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As a space project, in "Stem Cells" by the Japan Aerospace Exploration Agency (JAXA), frozen mouse ES cells were stored on the International Space Station (ISS) in the Minus Eighty Degree Laboratory Freezer for ISS (MELFI) for 1584 days. After taking these cells back to the ground, the cells were thawed and cultured, and their gene expressions were comprehensively analyzed using RNA sequencing in order to elucidate the early response of the cells to long-time exposure to space radiation consisting of various ionized particles. The comparisons of gene expression involved in double-stranded break (DSB) repair were examined. The expressions of most of the genes that were involved in homologous recombination (HR) and non-homologous end joining (NHEJ) were not significantly changed between the ISS-stocked cells and ground-stocked control cells. However, the transcription of Trp53inp1 (tumor protein 53 induced nuclear protein-1), Cdkn1a (p21), and Mdm2 genes increased in ISS-stocked cells as well as Fe ion-irradiated cells compared to control cells. This suggests that accumulated DNA damage caused by space radiation exposure would activate these genes, which are involved in cell cycle arrest for repair and apoptosis in a p53-dependent or -independent manner, in order to prevent cells with damaged genomes from proliferating and forming tumors. [ABSTRACT FROM AUTHOR]

While depression has been associated with alterations in the hypothalamic-pituitary adrenal (HPA) axis function, there is still controversy regarding the nature and extent of the dysfunction, such as in the debate about hypercortisolism vs. hypocortisolism. It may therefore be necessary to understand whether and how HPA axis function in depression is linked to mRNA expression of key genes regulating this system. We studied 163 depressed outpatients, most of whom were chronically ill, and 181 healthy controls. Blood mRNA expression levels of NR3C1 (including GRα , GRβ , and GR-P isoforms), FKBP4 , and FKBP5 were measured at baseline. HPA axis feedback sensitivity was measured by the dexamethasone (Dex)/corticotropin-releasing hormone (CRH) test. The association between mRNA expression levels and HPA axis feedback sensitivity was examined. Compared to controls, patients showed significantly higher expression of GRα and lower expression of FKBP5 , and higher post-Dex cortisol levels, even after controlling for age and sex. FKBP5 expression was significantly positively correlated with cortisol levels in patients, while GRα expression was significantly negatively correlated with cortisol levels in controls. Most patients were taking psychotropic medications. The large number of correlation tests may have caused type I errors. The tripartite relationship between depression, mRNA expression of GR and FKBP5 , and HPA axis function suggests that the altered gene expression affects HPA axis dysregulation and, as a result, impacts the development and/or illness course of depressive disorder. The combination of increased GRα expression and decreased FKBP5 expression may serve as a biomarker for chronic depression. • We examined blood mRNA expression of GR isoforms and FKBP4 / FKBP5 in depression. • Compared to controls, patients showed higher GRα and lower FKBP5 expression levels. • Compared to controls, patients had higher post-dexamethasone cortisol levels. • The differential gene expression was linked to altered HPA axis feedback sensitivity. • Gene expression affects HPA axis function, thereby possibly impacting depression. [ABSTRACT FROM AUTHOR]

Prostaglandins are bioactive compounds, and the activation of their receptors affects the expression of clock genes. However, the prostaglandin F receptor (Ptgfr) has no known relationship with biological rhythms. Here, we first measured the locomotor period lengths of Ptgfr-KO (B6.129-Ptgfrtm1Sna) mice and found that they were longer under constant dark conditions (DD) than those of wild-type (C57BL/6J) mice. We then investigated the clock gene patterns within the suprachiasmatic nucleus in Ptgfr-KO mice under DD and observed a decrease in the expression of the clock gene cryptochrome 1 (Cry1), which is related to the circadian cycle. Moreover, the expression of Cry1, Cry2, and Period2 (Per2) mRNA were significantly altered in the mouse liver in Ptgfr-KO mice under DD. In the wild-type mouse, the plasma prostaglandin F2α (PGF2α) levels showed a circadian rhythm under a 12 h cycle of light–dark conditions. In addition, in vitro experiments showed that the addition of PTGFR agonists altered the amplitude of Per2::luc activity, and this alteration differed with the timing of the agonist addition. These results lead us to hypothesize that the plasma rhythm of PGF2α is important for driving clock genes, thus suggesting the involvement of PGF2α- and Ptgfr-targeting drugs in the biological clock cycle. [ABSTRACT FROM AUTHOR]

