Publications by authors named "Kazuki Harada"

116 Publications

Complete Genome Sequence of an Possessing Enterobacter roggenkampii Strain Isolated from a Dog in Japan.

Microbiol Resour Announc 2021 Jul 29;10(30):e0042621. Epub 2021 Jul 29.

Department of Microbiology, Sapporo Medical University School of Medicine, Sapporo, Japan.

The complete genome sequence of -possessing Enterobacter roggenkampii En37, isolated from a dog in Japan, was determined. was located on a 70,277-bp IncFIB plasmid without any additional antimicrobial resistance genes.
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http://dx.doi.org/10.1128/MRA.00426-21DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8320466PMC
July 2021

The usefulness of postmortem computed tomography angiography for subdural hematoma caused by rupture of the cortical artery: A report of two autopsy cases and a literature review.

Leg Med (Tokyo) 2021 Jul 12;53:101941. Epub 2021 Jul 12.

Division of Legal Medicine, Department of Community Preventive Medicine, Niigata University Graduate School of Medical and Dental Sciences, Niigata, Japan; Center of Cause of Death Investigation, Niigata University Graduate School of Medical and Dental Sciences, Niigata, Japan.

Acute subdural hematoma (SDH) occurs following severe head trauma with brain contusion or rupture of bridging veins. Conversely, SDH caused by rupture of a cortical artery without trauma or with minor trauma is also possible. Although over 150 cases of the latter SDH have been reported, they were predominantly diagnosed only during surgery, and therefore, no adequate histological evaluation has been performed. Therefore, essential etiology of this SDH type has remained unclear. In addition, the scarcity of autopsy cases may be attributed to arterial rupture being missed if the macroscopic findings are too minimal to detect during autopsy. Here, we describe two autopsy cases of SDH of cortical artery origin. Extravasation on postmortem computed tomography angiography and arterial leakage on macroscopic observation during autopsy facilitated detection of the ruptured artery and allowed detailed histological evaluation of the ruptured artery and adjacent dura mater. The etiology of arterial rupture is briefly described on the basis of histopathological findings in this study and the available literature.
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http://dx.doi.org/10.1016/j.legalmed.2021.101941DOI Listing
July 2021

Strong association between insufficient plasma exchange and fatal outcomes in Japanese patients with immune-mediated thrombotic thrombocytopenic purpura.

Int J Hematol 2021 Oct 22;114(4):415-423. Epub 2021 Jul 22.

Department of Blood Transfusion Medicine, Nara Medical University, 840 Shijo-Cho, Kashihara, 634-8522, Japan.

Plasma exchange (PEX) using fresh frozen plasma has considerably reduced the mortality rate in patients with immune-mediated thrombotic thrombocytopenic purpura (iTTP). However, some patients still do not survive even with treatment, but little information is available regarding which treatment these patients received. This study was conducted to obtain this information in 240 patients who met the current iTTP diagnostic criteria and completed at least 30 days of follow-up except for deceased cases. These patients were divided into three groups: survivors (n = 195), TTP-related deaths (n = 32), and other cause of death (n = 13). In the TTP-related death group, 26 of 32 patients experienced sudden death, mostly following radical hypotension and bradycardia. The median follow-up time after admission was 5.0 days, and the median number of PEX sessions was 2.5. Nine patients underwent autopsy and had cardiac microvascular thrombi in arterioles. Levels of lactate dehydrogenase, total bilirubin, serum creatinine, and D-dimer were significantly higher in the TTP-related death group than in the survivors group. Frequent PEX (> 20 sessions) was not associated with TTP-related death. In the acute phase of iTTP, patients with substantial organ damage caused by microthrombi have a greater mortality risk, even after just a few PEX sessions.
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http://dx.doi.org/10.1007/s12185-021-03197-5DOI Listing
October 2021

Development of red genetically encoded biosensor for visualization of intracellular glucose dynamics.

Cell Chem Biol 2021 Jun 29. Epub 2021 Jun 29.

