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Ectopic induction of optogenetic actuators, such as channelrhodopsin, is a promising approach to restoring vision in the degenerating retina. However, the cell type-specific response of ectopic photoreception has not been well understood. It is limited to obtain efficient gene expression in a specifically targeted cell population by a transgenic approach. In the present study, we established a murine model with high efficiency of gene induction to retinal ganglion cells (RGCs)- and amacrine cells using an improved tetracycline transactivator-operator bipartite system (KENGE-tet system). To investigate the cell type-specific visual restorative effect, we expressed the channelrhodopsin gene into RGCs and amacrine cells using the KENGE-tet system. As a result, enhancement in the visual restorative effect was observed to RGCs and starburst amacrine cells. In conclusion, a photoresponse from amacrine cells may enhance the maintained response of RGCs and further increase/improve the visual restorative effect.

Organoid models provide powerful tools to study tissue biology and development in a dish. Presently, organoids have not yet been developed from mouse tooth. Here, we established tooth organoids (TOs) from early-postnatal mouse molar and incisor, which are long-term expandable, express dental epithelium stem cell (DESC) markers, and recapitulate key properties of the dental epithelium in a tooth-type-specific manner. TOs display in vitro differentiation capacity toward ameloblast-resembling cells, even more pronounced in assembloids in which dental mesenchymal (pulp) stem cells are combined with the organoid DESCs. Single-cell transcriptomics supports this developmental potential and reveals co-differentiation into junctional epithelium- and odontoblast-/cementoblast-like cells in the assembloids. Finally, TOs survive and show ameloblast-resembling differentiation also in vivo. The developed organoid models provide new tools to study mouse tooth-type-specific biology and development and gain deeper molecular and functional insights that may eventually help to achieve future human biological tooth repair and replacement. We are grateful to Evelyne Van Kerckhove (UHasselt), Marc Jans (UHasselt), Jeanine Santermans (UHasselt), and Veerle Vanslembrouck (KU Leuven) for valuable technical help. We thank Dr. Diether Lambrechts’ group (VIB,KULeuven) for technical assistance in 103 Genomics. Computational resources for transcriptome analyses were provided by the ‘‘Vlaams Supercomputer Centrum’’ (VSC), managed by the Fund for Scientific Research (FWO), Flanders (Belgium).We are also grateful to the ImagingCore (VIB,KULeuven) and the CIC (KU Leuven) for use ofmicroscopes and the Center for Brain & Disease Research (CBD) Histology unit (VIB, KU Leuven) for use of histology equipment. We acknowledge the use of the TEMplatforms at VIB-KU Leuven, UHasselt, and Tohoku University. The authors also thank Dr. Adrian Ranga (KU Leuven), Dr. Ronald Driesen(UHasselt), and other non-coauthor members of the Laboratory of Tissue Plasticity in Health and Disease (KU Leuven) for their input. Certain figures were created using BioRender.com or GraphPad Prism (v9.3.1) for macOS. This work was supported by grants from KU Leuven (Research Fund) and Fund for Scientific Research (FWO) Flanders. L.H. was an FWO PhD Fellow (1S84718N). C.B. is an FWO PhD Fellow (1129323N), while F.H. was supported by an FWO project grant (G061819FWO). Use of the Zeiss LSM 780 – SP Mai Tai HP DS is supported by Hercules AKUL/11/37 and FWO G.0929.15 funding to Dr. Pieter Vanden Berghe (CIC, KU Leuven), while the application of the JEOL FLASH 1400 TEM was supported by Hercules FWO funding I000220N to I.L.

Several lines of research suggest that BCL-XL-mediated anti-apoptotic effects may contribute to pathogenesis of myeloproliferative neoplasms driven by JAK2V617F and serve as therapeutic target. Here, we used a knock-in JAK2V617F mouse model and confirmed that Bcl-xl was overexpressed in erythroid progenitors. MPN-induced phenotype in the peripheral blood by conditional knockin of JAK2V617F was abrogated by conditional knockout of Bcl2l1, which presented anemia and thrombocytopenia independently of JAK2 mutation status. The Mx1-Cre Jak2V617W/VF /Bcl2l1f/f mice presented persistent splenomegaly as a result of extramedullary hematopoiesis and pro-apoptotic stimuli in terminally differentiated erythroid progenitors. The pan-BH3 mimetic inhibitor obatoclax showed superior cytotoxicity in JAK2V617F cell models, and reduced clonogenic capacity in ex vivo assay using Vav-Cre Jak2V617F bone marrow cells. Both ruxolitinib and obatoclax significantly reduced spleen weights in a murine Jak2V617F MPN model but did not show additive effect. The tumor burden reduction was observed with either ruxolitinib or obatoclax in terminal differentiation stage neoplastic cells but not in myeloid-erythroid precursors. Therefore, disrupting BCL2 balance is not sufficient to treat MPN at the stem cell level, but it is certainly an additional option for controlling the critical myeloid expansion of the disease.

BACKGROUND Postoperative prolonged air leakage is a frequent complication after lung resection. We have developed a new sealant based on a hydrophobically modified Alaska pollock-derived gelatin (ApGltn) sealant. The purpose of this study was to evaluate the adhesive strength of the ApGltn sealant in comparison with a fibrin sealant using a new spray system in ex vivo and in vivo models. METHODS Pleural defects in ex vivo and in vivo porcine models were created, to which the ApGltn sealant or the fibrin sealant was applied. The pressure resistance was assessed with a stepwise increase in airway pressure to confirm air leakage from the sealing site. Tissue samples covered with each sealant were obtained for histologic assessment. RESULTS In the ex vivo experiment, the leak pressures of the ApGltn sealant were significantly greater than those of the fibrin sealant (102.94 ± 15.6 cm H2O and 28.37 ± 5.1 cm H2O, respectively) (P < .01). In the in vivo experiment, the leak pressures of the ApGltn sealant were also significantly greater than those of the fibrin sealant (68.82 ± 18.04 cm H2O and 43.33 ± 7.13 cm H2O, respectively) (P = .043). The histologic examination confirmed that the ApGltn sealant adhered tightly to both the pleura and the surface of the pleural defect. CONCLUSIONS The ApGltn sealant has sufficiently high adhesive quality in ex vivo and in vivo porcine lungs, which could be considered suitable and effective for use in the prevention of air leakage from the lungs.

Background : : Febrile neutropenia (FN) remains an important complication in pediatric cancer patients. The present study compared the efficacy of meropenem (MEPM) and piperacillin/tazobactam (PIPC/TAZ) with or without intravenous immunoglobulin (IVIG) as second-line therapy for FN in pediatric patients. Methods : : As first-line treatment for FN, 394 episodes in 99 patients were randomly assigned to receive PIPC/TAZ (360 mg/kg/day, maximum 18 g/day) or MEPM (120 mg/kg/day, maximum 3 g/day). Eighty-four episodes in 42 patients were judged as failures, and, thus, were enrolled for second-line treatment. In second-line treatment, antibiotics were switched to MEPM or PIPC/TAZ, and episodes were further randomized for treatment with or without concomitant IVIG at 100 mg/kg/day (maximum 5 g/day) for 3 consecutive days. Results : : The total success rate of second-line treatment was 50.0% (52.0% in PIPC/TAZ and 47.2% in MEPM with or without IVIG, P=0.826). The success rates of patients treated with (IVIG+ group) and without IVIG (IVIG− group) were 53.8 and 46.7%, respectively (P=0.662). In the IVIG+ group, the success rate of patients younger than 8 years old was 78.6%, which was significantly higher than that of those aged 8 years and older (40.0%, P=0.043). Conclusion : : PIPC/TAZ and MEPM were equally effective as second-line treatment. Concomitant IVIG was also effective, particularly in patients younger than 8 years.

