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Article: Epstein-Barr virus-derived vector suitable for long-term expression in neurons.

Kiyosue, Kazuyuki / Miwa, Yoshihiro

2020 Volume 6, Issue 3, Page(s) e03504

Abstract: Exogenous gene expression is a fundamental and indispensable technique for testing gene function in neurons. Several ways to express exogenous genes in neurons are available, but each method has pros and cons. The lentivirus vector is useful for high ... ...

Abstract	Exogenous gene expression is a fundamental and indispensable technique for testing gene function in neurons. Several ways to express exogenous genes in neurons are available, but each method has pros and cons. The lentivirus vector is useful for high efficiency gene transfer to neurons and stabilizes gene expression via genome integration, but this integration may destroy the host genome. The Epstein-Barr virus (EBV)-derived vector (EB vector) is an accessible and useful vector in human cell lines because the vector is not integrated into the host genome but stays in the nucleus as an episome. However, there has been no report on this process in rodent neurons. We examined the usefulness of the EB vector for testing gene function in neurons. We found that EB vector-derived exogenous proteins such as green fluorescent protein (GFP) and GFP-tagged actin were easily detectable even after three weeks of transfection. Second, a tetracycline-induced gene expression system in the EB vector was active after three weeks of transfection, indicating that plasmids were retained in neurons for up to three weeks. Third, we determined that only Family of repeat element of the plasmid vector is essential for its long-term presence in neurons. These results show that the modified EB vector is a useful tool for examining gene function in neurons.
Language	English
Publishing date	2020-03-11
Publishing country	England
Document type	Journal Article
ZDB-ID	2835763-2
ISSN	2405-8440
ISSN	2405-8440
DOI	10.1016/j.heliyon.2020.e03504
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Article ; Online: Epstein-Barr virus-derived vector suitable for long-term expression in neurons

Kiyosue, Kazuyuki / Miwa, Yoshihiro

Heliyon. 2020 Mar., v. 6, no. 3 p.e03504-

2020

Abstract	Exogenous gene expression is a fundamental and indispensable technique for testing gene function in neurons. Several ways to express exogenous genes in neurons are available, but each method has pros and cons. The lentivirus vector is useful for high efficiency gene transfer to neurons and stabilizes gene expression via genome integration, but this integration may destroy the host genome. The Epstein-Barr virus (EBV)-derived vector (EB vector) is an accessible and useful vector in human cell lines because the vector is not integrated into the host genome but stays in the nucleus as an episome. However, there has been no report on this process in rodent neurons. We examined the usefulness of the EB vector for testing gene function in neurons. We found that EB vector-derived exogenous proteins such as green fluorescent protein (GFP) and GFP-tagged actin were easily detectable even after three weeks of transfection. Second, a tetracycline-induced gene expression system in the EB vector was active after three weeks of transfection, indicating that plasmids were retained in neurons for up to three weeks. Third, we determined that only Family of repeat element of the plasmid vector is essential for its long-term presence in neurons. These results show that the modified EB vector is a useful tool for examining gene function in neurons.
Keywords	Human gammaherpesvirus 4 ; actin ; gene expression ; genes ; green fluorescent protein ; humans ; plasmid vectors ; plasmids ; rodents ; transfection ; Cell biology ; Neuroscience ; Neurology ; Nervous system ; Neuronal death ; Electrophysiology ; Plasmid vector ; Episome ; Postmitotic cell ; Long-term exogenous gene expression ; Epstein-barr virus based vector
Language	English
Dates of publication	2020-03
Publishing place	Elsevier Ltd
Document type	Article ; Online
Note	Use and reproduction
ZDB-ID	2835763-2
ISSN	2405-8440
ISSN	2405-8440
DOI	10.1016/j.heliyon.2020.e03504
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Article ; Online: Efficient selection of knocked-in pluripotent stem cells using a dual cassette cellular elimination system.

Nakade, Koji / Tsukamoto, Satomi / Nakashima, Kenichi / An, Yuri / Sato, Iori / Li, Jingyue / Shimoda, Yuzuno / Hemmi, Yasuko / Miwa, Yoshihiro / Hayashi, Yohei

Cell reports methods

2023 Volume 3, Issue 12, Page(s) 100662

Abstract: Although recent advances in genome editing technology with homology-directed repair have enabled the insertion of various reporter genes into the genome of mammalian cells, the efficiency is still low due to the random insertion of donor vectors into the ...

