WileyActa Medica Okayama0385-24072025A Case of Netherton Syndrome/SPINK5-Syndromic Epidermal Differentiation Disorder Evaluated by Serial Tape-Stripping: Persistent Elevation of Serine Protease Activities Despite Clinical ImprovementENShinMorizaneDepartment of Dermatology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesAnriMoritaDepartment of Dermatology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesKoSunagawaDepartment of Dermatology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesHayatoNomuraDepartment of Dermatology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesKen‐IchiHasuiDepartment of Dermatology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesHiroyukiHondaDepartment of General Medicine, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesFumioOtsukaDepartment of General Medicine, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesMamoruOuchidaDepartment of Molecular Oncology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesNo potential conflict of interest relevant to this article was reported.WileyActa Medica Okayama0385-240752102025Biologics and Small‐Molecule Therapies in Netherton Syndrome: A Comprehensive Review14831493ENShinMorizaneDepartment of Dermatology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesTomoyukiMukaiDepartment of Immunology and Molecular Genetics, Kawasaki Medical SchoolKoSunagawaDepartment of Dermatology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesKen‐ichiHasuiDepartment of Dermatology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesAnriMoritaDepartment of Dermatology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesHayatoNomuraDepartment of Dermatology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesMamoruOuchidaDepartment of Molecular Oncology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesNetherton syndrome (NS) is a rare congenital ichthyosis caused by loss-of-function mutations in the SPINK5 gene, leading to defective expression of the serine protease inhibitor LEKTI. Dysregulated epidermal protease activity results in impaired skin barrier function and chronic inflammation, accompanied by complex immune profiles. NS patients commonly show activation of the inflammatory axis, centered on IL-17 and IL-36, in the skin and blood, and show a psoriasis-like shift to Th17. Conversely, the immune profile differs depending on the clinical type, with ichthyosis linearis circumflexa type characterized by complement activation and Th2-type allergic responses, and scaly erythroderma type characterized by a type I IFN signature and Th9-type allergic responses. While symptomatic treatments such as emollients and topical corticosteroids have been the mainstay of care, recent advances have opened new therapeutic avenues involving biologic agents and oral small-molecule immunomodulators. This review provides a comprehensive overview of the current clinical landscape and future directions of biologics (e.g., dupilumab, secukinumab, ustekinumab) and small-molecule therapies (e.g., JAK inhibitors such as tofacitinib, baricitinib, and upadacitinib) in the treatment of NS. Though evidence remains limited to case reports and small series, preliminary data suggest that cytokine-targeted interventions—particularly those inhibiting IL-4, IL-13, IL-17, IL-36, and JAK pathways—may offer tangible clinical benefits. Well-designed clinical trials and mechanistic investigations are crucial to establishing their place in NS management.No potential conflict of interest relevant to this article was reported.Elsevier B.V.Acta Medica Okayama0304-41651869122025The F54L mutation of Thioredoxin shows protein instability and increased fluctuations of the catalytic center130860ENTakumiBabaLife Science Research Infrastructure Group, Advanced Photon Technology Division, RIKEN SPring-8 CenterGoUenoLife Science Research Infrastructure Group, Advanced Photon Technology Division, RIKEN SPring-8 CenterChikaOheLife Science Research Infrastructure Group, Advanced Photon Technology Division, RIKEN SPring-8 CenterShukuSajiStructural Biology Division, Japan Synchrotron Radiation Research InstituteSachikoYamamotoStructural Biology Division, Japan Synchrotron Radiation Research InstituteMasakiYamamotoLife Science Research Infrastructure Group, Advanced Photon Technology Division, RIKEN SPring-8 CenterHiroshiNakagawaMaterials Sciences Research Center, Japan Atomic Energy AgencyNobuoOkazakiNeutron Science and Technology Center, Comprehensive Research Organization for Science and Society (CROSS)MamoruOuchidaDepartment of Molecular Oncology, Faculty of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityIoriKawasaki-OhmoriSection of Developmental Physiology and Pathology, Faculty of Education, Okayama UniversityKoheiTakeshitaLife Science Research Infrastructure Group, Advanced Photon Technology Division, RIKEN SPring-8 CenterThioredoxin is a ubiquitous redox protein that acts as an electron donor via its conserved dithiol motif (C32GPC35), catalyzing dithiol–disulfide exchange to regulate the redox state of target proteins. It supports antioxidant defense via peroxiredoxins, facilitates DNA synthesis by donating electrons to ribonucleotide reductase, and regulates redox-sensitive signaling pathways, including those controlling transcription and apoptosis. Neuronal degeneration and chronic kidney disease have been observed in Txn-F54L mutant rats; however, the details of why the Txn mutation causes these phenomena remain unknown. The present study aimed to elucidate the functional and structural changes caused by the F54L mutation. The Thioredoxin-F54L showed less insulin-reducing activity and more