Leaf mould caused by Fulvia fulva is a major disease of tomato in Japan. Resistance to several types of fungicides, including succinate dehydrogenase inhibitors (SDHIs), has been reported in isolates from Japan. In this study, we evaluated the sensitivity of F. fulva to SDHI fungicides, SDHI fungicide efficacy in controlling sensitive and resistant isolates, and the presence of mutations in the Sdh B gene, the target site of SDHI fungicides. The minimum inhibitory concentration (MIC) of six SDHI fungicides, excluding isofetamid, ranged from 0.1 to >100 μg/mL against mycelial growth of 45 F. fulva isolates collected in Nara prefecture, Japan, whereas that of isofetamid ranged from 0.1 to only 10 μg/mL. The peak MIC distribution of fluopyram and isofetamid was 10 μg/mL, that of inpyrfluxam was 100 μg/mL and was >100 μg/mL for the remaining SDHI fungicides. In a greenhouse inoculation experiment, only isofetamid controlled the highly resistant isolates with an MIC of 10 μg/mL and the remaining fungicides had MIC >100 μg/mL. In addition, moderately and highly resistant isolates were controlled by the application of isofetamid 6 h or 7 days before inoculation or 3 days after inoculation. Mutations in the Sdh B gene were not observed among sensitive, moderately resistant and highly resistant isolates, suggesting that mutations in other Sdh genes and/or changes in efflux pump expression were involved. Our results indicate the presence of cross‐resistance and different levels of sensitivity reduction among SDHI fungicides. Isofetamid showed the lowest resistance development among the tested SDHI fungicides and is considered effective in controlling tomato leaf mould. [ABSTRACT FROM AUTHOR]

Sarcomatoid hepatocellular carcinoma (SHCC) is a rare and highly lethal subtype of HCC. The present study aimed to explore the unique markers of SHCC using whole gene expression analysis. Subsequently, gene expression analysis was performed using five sarcomatoid and seven carcinomatoid components of seven tissues from patients with SHCC. The results demonstrated a significant downregulation of polybromo 1 (PBRM1) gene expression in the sarcomatoid components. Immunohistochemical staining also indicated a decreased expression of PBRM1 in the sarcomatoid components. Moreover, gene ontology enrichment analysis revealed that most of the 336 differentially expressed genes between the sarcomatoid and carcinomatoid components were involved in functions associated with DNA replication and histone methylation, which was consistent with the loss of function of PBRM1 which encodes Switch/sucrose-non-fermentable chromatin remodeling complex protein. Therefore, the results of the present study suggested that PBRM1 may be a candidate biomarker for the evaluation of SHCC. [ABSTRACT FROM AUTHOR]

5-azacytidine (AZA), a representative DNA-demethylating drug, has been widely used to treat myelodysplastic syndromes (MDS). However, it remains unclear whether AZA's DNA demethylation of any specific gene is correlated with clinical responses to AZA. In this study, we investigated genes that could contribute to the development of evidence-based epigenetic therapeutics with AZA. A DNA microarray identified that AZA specifically upregulated the expression of 438 genes in AZA-sensitive MDS-L cells but not in AZA-resistant counterpart MDS-L/CDA cells. Of these 438 genes, the ALOX12 gene was hypermethylated in MDS-L cells but not in MDS-L/CDA cells. In addition, we further found that (1) the ALOX12 gene was hypermethylated in patients with MDS compared to healthy controls; (2) MDS classes with excess blasts showed a relatively lower expression of ALOX12 than other classes; (3) a lower expression of ALOX12 correlated with higher bone marrow blasts and a shorter survival in patients with MDS; and (4) an increased ALOX12 expression after AZA treatment was associated with a favorable response to AZA treatment. Taking these factors together, an enhanced expression of the ALOX12 gene may predict favorable therapeutic responses to AZA therapy in MDS. [ABSTRACT FROM AUTHOR]