Abstract	Although recent advances in genome editing technology with homology-directed repair have enabled the insertion of various reporter genes into the genome of mammalian cells, the efficiency is still low due to the random insertion of donor vectors into the host genome. To efficiently select knocked-in cells without random insertion, we developed the "double-tk donor vector system," in which the expression units of the thymidine kinase of herpes simplex virus (HSV-tk) are placed on both outer sides of homology arms. This system is superior in enriching knocked-in human induced pluripotent stem cells (hiPSCs) than conventional donor vector systems with a single or no HSV-tk cassette. Using this system, we efficiently generated fluorescent reporter knockin hiPSCs targeting POU5F1 (OCT3/4), EEF1A1, H2BC21 (H2B clustered histone 21), ISL1, and MYH7 genes. These results indicate that the double-tk donor vector system enables efficient selection of knocked-in hiPSCs carrying reporter proteins.
MeSH term(s)	Animals ; Humans ; Induced Pluripotent Stem Cells/metabolism ; Simplexvirus ; Gene Editing ; Genes, Homeobox ; Mammals
Language	English
Publishing date	2023-12-11
Publishing country	United States
Document type	Journal Article
ISSN	2667-2375
ISSN (online)	2667-2375
DOI	10.1016/j.crmeth.2023.100662
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Article ; Online: Retrospective Analysis of the Risk Factors for Grade IV Neutropenia in Oesophageal Cancer Patients Treated with a Docetaxel, Cisplatin, and 5-Fluorouracil Regimen.

Naito, Masahito / Yamamoto, Tomoya / Shimamoto, Chikao / Miwa, Yoshihiro

Chemotherapy

2017 Volume 62, Issue 4, Page(s) 215–224

Abstract: Background: Previous Japanese trials of the docetaxel, cisplatin, and 5-fluorouracil regimen for oesophageal cancer have demonstrated that a large proportion of patients also develop grade IV neutropenia. Our aim was to examine the risk factors for ... ...

Abstract	Background: Previous Japanese trials of the docetaxel, cisplatin, and 5-fluorouracil regimen for oesophageal cancer have demonstrated that a large proportion of patients also develop grade IV neutropenia. Our aim was to examine the risk factors for neutropenia in patients treated with this regimen. Methods: We retrospectively analysed the risk factors for developing grade IV neutropenia in 66 patients with oesophageal cancer using a multivariate analysis. Results: After administering the docetaxel, cisplatin, and 5-fluorouracil regimen, 49 patients (74.2%) developed grade IV neutropenia. Grade IV neutropenia was significantly associated with platelet count (p < 0.01), alanine transaminase level (p = 0.05), and proton-pump inhibitor administration (p < 0.05). Receiver operating characteristic curve analysis confirmed a platelet count of 290 × 103/μL as the optimal diagnostic cut-off value for grade IV neutropenia. The receiver operating characteristic area for grade IV neutropenia was increased by including patients that were administered a proton-pump inhibitor and alanine transaminase level (updated model; sensitivity and specificity, 75.5 and 88.2%, respectively). Conclusions: Our findings suggest that a platelet count is the most significant predictor of grade IV neutropenia.
MeSH term(s)	Aged ; Alanine Transaminase/blood ; Antineoplastic Agents/adverse effects ; Antineoplastic Agents/therapeutic use ; Area Under Curve ; Blood Platelets/cytology ; Cisplatin/adverse effects ; Cisplatin/therapeutic use ; Drug Administration Schedule ; Esophageal Neoplasms/drug therapy ; Esophageal Neoplasms/pathology ; Female ; Fluorouracil/adverse effects ; Fluorouracil/therapeutic use ; Humans ; Male ; Middle Aged ; Neutropenia/etiology ; Platelet Count ; Proton Pump Inhibitors/administration & dosage ; ROC Curve ; Retrospective Studies ; Risk Factors ; Taxoids/adverse effects ; Taxoids/therapeutic use
Chemical Substances	Antineoplastic Agents ; Proton Pump Inhibitors ; Taxoids ; docetaxel (15H5577CQD) ; Alanine Transaminase (EC 2.6.1.2) ; Cisplatin (Q20Q21Q62J) ; Fluorouracil (U3P01618RT)
Language	English
Publishing date	2017
Publishing country	Switzerland
Document type	Journal Article
ZDB-ID	6708-8
ISSN	1421-9794 ; 0009-3157
ISSN (online)	1421-9794
ISSN	0009-3157
DOI	10.1159/000464273
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Article ; Online: Multicomponent high-performance liquid chromatography/tandem mass spectrometry analysis of ten chemotherapeutic drugs in wipe samples.