thermosensitivity to denaturation in the body temperature range compared to the wild type. The crystal structure revealed that F54 forms hydrophobic interactions with the surrounding hydrophobic amino acids. In addition, molecular dynamics simulation predicts increased fluctuations around the F54L mutation and a tendency for the distance between residues C32 and C35 at the catalytic center to be widened. The increased distance between residues C32 and C35 of the catalytic center may affect the reducing activity of the enzyme on the substrate. The finding that Thioredoxin-F54L is prone to denaturation at normal body temperature may reduce the normally functioning Thioredoxin. These molecular characteristics of Thioredoxin-F54L may be related to brain and kidney disease development in the Txn-F54L rats.No potential conflict of interest relevant to this article was reported.Frontiers MediaActa Medica Okayama1664-3224142023“Input/output cytokines” in epidermal keratinocytes and the involvement in inflammatory skin diseases1239598ENShinMorizaneDepartment of Dermatology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesTomoyukiMukaiDepartment of Immunology and Molecular Genetics, Kawasaki Medical SchoolKoSunagawaDepartment of Dermatology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesKotaTachibanaDepartment of Dermatology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesYoshioKawakamiDepartment of Dermatology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesMamoruOuchidaDepartment of Molecular Oncology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesConsidering the role of epidermal keratinocytes, they occupy more than 90% of the epidermis, form a physical barrier, and also function as innate immune barrier. For example, epidermal keratinocytes are capable of recognizing various cytokines and pathogen-associated molecular pattern, and producing a wide variety of inflammatory cytokines, chemokines, and antimicrobial peptides. Previous basic studies have shown that the immune response of epidermal keratinocytes has a significant impact on inflammatory skin diseases. The purpose of this review is to provide foundation of knowledge on the cytokines which are recognized or produced by epidermal keratinocytes. Since a number of biologics for skin diseases have appeared, it is necessary to fully understand the relationship between epidermal keratinocytes and the cytokines. In this review, the cytokines recognized by epidermal keratinocytes are specifically introduced as "input cytokines", and the produced cytokines as "output cytokines". Furthermore, we also refer to the existence of biologics against those input and output cytokines, and the target skin diseases. These use results demonstrate how important targeted cytokines are in real skin diseases, and enhance our understanding of the cytokines.No potential conflict of interest relevant to this article was reported.Elsevier BVActa Medica Okayama0969-99611752022Thioredoxin deficiency increases oxidative stress and causes bilateral symmetrical degeneration in rat midbrain105921ENIoriOhmoriSection of Developmental Physiology and Pathology, Faculty of Education, Okayama UniversityMamoruOuchidaDepartment of Molecular Oncology, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityHirohikoImaiDepartment of Systems Science, Kyoto University Graduate School of InformaticsSaekoIshidaDivision of Animal Genetics, Laboratory Animal Research Center, Institute of Medical Science, The University of TokyoShinyaToyokuniDepartment of Pathology and Biological Responses, Nagoya University Graduate School of MedicineTomojiMashimoDivision of Animal Genetics, Laboratory Animal Research Center, Institute of Medical Science, The University of TokyoThioredoxin, encoded by Txn1, acts as a critical antioxidant in the defense against oxidative stress by regulating the dithiol/disulfide balance of interacting proteins. The role of thioredoxin in the central nervous system (CNS) is largely unknown. A phenotype-driven study of N-ethyl-N-nitrosourea-mutated rats with wild-running seizures revealed the importance of Txn1 mutations in CNS degeneration. Genetic mapping identified Txn1-F54L in the epileptic rats. The insulin-reducing activity of Txn1-F54L was approximately one-third of that of the wild-type (WT). Bilateral symmetrical vacuolar degeneration in the midbrain, mainly in the thalamus and the inferior colliculus, was observed in the Txn1-F54L rats. The lesions displayed neuronal and oligodendrocytic cell death. Neurons in Txn1-F54L rats showed morphological changes in the mitochondria. Vacuolar degeneration peaked at five weeks of age, and spontaneous repair began at seven weeks. The TUNEL assay showed that fibroblasts derived from homozygotes were susceptible to cell death under oxidative stress. In five-week-old WT rats, energy metabolism in the thalamus was significantly higher than that in the cerebral cortex. In conclusion, in juvenile rats, Txn1 seems to play an essential role in reducing oxidative stress in the midbrains with high energy metabolism.No potential conflict of interest relevant to this article was reported.Elsevier BVActa Medica Okayama0923-181110712022Aberrant serine protease activities in atopic dermatitis27ENShinMorizaneDepartment of Dermatology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesKoSunagawaDepartment of Dermatology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesHayatoNomuraDepartment of Dermatology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesMamoruOuchidaDepartment of Molecular Oncology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesAtopic dermatitis (AD) is a