We investigated the effects of a Tankyrase (TNKS-1/2) inhibitor on mechanical stress-induced gene expression in human chondrocytes and examined TNKS-1/2 expression in human osteoarthritis (OA) cartilage. Cells were seeded onto stretch chambers and incubated with or without a TNKS-1/2 inhibitor (XAV939) for 12 h. Uni-axial cyclic tensile strain (CTS) (0.5 Hz, 8% elongation, 30 min) was applied and the gene expression of type II collagen a1 chain (COL2A1), aggrecan (ACAN), SRY-box9 (SOX9), TNKS-1/2, a disintegrin and metalloproteinase with thrombospondin motifs-5 (ADAMTS-5), and matrix metalloproteinase-13 (MMP-13) were examined by real-time PCR. The expression of ADAMTS-5, MMP-13, nuclear translocation of nuclear factor-κB (NF-κB), and β-catenin were examined by immunocytochemistry and Western blotting. The concentration of IL-1β in the supernatant was examined by enzyme-linked immunosorbent assay (ELISA). TNKS-1/2 expression was assessed by immunohistochemistry in human OA cartilage obtained at the total knee arthroplasty. TNKS-1/2 expression was increased after CTS. The expression of anabolic factors were decreased by CTS, however, these declines were abrogated by XAV939. XAV939 suppressed the CTS-induced expression of catabolic factors, the release of IL-1β, as well as the nuclear translocation of NF-κB and β-catenin. TNKS-1/2 expression increased in mild and moderate OA cartilage. Our results demonstrated that XAV939 suppressed mechanical stress-induced expression of catabolic proteases by the inhibition of NF-κB and activation of β-catenin, indicating that TNKS-1/2 expression might be associated with OA pathogenesis. [ABSTRACT FROM AUTHOR]

Background/Objectives: Esophageal achalasia is an archetypal esophageal motility disorder characterized by abnormal peristalsis of the esophageal body and impaired lower esophageal sphincter (LES) relaxation. Methods: In this study, the mRNA expression of docking proteins 1 and 2 (DOK1 and DOK2, respectively) were analyzed and the mechanisms underlying achalasia onset were investigated. Results: DOK1 and DOK2 mRNA levels significantly increased in the LES of patients with achalasia. Moreover, significant correlations were observed between IL-1β and DOK1, IL-1β and DOK2, ATG16L1 and DOK1, and HSV1-miR-H1-3p and DOK2 expression levels. However, a correlation between ATG16L1 and DOK2 or between HSV-miR-H1-3p and DOK1 expression was not observed. In addition, a positive correlation was observed between patient age and DOK1 expression. Microarray analysis revealed a significant decrease in the expression of hsa-miR-377-3p and miR-376a-3p in the LES muscle of patients with achalasia. Conclusions: These miRNAs possessed sequences targeting DOK. The upregulation of DOK1 and DOK2 expression induces IL-1β expression in the LES of achalasia patients, which may contribute to the development of esophageal motility disorder. [ABSTRACT FROM AUTHOR]

Objective: Interferon‐λ3 (IFNλ3) is a cytokine with antiviral functions on barrier surfaces, and it is associated with disease activity in autoimmune diseases. This study assessed the clinical significance of serum IFNλ3 levels in polymyositis/dermatomyositis (PM/DM)–associated interstitial lung disease (ILD). Methods: We measured serum IFNλ3 levels in 221 patients with PM/DM‐ILD (155 in the derivation cohort, 66 in the validation cohort) and 38 controls. We evaluated factors associated with mortality risk among 79 patients with anti‐melanoma differentiation–associated gene 5 (MDA5) antibody–positive DM‐ILD. Results: Serum IFNλ3 levels at diagnosis were significantly higher in patients with PM/DM‐ILD than in healthy controls. Remarkably, serum IFNλ3 levels were specifically increased in patients with anti‐MDA5 antibody–positive DM‐ILD in both the derivation and validation cohorts. In anti‐MDA5 antibody–positive DM‐ILD, patients with high IFNλ3 levels (>120 pg/mL) had significantly lower survival rates than those with low IFNλ3 levels (≤120 pg/mL). A multivariate analysis revealed that high IFNλ3 levels, as well as old age and low Pao2, were significantly associated with poor prognoses in patients with anti‐MDA5 antibody–positive DM‐ILD. In a classification analysis of patients with anti‐MDA5 antibody–positive DM‐ILD based on age, IFNλ3 level, and Pao2, patients with old age (>53 years), high IFNλ3 levels (>120 pg/mL), and low Pao2 (<75 mm Hg) had the worst survival. In lung pathologic analyses, IFNλ3‐positive staining was observed in macrophages, airway epithelial cells, the pleural region, and intrapulmonary veins in patients with anti‐MDA5 antibody–positive DM‐ILD. Conclusion: Serum IFNλ3 is a promising biomarker for identifying patients at high risk of poor outcomes in anti‐MDA5 antibody–positive DM‐ILD. [ABSTRACT FROM AUTHOR]