Maeda, Shinichiro / Miwa, Yoshihiro

Journal of chromatography. B, Analytical technologies in the biomedical and life sciences

2013 Volume 921-922, Page(s) 43–48

Abstract: Progress in chemotherapy leads to increased numbers and variety of chemotherapeutic drugs, and multicomponent analysis of these drugs is a necessary step. We used liquid chromatography-tandem mass spectrometry and developed a multicomponent analysis of ... ...

Abstract	Progress in chemotherapy leads to increased numbers and variety of chemotherapeutic drugs, and multicomponent analysis of these drugs is a necessary step. We used liquid chromatography-tandem mass spectrometry and developed a multicomponent analysis of ten drugs used in chemotherapy: vindesine, vincristine, vinblastine, doxorubicin, epirubicin, ifosfamide, cyclophosphamide, irinotecan, docetaxel, and paclitaxel. We selected five internal standards for each category of drug, because the ionization efficiencies of product ions varied widely. The total run time was 22min, applying a gradient elution of water and acetonitrile in the presence of 0.1% formic acid. The lower limit of quantification was 50ng/wipe samples for vindesine, vincristine, and vinblastine, and 5ng/wipe samples for the remaining seven drugs. Accuracy (88.6-112.9%, 85.2-111.7%) and precision (1.0-11.5%CV, 3.6-14.4%CV) in within-run and between-run assays of QC solutions were acceptable. Without outliers, in within-run and between-run assays of QC samples, accuracy was 90.6-113.9% and 91.1-130.4%, respectively, and precision was 2.2-19.0%CV and 4.8-14.9%CV, respectively. Accuracy and precision of High QC samples of irinotecan were deviated. Our analysis procedure has sufficient sensitivity and is convenient enough for regular monitoring.
MeSH term(s)	Antineoplastic Agents/analysis ; Antineoplastic Agents/isolation & purification ; Chromatography, High Pressure Liquid/methods ; Environmental Exposure/analysis ; Environmental Monitoring/methods ; Humans ; Sensitivity and Specificity ; Tandem Mass Spectrometry/methods
Chemical Substances	Antineoplastic Agents
Language	English
Publishing date	2013-03-15
Publishing country	Netherlands
Document type	Journal Article ; Research Support, Non-U.S. Gov't ; Validation Studies
ZDB-ID	1180823-8
ISSN	1873-376X ; 0378-4347 ; 1570-0232 ; 1387-2273
ISSN (online)	1873-376X
ISSN	0378-4347 ; 1570-0232 ; 1387-2273
DOI	10.1016/j.jchromb.2013.01.014
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Article ; Online: Hemoglobin Value Is the Most Important Factor in the Development of Hand-Foot Syndrome under the Capecitabine Regimen.

Naito, Masahito / Yamamoto, Tomoya / Hara, Shinsuke / Shimamoto, Chikao / Miwa, Yoshihiro

Chemotherapy

2017 Volume 62, Issue 1, Page(s) 23–29

Abstract: Background: Hand-foot syndrome (HFS) is a common side effect that has a high occurrence rate with capecitabine (Cape) chemotherapy. However, little is known about the risk factors of developing HFS under the Cape regimen. Our aim was to examine these ... ...