Department of Life Sciences, Graduate School of Arts and Sciences, The University of Tokyo, 3-8-1 Komaba, Meguro, Tokyo 153-8902, Japan; Department of Biological Sciences, Graduate School of Science, The University of Tokyo, 7-3-1 Hongo, Bunkyo, Tokyo 113-0033, Japan. Electronic address:

Glucose is the main source of energy for organisms, and it is important to understand the spatiotemporal dynamics of intracellular glucose. Single fluorescent protein-based glucose indicators, named "Red Glifons" have been developed that apply to live-cell and dual-color imaging. These indicators exhibited more than 3-fold increase in fluorescence intensity in the presence of 10 mM glucose. The two Red Glifons developed have different half-maximal effective concentration (EC) values for glucose (300 μM and 3,000 μM) and are able to monitor a wide range of glucose dynamics. Red Glifon combined with green indicators allowing visualization of the interplay between glucose and ATP, lactate, or pyruvate. Glucose influx in the pharyngeal muscle of Caenorhabditis elegans, enteroendocrine cells, and human iPS cell-derived cardiac myocytes was observed using the Red Glifons. Thus these red glucose indicators serve as a multi-color imaging toolkit for investigating complex interactions in energy metabolism.
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http://dx.doi.org/10.1016/j.chembiol.2021.06.002DOI Listing
June 2021

Complete Genome Sequence of an -Possessing Enterobacter asburiae Strain Isolated from a Cat in Japan.

Microbiol Resour Announc 2021 Jul 1;10(26):e0028121. Epub 2021 Jul 1.

Department of Microbiology, Sapporo Medical University School of Medicine, Sapporo, Japan.

We report the complete genome sequence of the -possessing strain Enterobacter asburiae En30, isolated from a cat in Japan. The genome sequence was obtained by using long- and short-read sequencing.
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http://dx.doi.org/10.1128/MRA.00281-21DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8248887PMC
July 2021

Antimicrobial resistance and multilocus sequence types of isolated from dogs and cats in Japan.

J Med Microbiol 2021 Apr;70(4)

Department of Veterinary Internal Medicine, Tottori University, Minami 4-101, Koyama, Tottori-shi, Tottori 680-8553, Japan.

As the representative multidrug-resistant pathogen, has multiple intrinsic and acquired resistances, including carbapenem resistance. In companion animals, the antimicrobial susceptibility and sequence types (STs) of are not well understood due to its limited isolation rate. We investigated the antimicrobial susceptibilities and multilocus sequence types (MLSTs) of 38 . strains isolated from dogs and cats in Japan. Prevalence of resistance was detected for imipenem (100 %), aztreonam (94.7 %), piperacillin (65.8 %), trimethoprim-sulfamethoxazole (65.8 %), and ceftazidime (60.5 %). Rates of resistances to chloramphenicol, minocycline, and levofloxacin were low (2.6-5.3 %). MLST analysis revealed that all 38 strains were assigned to 34 STs, including 11 previously reported STs and 23 newly identified STs. Phylogenetic analysis of MLSTs enabled categorization of 13 isolates (34.2 %) into genogroup 6, which is a major genogroup of human isolates. Multinational surveillance would be needed to clarify the significance of antimicrobial-resistant isolates from companion animals.
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http://dx.doi.org/10.1099/jmm.0.001344DOI Listing
April 2021

Results of repeat manipulation under ultrasound-guided cervical nerve root block with corticosteroid and local anaesthetic injection for recurrence of frozen shoulder.

J Orthop Surg Res 2020 Dec 7;15(1):586. Epub 2020 Dec 7.

Paku Pain Clinic, 6-1-20, Gokodori, Chuo-ku, Kobe, Hyogo, 651-0087, Japan.

Background: This study aimed to evaluate the clinical results of a repeat manipulation under ultrasound-guided cervical nerve root block (MUC) with corticosteroid and local anaesthetic injection for recurrence of idiopathic frozen shoulder after MUC.

Methods: A consecutive series of 42 shoulders in 39 patients with idiopathic frozen shoulder underwent MUC. All patients were assessed according to the American Shoulder Elbow Surgeon (ASES) score and shoulder range of motion (ROM) both before MUC and at 1 year thereafter. If patients continued to have pain and limited ROM at 3 months after the procedure, they were offered a repeat MUC. Such patients were also assessed before the procedure and at 3 months and 1 year thereafter.

