The Roles and Regulatory Mechanisms of Tight Junction Protein Cingulin and Transcription Factor Forkhead Box Protein O1 in Human Lung Adenocarcinoma A549 Cells and Normal Lung Epithelial Cells.

Tight junction (TJ) protein cingulin (CGN) and transcription factor forkhead box protein O1 (FOXO1) contribute to the development of various cancers. Histone deacetylase (HDAC) inhibitors have a potential therapeutic role for some cancers. HDAC inhibitors affect the expression of both CGN and FOXO1. However, the roles and regulatory mechanisms of CGN and FOXO1 are unknown in non-small cell lung cancer (NSCLC) and normal human lung epithelial (HLE) cells. In the present study, to investigate the effects of CGN and FOXO1 on the malignancy of NSCLC, we used A549 cells as human lung adenocarcinoma and primary human lung epithelial (HLE) cells as normal lung tissues and performed the knockdown of CGN and FOXO1 by siRNAs. Furthermore, to investigate the detailed mechanisms in the antitumor effects of HDAC inhibitors for NSCLC via CGN and FOXO1, A549 cells and HLE cells were treated with the HDAC inhibitors trichostatin A (TSA) and Quisinostat (JNJ-2648158). In A549 cells, the knockdown of CGN increased bicellular TJ protein claudin-2 (CLDN-2) via mitogen-activated protein kinase/adenosine monophosphate-activated protein kinase (MAPK/AMPK) pathways and induced cell migration, while the knockdown of FOXO1 increased claudin-4 (CLDN-4), decreased CGN, and induced cell proliferation. The knockdown of CGN and FOXO1 induced cell metabolism in A549 cells. TSA and Quisinostat increased CGN and tricellular TJ protein angulin-1/lipolysis-stimulated lipoprotein receptor (LSR) in A549. In normal HLE cells, the knockdown of CGN and FOXO1 increased CLDN-4, while HDAC inhibitors increased CGN and CLDN-4. In conclusion, the knockdown of CGN via FOXO1 contributes to the malignancy of NSCLC. Both HDAC inhibitors, TSA and Quisinostat, may have potential for use in therapy for lung adenocarcinoma via changes in the expression of CGN and FOXO1.

Number of lymph nodes dissected and upstaging rate of the N factor in robot-assisted thoracic surgery versus video-assisted thoracic surgery for patients with cN0 primary lung cancer.

PURPOSE: The accuracy of lymph node (LN) dissection in robotic surgery for lung cancer remains controversial. We compared the accuracy of LN dissection in robot-assisted thoracic surgery (RATS) vs. video-assisted thoracic surgery (VATS). METHODS: The subjects of this retrospective analysis were 226 patients with cN0 primary lung cancer who underwent robot-assisted or video-assisted thoracic lobectomy with LN dissection, in our department, between April, 2016 and February, 2021. We compared the numbers of all LNs and mediastinal LNs dissected, the time required for LN dissection, complications, and upstaging rates of the N factor between the groups. Furthermore, we performed an inverse probability of treatment weighting-adjusted analysis to reduce potential bias between the groups. RESULTS: The number of dissected LNs was higher in the RATS group in both the unweighted and weighted analyses. The time required for lymph node dissection was also longer in RATS. There was no significant difference in complications or in the upstaging rate of the N factor between the groups. CONCLUSION: More LNs were dissected with RATS. Thus, the usefulness of robot-assisted surgery for LN dissection needs to be investigated further.

[Invasive Mucinous Adenocarcinoma that Invades the Wall of a Cystic Lesion:Report of a Case].

Invasive mucinous adenocarcinoma (IMA) is a rare and special type of lung adenocarcinoma. We report a case of IMA presenting as a cystic lesion in the S10 of the right lung, diagnosed by surgical biopsy and treated with right lower lobectomy. The patient was a 60-year-old man who was found to have a 10-mm-sized frosted ground-glass opacity with a 10-mm-sized air space in the S10 of the right lung while undergoing follow-up after renal cancer surgery in 2018. The air space gradually enlarged and, in 2022, began to show a 40-mm-sized cyst, with partial wall thickening and nodularity on the caudal side. A thoracoscopic partial pneumonectomy was performed to confirm the diagnosis of IMA, and a thoracoscopic radical resection of the right remaining lower lobe was performed. It is important to recognize that adenocarcinoma may occur in patients with thin-wall cavity, as in this case. Additionally, it is necessary to determine the treatment strategy based on the assumption that the tumor may extend to the entire cavity wall, even if it is thin-walled.