Mutations in the OTOF gene are a common cause of hereditary hearing loss and the main cause of auditory neuropathy spectrum disorder (ANSD). Although it is reported that most of the patients with OTOF mutations have stable, congenital or prelingual onset severe-to-profound hearing loss, some patients show atypical clinical phenotypes, and the genotype–phenotype correlation in patients with OTOF mutations is not yet fully understood. In this study, we aimed to reveal detailed clinical characteristics of OTOF-related hearing loss patients and the genotype–phenotype correlation. Detailed clinical information was available for 64 patients in our database who were diagnosed with OTOF-related hearing loss. As reported previously, most of the patients (90.6%) showed a “typical” phenotype; prelingual and severe-to-profound hearing loss. Forty-seven patients (73.4%) underwent cochlear implantation surgery and showed successful outcomes; approximately 85–90% of the patients showed a hearing level of 20–39 dB with cochlear implant and a Categories of Auditory Performance (CAP) scale level 6 or better. Although truncating mutations and p.Arg1939Gln were clearly related to severe phenotype, almost half of the patients with one or more non-truncating mutations showed mild-to-moderate hearing loss. Notably, patients with p.His513Arg, p.Ile1573Thr and p.Glu1910Lys showed “true” auditory neuropathy-like clinical characteristics. In this study, we have clarified genotype–phenotype correlation and efficacy of cochlear implantation for OTOF-related hearing loss patients in the biggest cohort studied to date. We believe that the clinical characteristics and genotype–phenotype correlation found in this study will support preoperative counseling and appropriate intervention for OTOF-related hearing loss patients.

Foetal hypoxia–ischaemia is a key trigger of meconium aspiration syndrome (MAS). However, many neonates develop MAS without evidence of hypoxia–ischaemia, suggesting the presence of covert but important risk variables. We evaluated the association of MAS with clinical variables, placental histopathologic findings, and inflammatory biomarkers at birth. Of 1336 symptomatic and asymptomatic term singleton neonates with meconium-stained amniotic fluid, 88 neonates (6.6%) developed MAS. Univariate analysis showed that MAS development was associated with low 1- and 5-min Apgar scores, low cord blood pH, funisitis, higher α1-acid glycoprotein levels, and higher haptoglobin levels (all p 1-acid glycoprotein levels (OR 1.02; 95% Cl 1.01–1.03; p = 0.001). Our data from a large cohort of neonates suggested that intrauterine inflammation is one of the key independent variables of MAS development, together with foetal hypoxia–ischaemia.

Viruses belonging to the genus Quaranjavirus in the family Orthomyxoviridae are known as argasid tick-borne viruses. Some viruses in this genus or an unassigned quaranjavirus-like virus have the ability to infect humans although little is known about their pathogenicity. During the surveillance of tick-borne viruses in ixodid ticks in Ehime Prefecture, Japan, novel quaranjavirus-like sequences were detected in three pooled samples of Haemaphysalis histricis nymphs. Phylogenetic analysis revealed that the detected viruses formed a cluster with quaranjaviruses and other related viruses. Specifically, the viruses were closely related to Zambezi tick virus 1 and Uumaja virus, which are quaranjavirus-like viruses recently discovered in ixodid ticks in Africa and Europe, respectively. These findings indicate that the viruses detected in this study were probably a new member of the Quaranjavirus genus or a related group. The viruses were tentatively named Ohshima virus even though only limited sequences of their genome were available. This is the first report on the detection of a quaranjavirus-like virus in the East Asian region. Further investigations are needed to discern its infectivity and pathogenicity against humans or other animals and to determine the potential risk of an emerging tick-borne viral disease.

Alveolar bone defects caused by tooth loss often lead to challenges in implant dentistry, with a need for development of optimal bone biomaterials to predictably rebuild these tissues. To address this problem, we fabricated a novel bone block using platelet-rich fibrin (PRF) and Deproteinized Bovine Bone Mineral (DBBM), and characterized their mechanical and biological properties. The bone block was prepared by mixing DBBM, Liquid-PRF, and Solid-PRF fragments in various combinations as follows: (1) BLOCK-1 made with Solid-PRF fragments + DBBM, (2) BLOCK-2 made with Liquid-PRF + DBBM, (3) BLOCK-3 made with Solid-PRF fragments + Liquid-PRF + DBBM. The time for solidification and the degradation properties were subsequently recorded. Scanning electron microscopy (SEM) and tensile tests were carried out to investigate the microstructure and mechanical properties of each block. The bioactivity of the three groups towards osteoblast differentiation was also evaluated by culturing cells with the conditioned medium from each of the three groups including cell proliferation assay, cell migration assay, alkaline phosphatase (ALP) staining, and alizarin red staining (ARS), as well as by real-time PCR for genes encoding runt-related transcription factor 2 (RUNX2), ALP, collagen type I alpha1(COL1A) and osteocalcin (OCN). BLOCK-3 made with Solid-PRF fragments + Liquid-PRF + DBBM had by far the fastest solidification period (over a 10-fold increase) as well as the most resistance to degradation. SEM and tensile tests also revealed that the mechanical properties of BLOCK-3 were superior in strength when compared to all other groups and further induced the highest osteoblast migration and osteogenic differentiation confirmed by ALP, ARS and real-time PCR. PRF bone blocks made through the combination of Solid-PRF fragments + Liquid-PRF + DBBM had the greatest mechanical and biological properties when compared to either used alone. Future clinical studies are warranted to further support the clinical application of PRF bone blocks in bone regeneration procedures.

Perillaldehyde (PAH), one of the active ingredients of the traditional Chinese medicine (TCM) plant Perilla frutescens, is widely used and exerts crucial anti-cancer activities. The aim of current study is to illustrate the potential mechanisms of PAH-mediated regulation of bone metastasis and osteoclastogenesis in prostate cancer (PCa) cell lines. Effects of PAH on proliferation, invasion and migration of PC-3 cells were assessed with the Cell Counting Kit-8 (CCK-8) assay and Transwell assays, respectively. Effects of PAH on stem cell characteristics of PC-3 cells were evaluated by cell-matrix adhesion assay, colony formation assay, spheroid formation assay, as well as western blot . The anti-metastasis and anti-osteoclastogenesis activity of PAH in RAW264.7 cells was examined by osteoclast differentiation assay and western blot. The protein levels of CD133 and CD44 in PC-3 cells and the activity of nuclear factor kappa B (NF-κB) signaling pathway in RAW264.7 cells were measured by western blot. PAH suppressed proliferation, invasion and migration of PC-3 cells, prevented stem cell characteristics including cell-matrix adhesion, colony formation, spheroid formation as well as CD133 and CD44 expression. PAH inhibited bone metastasis and osteoclastogenesis via repressing the activation of NF-κB pathway as well as (RANKL)- and cancer cell- induced osteoclastogenesis in PCa cells. These findings suggested the potential therapeutic effects of PAH on the metastasis of patients with PCa.

The spread of multidrug-resistant and virulent Staphylococcus aureus among children is a public health concern, but the actual conditions in Myanmar have not been characterized. In this study, a to...

