistent with previous reports in other cell types, blockade of TNFalpha-induced ROS by treatment with N-acetylcysteine, diphenylene iodonium or NADPH oxidase 4 (NOX4) siRNA suppressed TNFalpha-induced ICAM-1 expression and subsequent monocytic adhesion, indicating that TNFalpha mediates pro-inflammatory signals via NOX4-dependent ROS generation in human endothelial cells. Finally, pretreatment with PPARgamma agonists significantly suppressed TNFalpha-induced increases of intracellular ROS. Our results collectively suggest that PPARgamma agonists might exert an anti-inflammatory effect on endothelial cells in a ROS-dependent manner.