chronic inflammatory skin disease; the three major factors responsible for AD, i.e., epidermal barrier dysfunction, allergic inflammation, and itching, interact with each other to form a pathological condition. Excessive protease activities are characteristic abnormalities that affect the epi-dermal barrier in patients with AD. In normal skin, epidermal serine protease activities are controlled by kallikrein-related peptidases (KLKs) and their inhibitors, including lympho-epithelial Kazal-type-related inhibitor (LEKTI). In AD lesions, KLKs are excessively expressed, which results in the enhancement of epi-dermal serine protease activities and facilitates the invasion by allergens and microorganisms. In addition, some KLKs can activate protease-activated receptor 2 (PAR2) in epidermal keratinocytes and peripheral nerves, resulting in the induction of inflammation and itching. Furthermore, in AD patients with single nucleotide polymorphism (SNP) such as E420K and D386N of SPINK5 which encodes LEKTI, LEKTI function is attenuated, resulting in the activation of KLKs and easy invasion by allergens and microorganisms. Further analysis is needed to elucidate the detailed mechanism underlying the control of serine protease activities, which may lead to the development of new therapeutic and prophylactic agents for AD.(c) 2022 Japanese Society for Investigative Dermatology. Published by Elsevier B.V. All rights reserved.No potential conflict of interest relevant to this article was reported.MDPIActa Medica Okayama1422-006722232021Multifaceted Analysis of IL-23A-and/or EBI3-Including Cytokines Produced by Psoriatic Keratinocytes12659ENKotaTachibanaDepartment of Dermatology, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical ScienceNinaTangDepartment of Dermatology, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical ScienceHitoshiUrakamiDepartment of Dermatology, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical ScienceAiKajitaDepartment of Dermatology, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical ScienceMinaKobashiDepartment of Dermatology, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical ScienceHayatoNomuraDepartment of Dermatology, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical ScienceMinoriSasakuraDepartment of Dermatology, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical ScienceSatoruSugiharaDepartment of Dermatology, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical ScienceFanJiangDepartment of Cell Biology, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical ScienceNahokoTomonobuDepartment of Cell Biology, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical ScienceMasakiyoSakaguchiDepartment of Cell Biology, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical ScienceMamoruOuchidaDepartment of Molecular Oncology, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical ScienceShinMorizaneDepartment of Dermatology, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical ScienceInterleukin (IL) 23 (p19/p40) plays a critical role in the pathogenesis of psoriasis and is upregulated in psoriasis skin lesions. In clinical practice, anti-IL-23Ap19 antibodies are highly effective against psoriasis. IL-39 (p19/ Epstein-Barr virus-induced (EBI) 3), a newly discovered cytokine in 2015, shares the p19 subunit with IL-23. Anti-IL-23Ap19 antibodies may bind to IL-39; also, the cytokine may contribute to the pathogenesis of psoriasis. To investigate IL23Ap19- and/or EBI3-including cytokines in psoriatic keratinocytes, we analyzed IL-23Ap19 and EBI3 expressions in psoriasis skin lesions, using immunohistochemistry and normal human epidermal keratinocytes (NHEKs) stimulated with inflammatory cytokines, using quantitative real-time polymerase chain reaction (RT-PCR), enzyme-linked immunosorbent assay (ELISA), and liquid chromatography-electrospray tandem mass spectrometry (LC-Ms/Ms). Immunohistochemical analysis showed that IL-23Ap19 and EBI3 expressions were upregulated in the psoriasis skin lesions. In vitro, these expressions were synergistically induced by the triple combination of tumor necrosis factor (TNF)-alpha, IL-17A, and interferon (IFN)-gamma, and suppressed by dexamethasone, vitamin D3, and acitretin. In ELISA and LC-Ms/Ms analyses, keratinocyte-derived IL-23Ap19 and EBI3, but not heterodimeric forms, were detected with humanized anti-IL-23Ap19 monoclonal antibodies, tildrakizumab, and anti-EBI3 antibodies, respectively. Psoriatic keratinocytes may express IL-23Ap19 and EBI3 proteins in a monomer or homopolymer, such as homodimer or homotrimer.No potential conflict of interest relevant to this article was reported.ElsevierActa Medica Okayama0197-01861412020Scn1a and Cacna1a mutations mutually alter their original phenotypes in rats104859ENIoriOhmoriGraduate School of Education, Okayama UniversityKiyokaKobayashiDepartment of Physiology, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityMamoruOuchidaDepartment of Molecular Oncology, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityThis study aimed to examine the effects of Cacna1a mutation on the phenotype of Scn1a-associated epilepsy in rats. We used rats with an N1417H missense mutation in the Scn1a gene and others with an M251K mutation in the Cacna1a gene. Scn1a/Cacna1a double mutant rats were generated by mating both Scn1a and Cacna1a mutants. We investigated general health and the epileptic phenotype in all these genotypes. The onset threshold of hyperthermia-induced seizures was examined at 5 weeks and spontaneous seizures were monitored using video-EEG recordings from 6 to 12 weeks of age. Scn1a/Cacna1a double mutants showed significantly