Results: The initial MUC was successful in 31 shoulders (single group). Repeat MUC was required in 11 shoulders (repeat group). Patients in the single group showed significant improvement in ROM and ASES score at 1 year after the procedure (p < 0.001); similarly, patients in the repeat group had significant improvement in ROM and ASES score at 3 months and 1 year after the procedure (p < 0.001). Patients in the repeat group had had significantly more severely limited ROM (p < 0.01) and decreased ASES score (p < 0.001) before the procedure compared with those in the single group.

Conclusions: A repeat MUC with corticosteroid and local anaesthetic injection is a valuable option before proceeding to surgery for recurrence of idiopathic frozen shoulder. When there is severely limited ROM and decreased ASES score before the MUC, a repeat MUC may be necessary, which would require the patient's informed consent.

Trial Registration: Retrospectively registered.
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http://dx.doi.org/10.1186/s13018-020-02120-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7720550PMC
December 2020

Green fluorescent protein-based lactate and pyruvate indicators suitable for biochemical assays and live cell imaging.

Sci Rep 2020 11 11;10(1):19562. Epub 2020 Nov 11.

Department of Life Sciences, Graduate School of Arts and Sciences, The University of Tokyo, 3-8-1 Komaba, Meguro, Tokyo, 153-8902, Japan.

Glycolysis is the metabolic pathway that converts glucose into pyruvate, whereas fermentation can then produce lactate from pyruvate. Here, we developed single fluorescent protein (FP)-based lactate and pyruvate indicators with low EC for trace detection of metabolic molecules and live cell imaging and named them "Green Lindoblum" and "Green Pegassos," respectively. Green Lindoblum (EC of 30 µM for lactate) and Green Pegassos (EC of 70 µM for pyruvate) produced a 5.2- and 3.3-fold change in fluorescence intensity in response to lactate and pyruvate, respectively. Green Lindoblum measured lactate levels in mouse plasma, and Green Pegassos in combination with D-serine dehydratase successfully estimated D-serine levels released from mouse primary cultured neurons and astrocytes by measuring pyruvate level. Furthermore, live cell imaging analysis revealed their utility for dual-colour imaging, and the interplay between lactate, pyruvate, and Ca in human induced pluripotent stem cell-derived cardiomyocytes. Therefore, Green Lindoblum and Green Pegassos will be useful tools that detect specific molecules in clinical use and monitor the interplay of metabolites and other related molecules in diverse cell types.
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http://dx.doi.org/10.1038/s41598-020-76440-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7659002PMC
November 2020

Improvement Effect of 5-Aminolevulinic Acid on Hyperlipidemia in Miniature Schnauzer Dogs: An Open Study in 5 Cases of One Pedigree.

Yonago Acta Med 2020 Aug 10;63(3):234-238. Epub 2020 Jul 10.

Department of Veterinary Internal Medicine, Faculty of Agriculture, Tottori University, Tottori 680-8550, Japan.

This is the first study to examine the long-term effect of 5-aminolevulinic acid mainly on serum lipoproteins in dogs with hyperlipidemia. We studied 5 Miniature Schnauzer cases whose fasting serum total triglyceride and very-low-density lipoprotein of triglyceride levels were extremely high (635 ± 116 and 520 ± 92 mg/dL, respectively). Although the total cholesterol values were normal, the very-low-density lipoprotein of cholesterol level was high (49 ± 7 mg/dL). Each dog received a 5-aminolevulinic acid supplement (5 mg/day) orally for 6 months. The mean values of total triglyceride, very-low-density lipoprotein of both triglyceride and cholesterol decreased significantly after the treatment period (319 ± 29, 245 ± 18, and 27 ± 2 mg/dL, respectively, < 0.05). Our present results may present evidence that 5-ALA administration contributes to improvement of hyperlipidemia in Miniature Schnauzer.
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http://dx.doi.org/10.33160/yam.2020.08.006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7435120PMC
August 2020

A Pilot Study on Efficacy of Lipid Bubbles for Theranostics in Dogs with Tumors.