Uterine leiomyosarcoma (ULMS) is a rare malignant tumor, which is aggressive, and has a poor prognosis even during its early stages. Extracellular vesicles (EVs) carry cargo, such as microRNAs (miRNAs), which are involved in intercellular communication in the tumor microenvironment and other processes. Because there are no studies on EV-related miRNAs in ULMS, we identified EV-related miRNAs in ULMS and examined their function. Small EVs (sEVs) and medium/large EVs (m/lEVs) were extracted from ULMS cells by ultracentrifugation and their basic characteristics were evaluated. Then, small RNA sequencing was done to obtain EV-related miRNA profiles. Next, miRNA expression levels in sera and tissues of ULMS patients were compared with those of myoma patients. miR-654-3p and miR-369-3p were indicated to be highly expressed in both sera and tissues of ULMS patients. These two miRNAs are also highly expressed in ULMS cell lines and ULMS-derived EVs. Some cancer-associated fibroblast (CAF) markers were increased when fibroblasts were treated with ULMS-derived EVs. Furthermore, fibroblasts took up EVs derived from ULMS as determined by confocal laser microscopy. In addition, the transfection of the two candidate miRNAs into fibroblasts significantly increased some CAF markers, particularly ACTA2. miR-654-3p and miR-369-3p are highly expressed in ULMS-derived EVs, indicating that these EV-related miRNAs induce the formation of cancer-associated fibroblasts. • Small RNA sequencing revealed uterine leiomyosarcoma-specific miRNA profiles. • Uterine leiomyosarcoma cells secrete extracellular vesicles with specific miRNAs. • The extracellular vesicles induce the features of cancer-associated fibroblasts. [ABSTRACT FROM AUTHOR]

Current differentiation protocols for human induced pluripotent stem cells (hiPSCs) produce heterogeneous cardiomyocytes (CMs). Although chamber-specific CM selection using cell surface antigens enhances biomedical applications, a cell surface marker that accurately distinguishes between hiPSC-derived atrial CMs (ACMs) and ventricular CMs (VCMs) has not yet been identified. We have developed an approach for obtaining functional hiPSC-ACMs and -VCMs based on CD151 expression. For ACM differentiation, we found that ACMs are enriched in the CD151low population and that CD151 expression is correlated with the expression of Notch4 and its ligands. Furthermore, Notch signaling inhibition followed by selecting the CD151low population during atrial differentiation leads to the highly efficient generation of ACMs as evidenced by gene expression and electrophysiology. In contrast, for VCM differentiation, VCMs exhibiting a ventricular-related gene signature and uniform action potentials are enriched in the CD151high population. Our findings enable the production of high-quality ACMs and VCMs appropriate for hiPSC-derived chamber-specific disease models and other applications. The authors demonstrate that CD151 expression distinguishes atrial from ventricular cardiomyocytes derived from induced pluripotent stem cells, while further showing the role of Notch signaling in mediating the process. [ABSTRACT FROM AUTHOR]