reduced threshold for hyperthermia-sensitive seizures onset compared with the Scn1a mutants and had absence seizures having 6–7 c/s spike-wave bursts with changes in the spike-wave pattern, whereas Cacna1a mutants had regular 6–7 c/s spike-wave bursts. In Scn1a/Cacna1a double mutants, 6–7 c/s spike-wave bursts were accompanied with eyelid myoclonia and continuously shifting generalized clonic seizures, which were not observed in either Scn1a or Cacna1a mutants. Although a curvature of the spine was observed in rats of all these genotypes, the degree of curvature was more pronounced in Scn1a/Cacna1a double mutants, followed by Cacna1a and Scn1a mutants. Our results indicate that Cacna1a and Scn1a mutations mutually alter their original phenotypes in rats. The phenotype of absence seizures with eyelid myoclonia, generalized clonic seizures, and of spine curvature in the Scn1a/Cacna1a double mutants were similar to that observed in patients with Dravet syndrome.No potential conflict of interest relevant to this article was reported.BlackwellActa Medica Okayama0013-95804932007Rasmussen encephalitis associated with SCN1A mutation521526ENIoriOhmoriDepartments of Cellular Physiology, Graduate School of Medicine,Dentistry and Pharmaceutical Sciences, Okayama UniversityMamoruOuchidaDepartments of Molecular Genetics, Graduate School of Medicine,Dentistry and Pharmaceutical Sciences, Okayama UniversityKatsuhiroKobayashiDepartments of Child Neurology, Graduate School of Medicine,Dentistry and Pharmaceutical Sciences, Okayama UniversityYoshimiJitsumoriTakushiInoueDepartments of Child Neurology, Graduate School of Medicine,Dentistry and Pharmaceutical Sciences, Okayama UniversityKenjiShimizuDepartments of Molecular Genetics, Graduate School of Medicine,Dentistry and Pharmaceutical Sciences, Okayama UniversityHidekiMatsuiDepartments of Cellular Physiology, Graduate School of Medicine,Dentistry and Pharmaceutical Sciences, Okayama UniversityYokoOhtsukaDepartments of Child Neurology, Graduate School of Medicine,Dentistry and Pharmaceutical Sciences, Okayama UniversityYoshihiroMaegakiDivision of Child Neurology, Institute of Neurological Sciences, Faculty of Medicine, Tottori University Mutations in the SCN 1 A gene, encoding the neuronal voltage-gated sodium channel alpha1 subunit, cause SMEI, GEFS+, and related epileptic syndromes. We herein report the R1575C-SCN 1 A mutation identified in a patient with Rasmussen encephalitis. R1575C were constructed in a recombinant human SCN 1 A and then heterologously expressed in HEK293 cells along with the human beta1 and beta2 sodium channel accessory subunits. Whole-cell patch-clamp recording was used to define biophysical properties. The R1575C channels exhibited increased channel availability and an increased persistent sodium current in comparison to the wild-type. These defects of electrophysiological properties can result in neuronal hyperexitability. The seizure susceptibility allele may influence the pathogenesis of Rasmussen encephalitis in this case.No potential conflict of interest relevant to this article was reported.Elsevier ScienceActa Medica Okayama092012111051-22013Inhalation of 10% carbon dioxide rapidly terminates Scn1a mutation-related hyperthermia-induced seizures220224ENIoriOhmoriDepartment of Physiology, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityKeiichiroHayashiDepartment of Physiology, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityHaijiaoWangDepartment of Physiology, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityMamoruOuchidaDepartment of Molecular Genetics, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityNaohiroFujitaDepartment of Physiology, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityTakushiInoueDepartment of Child Neurology, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityHiroyukiMichiueDepartment of Physiology, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityTeiichiNishikiDepartment of Physiology, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityHidekiMatsuiDepartment of Physiology, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama University The aim of this study was to assess the anticonvulsant effect of carbon dioxide (CO2) on Scn1a mutation-related febrile seizures. We examined physiological changes in the blood gas levels after the induction of hyperthermia-induced seizures (HISs), which were associated with the Scn1a missense mutation. We determined the efficacy of inhalation of 5% or 10% CO2 to treat HISs. HISs were evoked in Scn1a mutant and wild-type (WT) rats by hot water baths. To determine the anticonvulsant effect of CO2 inhalation, rats were placed in a chamber filled with air or mixed gas containing 5% CO2 or 10% CO2 for 3 min, immediately after the induction of HISs. We also analyzed the blood gas levels at the end of inhalation of CO2. Hot water bathing induced a significant reduction in the partial pressure of CO2 (pCO2) and respiratory alkalosis in the WT and Scn1a mutant rats. HISs were evoked in 100% of the Scn1a mutant rats within 5 min, but in none of the WT rats. The Scn1a mutant rats demonstrated a higher HISs susceptibility associated with respiratory alkalosis than the WT rats. Inhalation of 10% CO2 shortened the seizure duration from 62.6±12.1 s to 15.5±1.0 s. Blood gas analysis after the inhalation of 10% CO2 demonstrated an elevated pCO2 level and respiratory acidosis. Inhalation of 10% CO2 demonstrated a potent and fast-acting anticonvulsant effect against HISs.No potential conflict of interest relevant to this article was reported.Academic PressActa Medica Okayama09699961502013CACNA1A variants may modify