Cancers (Basel) 2020 Aug 26;12(9). Epub 2020 Aug 26.

Joint Department of Veterinary Clinical Medicine, Faculty of Agriculture, Tottori University, 4-101 Koyama-Minami, Tottori 680-8553, Japan.

The combined administration of microbubbles and ultrasound (US) is a promising strategy for theranostics, i.e., a combination of therapeutics and diagnostics. Lipid bubbles (LBs), which are experimental theranostic microbubbles, have demonstrated efficacy in vitro and in vivo for both contrast imaging and drug delivery in combination with US irradiation. To evaluate the clinical efficacy of LBs in combination with US in large animals, we performed a series of experiments, including clinical studies in dogs. First, contrast-enhanced ultrasonography using LBs (LB-CEUS) was performed on the livers of six healthy Beagles. The hepatic portal vein and liver tissue were enhanced; no adverse reactions were observed. Second, LB-CEUS was applied clinically to 21 dogs with focal liver lesions. The sensitivity and specificity were 100.0% and 83.3%, respectively. These results suggested that LB-CEUS could be used safely for diagnosis, with high accuracy. Finally, LBs were administered in combination with therapeutic US to three dogs with an anatomically unresectable solid tumor in the perianal and cervical region to determine the enhancement of the chemotherapeutic effect of liposomal doxorubicin; a notable reduction in tumor volume was observed. These findings indicate that LBs have potential for both therapeutic and diagnostic applications in dogs in combination with US irradiation.
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http://dx.doi.org/10.3390/cancers12092423DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7564957PMC
August 2020

Optimal Aspergillus fumigatus and Asp f 1 serum IgG cut-offs for the diagnosis of allergic bronchopulmonary aspergillosis.

Allergol Int 2021 Jan 16;70(1):74-80. Epub 2020 Aug 16.

Clinical Research Center for Allergy and Rheumatology, National Hospital Organization Sagamihara National Hospital, Kanagawa, Japan; Course of Allergy and Clinical Immunology, Juntendo University Graduate School of Medicine, Tokyo, Japan; Center for Immunology and Allergology, Shonan Kamakura General Hospital, Kanagawa, Japan.

Background: The presence of IgG antibodies (Abs) to Aspergillus fumigatus (Af) is a crucial diagnostic criterion for allergic bronchopulmonary aspergillosis (ABPA). Although precipitation is traditionally used to document IgG Abs, anti-Af serum IgG levels can also be measured by enzyme immunoassay (EIA). However, there are insufficient data on the optimal cut-offs to assess diagnostic performance of the EIA method. This study aimed to determine cut-off levels of IgG binding crude Af extracts or recombinant Asp f 1 (by ImmunoCAP®) and to compare their efficacy for ABPA diagnosis with Af-precipitating Abs.

Methods: The age distribution of levels of IgG to crude extracts of Af (Af-IgG) and recombinant Asp f 1 (Asp f 1-IgG) was established using sera from 694 healthy controls (HC). Receiver operating characteristic analysis for Af-IgG and Asp f 1-IgG levels for the purpose of ABPA diagnosis was performed in 306 Af-sensitized asthma patients (including 49 ABPA), and cut-offs were determined.

Results: An age-dependent decline in the levels of Af-IgG was observed in HC. Thus, cut-offs for Af-IgG levels were determined separately by age as 60 mg/L for patients aged <55 years, and 45 mg/L for those aged ≥55 years. For Asp f 1-IgG, 6.6 mg/L was set as the cut-off regardless of age. Although such IgG testing by EIA allowed a sufficiently good diagnostic performance, Af-precipitating Abs had better diagnostic applicability for ABPA.

Conclusions: We determined cut-offs for Af-IgG and Asp f 1-IgG measured by EIA, which can be useful in clinical settings where precipitating Abs are unavailable.
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http://dx.doi.org/10.1016/j.alit.2020.07.006DOI Listing
January 2021

An autopsy case: Unusual characteristics of stab wounds inflicted by a survival knife.

Leg Med (Tokyo) 2020 Nov 20;47:101762. Epub 2020 Jul 20.

Department of Forensic Medicine, National Defense Medical College, Tokorozawa, Japan.