Mitochondria are vital organelles regulating essential cellular functions. Human mitochondrial DNA (mtDNA) consists of 37 genes, 13 of which encode mitochondrial proteins, and the remaining 24 genes encode two ribosomal RNAs and 22 transfer RNAs needed for the translation of the mtDNA‐encoded 13 proteins. However, mtDNA often impairs the expression and function of these genes due to various mutations, ultimately causing mitochondrial dysfunction. To recover from this desperate condition, developing the technology to supply all mitochondrial proteins encoded by mtDNA at once is an urgent task, but there is no established strategy for this purpose. In this study, a simple yet effective mitochondrial gene delivery system is proposed comprising an artificial peptide inspired by a transmembrane mitochondrial membrane protein. The designed mitochondria‐targeting peptides presented on the carrier surface effectively guide the encapsulated plasmid to the mitochondria, facilitating mitochondrial uptake and gene expression. The developed system successfully delivers exogenous mtDNA to mtDNA‐depleted cells and leads to simultaneous multigene expression, ultimately restoring mitochondrial functions, including the mitochondrial respiration rate. The established multiple gene expression system in each mitochondrion is a game‐changing technology that can accelerate the development of mitochondrial engineering technologies as well as clinical applications for mitochondrial diseases. [ABSTRACT FROM AUTHOR]

Background: Psychological stress is associated with changes in salivary flow and composition. However, studies to show the effect of psychological stress on the transcriptome of the salivary gland are limited. This study aims to perform a transcriptomic analysis of the submandibular gland under psychological stress using a chronic restraint stress model of rats. Methods: Sprague‐Dawley rats were divided into stress groups and control groups. Psychological stress was induced in the stress group rats by enclosing them in a plastic tube for 4 h daily over 6 weeks. RNA sequencing was performed on RNA extracted from the submandibular gland. The differentially expressed genes were identified, and the genes of interest were further validated using qRT‐PCR, immunofluorescence, and western blot. Results: A comparison between control and stress groups showed 45 differentially expressed genes. The top five altered genes in RNA sequencing data showed similar gene expression in qRT‐PCR validation. The most downregulated gene in the stress group, FosB, was a gene of interest and was further validated for its protein‐level expression using immunofluorescence and western blot. The genesets for gene ontology cellular component, molecular function, and KEGG showed that pathways related to ribosome biosynthesis and function were downregulated in the stress group compared to the control. Conclusion: Psychological stress showed transcriptomic alteration in the submandibular gland. The findings may be important in understanding stress‐related oral diseases. [ABSTRACT FROM AUTHOR]

The aim of this study was to clarify DNA methylation profiles determining the clinicopathological diversity of urothelial carcinomas. Genome-wide DNA methylation analysis was performed using the Infinium HumanMethylation450 BeadChip in 46 paired samples of non-cancerous urothelium (N) and corresponding cancerous tissue (T), and 26 samples of normal control urothelium obtained from patients without urothelial carcinomas (C). For genes of interest, correlation between DNA methylation and mRNA expression was examined using the Cancer Genome Atlas database. In addition, the role of a selected target for cancer-relevant endpoints was further examined in urothelial carcinoma cell lines. The genes showing significant differences in DNA methylation levels between papillary carcinomas and more aggressive non-papillary (nodular) carcinomas were accumulated in signaling pathways participating in cell adhesion and cytoskeletal remodeling. Five hundred ninety-six methylation sites showed differences in DNA methylation levels between papillary and nodular carcinomas. Of those sites, that were located in CpG-islands around transcription start site, 5′-untranslated region or 1st exon, 16 genes exhibited inverse correlations between DNA methylation and mRNA expression levels. Among the latter, only the KLF11 gene showed papillary T sample-specific DNA hypermethylation in comparison to C and N samples. The DNA methylation levels of KLF11 were not significantly different between T samples and N samples or T samples and C samples for patients with papillo-nodular or nodular carcinomas. Knockdown experiments using the urothelial carcinoma cell lines HT1376 and 5637, which are considered models for papillary carcinoma, revealed that KLF11 participates in altering the adhesiveness of cells to laminin-coated dishes, although cell growth was not affected. These data indicate that DNA hypermethylation of KLF11 may participate in the generation of papillary urothelial carcinomas through induction of aberrant cancer cell adhesion to the basement membrane. [Display omitted] [ABSTRACT FROM AUTHOR]