[Inflammatory Polyp due to Peanut Aspiration, Needed to Differentiate from Malignant Tumor: Report of a Case].

The patient was in his 70s. He was addmitted to our hospital because of obstructive pneumonia for 3 months. Chest computed tomography( CT) showed a nodule at the base of the right B8, obstructing the basal branch, with consolidation of the peripheral lung. Bronchoscopy revealed the right basal trunk obstruction by a tumorous lesion. FDG-PET showed heterogeneous FDG uptake at the right hilum and the lower lobe suggesting malignancy, and a thoracoscopic right lower lobectomy was performed. Pathology showed a granulation-like nodule and a brown oval foreign body incarcerated in the peripheral bronchus, which was later revealed to be a peanut, and no obvious malignant findings were observed.

Microbiome analysis in women with endometriosis: Does a microbiome exist in peritoneal fluid and ovarian cystic fluid?

Purpose: To investigate the relationship between the microbiome of the female genital tract and endometriosis. Methods: This prospective cohort study included 36 women who underwent laparoscopic surgery for ovarian tumor from July 2019 to April 2020. Of them, 18 had endometriosis, and 18 did not have endometriosis. Vaginal secretions, endometrial fluid, peritoneal fluid, and ovarian cystic fluid were collected during surgery. Next-generation sequencing of bacterial 16S rRNA was performed to characterize the microbiome. Results: Specific microbiomes were not detected in either peritoneal fluid or ovarian cystic fluid regardless of the presence or absence of endometriosis and the type of cyst. When the cutoff value of infectious bacterial abundance in the vagina was set as 64.3%, there were many cases more than a cutoff value in the endometriosis group significantly (p = 0.01). When the cutoff value of infectious bacterial abundance in the endometrium was set as 18.6%, there were many cases more than a cutoff level in the endometriosis cases significantly (p = 0.02). Conclusion: Peritoneal fluid and ovarian cystic fluid are almost sterile, although dysbiosis may occur in the vaginal and endometrial microbiome in women with endometriosis.

Effects of histone deacetylase inhibitors Tricostatin A and Quisinostat on tight junction proteins of human lung adenocarcinoma A549 cells and normal lung epithelial cells.

Histone deacetylase (HDAC) inhibitors have a potential therapeutic role for non-small cell lung cancer (NSCLC). However, more preclinical studies of HDAC inhibitors in NSCLC and normal lung epithelial cells are required to evaluate their antitumor activities and mechanisms. The bicellular tight junction molecule claudin-2 (CLDN-2) is highly expressed in lung adenocarcinoma tissues and increase the proliferation of adenocarcinoma cells. Downregulation of the tricellular tight junction molecule angulin-1/LSR induces malignancy via EGF-dependent CLDN-2 and TGF-ß-dependent cellular metabolism in human lung adenocarcinoma cells. In the present study, to investigate the detailed mechanisms of the antitumor activities of HDAC inhibitors in lung adenocarcinoma, human lung adenocarcinoma A549 cells and normal lung epithelial cells were treated with the HDAC inibitors Trichostatin A (TSA) and Quisinostat (JNJ-2648158) with or without TGF-ß. Both HDAC inhibitors increased anguin-1/LSR, decrease CLDN-2, promoted G1 arrest and prevented the migration of A549 cells. Furthermore, TSA but not Quisinostat with or without TGF-ß induced cellular metabolism indicated as the mitochondrial respiration measured using the oxygen consumption rate. In normal human lung epithelial cells, treatment with TSA and Quisinostat increased expression of LSR and CLDN-2 and decreased that of CLDN-1 with or without TGF-ß in 2D culture. Quisinostat but not TSA with TGF-ß increased CLDN-7 expression in 2D culture. Both HDAC inhibitors prevented disruption of the epithelial barrier measured as the permeability of FD-4 induced by TGF-ß in 2.5D culture. TSA and Quisinostat have potential for use in therapy for lung adenocarcinoma via changes in the expression of angulin-1/LSR and CLDN-2.

Next-generation sequencing of 16S rRNA for identification of invasive bacterial pathogens in a formalin-fixed paraffin-embedded placental specimen: a case report of perinatal fulminant Streptococcus pyogenes infection.