A woman was stabbed 10 times and killed. The suspect confessed that he had used a thick (0.7 cm), single-edged survival knife with a serrated spine. However, the bilateral ends of the stab wounds appeared sharp. To address this discrepancy, we used pig skin to examine whether the knife could have been used to make the thin stab wounds observed. The widths of the pig skin wounds were approximately half the width of the knife and the ends of some wounds appeared sharp, particularly when the knife was held at a right angle. It was concluded that the suspect could have killed the victim using the survival knife. Similar pig skin experiments could be useful for forensic pathologists to determine whether a weapon of interest might have been used to inflict certain injuries.
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http://dx.doi.org/10.1016/j.legalmed.2020.101762DOI Listing
November 2020

Microdiamond in a low-grade metapelite from a Cretaceous subduction complex, western Kyushu, Japan.

Sci Rep 2020 07 15;10(1):11645. Epub 2020 Jul 15.

Institute of Liberal Arts and Science, Kanazawa University, Kakuma, Kanazawa, 920-1164, Japan.

Microdiamonds in metamorphic rocks are a signature of ultrahigh-pressure (UHP) metamorphism that occurs mostly at continental collision zones. Most UHP minerals, except coesite and microdiamond, have been partially or completely retrogressed during exhumation; therefore, the discovery of coesite and microdiamond is crucial to identify UHP metamorphism and to understand the tectonic history of metamorphic rocks. Microdiamonds typically occur as inclusions in minerals such as garnet. Here we report the discovery of microdiamond aggregates in the matrix of a metapelite from the Nishisonogi unit, Nagasaki Metamorphic Complex, western Kyushu, Japan. The Nishisonogi unit represents a Cretaceous subduction complex which has been considered as an epidote-blueschist subfacies metamorphic unit, and the metapelite is a member of a serpentinite mélange in the Nishisonogi unit. The temperature condition for the Nishisonogi unit is 450 °C, based on the Raman micro-spectroscopy of graphite. The coexistence of microdiamond and Mg-carbonates suggests the precipitation of microdiamond from C-O-H fluid under pressures higher than 2.8 GPa. This is the first report of metamorphic microdiamond from Japan, which reveals the hidden UHP history of the Nishisonogi unit. The tectonic evolution of Kyushu in the Japanese Archipelago should be reconsidered based on this finding.
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http://dx.doi.org/10.1038/s41598-020-68599-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7363827PMC
July 2020

Urinary Pharmacokinetic and Pharmacodynamic Profiles of Fosfomycin against Extended-Spectrum β-Lactamase-Producing with Canine Ex Vivo Modeling: A Pilot Study.

Antibiotics (Basel) 2020 May 5;9(5). Epub 2020 May 5.

Anicom Specialty Medical Institute Inc., Kanagawa 231-0033, Japan.

Fosfomycin is a candidate drug for extended-spectrum β-lactamase (ESBL)-producing bacteria, but its efficacy is yet to be investigated in dogs. This study investigated the urinary pharmacokinetic/pharmacodynamic (PK/PD) profile of fosfomycin orally administered at 80 mg/kg to six healthy dogs to assess its efficacy for canine urinary tract infections (UTIs) caused by ESBL-producing bacteria. Four strains of ESBL-producing (ESBL-EC) characterized by fosfomycin minimum inhibitory concentrations (MICs) of 0.5, 1, 2, and 32 µg/mL were used. Urine samples for the measurement of urinary drug concentrations and urinary bactericidal titers (UBTs) were obtained after drug administration. The urinary concentrations (µg/mL, mean ± SE) were 1348.2 ± 163.5, 1191.6 ± 260.2, and 661.1 ± 190.4 at 0-4, 4-8, and 8-12 h, respectively, after drug administration. The mean urinary area under the curve during the test period (AUC) of fosfomycin was estimated to be 12,803.8 µg·h/mL. The UBTs for all tested strains fluctuated closely with urine concentration during the test period (r = 0.944-1.000), and the area under the UBT-versus-time curve correlated with the urinary AUC/MIC of each strain (r = 0.991). According to the optimal urinary PK/PD target value, fosfomycin at 80 mg/kg twice daily may be suitable for the treatment of canine UTIs caused by ESBL-EC presenting MIC ≤ 128 µg/mL.
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http://dx.doi.org/10.3390/antibiotics9050230DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7277591PMC
May 2020

Assessment of the Usefulness of Cefapirin and Cefalonium Disks for Susceptibility Testing of Isolates from Bovine Mastitis.