the epileptic phenotype of Dravet syndrome209217ENIoriOhmoriDepartment of Physiology, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityMamoruOuchidaDepartment of Molecular Genetics, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityKatsuhiroKobayashiYoshimiJitsumoriAkikoMoriDepartment of Physiology, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityHiroyukiMichiueDepartment of Physiology, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityTeiichiNishikiDepartment of Physiology, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama UniversityYokoOhtsukaHidekiMatsuiDepartment of Physiology, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama University Dravet syndrome is an intractable epileptic syndrome beginning in the first year of life. De novo mutations of SCN1A, which encode the Na(v)1.1 neuronal voltage-gated sodium channel, are considered the major cause of Dravet syndrome. In this study, we investigated genetic modifiers of this syndrome. We performed a mutational analysis of all coding exons of CACNA1A in 48 subjects with Dravet syndrome. To assess the effects of CACNA1A variants on the epileptic phenotypes of Dravet syndrome, we compared clinical features in two genotype groups: 1) subjects harboring SCN1A mutations but no CACNA1A variants (n=20) and 2) subjects with SCN1A mutations plus CACNA1A variants (n=20). CACNA1A variants detected in patients were studied using heterologous expression of recombinant human Ca(v)2.1 in HEK 293 cells and whole-cell patch-clamp recording. Nine CACNA1A variants, including six novel ones, were detected in 21 of the 48 subjects (43.8%). Based on the incidence of variants in healthy controls, most of the variants seemed to be common polymorphisms. However, the subjects harboring SCN1A mutations and CACNA1A variants had absence seizures more frequently than the patients with only SCN1A mutations (8/20 vs. 0/20, p=0.002). Moreover, the former group of subjects exhibited earlier onset of seizures and more frequent prolonged seizures before one year of age, compared to the latter group of subjects. The electrophysiological properties of four of the five novel Ca(v)2.1 variants exhibited biophysical changes consistent with gain-of-function. We conclude that CACNA1A variants in some persons with Dravet syndrome may modify the epileptic phenotypes.No potential conflict of interest relevant to this article was reported.Okayama University Medical SchoolActa Medica Okayama0386-300X7112017Human RAD 17 Polymorphism at Codon 546 Is Associated with the Risk of Colorectal Cancer5968ENYukikoYasudaDepartment of Molecular Oncology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesAkikoSakaiDepartment of Molecular Oncology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesSachioItoDepartment of Molecular Oncology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesKaoriSasaiDepartment of Molecular Oncology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesHiromasaYamamotoDepartment of General Thoracic Surgery and Breast and Endocrinological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesNagahideMatsubaraDepartment of Gastroenterological Surgery, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesMamoruOuchidaDepartment of Molecular Oncology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesHiroshiKatayamaDepartment of Molecular Oncology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesKenjiShimizuDepartment of Molecular Oncology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical SciencesOriginal Article10.18926/AMO/54826Human RAD17 acts as an activator of checkpoint signals in response to DNA damage. Here we evaluated the association of hRAD17 Leu546Arg (rs1045051), a missense single nucleotide polymorphism, with the risk of colorectal cancer (CRC) in relation to smoking and alcohol consumption habits in 212 CRC patients and 1,142 cancer-free controls in a case-control study conducted in Japan. The results showed that the hRAD17 Leu/Arg genotype compared to the Leu/Leu genotypes was significantly associated with the protective effect on CRC risk with the adjusted odds ratio (OR) of 0.68 [95% confidence interval (CI): 0.49−0.95, p=0.024], and the males with the Arg/Arg genotype had a greater risk of CRC compared to those with the Leu/Leu and Leu/Arg genotypes (OR=1.87, 95%CI 1.03−3.40, p=0.04). In stratified studies, the protective effect of the Leu/Arg genotype on CRC risk was markedly higher in the light smokers (< 20 pack years) (OR=0.61, 95%CI 0.40−0.94, p=0.024) and the rectal cancer patients (OR=0.49, 95%CI 0.31−0.78, p=0.003). The risk of the Arg/Arg genotype was associated with heavy smoking (≥ 20 pack-years) (OR=2.24, 95%CI 1.09−4.61, p=0.03). These findings suggest that the genetic variant of hRAD17 Leu546Arg polymorphism has a significant effect on CRC susceptibility in Japanese.No potential conflict of interest relevant to this article was reported.Acta Medica Okayama0013-95805252011Therapy for hyperthermia-induced seizures in Scn1a mutant rats10101017ENKeiichiroHayashiSatoshiUeshimaMamoruOuchidaTomojiMashimoTeiichiNishikiToshiakiSendoTadaoSerikawaHidekiMatsuiIoriOhmoriPurpose: Mutations in the SCN1A gene, which encodes the alpha 1 subunit of voltage-gated sodium channels, cause generalized epilepsy with febrile seizures plus (GEFS+) and severe myoclonic epilepsy of infancy (SMEI). N1417H-Scn1a mutant rats are considered to be an animal model of human FS+ or GEFS+. To assess the pharmacologic validity of this model, we compared the efficacies of eight different antiepileptic drugs (AEDs) for the treatment of hyperthermia-induced seizures using N1417H-Scn1a mutant rats.