Intrauterine infection is one of the most important causes of maternal death. In perinatal emergency, we often miss an opportunity to obtain culture specimens. In this study, we tried to examine whether we investigated whether bacteria causing infection can be detected from a formalin-fixed paraffin-embedded (FFPE) placental specimen. We examined the placenta from a maternal invasive infection that resulted in infectious abortion at 18 weeks of gestation. The case was diagnosed by acute fever and abdominal pain, and the patient was cured after 3 weeks of intensive antimicrobial treatment. Four Streptococcus pyogenes strains were isolated from vaginal fluid and blood cultures of the patient. All of the strain types were emm1/ST28. We amplified the V1-V2 region of 16S rRNA from an FFPE placental specimen and sequencing was performed using a next-generation sequencer (NGS). Taxonomic analysis was then performed for sequenced data. We succeeded in detecting causative pathogens from the FFPE placenta: 69.1% of the predominantly identified bacteria were S. pyogenes and other small populations of bacteria were detected. Our results revealed the utility of NGS for 16S rRNA analysis of an FFPE placenta. This method may reveal previous perinatal invasive infections of unknown origin retrospectively.

[Congenital Pericardial Defect Incidentally Found at Thoracoscopic Left Upper Lobe Resection:Report of a Case].

We report a rare case of a congenital pericardial defect that was incidentally found at thoracoscopic left upper lobe resection in a patient with lung cancer. A 75-year-old man with a left upper lobe lung cancer was referred to our hospital. We performed thoracoscopic left upper lobectomy and incidentally found a pericardial defect intraoperatively. Careful lymph node dissection was necessary to avoid injury of phrenic nerve and pulmonary artery. Surgery for lung cancer was completed without pericardial repair. After surgery, no complications associated with the pericardial defect has not been encountered.

[Spinal Cord Infarction after Right Upper Lobectomy Combined with Chest Wall Resection].

A 42-year-old man presented with a one-month history of back pain. Chest computed tomography revealed a mass (7.6×5.7 cm) in the right upper lobe, suspicious of chest wall invasion. We performed right upper lobectomy combined with chest wall resection. Partial dissections of the second to sixth ribs and the third and fourth vertebral bodies were conducted. Postoperatively, motor paralysis of the right lower extremity was observed and a diagnosis of spinal infarction was made. After cerebrospinal fluid drainage and administration of edaravone with early rehabilitation, he was able to walk with a brace and was discharged from the hospital.

[Probable Immunoglobulin G4 (IgG4)-related Periaortitis Requiring Differentiation from Mediastinal Tumor].

A 42-year-old man with a history of suspected of Behcet's disease underwent oral steroid treatment. During follow-up, chest X-ray revealed an abnormal shadow of the mediastinum. Chest computed tomography(CT) showed a circumferential tumor around the descending thoracic aorta. Enhanced CT showed a lowly and uniformly enhanced tumor at delay phase. A mediastinal tumor was suspected, which prompted a biopsy of the periaortic tumor by video-assisted thoracic surgery (VATS). Histopathological diagnosis showed numerous immunogloblin G4 (IgG4)-positive plasma cells suggesting the possibility of IgG4-related periaortitis. However, based on the diagnostic criteria, the case was comprehensively diagnosed as probable IgG4-related periaortitis, steroid treatment may have affected blood IgG4-positive cells and tissues.

Effects of fatty acids on inducing endoplasmic reticulum stress in bovine mammary epithelial cells.