Abstract	Background: Hand-foot syndrome (HFS) is a common side effect that has a high occurrence rate with capecitabine (Cape) chemotherapy. However, little is known about the risk factors of developing HFS under the Cape regimen. Our aim was to examine these risk factors. Methods: A univariate analysis was used to determine the risk factors associated with developing HFS, and we calculated the effect sizes between the patients who developed HFS compared to those who did not. Results: Of the 52 patients enrolled in our research, 24 (46.2%) developed HFS. This group was significantly associated with hemoglobin (Hb) values (p < 0.001), and the effect size (1.21) was more than moderate. The receiver operating characteristic curve analysis confirmed 12 mg/dl Hb as the best diagnostic cut-off value for developing HFS. The sensitivity and specificity were 75.5 and 88.2%, respectively. Patients who had Hb values of 12 or below who developed HFS had longer median times without HFS compared to patients with high Hb values (115 vs. 75 days, p = 0.30, hazard ratio = 1.42, 95% CI 0.73-2.76) and a greater area under the Kaplan-Meier curves (p < 0.05). Conclusion: This research suggests that the Hb value is an important factor for developing HFS.
MeSH term(s)	Antimetabolites, Antineoplastic/adverse effects ; Antimetabolites, Antineoplastic/therapeutic use ; Capecitabine/adverse effects ; Capecitabine/therapeutic use ; Female ; Follow-Up Studies ; Hand-Foot Syndrome/etiology ; Hemoglobins/analysis ; Humans ; Kaplan-Meier Estimate ; Male ; Middle Aged ; Neoplasms/drug therapy ; Neoplasms/mortality ; Odds Ratio ; Proportional Hazards Models ; Risk Factors
Chemical Substances	Antimetabolites, Antineoplastic ; Hemoglobins ; Capecitabine (6804DJ8Z9U)
Language	English
Publishing date	2017
Publishing country	Switzerland
Document type	Journal Article
ZDB-ID	6708-8
ISSN	1421-9794 ; 0009-3157
ISSN (online)	1421-9794
ISSN	0009-3157
DOI	10.1159/000445866
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Article: Retrospective Analysis of the Risk Factors for Grade IV Neutropenia in Oesophageal Cancer Patients Treated with a Docetaxel, Cisplatin, and 5-Fluorouracil Regimen

Naito, Masahito / Yamamoto, Tomoya / Shimamoto, Chikao / Miwa, Yoshihiro

Chemotherapy

2017 Volume 62, Issue 4, Page(s) 215–224

Institution	Osaka University of Pharmaceutical Sciences Department of Pharmacy, Osaka University Hospital, and Osaka University Graduate School of Medicine and Health Science, Osaka, Japan
Abstract	Background: Previous Japanese trials of the docetaxel, cisplatin, and 5-fluorouracil regimen for oesophageal cancer have demonstrated that a large proportion of patients also develop grade IV neutropenia. Our aim was to examine the risk factors for neutropenia in patients treated with this regimen. Methods: We retrospectively analysed the risk factors for developing grade IV neutropenia in 66 patients with oesophageal cancer using a multivariate analysis. Results: After administering the docetaxel, cisplatin, and 5-fluorouracil regimen, 49 patients (74.2%) developed grade IV neutropenia. Grade IV neutropenia was significantly associated with platelet count (p < 0.01), alanine transaminase level (p = 0.05), and proton-pump inhibitor administration (p < 0.05). Receiver operating characteristic curve analysis confirmed a platelet count of 290 × 103/μL as the optimal diagnostic cut-off value for grade IV neutropenia. The receiver operating characteristic area for grade IV neutropenia was increased by including patients that were administered a proton-pump inhibitor and alanine transaminase level (updated model; sensitivity and specificity, 75.5 and 88.2%, respectively). Conclusions: Our findings suggest that a platelet count is the most significant predictor of grade IV neutropenia.
Keywords	Docetaxel ; Neutropenia ; Oesophageal cancer ; Platelet count
Language	English
Publishing date	2017-04-19
Publisher	S. Karger AG
Publishing place	Basel, Switzerland
Document type	Article
Note	Anticancer Section / Original Paper
ZDB-ID	6708-8
ISSN	1421-9794 ; 0009-3157
ISSN (online)	1421-9794
ISSN	0009-3157
DOI	10.1159/000464273
Database	Karger publisher's database

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Article ; Online: Non-invasive in vivo imaging of UCP1 expression in live mice via near-infrared fluorescent protein iRFP720.