Ochratoxin A (OTA) is a renal carcinogen in rats, and repeated administration induces karyomegaly in proximal tubular epithelial cells (PTECs) of the outer stripe of the outer medulla (OSOM) before inducing proliferative lesions. To investigate whether OTA induces micronuclei (MN) in PTECs, we performed an in vitro MN assay using rat renal NRK-52E PTECs after treatment for ≤21 days, and an in vivo OSOM MN assay in rats treated with OTA, other renal carcinogens, or non-carcinogenic renal toxicants for 4 or 13 weeks. The in vitro assay revealed an increased frequency of micronucleated cells from the acceptable dose level for cell viability, even after 21 days of treatment. The in vivo assay also revealed a dose- and treatment period-dependent increase in PTECs with γ-H2AX+ MN. OTA-specific gene expression profiling by OSOM RNA sequencing after week 13 revealed the altered expression of genes related to microtubule–kinetochore binding, the kinesin superfamily, centriole assembly, DNA damage repair, and cell cycle regulation. MN formation was also observed with other renal carcinogens that induce karyomegaly similarly to OTA. These results imply that γ-H2AX+ MN formation by OTA treatment is related to the induction of chromosomal instability accompanying karyomegaly formation before proliferative lesions form, providing a new insight into the carcinogenic mechanism that may be relevant to humans. • OTA induced in vitro MN formation in rat renal tubular epithelial NRK-52E cells. • OTA induced γ-H2AX+ MN formation dose- and time-dependently in rat OSOM PTECs. • Other karyomegaly-inducing carcinogens also induced MN formation in rat OSOM PTECs. • OTA-specific gene expression profiling revealed CIN-related genes in the rat OSOM. [ABSTRACT FROM AUTHOR]

In the article "Design of an Artificial Peptide Inspired by Transmembrane Mitochondrial Protein for Escorting Exogenous DNA into the Mitochondria to Restore their Functions by Simultaneous Multiple Gene Expression," researchers Keiji Numata and his team discuss their achievement in using an artificial peptide to deliver genes to the mitochondria. This peptide-based delivery carrier selectively targets the mitochondria and allows for the efficient expression of multiple genes in cells with mitochondrial dysfunction. The study contributes to the advancement of mitochondrial engineering technologies and their potential application in treating mitochondrial diseases. [Extracted from the article]

To clarify the involvement of clock genes in the production of inflammatory mediators from RA-FLS, we examined the role of Bmal1, one of the master clock genes. RA-FLSs were stimulated with IL-1β (0, 20 ng/mL), IL-6 (0, 20 ng/mL), IL-17 (0, 20 ng/mL), TNF-α (0, 20 ng/mL) or IFN-γ (0, 20 ng/mL) to examine the expression of Bmal1, MMP-3, CCL2, IL-6, IL-7 and IL-15 by qPCR and immunofluorescence staining. After silencing Bmal1, RA-FLSs were stimulated with IL-1β (0, 20 ng/mL), TNF-α (0, 20 ng/mL) or IFN-γ (0, 20 ng/mL) to examine the expressions of inflammatory mediators; MMP-3, CCL2, IL-6 and IL-15 by qPCR, ELISA and immunofluorescence staining. Bmal1 expressions were increased by IL-1β, TNF-α and IFN-γ stimulations. Under stimulations with TNF-α, IL-1β, and IFN-γ, mRNA and protein expressions of MMP-3, CCL2 and IL-6 were suppressed by siBmal1. Results indicate that Bmal1 contributes the production of MMP-3, CCL2, and IL-6 from RA-FLS, implying Bmal1 is involved in the pathogenesis of RA by regulating the inflammation. • IL-1β, TNF-α and IFN-γ increased the expressions of Bmal1, MMP-3, CCL2, IL-6 and IL-15 in RA-FLS. • The clock gene, Bmal1 controls the expressions of MMP-3, CCL2 and IL-6 in RA-FLS. • Bmal1 might be a novel therapeutic target of RA to attenuate the inflammatory mediators such as MMP-3, CCL2 and IL-6. [ABSTRACT FROM AUTHOR]