Methods: AEDs used in this study included valproate, carbamazepine (CBZ), phenobarbital, gabapentin, acetazolamide, diazepam (DZP), topiramate, and potassium bromide (KBr). The effects of these AEDs were evaluated using the hot water model, which is a model of experimental FS. Five-week-old rats were pretreated with each AED and immersed in water at 45 degrees C to induce hyperthermia-induced seizures. The seizure manifestations and video-electroencephalographic recordings were evaluated. Furthermore, the effects of each AED on motor coordination and balance were assessed using the balance-beam test.
Key Findings: KBr significantly reduced seizure durations, and its anticonvulsant effects were comparable to those of DZP. On the other hand, CBZ decreased the seizure threshold. In addition, DZP and not KBr showed significant impairment in motor coordination and balance.
Significance: DZP and KBr showed potent inhibitory effects against hyperthermia-induced seizures in the Scn1a mutant rats, whereas CBZ exhibited adverse effects. These responses to hyperthermia-induced seizures were similar to those in patients with GEFS+ and SMEI. N1417H-Scn1a mutant rats may, therefore, be useful for testing the efficacy of new AEDs against FS in GEFS+ and SMEI patients.No potential conflict of interest relevant to this article was reported.Public Library ScienceActa Medica Okayama1932-62038102013Truncated SSX Protein Suppresses Synovial Sarcoma Cell Proliferation by Inhibiting the Localization of SS18-SSX Fusion ProteinENYasushiYonedaSachioItoToshiyukiKunisadaYukiMorimotoHirotakaKanzakiAkiYoshidaKenjiShimizuToshifumiOzakiMamoruOuchidaSynovial sarcoma is a relatively rare high-grade soft tissue sarcoma that often develops in the limbs of young people and induces the lung and the lymph node metastasis resulting in poor prognosis. In patients with synovial sarcoma, specific chromosomal translocation of t(X; 18) (p11.2; q11.2) is observed, and SS18-SSX fusion protein expressed by this translocation is reported to be associated with pathogenesis. However, role of the fusion protein in the pathogenesis of synovial sarcoma has not yet been completely clarified. In this study, we focused on the localization patterns of SS18-SSX fusion protein. We constructed expression plasmids coding for the full length SS18-SSX, the truncated SS18 moiety (tSS18) and the truncated SSX moiety (tSSX) of SS18-SSX, tagged with fluorescent proteins. These plasmids were transfected in synovial sarcoma SYO-1 cells and we observed the expression of these proteins using a fluorescence microscope. The SS18-SSX fusion protein showed a characteristic speckle pattern in the nucleus. However, when SS18-SSX was co-expressed with tSSX, localization of SS18-SSX changed from speckle patterns to the diffused pattern similar to the localization pattern of tSSX and SSX. Furthermore, cell proliferation and colony formation of synovial sarcoma SYO-1 and YaFuSS cells were suppressed by exogenous tSSX expression. Our results suggest that the characteristic speckle localization pattern of SS18-SSX is strongly involved in the tumorigenesis through the SSX moiety of the SS18-SSX fusion protein. These findings could be applied to further understand the pathogenic mechanisms, and towards the development of molecular targeting approach for synovial sarcoma.No potential conflict of interest relevant to this article was reported.Wiley-BlackwellActa Medica Okayama0919-65443332013Proteomics-based analysis of invasion-related proteins in malignant gliomas264275ENTomokoMaruoTomotsuguIchikawaHirotakaKanzakiSatoshiInoueKazuhikoKurozumiManabuOnishiKoichiYoshidaHirokazuKambaraMamoruOuchidaKenjiShimizuSeijiTamaruE. AntonioChioccaIsaoDateOne of the insidious biological features of gliomas is their potential to extensively invade normal brain tissue, yet molecular mechanisms that dictate this locally invasive behavior remain poorly understood. To investigate the molecular basis of invasion by malignant gliomas, proteomic analysis was performed using a pair of canine glioma subclones - J3T-1 and J3T-2 - that show different invasion phenotypes in rat brains but have similar genetic backgrounds. Two-dimensional protein electrophoresis of whole-cell lysates of J3T-1 (angiogenesis-dependent invasion phenotype) and J3T-2 (angiogenesis-independent invasion phenotype) was performed. Twenty-two distinct spots were recognized when significant alteration was defined as more than 1.5-fold change in spot intensity between J3T-1 and J3T-2. Four proteins that demonstrated increased expression in J3T-1, and 14 proteins that demonstrated increased expression in J3T-2 were identified using liquid chromatography-mass spectrometry analysis. One of the proteins identified was annexin A2, which was expressed at higher levels in J3T-1 than in J3T-2. The higher expression of annexin A2 in J3T-1 was corroborated by quantitative RT-PCR of the cultured cells and immunohistochemical staining of the rat brain tumors. Moreover, immunohistochemical analysis of human glioblastoma specimens showed that annexin A2 was expressed at high levels in the tumor cells that formed clusters around dilated vessels. These results reveal differences in the proteomic profiles between these two cell lines that might correlate with their different invasion profiles. Thus, annexin