Fukuda, Aya / Honda, Shiho / Fujioka, Norie / Sekiguchi, Yuya / Mizuno, Seiya / Miwa, Yoshihiro / Sugiyama, Fumihiro / Hayashi, Yohei / Nishimura, Ken / Hisatake, Koji

PloS one

2019 Volume 14, Issue 11, Page(s) e0225213

Abstract: Uncoupling protein 1 (UCP1) is a mitochondrial protein that is expressed in both brown and beige adipocytes. UCP1 uncouples the mitochondrial electron transport chain from ATP synthesis to produce heat via non-shivering thermogenesis. Due to their ... ...

Abstract	Uncoupling protein 1 (UCP1) is a mitochondrial protein that is expressed in both brown and beige adipocytes. UCP1 uncouples the mitochondrial electron transport chain from ATP synthesis to produce heat via non-shivering thermogenesis. Due to their ability to dissipate energy as heat and ameliorate metabolic disorders, UCP1-expressing adipocytes are considered as a potential target for anti-obesity treatment. To monitor the expression of UCP1 in live mice in a non-invasive manner, we generated the Ucp1-iRFP720 knock-in (Ucp1-iRFP720 KI) mice, in which the gene encoding a near-infrared fluorescent protein iRFP720 is inserted into the Ucp1 gene locus. Using the heterozygous Ucp1-iRFP720 KI mice, we observed robust iRFP fluorescence in the interscapular region where brown adipose tissue is located. Moreover, the iRFP fluorescence was clearly observable in inguinal white adipose tissues in live mice administered with β3-adrenergic receptor agonist CL316,243. We also found that the homozygous Ucp1-iRFP720 KI mice, which are deficient in UCP1, displayed prominent iRFP fluorescence in the inguinal regions at the standard housing temperature. Consistent with this, the mice exhibited expanded populations of beige-like adipocytes in inguinal white adipose tissue, in which the Ucp1 promoter was dramatically activated. Thus, the Ucp1-iRFP720 KI mice provide a convenient model for non-invasive in vivo imaging of UCP1 expression in both brown and beige adipocytes in live mice.
MeSH term(s)	Adipocytes, Beige/metabolism ; Animals ; Cell Line ; Gene Expression ; Gene Targeting ; Genetic Loci ; Genotype ; Humans ; Luminescent Proteins/genetics ; Luminescent Proteins/metabolism ; Mice ; Mice, Knockout ; Mice, Transgenic ; Molecular Imaging/methods ; Spectroscopy, Near-Infrared ; Uncoupling Protein 1/genetics ; Uncoupling Protein 1/metabolism ; Red Fluorescent Protein
Chemical Substances	Luminescent Proteins ; UCP1 protein, human ; Uncoupling Protein 1
Language	English
Publishing date	2019-11-15
Publishing country	United States
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	2267670-3
ISSN	1932-6203 ; 1932-6203
ISSN (online)	1932-6203
ISSN	1932-6203
DOI	10.1371/journal.pone.0225213
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Article ; Online: The Metabolism of Methazolamide in Immortalized Human Keratinocytes, HaCaT Cells.

Sasabe, Tetsuo / Maeda, Shinichiro / Kishida, Kenichi / Yamano, Mariko / Miwa, Yoshihiro / Sugiyama, Toshihiro

Drug metabolism letters

2017 Volume 10, Issue 4, Page(s) 295–305

Abstract: Objective: Drug therapy is occasionally accompanied by an idiosyncratic severe toxicity, which occurs very rarely, but can lead to patient mortality. Methazolamide, an anti-glaucomatous agent, could cause severe skin eruptions called Stevens-Johnson ... ...