Tumor heterogeneity underlies resistance to tyrosine kinase inhibitors (TKI) in lung cancers harboring EGFR mutations. Previous evidence suggested that subsets of preexisting resistant cells are selected by EGFR-TKI treatment, or alternatively, that diverse acquired resistance mechanisms emerge from drug-tolerant persister (DTP) cells. Many studies have used bulk tumor specimens or subcloned resistant cell lines to identify resistance mechanism. However, intratumoral heterogeneity can result in divergent responses to therapies, requiring additional approaches to reveal the complete spectrum of resistance mechanisms. Using EGFR-TKI-resistant cell models and clinical specimens, we performed single-cell RNA-seq and single-cell ATAC-seq analyses to define the transcriptional and epigenetic landscape of parental cells, DTPs, and tumor cells in a fully resistant state. In addition to AURKA, VIM, and AXL, which are all known to induce EGFR-TKI resistance, CD74 was identified as a novel gene that plays a critical role in the drug-tolerant state. In vitro and in vivo experiments demonstrated that CD74 upregulation confers resistance to the EGFR-TKI osimertinib and blocks apoptosis, enabling tumor regrowth. Overall, this study provides new insight into the mechanisms underlying resistance to EGFR-TKIs. Significance: Single-cell analyses identify diverse mechanisms of resistance as well as the state of tolerant cells that give rise to resistance to EGFR tyrosine kinase inhibitors.

The cell line NISES-AnPe-428 (AnPe), derived from the Chinese oak silkworm Antheraea pernyi, was characterized for its permissiveness and productivity for six different nucleopolyhedrovirus (NPV) species. These NPVs included homologous Antheraea pernyi NPV (AnpeNPV) and heterologous Autographa californica multiple NPV (AcMNPV), Bombyx mori NPV (BmNPV), Hyphantria cunea MNPV (HycuMNPV), Spodoptera exigua MNPV (SeMNPV), and Lymantria dispar MNPV (LdMNPV), representing viruses that had been isolated from insect species belonging to five different families (Saturniidae, Noctuidae, Bombycidae, Arctiidae, and Lymantriidae). We found that AnPe cells supported productive replication of AnpeNPV, AcMNPV, BmNPV, HycuMNPV, and SeMNPV to varying degrees. Upon infection with SeMNPV, a subset of AnPe cell population in the culture underwent apoptosis, while remaining cells produced limited amounts of progeny virions and polyhedra. AnPe cells were refractory to LdMNPV infection and failed to support replication of viral DNA, indicating that viral replication was restricted at or prior to the step of viral DNA replication. These results indicated that AnPe cells have the potential to provide excellent systems for studying the molecular mechanisms underlying cellular permissiveness for NPV replication and host-range determination of NPVs.

Background: We previously reported an increased prevalence of asthma among patients who had lived in temporary housing after the 2011 Great East Japan Earthquake. We investigated the prognosis of asthma in former residents of temporary housing after allergen avoidance.Methods: Asthma was diagnosed in adults ≥ 15 years from 2014 to 2019 who had lived in temporary housing in Ishinomaki City for at least 1 year. The disease prognosis after the intervention of allergen avoidance in cases that were followed for more than 3 years during the 6-year study period was analyzed. We measured the Dermatophagoides farinae -specific immunoglobulin E (IgE) levels in serum, and the amount of Dermatophagoides group 1 (Der 1) antigen on their futons or mattresses. We instructed residents in an allergen avoidance strategy that included 32 tasks, including using microfiber bedding covers. Results: Of the 202 examinees who were followed for at least 3 years during the 6-year study period, 72(35.6%) were asthmatic during at least one examination. Of these 72 asthmatics, 55(76.4%) developed the disease after the earthquake, and more than half of the cases that we diagnosed at the examination were mild intermittent asthma. After the allergen-avoidance intervention, both the Der 1 level on the futons or mattresses of residents who were diagnosed with asthma but who were nonasthmatic at the final screening and their serum Der f-specific IgE levels significantly decreased (P < 0.01) at the final examination.Conclusion: Antigen avoidance ameliorated mild asthma that was prevalent among residents of temporary housing after the earthquake.

Epithelial–mesenchymal transition (EMT) in retinal pigment epithelial (RPE) cells plays a key role in proliferative retinal diseases such as age-related macular degeneration by contributing to subretinal fibrosis. To investigate the potential role of retinoic acid receptor-α (RAR-α) signaling in this process, we have now examined the effects of the RAR-α agonist Am580 on EMT induced by transforming growth factor-β2 (TGF-β2) in primary mouse RPE cells cultured in a three-dimensional type I collagen gel as well as on subretinal fibrosis in a mouse model. We found that Am580 inhibited TGF-β2-induced collagen gel contraction mediated by RPE cells. It also attenuated the TGF-β2-induced expression of the mesenchymal markers α-smooth muscle actin, fibronectin, and collagen type I; production of pro-matrix metalloproteinase 2 and interleukin-6; expression of the focal adhesion protein paxillin; and phosphorylation of SMAD2 in the cultured RPE cells. Finally, immunofluorescence analysis showed that Am580 suppressed both the TGF-β2-induced translocation of myocardin-related transcription factor-A (MRTF-A) from the cytoplasm to the nucleus of cultured RPE cells as well as subretinal fibrosis triggered by laser-induced photocoagulation in a mouse model. Our observations thus suggest that RAR-α signaling inhibits EMT in RPE cells and might attenuate the development of fibrosis associated with proliferative retinal diseases.

Cardiac muscle contraction involves a plethora of molecular actors. In cardiomyocytes, the sensor molecule troponin responds by a conformational change to a signal, calcium binding, which triggers the next steps leading to muscle contraction. This work explores the structural details of calcium binding, structural rearrangements and key interactions within this molecular sensor system., Understanding the regulation of cardiac muscle contraction at a molecular level is crucial for the development of therapeutics for heart conditions. Despite the availability of atomic structures of the protein components of cardiac muscle thin filaments, detailed insights into their dynamics and response to calcium are yet to be fully depicted. In this study, we used molecular dynamics simulations of the core domains of the cardiac muscle protein troponin to characterize the equilibrium dynamics of its calcium‐bound and calcium‐free forms, with a focus on elements of cardiac muscle contraction activation and deactivation, that is, calcium binding to the cardiac troponin Ca2+‐binding subunit (TnC) and the release of the switch region of the troponin inhibitory subunit (TnI) from TnC. The process of calcium binding to the TnC binding site is described as a three‐step process commencing with calcium capture by the binding site residues, followed by cooperative residue interplay bringing the calcium ion to the binding site, and finally, calcium–water exchange. Furthermore, we uncovered a set of TnC–TnI interdomain interactions that are critical for TnC N‐lobe hydrophobic pocket dynamics. Absence of these interactions allows the closure of the TnC N‐lobe hydrophobic pocket while the TnI switch region remains expelled, whereas if the interactions are maintained, the hydrophobic pocket remains open. Modification of these interactions may fine‐tune the ability of the TnC N‐lobe hydrophobic pocket to close or remain open, modulate cardiac contractility and present potential therapy‐relevant targets.

Pontin and Reptin are closely related proteins belonging to the AAA+ (ATPases Associated with various cellular Activities) family. They form a hetero-oligomeric complex, Pontin/Reptin, which is involved in protein stability and assembly of the protein complexes as a molecular chaperone. Overexpression of Pontin and Reptin in tumor cells has been reported and is implicated in the development of various cancers. However, the molecular mechanism of Pontin/Reptin function in oral squamous cell carcinoma (OSCC) development remains unclear. Here, we identify HEAT repeat-containing protein 1 (HEATR1) as a novel binding factor of Pontin/Reptin. Functionally, HEATR1 stabilizes Pontin/Reptin and positively regulates OSCC cell proliferation by activating mTOR and pre-rRNA synthesis. We also find that HEATR1 expression is markedly upregulated in tumor region of OSCC tissue. Hence, we propose that HEATR1 is involved in the regulation of mTOR and ribosome biogenesis as a potential protein stabilizer of Pontin/Reptin in OSCC.