Antibiotics (Basel) 2020 Apr 21;9(4). Epub 2020 Apr 21.

Laboratory of Veterinary Microbiology, Nippon Veterinary and Life Science University, Tokyo 180-8602, Japan.

Cefapirin (CEP) and cefalonium (CNM) are first-generation cephalosporins widely used to treat bovine mastitis caused by Gram-positive bacteria including staphylococci. However, disks for susceptibility testing of those drugs in causative bacteria are not available. This study evaluated the efficacy of 10 µg and 30 µg pilot disks of CEP (CEP10 and CEP30) and CNM (CNM10 and CNM30) against 130 isolates from bovine mastitis. Scattergrams of minimum inhibitory concentrations (MICs) and zone diameters (ZDs) illustrated significant correlations between the MICs and ZDs of CEP10 ( = -0.912), CEP30 ( = -0.933), CNM10 ( = -0.847), and CNM30 ( = -0.807). The analysis by Normalized Resistance Interpretation indicated that the epidemiolocal cut-off value (ECV) of MIC for both cefapirin and cefalonium is ≤ 0.5 µg/mL, and the ECV of ZD for CEP10, CEP30, CNM10, and CNM30 were ≥ 22 mm, ≥ 25 mm, ≥ 22 mm, and ≥ 29 mm, respectively. We believe that both 10 μg and 30 μg CEP and CNM susceptibility disks will be helpful for guiding the appropriate use of these antibiotics for bovine mastitis. Further studies toward the establishment of clinical breakpoint of CEP and CNM would be needed for their routine use.
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http://dx.doi.org/10.3390/antibiotics9040197DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7235829PMC
April 2020

Glutamine-induced signaling pathways via amino acid receptors in enteroendocrine L cell lines.

J Mol Endocrinol 2020 04;64(3):133-143

Department of Life Sciences, Graduate School of Arts and Sciences, The University of Tokyo, Meguro, Tokyo, Japan.

Glucagon-like peptide-1 (GLP-1), secreted by gastrointestinal enteroendocrine L cells, induces insulin secretion and is important for glucose homeostasis. GLP-1 secretion is induced by various luminal nutrients, including amino acids. Intracellular Ca2+ and cAMP dynamics play an important role in GLP-1 secretion regulation; however, several aspects of the underlying mechanism of amino acid-induced GLP-1 secretion are not well characterized. We investigated the mechanisms underlying the L-glutamine-induced increase in Ca2+ and cAMP intracellular concentrations ([Ca2+]i and [cAMP]i, respectively) in murine enteroendocrine L cell line GLUTag cells. Application of L-glutamine to cells under low extracellular [Na+] conditions, which inhibited the function of the sodium-coupled L-glutamine transporter, did not induce an increase in [Ca2+]i. Application of G protein-coupled receptor family C group 6 member A and calcium-sensing receptor antagonist showed little effect on [Ca2+]i and [cAMP]i; however, taste receptor type 1 member 3 (TAS1R3) antagonist suppressed the increase in [cAMP]i. To elucidate the function of TAS1R3, which forms a heterodimeric umami receptor with taste receptor type 1 member 1 (TAS1R1), we generated TAS1R1 and TAS1R3 mutant GLUTag cells using the CRISPR/Cas9 system. TAS1R1 mutant GLUTag cells exhibited L-glutamine-induced increase in [cAMP]i, whereas some TAS1R3 mutant GLUTag cells did not exhibit L-glutamine-induced increase in [cAMP]i and GLP-1 secretion. These findings suggest that TAS1R3 is important for L-glutamine-induced increase in [cAMP]i and GLP-1 secretion. Thus, TAS1R3 may be coupled with Gs and related to cAMP regulation.
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http://dx.doi.org/10.1530/JME-19-0260DOI Listing
April 2020

Nivolumab-induced fulminant type 1 diabetes with precipitous fall in C-peptide level.