Fatty acids play important roles in the regulation of endoplasmic reticulum (ER) stress-induced apoptosis in different cells. Currently, the effects of fatty acids on bovine mammary epithelial cells (MEC) remain unknown. Our study examined bovine MEC viability and measured unfolded protein response (UPR)-related gene and protein expressions following fatty acid treatments. To evaluate the role of fatty acids, we treated MAC-T cells (a line of MEC) with 100 to 400 µM of saturated (palmitic and stearic acid) and unsaturated (palmitoleic, oleic, linoleic, and linolenic acid) fatty acids and 1 to 5 mM of short- and medium-chain fatty acids (acetic, propionic, butyric, and octanoic acid). Thereafter, we determined UPR-related gene expression using quantitative real-time PCR. Palmitic acid stimulated expression of XBP1s, ATF4, ATF6A, and C/EBP homologous protein (CHOP). Stearic acid increased expression of XBP1s and CHOP and decreased expression of ATF4 and ATF6A. Results of Western blot analysis and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay revealed that palmitic and stearic acid reduced MAC-T cell viability and induced extreme ER stress by increasing the protein expression of ER stress markers, such as phospho-PKR-like endoplasmic reticulum kinase, phospho-eIF2α, cleaved CASP-3, and CHOP. Among unsaturated long-chain fatty acids, palmitoleic acid increased expression of ATF4 and ATF6A. Oleic acid increased expression of XBP1s, ATF4, and ATF6A. Linoleic and linolenic acids increased expression of XBP1s, ATF4, and ATF6A but decreased expression of XBP1s and ATF6A at the highest dose. Although palmitoleic, oleic, and linoleic acid decreased CHOP expression, only palmitoleic acid increased MAC-T cell viability. Therefore, unsaturated long-chain fatty acids did not induce severe ER stress. Acetic, propionic, and butyric acids decreased expression of ATF4, ATF6A, and CHOP and increased XBP1s expression. Although only octanoic acid increased ATF4 and ATF6A expressions, it lowered expression of XBP1s and CHOP. Although fatty acid treatment did not increase the levels of ER stress proteins, butyric and octanoic acids reduced cell viability, possibly because of early differentiation. These results suggest that saturated fatty acids play important roles in MEC viability by inducing severe ER stress compared with unsaturated fatty acids. In addition, acetic and propionic acids (short- and medium-chain fatty acids) reduced ER stress. Therefore, the present study reflects the new insight that serum fatty acid concentration plays an important role in maintaining the lactation physiology of dairy cows.

Pulmonary vessels and bronchial anatomy of the left lower lobe.

PURPOSE: To identify and clarify the comprehensive anatomic patterns in the left lower lobe (LLL). METHODS: Using computed tomography (CT) imaging data, including that obtained using three-dimensional CT, we reviewed the anatomic patterns of the pulmonary vessels and bronchi in the left lungs of 539 patients, focusing on the LLL. RESULTS: The two-stem type in A6 was observed in 131 (24.7%) patients and the three-stem type in A6 was observed in 11 (2.1%) patients. The independent two-stem type in B6 was observed in four (0.75%) patients. The B7 with independent branching from the basal bronchi was observed in 42 (7.9%) patients. B* was observed in 129 (24.0%) patients and B* was accompanied by A* in all patients. An extrapericardial common trunk of the left pulmonary veins was identified in five patients (0.93%). CONCLUSION: We identified various bronchovascular patterns in the LLL of a large number of patients. Our results provide useful information for anatomic pulmonary resection, especially segmentectomy.

[Left Upper Lobectomy Followed by Left Lower Pulmonary Vein Reconstruction after Miscutting the Common Trunk of the Left Pulmonary Veins].

A 65-year-old woman was diagnosed with lung cancer on the left upper lobe. During thoracoscopic left upper lobectomy, the common trunk of pulmonary vein was mistaken for the left upper pulmonary vein and divided incorrectly. Instead of left pneumonectomy, we successfully performed pulmonary vein reconstruction. As a result of anticoagulant therapy for 1 month, postoperative course was uneventful.

[Successful Robotic Resection of Left Upper Mediastinal Tumor].

A man was diagnosed with a left upper mediastinal mass. The mass was located near the left subclavian vein, phrenic nerve, vagus nerve, left subclavian artery, and left brachiocephalic vein. He underwent a robotic surgery without additional approaches such as cervical approach on transmanubrial approach. Robotic surgery enabled to remove the tumor safely due to the highly flexible robot forceps under a 3-dimensional visual field. Robotic surgery may be effective for tumors in the upper mediastinum, where important blood vessels and nerves are closely present.

Characterization of Microbiota in Endometrial Fluid and Vaginal Secretions in Infertile Women with Repeated Implantation Failure.

Studies suggest that persisting intrauterine bacterial infectious conditions such as chronic endometritis potentially impair the embryo implantation process. The microbial environment in the female reproductive tract, however, remains largely undetermined in infertile patients with a history of repeated implantation failure (RIF). Using next-generation sequencing, we aimed to characterize the microbiota in the endometrial fluid (EF) and vaginal secretions (VS) in women with RIF. Twenty-eight infertile women with a history of RIF and eighteen infertile women undergoing the first in vitro fertilization-embryo transfer attempt (the control group) were enrolled in the study. On days 6-8 in the luteal phase of the natural, oocyte-pickup, or hormone replacement cycle, the paired EF and VS samples were obtained separately. Extracted genomic DNA was pyrosequenced for the V4 region of 16S ribosomal RNA using a next-generation sequencer. The EF microbiota had higher α-diversity and broader bacterial species than the VS microbiota both in the RIF and control groups. The analysis of the UniFrac distance matrices between EF and VS also revealed significantly different clustering. Additionally, the EF microbiota, but not the VS microbiota, showed significant variation in community composition between the RIF group and the control group. Burkholderia species were not detected in the EF microbiota of any samples in the control group but were detectable in a quarter of the RIF group. To our best knowledge, this is the first study investigating the microbiota in the paired EF and VS samples in infertile women with RIF.