A2 may be related to angiogenesis-dependent invasion.No potential conflict of interest relevant to this article was reported.Okayama University Medical SchoolActa Medica Okayama0386-300X6662012Eradication of Hepatitis C Virus Subgenomic Replicon by Interferon Results in Aberrant Retinol-Related Protein Expression461468ENKazukoKoikeAkinobuTakakiNobuyukiKatoMamoruOuchidaHirotakaKanzakiTetsuyaYasunakaHidenoriShirahaYasuhiroMiyakeKazuhideYamamotoOriginal Article10.18926/AMO/49042Hepatitis C virus (HCV) infection induces several changes in hepatocytes, such as oxidative stress, steatosis, and hepatocarcinogenesis. Although considerable progress has been made during recent years, the mechanisms underlying these functions remain unclear. We employed proteomic techniques in HCV replicon-harboring cells to determine the effects of HCV replication on host-cell protein expression. We examined two-dimensional electrophoresis (2-DE) and mass spectrometry to compare and identify differentially expressed proteins between HCV subgenomic replicon-harboring cells and their “cured” cells. One of the identified proteins was confirmed using enzyme-linked immunosorbent assay (ELISA) and Western blot analysis. Full-length HCV genome RNA replicating and cured cells were also assessed using ELISA. Replicon-harboring cells showed higher expression of retinal dehydrogenase 1 (RALDH-1), which converts retinol to retinoic acid, and the cured cells showed higher expression of retinol-binding protein (RBP), which transports retinol from the liver to target tissues. The alteration in RBP expression was also confirmed by ELISA and Western blot analysis. We conclude that protein expression profiling demonstrated that HCV replicon eradication affected retinol-related protein expression.No potential conflict of interest relevant to this article was reported.Okayama University Medical SchoolActa Medica Okayama0386-300X6632012Antinociceptive Effects of Intrathecal Landiolol Injection in a Rat Formalin Pain Model285289ENSatoshiMizobuchiYoshikazuMatsuokaNorihikoObataRyujiKakuYoshitaroItanoNaotoTomotsukaArataTaniguchiHiroyukiNishieHirotakaKanzakiMamoruOuchidaKiyoshiMoritaOriginal Article10.18926/AMO/48569Perioperative beta-blocker administration has recently been recommended for patients undergoing cardiac or other surgery due to the beneficial cardiovascular effects of these agents. In addition, some studies have reported that perioperatively administered beta-blockers also have analgesic effects. In this study, to investigate the antinociceptive effects and the analgesic profile of landiolol, we examined the effects of intrathecal landiolol administration on nociceptive pain behavior and c-fos mRNA expression (a neural marker of pain) in the spinal cord using a rat formalin model. We found that pain-related behavior was inhibited by intrathecal landiolol administration. Moreover, the increase in c-fos mRNA expression on the formalin-injected side was less pronounced in rats administered landiolol
than in saline administered controls. Thus, intrathecal administration of landiolol exhibited antinociceptive
effects. Further investigation of the antinociceptive mechanism of landiolol is required.No potential conflict of interest relevant to this article was reported.Okayama University Medical SchoolActa Medica Okayama0386-300X6622012Different Responses to 5-fluoraouracil in Mutagenicity and Gene Expression between Two Human Lymphoblastoid Cell Lines with or without TP53 Mutation119129ENHiroakiOkaMamoruOuchidaTakuyaKondoFumioMoritaKenjiShimizuOriginal Article10.18926/AMO/48262Human lymphoblastoid TK6 and WTK-1 cells are widely used to detect mutagens in vitro. TK6 cells have wild-type TP53 alleles, while WTK-1 cells have one allele of mutated TP53. Both cells were treated with 5-fluorouracil (5-FU), and gene mutation assay and micronucleus assay were performed to clarify the differential response related to the TP53 gene status. The effects of 5-FU on gene expression were assessed by microarray and quantitative RT-PCR analyses. In WTK-1 cells, 5-FU increased the frequency of cells with micronucleus and mutation. In TK6 cells, frequency of cells with micronucleus was increased but the mutation frequency was not. The cytotoxicity induced by 5-FU was more prominent in TK6 cells than in WTK-1 cells. Analysis of gene expression showed that the genes involved in the TP53 pathway were up-regulated in TK6 cells but not in WTK-1 cells. The differential
responses to 5-FU between these cell lines appeared to be due to the difference in the TP53 gene status, thus providing a molecular basis for the bioassays using these cell lines in the toxicology field. Our results indicate that the clinical efficacy of 5-FU chemotherapy may depend on the TP53 genotype.No potential conflict of interest relevant to this article was reported.Wiley-Liss, Inc.Acta Medica Okayama0020-713611212004Tumor-specific exon creation of the HELLS/SMARCA6 gene in non-small cell lung cancer813ENMasaakiYanoMamoruOuchidaHisayukiShigematsuNoriyoshiTanakaKoichiIchimuraKazuyasuKobayashiYasuhikoInakiShinichiToyookaKazunoriTsukudaNobuyoshiShimizuKenjiShimizuIn an attempt to identify tumor suppressor genes on chromosome 10 in non-small cell lung
cancers, we isolated 10 types of splicing variants of the HELLS/ SMARCA6 gene transcripts.