J Diabetes Investig 2020 May 6;11(3):748-749. Epub 2019 Oct 6.

Division of Nephrology, Endocrinology and Metabolism, Department of Internal Medicine, Tokai University School of Kyodo Hospital, Isehara, Japan.

We describe here a case of nivolumab-induced type 1 diabetes, which developed within 9 days of treatment. The case highlights the importance of frequent monitoring of glucose after initiation of nivolumab treatment.
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http://dx.doi.org/10.1111/jdi.13143DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7232357PMC
May 2020

Prevalence of 16S rRNA methylases in Gram-negative bacteria derived from companion animals and livestock in Japan.

J Vet Med Sci 2019 Jun 2;81(6):874-878. Epub 2019 May 2.

Laboratory of Food Microbiology and Food Safety, School of Veterinary Medicine, Rakuno Gakuen University, Ebetsu, Hokkaido 069-8501, Japan.

The emergence and spread of aminoglycoside-resistant bacteria are a public health concern. The acquisition of the genes encoding 16S rRNA methylases, such as armA, rmtA, and rmtB, confers high-level resistance to aminoglycosides. However, the prevalence has not been well investigated in Japanese veterinary fields. To determine the prevalence of 16S rRNA methylases in animals, we detected 16S rRNA methylases genes in Gram-negative bacteria from animals. Here, we report the isolation of rmtB amd armA from two of the 446 Escherichia coli (0.5%) and one of the 103 Klebsiella spp. isolates (1.0%) from companion animals, respectively. However, none of the isolations were observed from 2445 E. coli isolates derived from livestock in Japan. The prevalence of 16S rRNA methylases in animals, especially in companion animals, should be carefully monitored in Japanese veterinary fields to avoid the spreading of the genes.
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http://dx.doi.org/10.1292/jvms.19-0144DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6612502PMC
June 2019

Sexually Dimorphic Regulation of Behavioral States by Dopamine in .

J Neurosci 2019 06 15;39(24):4668-4683. Epub 2019 Apr 15.

Department of Life Sciences, Graduate School of Arts and Sciences, University of Tokyo, Tokyo, 153-8902, Japan.

Sex differences in behavior allow animals to effectively mate and reproduce. However, the mechanism by which biological sex regulates behavioral states, which underlie the regulation of sex-shared behaviors, such as locomotion, is largely unknown. In this study, we studied sex differences in the behavioral states of and found that males spend less time in a low locomotor activity state than hermaphrodites and that dopamine generates this sex difference. In males, dopamine reduces the low activity state by acting in the same pathway as polycystic kidney disease-related genes that function in male-specific neurons. In hermaphrodites, dopamine increases the low activity state by suppression of octopamine signaling in the sex-shared SIA neurons, which have reduced responsiveness to octopamine in males. Furthermore, dopamine promotes exploration both inside and outside of bacterial lawn (the food source) in males and suppresses it in hermaphrodites. These results demonstrate that sexually dimorphic signaling allows the same neuromodulator to promote adaptive behavior for each sex. The mechanisms that generate sex differences in sex-shared behaviors, including locomotion, are not well understood. We show that there are sex differences in the regulation of behavioral states in the model animal Dopamine promotes the high locomotor activity state in males, which must search for mates to reproduce, and suppresses it in self-fertilizing hermaphrodites through distinct molecular mechanisms. This study demonstrates that sex-specific signaling generates sex differences in the regulation of behavioral states, which in turn modulates the locomotor activity to suit reproduction for each sex.
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http://dx.doi.org/10.1523/JNEUROSCI.2985-18.2019DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6561698PMC
June 2019

Assessment of urinary pharmacokinetic and pharmacodynamic profiles of faropenem against extended-spectrum -lactamase-producing with canine modelling: a pilot study.

Access Microbiol 2019 20;1(1):e000004. Epub 2019 Mar 20.

Department of Veterinary Internal Medicine, Tottori University, Minami 4-101, Koyama-Cho, Tottori 680-8553, Japan.