Abstract	Objective: Drug therapy is occasionally accompanied by an idiosyncratic severe toxicity, which occurs very rarely, but can lead to patient mortality. Methazolamide, an anti-glaucomatous agent, could cause severe skin eruptions called Stevens-Johnson syndrome/toxic epidermal necrolyis (SJS/TEN). Its precise etiology is still uncertain. In this study, the metabolism of methazolamide was investigated in immortalized human keratinocytes to reveal the possible mechanism which causes SJS/TEN. Methods: The metabolism of methazolamide was studied using immortalized human keratinocytes, HaCaT cells. HPLC was used to isolate a metabolite from the culture medium. Mass spectrometry (LCMS/ MS) was employed for its characterization. Three typical chemical inducers were assessed for the inducibility of cytochrome P450, and methimazole was used as the inhibitor of flavin-containing monooxygenase (FMO). Results: A sulfonic acid, N-[3-methyl-5-sulfo-1,3,4-thiadiazol-2(3H)-ylidene]acetamide (MSO) was identified as the final metabolite. Dexamethasone and β-naphthoflavone behaved as an inducer of cytochrome P450 in the metabolism, but isoniazid did not. The effect of methimazole was not consistent. We did not detect any glucuronide nor any mercapturic acid (N-acetylcysteine conjugate). Conclusion: N-[3-methyl-5-sulfo-1,3,4-thiadiazol-2(3H)-ylidene]acetamide (MSO) is not considered to be a direct product of an enzymatic reaction, but rather an auto-oxidation product of N-[3-methyl-5- sulfe-1,3,4-thiadiazol-2(3H)-ylidene]acetamide, a chemically unstable sulfenic acid, which is produced by cytochrome P450 from the β-lyase product of cysteine conjugate of methazolamide. MSO is considered to be susceptible to glutathione and to return to glutathione conjugate of methazolamide, forming a futile cycle. A hypothetical scenario is presented as to the onset of the disease.
Language	English
Publishing date	2017
Publishing country	United Arab Emirates
Document type	Journal Article
ISSN	1874-0758
ISSN (online)	1874-0758
DOI	10.2174/1872312811666170127160931
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Article ; Online: Albino mice with the point mutation at the tyrosinase locus show high cholesterol diet-induced NASH susceptibility.

Kulathunga, Kaushalya / Wakimoto, Arata / Hiraishi, Yukiko / Yadav, Manoj Kumar / Gentleman, Kyle / Warabi, Eiji / Sakasai, Tomoki / Miwa, Yoshihiro / Mizuno, Seiya / Takahashi, Satoru / Hamada, Michito

Scientific reports

2021 Volume 11, Issue 1, Page(s) 21827

Abstract: Non-alcoholic fatty liver disease (NAFLD) constitutes a metabolic disorder with high worldwide prevalence and increasing incidence. The inflammatory progressive state, non-alcoholic steatohepatitis (NASH), leads to liver fibrosis and carcinogenesis. Here, ...

Abstract	Non-alcoholic fatty liver disease (NAFLD) constitutes a metabolic disorder with high worldwide prevalence and increasing incidence. The inflammatory progressive state, non-alcoholic steatohepatitis (NASH), leads to liver fibrosis and carcinogenesis. Here, we evaluated whether tyrosinase mutation underlies NASH pathophysiology. Tyrosinase point-mutated B6 (Cg)-Tyr
MeSH term(s)	Albinism, Oculocutaneous/complications ; Albinism, Oculocutaneous/enzymology ; Albinism, Oculocutaneous/genetics ; Animals ; Cholesterol/metabolism ; Cholesterol, Dietary/administration & dosage ; Cholesterol, Dietary/adverse effects ; Diet, High-Fat/adverse effects ; Disease Models, Animal ; Disease Susceptibility ; Humans ; Intestine, Small/metabolism ; Intestine, Small/pathology ; Lipoproteins/blood ; Liver/metabolism ; Liver/pathology ; Male ; Mice ; Mice, Inbred BALB C ; Mice, Inbred C57BL ; Mice, Mutant Strains ; Monophenol Monooxygenase/genetics ; Non-alcoholic Fatty Liver Disease/enzymology ; Non-alcoholic Fatty Liver Disease/etiology ; Non-alcoholic Fatty Liver Disease/genetics ; Point Mutation
Chemical Substances	Cholesterol, Dietary ; Lipoproteins ; Cholesterol (97C5T2UQ7J) ; Monophenol Monooxygenase (EC 1.14.18.1)
Language	English
Publishing date	2021-11-08
Publishing country	England
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	2615211-3
ISSN	2045-2322 ; 2045-2322
ISSN (online)	2045-2322
ISSN	2045-2322
DOI	10.1038/s41598-021-00501-5
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