IRE1-XBP1 Pathway of the Unfolded Protein Response Is Required during Early Differentiation of C2C12 Myoblasts.

In skeletal muscle, myoblast differentiation results in the formation of multinucleated myofibers. Although recent studies have shown that unfolded protein responses (UPRs) play an important role in intracellular remodeling and contribute to skeletal muscle differentiation, the involvement of IRE1-XBP1 signaling, a major UPR signaling pathway, remains unclear. This study aimed to investigate the effect of the IRE1-XBP1 pathway on skeletal muscle differentiation. In C2C12 cells, knockdown of IRE1 and XBP1 in cells remarkably suppressed differentiation. In addition, apoptosis and autophagy were dramatically enhanced in the XBP1-knockdown cells, highlighting the participation of IRE1-XBP1 in cell survival maintenance with differentiation stimuli during skeletal muscle differentiation. In myogenic cells, we demonstrated that the expression of CDK5 (cyclin-dependent kinase 5) is regulated by XBP1s, and we propose that XBP1 regulates the expression of MyoD family genes via the induction of CDK5. In conclusion, this study revealed that IRE1-XBP1 signaling plays critical roles in cell viability and the expression of differentiation-related genes in predifferentiated myoblasts and during the early differentiation phase.

[Paraffinoma with Spinal Paralysis 40 Years After Pleural Plombage for Tuberculosis].

Paraffin had been used for pleural plombage in the treatment of tuberculosis. However, paraffin use has been reported to cause late postoperative complications. A 79-year-old man was presented with an extramedullary tumor and spinal paralysis. Forty-seven years ago, he had undergone pleural plombage using paraffin for the treatment of pulmonary tuberculosis. Since the extramedullary tumor was found to be paraffinoma, paraffin in the vertebral canal and thoracic cavity was removed surgically. All the paraffin in the vertebral canal and thoracic cavity was removed. After surgery, the patient remains well, without spinal paralysis.

[Surgical Treatment for Pneumopyothorax Secondary to Pancreatic Fistula].

A 46-year-old woman with cervical cancer with multiorgan metastasis visited our hospital. She underwent a total gastrectomy, splenectomy, distal pancreatectomy, left adrenalectomy, and left partial diaphragmatic resection. Postoperatively, she developed pleural effusion with high level of amylase secondary to a pancreatic fistula, consequently causing left-sided empyema. She developed acute respiratory distress syndrome. Urgent surgical treatment was scheduled, and left lower lobectomy, with diaphragmatic partial resection were performed under the venovenous extracorporeal membrane oxygenation. After surgery, intensive care for 45 days was necessary and she was discharged home 6 months post operatively.

MAPLE 2.3.0: an improved system for evaluating the functionomes of genomes and metagenomes.

MAPLE is an automated system for inferring the potential comprehensive functions harbored by genomes and metagenomes. To reduce runtime in MAPLE analyzing the massive amino acid datasets of over 1 million sequences, we improved it by adapting the KEGG automatic annotation server to use GHOSTX and verified no substantial difference in the MAPLE results between the original and new implementations.

Polysaccharide hydrolase of the hadal zone amphipods Hirondellea gigas.

Hirondellea species are common inhabitants in the hadal region deeper than 7,000 m. We found that Hirondellea gigas thrived in the Challenger Deep possessed polysaccharide hydrolases as digestive enzymes. To obtain various enzymes of other H. gigas, we captured amphipods from the Japan Trench, and Izu-Ogasawara (Bonin) Trench. A phylogenetic analysis based on the cytochrome oxidase I gene showed close relationships among amphipods, despite the geographic distance between the localities. However, several differences in enzymatic properties were observed in these H. gigas specimens. We also carried out RNA sequencing of H. gigas from the Izu-Ogasawara Trench. The cellulase gene of H. gigas was highly homologous to cellobiohydrolase of Glucosyl Hydrolase family 7 (GH7). On the other hand, enzymatic properties of H. gigas's cellulase were different from those of typical GH7 cellobiohydrolase. Thus, these results indicate that hadal-zone amphipod can be good candidates as the new enzyme resource.