Background Acute ischemic stroke (AIS) is a serious cause of mortality and disability. AIS is a serious cause of mortality and disability. Early diagnosis of atherosclerosis, which is the major cause of AIS, allows therapeutic intervention before the onset, leading to prevention of AIS. Methods Serological identification by cDNA expression cDNA libraries and the protein array method were used for the screening of antigens recognized by serum IgG antibodies in patients with atherosclerosis. Recombinant proteins or synthetic peptides derived from candidate antigens were used as antigens to compare serum IgG levels between healthy donors (HDs) and patients with atherosclerosis-related disease using the amplified luminescent proximity homogeneous assay-linked immunosorbent assay. Results The first screening using the protein array method identified death-inducer obliterator 1 (DIDO1), forkhead box J2 (FOXJ2), and cleavage and polyadenylation specificity factor (CPSF2) as the target antigens of serum IgG antibodies in patients with AIS. Then, we prepared various antigens including glutathione S-transferase-fused DIDO1 protein as well as peptides of the amino acids 297–311 of DIDO1, 426–440 of FOXJ2, and 607–621 of CPSF2 to examine serum antibody levels. Compared with HDs, a significant increase in antibody levels of the DIDO1 protein and peptide in patients with AIS, transient ischemic attack (TIA), and chronic kidney disease (CKD) but not in those with acute myocardial infarction and diabetes mellitus (DM). Serum anti-FOXJ2 antibody levels were elevated in most patients with atherosclerosis-related diseases, whereas serum anti-CPSF2 antibody levels were associated with AIS, TIA, and DM. Receiver operating characteristic curves showed that serum DIDO1 antibody levels were highly associated with CKD, and correlation analysis revealed that serum anti-FOXJ2 antibody levels were associated with hypertension. A prospective case–control study on ischemic stroke verified that the serum antibody levels of the DIDO1 protein and DIDO1, FOXJ2, and CPSF2 peptides showed significantly higher odds ratios with a risk of AIS in patients with the highest quartile than in those with the lowest quartile, indicating that these antibody markers are useful as risk factors for AIS. Conclusions Serum antibody levels of DIDO1, FOXJ2, and CPSF2 are useful in predicting the onset of atherosclerosis-related AIS caused by kidney failure, hypertension, and DM, respectively.

Social behaviour is a complex construct that is reported to include several components of social approach, interaction and recognition memory. Alzheimer’s disease (AD) is mainly characterized by progressive dementia and is accompanied by cognitive impairments, including a decline in social ability. The cholinergic system is a potential constituent for the neural mechanisms underlying social behaviour, and impaired social ability in AD may have a cholinergic basis. However, the involvement of cholinergic function in social behaviour has not yet been fully understood. Here, we performed a selective elimination of cholinergic cell groups in the basal forebrain in mice to examine the role of cholinergic function in social interaction and social recognition memory by using the three-chamber test. Elimination of cholinergic neurons in the medial septum (MS) and vertical diagonal band of Broca (vDB) caused impairment in social interaction, whereas ablating cholinergic neurons in the nucleus basalis magnocellularis (NBM) impaired social recognition memory. These impairments were restored by treatment with cholinesterase inhibitors, leading to cholinergic system activation. Our findings indicate distinct roles of MS/vDB and NBM cholinergic neurons in social interaction and social recognition memory, suggesting that cholinergic dysfunction may explain social ability deficits associated with AD symptoms.

Background/purpose Conditioned media of cultured mesenchymal stem cells (MSCs) contain numerous kinds of secretomes such as cytokines and chemokines. We previously reported that conditioned media of bone marrow-derived MSCs (MSC-CM) promote bone formation. Recently, macrophage phenotype switching from the pro-inflammatory M1 type to the anti-inflammatory M2 type has been reported to be an important phenomenon during tissue regeneration. Some studies reported that this phenotype switching is regulated by secretomes. In this study, macrophage phenotype during bone formation by MSC-CM was investigated. Materials and methods Human MSCs (hMSCs) were cultured in serum-free medium and the collected medium was defined as MSC-CM. Macrophage-related gene expressions in hMSCs cultured with MSC-CM were evaluated by quantitative real-time polymerase chain reaction. MSC-CM was implanted and the evaluations by micro-CT and immunohistochemistry were performed using a rat the calvaria bone defect model. Results Two and four weeks after implantation, the MSC-CM group demonstrated enhanced bone regeneration. Gene expressions of C–C motif chemokine 2 (CCL2), colony-stimulating factor 2 (CSF2) and CD163 was significantly upregulated in cells exposed to MSC-CM. Immunohistochemical staining revealed that iNOS-positive M1 macrophages were reduced, while CD204-positive M2 macrophages were increased in the MSC-CM group at 72 h after implantation, and the M2/M1 ratio increased only in the MSC-CM group. Conclusion MSC-CM enhances macrophage migration and induces M1 to M2 type macrophage switching at an early stage of osteogenesis. Such phenotype switching provides a favorable environment for angiogenesis, cellular migration, and osteogenesis and contributes to MSC-CM-induced early bone formation.

Nuclear pore complexes (NPC) regulate molecular traffics on nuclear envelope, which plays crucial roles during cell fate specification and diseases. The viral accessory protein NSP9 of SARS-CoV-2 is reported to interact with nucleoporin 62 (NUP62), a structural component of the NPC, but its biological impact on the host cell remain obscure. Here, we established new cell line models with ectopic NSP9 expression and determined the subcellular destination and biological functions of NSP9. Confocal imaging identified NSP9 to be largely localized in close proximity to the endoplasmic reticulum. In agreement with the subcellular distribution of NSP9, association of NSP9 with NUP62 was observed in cytoplasm. Furthermore, the overexpression of NSP9 correlated with a reduction of NUP62 expression on the nuclear envelope, suggesting that attenuating NUP62 expression might have contributed to defective NPC formation. Importantly, the loss of NUP62 impaired translocation of p65, a subunit of NF-κB, upon TNF-α stimulation. Concordantly, NSP9 over-expression blocked p65 nuclear transport. Taken together, these data shed light on the molecular mechanisms underlying the modulation of host cells during SARS-CoV-2 infection.

Recent evidence has shown that gut microbiota dysbiosis is associated with development of gestational diabetes mellitus (GDM). However, the gut microbiota composition of non-obese women with GDM, which accounts for a relatively large percentage of Asian GDM, is unknown. We investigated the characteristics of gut microbiota of Japanese pregnant women with GDM. Fecal samples from Japanese pregnant women with GDM (n=20) and normal glucose tolerance (NGT, n=16) were collected at the time of GDM diagnosis (T1), at 35–37 weeks of gestation (T2), and at 4 weeks postpartum (T3). Gut microbiota composition was characterized from fecal DNA by sequencing of 16S rRNA genes. Serum samples were collected late in the third trimester, and the circulating levels of adiponectin and IL-6 were measured by ELISA. At the genus level, Peptostreptococcaceae Romboutsia was enriched in GDM women at T1 (p=0.008) and T2 (p=0.047). The women with lower serum adiponectin tended to have more Romboutsia. The Shannon index was significantly lower in the GDM women at T3 than in the NGT women (p=0.008), and that of the GDM women decreased significantly from T2 to T3 (p=0.02). No significant difference in bacterial community structure was found in a beta diversity analysis. The non-obese GDM women (body mass index

Soluble immune checkpoint molecules are emerging novel mediators of immune regulation. However, it is unclear whether soluble immune checkpoint proteins affect the development of hepatocellular carcinoma (HCC) during nucleos(t)ide analogue (NA) treatment in patients with chronic hepatitis B virus infection. This study included 122 NA-naïve patients who received NA therapy. We assessed the associations of clinical factors, including soluble immune checkpoint proteins, with HCC development during NA treatment. The baseline serum concentrations of 16 soluble immune checkpoint proteins were measured using multiplexed fluorescent bead-based immunoassay. In total, 13 patients developed HCC during the follow-up period (median duration, 4.3 years). Of the 16 proteins, soluble inducible T-cell co-stimulator (≥ 164.71 pg/mL; p = 0.014), soluble programmed cell death-1 (sPD-1) (≤ 447.27 pg/mL; p = 0.031), soluble CD40 (≤ 493.68 pg/mL; p = 0.032), and soluble herpes virus entry mediator (≤ 2470.83 pg/mL; p = 0.038) were significantly associated with HCC development (log-rank test). In multivariate analysis, an sPD-1 level ≤ 447.27 pg/mL (p = 0.014; hazard ratio [HR], 4.537) and α-fetoprotein level ≥ 6.4 ng/mL (p = 0.040; HR, 5.524) were independently and significantly associated with HCC development. Pre-treatment sPD-1 is a novel predictive biomarker for HCC development during NA treatment.