HELLS/SMARCA6 is a novel member of SNF2 family, which is implicated in cellular function like chromatin remodeling. Variant 1 was an alternatively spliced isoform containing
an insertion of a 44-ntd intronic sequence between exons 3 and 4, giving rise to a premature
termination of translation. The expression of the variant 1 was detected exclusively in the
lung cancer specimens (11 of 43 cases, 26%), but was not detected in corresponding normal tissues. D10S520 marker in the proximity of the HELLS/SMARCA6 gene showed prevalent allelic loss (41%) as compared with flanking markers (25-31%). These results suggest that loss of function of HELLS/SMARCA6 by allelic loss and aberrant proteins by tumor-specific exon creation may result in epigenetic deregulation, leading the lung cells to malignancy or its progression.No potential conflict of interest relevant to this article was reported.Elsevier Ireland Ltd.Acta Medica Okayama0304-383522422005Significant growth suppression of synovial sarcomas by the histone deacetylase inhibitor FK228 in vitro and in vivo311319ENTatsuoItoMamoruOuchidaYukiMorimotoAkiYoshidaYoshimiJitsumoriToshifumiOzakiHiroshiSonobeHajimeInoueKenjiShimizuAbout 97% of synovial sarcomas harbor the SYT-SSX fusion gene by chromosomal
translocation. We found that the histone deacetylase (HDAC) inhibitor FK228
significantly suppressed the growth of synovial sarcoma cells as compared with that of
osteosarcoma. The 50% growth inhibition IC50 value we obtained for FK228 was 0.02-0.2 nM, and it indicates that its suppression effect on synovial sarcoma cells is the highest of any of the HDAC inhibitors yet reported. It was not likely that the growth suppression of FK228 depends on the doubling time of these cells. Introduction of SYT-SSX cDNA into HEK293 cells enhanced the sensitivity of the cells for FK228.
Immunostaining of the FK228-treated cells using an anti-acetyl-histone H3 antibody showed that FK228 inhibits deacetylation of histone. In a mice assay, the growth of synovial sarcoma cells was markedly inhibited by FK228 treatment, and the invasion of tumors into surrounding tissues was suppressed. These results suggest that FK228 may be useful in developing therapeutic strategies to treat synovial sarcoma.No potential conflict of interest relevant to this article was reported.岡山医学会Acta Medica Okayama0030-155812132009Dravet 症候群患者に認められたカルシウムチャネル 機能異常を引き起こす CACNB4 遺伝子変異149156ENIoriOhmoriMamoruOuchidaNobuyoshiMimakiTeiichiNishikiKazuhitoTomizawaHidekiMatsuiNo potential conflict of interest relevant to this article was reported.岡山医学会Acta Medica Okayama0030-155812032008EGFR 変異と Uracil-Tegafur による肺腺癌術後補助療法の関連性についての検討265269ENHiroshiSuehisaShinichiToyookaKatsuyukiHottaAkikoUchidaJunichiSohYoshiroFujiwaraKeitaroMatsuoMamoruOuchidaMinoruTakataKatsuyukiKiuraHiroshiDateNo potential conflict of interest relevant to this article was reported.岡山医学会Acta Medica Okayama0030155811922007食道扁平上皮癌の発癌過程におけるDNAメチル化の役割113117ENTatsuhiroIshiiJunMurakamiKenjiNotoharaHiromiSasamotoTakeshiKambaraYasuhiroShirakawaYoshioNaomotoMamoruOuchidaKenjiShimizuR. JassJeremyNagahideMatsubaraNoriakiTanakaNo potential conflict of interest relevant to this article was reported.