La Trobe University Faculty of Science, Technology and Engineering Murray Darling Freshwater Research CentreMDFRC item.Density-size distributions of a river zooplankton community were examined about biweekly over a year in the upper tidal freshwater portion of a regulated Australian river, specifically the Hawkesbury-Nepean, Results were compared with those of similar studies in lakes (and reservoirs) to characterize the similarities and differences in the structure and grazing function of river and lake zooplankton communities. The density-size distributions of the river zooplankton community were similar to those of lake plankton communities, in terms of a marked temporal variation in shape and overall bimodal shape. They differed in terms of the truncation of the upper body size and the absence of a significant relationship between the slope (regression coefficient) of a log-linear density-size model and environmental variables. The river zooplankton community appeared to have a high average rate of biomass increase when measured against body-mass class and a low mass-specific grazing rate, compared with those of lake communities. Because of methodological differences between studies, intersystem comparisons of size spectra and grazing rates need cautious interpretations and generalizations.

Murray-Darling Freshwater Research CentreMDFRC item.Grazing rates by a zooplankton community were measured in situ by a radiotracer cellmethod at depths of 1 m and 4 m at the upper tidal freshwater portion of a regulated river over a year.The objectives were to evaluate the likely grazing impact on the river phytoplankton community and toproduce predictive models by regressing the measured grazing rates against zooplankton biomass,temperature and food concentrations (represented by chlorophyll a). Grazing attained rates (overallaverage 0.2 day-', range 0.01-0.59 day-', expressed as instantaneous mortality rates of algal cells)comparable to those reported for lentic zooplankton communities. The measured community grazingrates were predictable largely as a function of total biomass or rotifer biomass and surface temperaturefor 1 m depth, and as a function of total biomass or juvenile copepod biomass and surface temperaturefor 4 m depth, with all-positive regression coefficients in the models. Owing to the predominance ofmicrozooplankton in the river, the impact of zooplankton community grazing appears likely to be linkedto a small-size fraction of the phytoplankton community all year. Management strategies for river waterquality may need to take account of possible functional demarcation of grazing by river zooplankton.

Mouse telomerase and the DNA polymerase alpha-primase complex elongate the leading and lagging strands of telomeres, respectively. To elucidate the molecular mechanism of lagging strand synthesis, we investigated the interaction between DNA polymerase alpha and two paralogs of the mouse POT1 telomere-binding protein (POT1a and POT1b). Yeast two-hybrid analysis and a glutathione S-transferase pull-down assay indicated that the C-terminal region of POT1a/b binds to the intrinsically disordered N-terminal region of p180, the catalytic subunit of mouse DNA polymerase alpha. Subcellular distribution analyses showed that although POT1a, POT1b, and TPP1 were localized to the cytoplasm, POT1a-TPP1 and POT1b-TPP1 coexpressed with TIN2 localized to the nucleus in a TIN2 dose-dependent manner. Coimmunoprecipitation and cell cycle synchronization experiments indicated that POT1b-TPP1-TIN2 was more strongly associated with p180 than POT1a-TPP1-TIN2, and this complex accumulated during the S phase. Fluorescence in situ hybridization and proximity ligation assays showed that POT1a and POT1b interacted with p180 and TIN2 on telomeric chromatin. Based on the present study and a previous study, we propose a model in which POT1a/b-TPP1-TIN2 translocates into the nucleus in a TIN2 dose-dependent manner to target the telomere, where POT1a/b interacts with DNA polymerase alpha for recruitment at the telomere for lagging strand synthesis.

Primary vaginal neuroendocrine carcinoma (NEC) is extremely rare among female genital tract tumors. Here, we report 2 cases of vaginal small cell NEC (SCNEC) using immunohistochemistry and next-generation sequencing (NGS) analysis. The 2 patients were in their mid-to-late 70s, presented with abnormal vaginal bleeding and had a vaginal submucosal mass. The biopsied or resected tumors showed a typical neuroendocrine morphology consisting of solid nests of atypical tumor cells, with no specific organoid patterns, and proliferating in the vaginal submucosa. Immunohistochemical analysis showed strong and diffuse expression of chromogranin A, synaptophysin, and p16, but no thyroid transcription factor 1 expression. Additionally, both cases were positive for human papillomavirus (HPV) 18. An NGS-based cancer panel analysis revealed that the tumors carried NF1 and AR mutations, but no major driver mutations were detected. The results of this study suggested that HPV18 infection is linked to vaginal SCNEC.

Objectives: Pediatric acute-onset neuropsychiatric syndrome (PANS) is a clinical diagnosis in children who have an acute manifestation of varied neuropsychiatric symptoms, including obsessive compulsive disorder, eating disorders, tics, anxiety, irritability, and problems with attention/concentration. PANS may develop as a result of a postinfectious syndrome and may represent a new form of postinfectious autoimmunity. To test the hypothesis that multiple, consecutive infusions of intravenous immunoglobulin (IVIG) for PANS can be efficacious, a multisite, open-label study was designed. Methods: The primary endpoint was evaluation of the efficacy of IVIG [Octagam 5%] in PANS over a period of 6 months (six infusions) based on mean changes in psychological evaluation scores using 6 different assessments, including the Children's Yale-Brown Obsessive Compulsive Scale (CY-BOCS), Clinical Global Impression of Severity, and the Parent-Rated Pediatric Acute Neuropsychiatric Symptom Scale (PANS Scale). Results: The final cohort consisted of 21 subjects (7 per site) with moderate to severe PANS. The mean age was 10.86 years (range: 4–16 years). Results demonstrated statistically significant reductions in symptoms from baseline to end of treatment in all six assessments measured. CY-BOCS results demonstrated statistically significant reductions in obsessive compulsive symptoms (p 50% improvement sustained for at least 8 weeks after the final infusion and up to 46 weeks in a subset of subjects. Conclusions: In PANS, which may be associated with an underlying immune dysregulation, sequential infusions of IVIG [Octagam 5%] successfully ameliorated psychological symptoms and dysfunction, with sustained benefits for at least 8 weeks, and up to 46 weeks in a subset of subjects. In addition, baseline immune and autoimmune profiles demonstrated significant elevations in a majority of subjects, which requires further evaluation, characterization, and study to clarify the potential immune dysfunction by which PANS manifests and progresses.

Introduction The need for antiserum for immunohistochemical detection (IHC) of enterovirus (EV) in formaldehyde fixed and paraffin-embedded (FFPE) samples is increasing. The gold standard monoclonal antibody (clone 5D8/1) against EV-envelope protein (VP1) was proven to cross-react with other proteins. Another candidate marker of EV proteins is 2A protease (2Apro ), which is encoded by EV gene and translated by the host cells during EV replication and participates processing proproteins to viral capsid proteins. Materials and methods We raised polyclonal antiserum by immunizing a rabbit with an 18-mer peptide of Coxsackievirus B1 (CVB1)-2Apro and examined the specificity and sensitivity for EV on FFPE tissue samples. Results ELISA study showed a high titer of antibody for 18-mer peptide of CVB1-2Apro , cross reacting with CVB3-2Apro peptide. IHC demonstrated that antiserum against 2Apro reacted with CVB1-infected and VP1-positive Vero-cells. Confocal laser scanning microscopy demonstrated that antigen stained by the 2Apro antibody located in the same cell with VP1 stained by 5D8/1. IHC using 2Apro antiserum showed dense staining in the islets of EV-associated fulminant type 1 diabetic pancreas and that located in the same cell stained positive for VP1 (5D8/1). Specificity of 2Apro antiserum by IHC staining was confirmed by negative 2Apro in 14 VP1-negative non-diabetic control pancreases. Conclusion Our study provides a new polyclonal antiserum against CVB1-2Apro which may be useful for IHC of EV-infected human tissues stored as archive of FFPE tissue samples.

Secreted acid phosphatases (APases) dephosphorylate extracellular organic phosphate compounds to supply inorganic phosphate (Pi) to maintain cellular functions. Here, we show that APases are necessary to maintain a normal replicative lifespan in Saccharomyces cerevisiae. Deletion of all four APase genes shortened the lifespan in yeast strains on synthetic media (irrespective of the concentrations of Pi in the media), but it did not affect the intracellular ortho- and polyphosphate levels. Deletion of IPK1, which encodes inositol-pentakisphosphate 2-kinase, restored the lifespan in APase-null mutants, and IPK1 overexpression shortened the lifespan in wild-type strains. Overexpression of inositol hexakisphosphate (IP6 ) and heptakisphosphate kinases, KCS1 and VIP1, recovered the lifespan in APase-null mutants. Thus, yeast APases modulate the replicative lifespan, probably through dephosphorylation of intracellular IP6 .

Heat shock proteins (HSPs) play an essential role as molecular chaperones in proteostasis. Small HSPs are a group of low-molecular-weight HSPs in the range of 12- 43 kDa and are classified as HSPB. Within the ten members of the family, HSPB1(HSP27), HSPB5 (αB-crystallin), HSPB6 (HSP20), and HSPB8 (HSP22) ubiquitously exist in various tissues, including liver tissue. These small HSPs undergo phosphorylation as a post-translational modification, and their functions are modulated. Hepatocellular carcinoma (HCC) is one of the most frequent cancers and the fourth leading cause of cancer-related death worldwide. HSPs play a cytoprotective role as molecular chaperones. Thus, HSPB has been generally considered to protect HCC cells and help the progression of HCC. On the other hand, recent studies from our laboratories have demonstrated suppressive roles of phospho-HSPB1, HSPB6, and HSPB8 in the progression of HCC. These findings may provide a basis for a novel defense system by HSPB against HCC progression. This review focuses on the cellular functions of HSPB in HCC and summarizes the current research.

The objective of this study was to evaluate the effects of oral administration of fiber from the first week of life on the growth and hindgut environment of preweaning calves. Twenty newborn female Holstein calves were divided into 2 groups as control and treatment. Calves in both groups were reared under the same feeding program except for oral fiber administration. Timothy hay and psyllium were mixed at a 50-to-6 ratio as a treatment diet for oral fiber administration. Calves in the treatment group were orally administered 50 g of fiber daily from 3 to 7 d of age and 100 g of fiber from 8 d of age until weaning. Feed intake and occurrence of diarrhea were recorded daily, and body weight (BW) was recorded weekly for the individual calf. Fresh fe-ces were collected from calves at 7, 21, 35, 49, and 56 d of age to analyze fermentation parameters and microbiota to characterize the hindgut environment. Higher fiber intake in the treatment group due to oral administration of timothy and psyllium did not affect the starter intake and achieved higher BW at 21 d of age. The fecal pH, total volatile fatty acid, lactate, and ammonia nitrogen concentrations were not affected by oral fiber administration; meanwhile, the molar propor-tion of propionate was higher in the treatment group at 7 d of age. The difference in fecal microbiota in the calves subjected to the oral administration of fiber was observed within 21 d of life; Lactobacillus spp. and Prevotella spp. showed higher abundance, whereas that of Clostridium perfringens was decreased. These higher abundances of beneficial bacteria and lower abundance of pathogenic bacteria during early life may partly ex-plain the higher BW of calves in the treatment group at 21 d of age. Furthermore, no adverse effect was ob-served for the BW and health status in the treatment group throughout the preweaning period. Therefore, early fiber feeding via oral administration potentially contributes to improving the hindgut environment in newborn calves, which leads to better growth of calves during the early stage of life.

Key message Tolerance and acclimation of photosynthesis to warmer growing conditions among nine urban tree species was species-specific indicating that individual screening is needed to prepare urban greenspace for a warmer future. Abstract To prepare and manage urban greenspace for a warmer future, we must select trees that can tolerate or acclimate to warmer growing conditions. Here, we compared tolerance and acclimation of photosynthesis to warmer growing conditions among urban trees species in Japan. Two paired experiments were installed where saplings of nine species were grown outside at two locations (warm- and cool-temperate climates) and in a greenhouse with and without passive warming. We compared the temperature where quantum yield declined to 50% of maximum value (T50) due to excess heat (50–60 °C) and the thermal optimum for photosynthesis (Topt) among species. For two deciduous and three evergreen species, T50 was higher for saplings grown in warm- than cool-temperate climate. T50 was also higher under passive warming for three species, of which two evergreen species (Morella rubra, Quercus myrsinifolia) showed marked increases (5–7 °C). Topt was higher under passive warming in three of six species examined, but the acclimation response of photosynthesis and stomatal conductance to warming was highly species-specific. We inferred that M. rubra and Q. myrsinifolia, which acclimated consistently to warmer growing conditions in both experiments, are heat-hardy species. Our results also suggest warm-temperate evergreen species are not necessarily more tolerant of warmer growing conditions than cool-temperate deciduous species. For example, despite its warm-temperate origin, Machilus thumbergii showed no acclimation response and photosynthetic rates were lower under passive warming. Our results indicate species need to be screened individually to assess their physiological tolerance and acclimation potential to warmer climate.

Yoichi Nishii,1 Kazuki Furuhashi,1 Saki Nakamura,1 Miho Nishio,1 Yuki Nakamura,1 Kengo Ushiro,1 Kentaro Ito,1 Tadashi Sakaguchi,1 Yuta Suzuki,1 Kentaro Fujiwara,1 Taro Yasuma,2 Tetsu Kobayashi,3 Corina D’Alessandro-Gabazza,2 Esteban C Gabazza,2 Osamu Taguchi,1 Osamu Hataji1 1Respiratory Center, Matsusaka Municipal Hospital, Matsusaka, Mie, Japan; 2Department of Immunology, Faculty and Graduate School of Medicine, Mie University, Tsu-city, Mie, Japan; 3Department of Pulmonary and Critical Care Medicine, Faculty and Graduate School of Medicine, Mie University, Tsu-city, Mie, JapanCorrespondence: Esteban C GabazzaDepartment of Immunology, Mie University Graduate School of Medicine, Edobashi 2-174, Tsu-city, Mie, 514-8507, JapanTel +81 59 231 5017Fax +81 59 231 5225Email gabazza@doc.medic.mie-u.ac.jpIntroduction: Many patients with nontuberculous mycobacteria pulmonary disease are asymptomatic. The disease diagnosis is confirmed in only a small proportion of patients with radiological findings suspicious for nontuberculous mycobacteria pulmonary disease. Thus, many patients remained undiagnosed. Here, we evaluated the diagnostic value of digital polymerase chain reaction (PCR) in nontuberculous mycobacteria pulmonary disease.Methods: We prospectively evaluated 123 patients with radiological findings suspicious for nontuberculous mycobacteria pulmonary disease. Digital PCR was performed using bronchial lavage fluid, sputum, saliva, blood, and urine.Results: The culture of bronchial washing fluid was positive for nontuberculous mycobacteria in 53 patients and negative in 70. The positive detection rate of nontuberculous mycobacteria by digital PCR in patients with positive culture (n = 53) was as follows: bronchial lavage fluid 100%, sputum 62.9%, saliva 41.5%, blood 7.5%, and urine 3.8%. All patients with two or more positive partitions for nontuberculous mycobacteria in the digital PCR of bronchial lavage fluid showed nontuberculous mycobacteria growth in the bronchial lavage fluid culture. The digital PCR analysis of the bronchial lavage fluid showed a high sensitivity (100%), specificity (85.7%), positive predictive value (84.1%), negative predictive value (100%), and a high concordance rate (91.9%) with the bronchial lavage fluid culture results. In addition, the culture of bronchial lavage fluid was positive for nontuberculous mycobacteria in patients with two or more positive partitions in the digital PCR of sputum and saliva with a combined positive predictive value of 81.1%.Conclusion: Digital PCR analysis of nontuberculous mycobacteria in bronchial lavage fluid shows a high concordance rate with the bronchial lavage fluid culture results and a high positive predictive value using both sputum and saliva, suggesting the potential usefulness of dPCR for diagnosis of nontuberculous mycobacteria pulmonary disease in clinical practice.Keywords: digital PCR, nontuberculous mycobacteria pulmonary disease, diagnosis, bronchial lavage fluid culture, bronchoscopy

Angiotensin I converting enzyme (ACE) inhibition is one of the key factors for cardiovascular disease, and inhibition of ACE activity is related to the prevention of high blood pressure. Until now, some ACE inhibitory peptides have been found in the protein hydrolysates of red algae, and most of them were derived from phycobiliproteins and rubisco. In this study, we modified the preparation method to evaluate the potential of ACE inhibitory activity from protein hydrolysate of red algaMazzaella japonicaexcept for phycobiliproteins and rubisco. As a result, we identified 11 peptides (YRD, VSEGLD, TIMPHPR, GGPAT, SSNDYPI, SRIYNVKSNG, VDAHY, CPYDWV, YGDPDHY, NLGN and DFGVPGHEP) in the hydrolysate by the reversed phase-HPLC and MALDI-TOF/MS/MS. Among them, YRD would be derived from phycoerythrin alpha-subunit. However, the others would be from other proteins encoded in chloroplast or nuclear genomes. This study revealed that not only phycobiliproteins and rubisco but also the other proteins in red algae have the potential for the source of ACE inhibitory peptide.

Many of genes specifically expressed in peri-implant soft tissue (PIST) selected by microarray analysis are involved in the inflammatory response. This study investigated the gene expression and localization of PIST-specific inflammatory markers in PIST during wound healing. Pure titanium implants were implanted into the rat upper mandibular socket to create PIST. Samples were harvested from PIST as an experimental group, and tooth extracted area of oral mucosa tissue (OMT) and healthy periodontal tissue (PT) as control groups. The gene expressions of four standard inflammatory markers and nine PIST-specific inflammatory markers including chemokine (C-X-C motif) ligand 2 (CXCL2) during wound healing were examined. Immunoreactions of CXCL2 and immune cells in PIST and control tissues were compared. During wound healing, gene expression of PIST-specific inflammatory markers was higher in PIST than in OMT (p

A bis(μ-oxo)diiron(IV,IV) complex as a model for intermediate Q in the methane monooxygenase reaction cycle has been prepared. The precursor complex with a [FeIIIFeIV(μ-O)2] core was fully characterized by X-ray crystallography and other spectroscopic analyses and was converted to the [FeIV2(μ-O)2] complex via electrochemical oxidation at 1000 mV (vs Ag/Ag+) in acetone at 193 K. The UV-vis spectral features, Mossbauer parameters (ΔEQ = 2.079 mm/s and δ = -0.027 mm/s), and EXAFS analysis (Fe-O/N = 1.73/1.96 A and Fe···Fe = 2.76 A) support the structure of the low-spin (S = 1, for each Fe) [FeIV2(μ-O)2] core. The rate constants of the hydrogen abstraction reaction from 9,10-dihydroanthracene at 243 K suggest the high reactivity of these synthetic bis(μ-oxo)diiron complexes supported by simple N4 tripodal ligand.

Lung cancer is one of the most aggressive tumour types. Targeted therapies stratified by oncogenic drivers have substantially improved therapeutic outcomes in patients with non-small-cell lung cancer (NSCLC)1. However, such oncogenic drivers are not found in 25–40% of cases of lung adenocarcinoma, the most common histological subtype of NSCLC2. Here we identify a novel fusion transcript of CLIP1 and LTK using whole-transcriptome sequencing in a multi-institutional genome screening platform (LC-SCRUM-Asia, UMIN000036871). The CLIP1–LTK fusion was present in 0.4% of NSCLCs and was mutually exclusive with other known oncogenic drivers. We show that kinase activity of the CLIP1–LTK fusion protein is constitutively activated and has transformation potential. Treatment of Ba/F3 cells expressing CLIP1–LTK with lorlatinib, an ALK inhibitor, inhibited CLIP1–LTK kinase activity, suppressed proliferation and induced apoptosis. One patient with NSCLC harbouring the CLIP1–LTK fusion showed a good clinical response to lorlatinib treatment. To our knowledge, this is the first description of LTK alterations with oncogenic activity in cancers. These results identify the CLIP1–LTK fusion as a target in NSCLC that could be treated with lorlatinib. Whole-transcriptome sequencing of a subset of 75 non-small-cell lung cancer specimens in a multi-institutional genome screening study identified a fusion of the CLIP1 and LTK genes with transformational potential due to constitutive LTK kinase activity.

Fine bubble (FB) and ultrafine bubble (UFB) generators have been installed in hydroponic systems to promote plant growth. However, their application is limited because the mechanism of the effect o...

This study aimed to clarify the genetic features of endometrioid-type endometrial cancer arising in adenomyosis (EC-AIA) using targeted sequencing and immunohistochemistry (IHC) for both carcinoma and adjacent adenomyosis tissues. We identified three endometrioid-type EC-AIAs in 689 patients with endometrial cancer; two exhibited grade 3 endometrioid carcinoma. IHC revealed retained expression of PMS2, MSH6, ARID1A, and PAX2. Two of them showed diffuse strong p53 expression only in the carcinoma. PTEN expression was lost in carcinoma of only one of these cases. Carcinoma had many gene mutations than adjacent adenomyosis in all cases. KRAS and TP53 mutations were found in two of them. The other patient had mutations in KRAS, PIK3CA, and PPP2R1A. They were classified as two "p53-mutated" and one "non-specific molecular profile." These molecular alterations in endometrioid-type EC-AIA imply similar carcinogenesis to a subset of endometrial endometrioid carcinoma and might be used as targets of liquid biopsy after further validation.

Two new species of Rhinogobius found in streams on central part of Palawan Island, Philippines are described. The two new species, Rhinogobius estrellae and Rhinogobius tandikan share unique transverse rows of sensory papillae on the cheek with Rhinogobius similis Gill, 1859, but differ from the latter in fin ray counts, arrangement of the scales, etc. The two new species are distinguished from each other by the pectoral-fin ray count, the longitudinal- and predorsal-scale counts, and colouration of the body. Rhinogobius estrellae new species and R. tandikan new species have been found allopatrically in a stream within Malatgao River system flowing into the Sulu Sea and in the Cayulo River flowing into the South China Sea, respectively. The Malatgao River system is the southernmost habitat of the genus Rhinogobius. Rhinogobius similis had been considered as the only member of the most basal lineage of this genus, but our mitochondrial genome analysis suggested that the two new species are additional members of this lineage. They are considered to be relicts of their common ancestor with R. similis, which probably had a wider distribution.

A 70-year-old Japanese woman who was treated for interstitial pneumonia (IP) with steroid therapy developed cholecystitis. A serial computed-tomography (CT) imaging showed irregular thickness of the fundus wall of the gallbladder and two rapidly enlarged lymph nodes (LNs): number (no.) 12 and no. 8a. Positron-emission tomography-computed tomography (PET-CT) scan showed an abnormal uptake at the site of the gallbladder tumor and those LNs. We subsequently performed open radical cholecystectomy and LN dissection of the no. 12 and 8a LNs, following complete remission of IP. The histology showed gallbladder adenocarcinoma, with a single focus of neuroendocrine carcinoma (NEC) component of less than 30%; Ki-67 index > 80%, synaptophysin (Syn) (+), chromogranin A (CgA) (+), and clusters of differentiation (CD) 56 (+) (T2bN1M0, Stage IIIB). LN no. 8a was diffusely metastatic with NEC components. LN no. 12c, which was adjacent to the cystic duct, revealed necrosis without apparent tumor cells, but was highly suspicious for tumor necrosis. The final diagnosis was adenocarcinoma of the gallbladder with focal NEC (

Objective. Coil embolization (CE) for coronary artery perforation (CAP) has not been thoroughly evaluated. This study aimed to evaluate the extent of myocardial damage and impact on cardiac function after CE for CAP. Methods. A total of 110 consecutive patients treated with CE for CAP were retrospectively identified. The degree of myocardial damage and impact on cardiac function were evaluated. Results. Forty-nine (44.5%) cases involved chronic total occlusions. A guidewire was the cause of perforation in 97 (88.2%) patients. The success rate of CE was 98.2%. Almost all patients were prescribed either antiplatelet drugs or anticoagulant medication or both. Patients with perforation types III and IV were found to be prone to creatinine kinase (CK) elevation and epicardial main vessel perforation, thereby causing myocardial damage. No changes were noted in the ejection fraction (EF) in patients with type V distal perforation and collateral channel perforation, while patients with perforation of the epicardial main vessel may show impaired cardiac function afterward. Conclusions. CE is safe and effective for treating CAP, especially when collateral channels and distal vessels are involved. Meanwhile, efforts should be taken to prevent CAP in epicardial main vessels since it may be difficult to treat with CS and cause myocardial damage when bailed out with CE leading to vessel sacrifice. We found that it was not necessary to change the anticoagulant regimen after CE owing to its ability to achieve robust hemostasis.

Introduction GATA1 mutation plays an important role in initiating transient abnormal myelopoiesis (TAM) and in the clonal evolution towards acute megakaryoblastic leukaemia (AMKL) associated with Down syndrome (DS). This study aimed to develop and validate the clinical utility of a complementary DNA (cDNA) analysis in parallel with the conventional genomic DNA (gDNA) Sanger sequencing (Ss), as an initial screening test for GATA1 mutations. Methods GATA1 mutations were evaluated using both gDNA and cDNA in 14 DS patients using Ss and fragment analysis (FA), respectively. Results The detection sensitivity of conventional gDNA sequencing was limited in low blast percentage TAM (LBP-TAM); however, cDNA-based Ss readily detected all the pathognomonic GATA1 mutations. The cDNA-based FA readily detected GATA1 frameshift mutation with a reliable sensitivity ranging from 0.005% to 0.01% of clonal cells. Conclusions GATA1 mutations are heterogeneous; therefore, we would like to propose a dual cDNA and gDNA analysis as a standard diagnostic approach, especially for LBP-TAM. cDNA-based FA promises an excellent sensitivity for detecting frameshift GATA1 mutations in the longitudinal clonal evolution towards AMKL without using a patient specific primer.

Isocitrate dehydrogenase 1 (IDH1) is a key enzyme in cellular metabolism. IDH1 mutation (IDH1mut) is the most important genetic alteration in lower grade glioma, whereas glioblastoma (GB), the most common malignant brain tumor, often has wild‐type IDH1 (IDH1wt). Although there is no effective treatment yet for neither IDH1wt nor IDHmut GB, it is important to note that the survival span of IDH1wt GB patients is significantly shorter than those with IDH1mut GB. Thus, understanding IDH1wt GB biology and developing effective molecular‐targeted therapies is of paramount importance. Fatty acid‐binding protein 7 (FABP7) is highly expressed in GB, and its expression level is negatively correlated with survival in malignant glioma patients; however, the underlying mechanisms of FABP7 involvement in tumor proliferation are still unknown. In this study, we demonstrate that FABP7 is highly expressed and localized in nuclei in IDH1wt glioma. Wild‐type FABP7 (FABP7wt) overexpression in IDH1wt U87 cells increased cell proliferation rate, caveolin‐1 expression, and caveolae/caveosome formation. In addition, FABP7wt overexpression increased the levels of H3K27ac on the caveolin‐1 promoter through controlling the nuclear acetyl‐CoA level via the interaction with ACLY. Consistent results were obtained using a xenograft model transplanted with U87 cells overexpressing FABP7. Interestingly, in U87 cells with mutant FABP7 overexpression, both in vitro and in vivo phenotypes shown by FABP7wt overexpression were disrupted. Furthermore, IDH1wt patient GB showed upregulated caveolin‐1 expression, increased levels of histone acetylation, and increased levels of acetyl‐CoA compared with IDH1mut patient GB. Taken together, these data suggest that nuclear FABP7 is involved in cell proliferation of GB through caveolae function/formation regulated via epigenetic regulation of caveolin‐1, and this mechanism is critically important for IDH1wt tumor biology., In this study, we observed high expression and nuclear localization of FABP7 in IDH1wt glioblastoma (GB). Using GB cell lines, we investigated that nuclear FABP7 interacted with ATP citrate lyase (ACYL), as well as the associated increase of nuclear acetyl‐CoA levels, which led to an increase of histone acetylation at the caveolin‐1 promoter. Consequently, the increase in caveolin‐1 expression enhanced caveolae/caveosome function and formation, as well as the intracellular signalling activity in IDHwt cells related to GB tumour proliferation.

Background The ceftriaxone-resistant Neisseria gonorrhoeae FC428 clone was first discovered in Japan in 2015. Objectives We investigated the possibility of horizontal gene transfer from Neisseria subflava harbouring the mosaic-like PBP-2 in the emergence of the FC428 clone. We also analysed whether there were fitness costs associated with the sustained international dissemination of the clone. Methods Sequencing of the penA gene in ceftriaxone-resistant N. subflava strains was performed. For transformation experiments between donor N. subflava and ciprofloxacin-resistant wild-type penA N. gonorrhoeae recipient, the full-length PCR amplification product of the penA gene, including DUS regions, was used as the donor DNA. Biological fitness of the transformants was measured by growth competition assays. The impact of QRDR and mtrR mutations, which have been reported as compensatory mutations, on fitness was also assessed. Results The penA mosaic allele of the FC428 clone showed 100%, 91.8%, and 89.8% homology, respectively, with penA genes of three ceftriaxone-resistant N. subflava strains, No. 30, No. 9 and No. 14. Results were consistent with homologous recombination with the donated penA mosaic allele. In co-cultures with the parent strain, transformants showed comparable growth indicating that a gyrA mutation compensates for the fitness cost of mosaic penA alleles. Conclusions Our findings support the hypothesis that the FC428 clone was generated by transformation of the mosaic penA allele from oropharyngeal N. subflava to N. gonorrhoeae. Furthermore, it suggests that mutations in the gyrA QRDR region compensate for fitness costs and contribute to the continued transmission of the FC428 clone.