This study was carried out to investigate the urinary pharmacokinetics and pharmacodynamics of faropenem administered orally at 5 mg kg in six healthy dogs to assess the efficacy of the drug for canine urinary tract infections (UTIs) with extended-spectrum -lactamase (ESBL)-producing bacteria. Six strains of ESBL-producing (ESBL-EC) with the following faropenem minimum inhibitory concentrations (MICs) were used: 1 µg ml (=2), 2 µg ml (=2), 4 µg ml (=1) and 16 µg ml (=1). Urine samples were obtained every 4 h for the first 12 h after administration to measure urinary drug concentration and urinary bactericidal titres (UBTs). Both the urine concentration of faropenem and the UBTs for all tested strains peaked at 0-4 h after administration, and decreased markedly at 8-12 h. The mean urinary concentration of faropenem at 8-12 h (23±5.2 µg ml) exceeded the MIC of 1 µg ml by fourfold, which is required to inhibit the growth of 90  % of ESBL-EC. These findings indicate that faropenem administered twice daily at a dose of 5 mg kg is acceptable for the treatment of most dogs with ESBL-EC-related UTIs.
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http://dx.doi.org/10.1099/acmi.0.000004DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7470355PMC
March 2019

Green Fluorescent Protein-Based Glucose Indicators Report Glucose Dynamics in Living Cells.

Anal Chem 2019 04 22;91(7):4821-4830. Epub 2019 Mar 22.

Laboratory for Chemistry and Life Science, Institute of Innovative Research , Tokyo Institute of Technology , 4259 Nagatsuta-cho, Midori-ku , Yokohama , Kanagawa 226-8503 , Japan.

Glucose is the most important energy source for living animals. Here, we developed a series of single fluorescent protein (FP)-based glucose indicators, named as "Green Glifons", to understand the hierarchal and mutual relationships between molecules involved in energy metabolism. Three indicators showed a different EC for glucose (50, 600, and 4000 μM), producing a ∼7-fold change in fluorescence intensity in response to glucose. The indicators could visualize glucose dynamics in the cytoplasm, plasma membrane, nucleus and mitochondria of living HeLa cells and in vivo, in the pharyngeal muscle of C. elegans and could measure murine blood glucose levels. Finally, the indicators were applicable to dual-color imaging, revealing the dynamic interplay between glucose and Ca in mouse pancreatic MIN6 m9 β cells. We propose that these indicators will facilitate and contribute to in vivo and multicolor imaging of energy metabolism.
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http://dx.doi.org/10.1021/acs.analchem.9b00447DOI Listing
April 2019

Characterization of Antimicrobial Resistance in spp. and spp. Isolates from Companion Animals in Japan: Nosocomial Dissemination of Extended-Spectrum Cephalosporin-Resistant .

Microorganisms 2019 Feb 28;7(3). Epub 2019 Feb 28.

Sanritsu Zelkova Laboratory, Tokyo 135-0011, Japan.

In many countries including Japan, the status of emerging antimicrobial resistance among spp. and spp. in companion animals remains unknown because these genera are rarely isolated from animals. In this study, 30 spp. and 23 spp. isolates from companion animals underwent susceptibility testing for 10 antimicrobials. Phenotypic and genetic approaches were used to identify the mechanisms of extended-spectrum cephalosporins (ESC). Subsequently, ESC-resistant spp. strains underwent multilocus sequence typing and pulsed-field gel electrophoresis (PFGE). A significantly higher rate (34.8%) of ESC resistance was observed in spp. isolates than in spp. isolates (0%). ESC resistance was detected in five strains, two strains, and one strain. All of the ESC-resistant spp. strains harbored CMY-type and/or DHA-type AmpC β-lactamases. Three strains harbored the CTX-M-3-type extended-spectrum β-lactamases. Notably, the three CTX-3-producing and two CMY-117-bearing strains (obtained from different patients in one hospital) had the same sequence type (ST156 and ST18, respectively) and similar PFGE profiles. We believe that ESC-resistant spp. are important nosocomial pathogens in veterinary medicine. Therefore, infection control in animal hospitals is essential to prevent dissemination of these resistant pathogens.
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http://dx.doi.org/10.3390/microorganisms7030064DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6462910PMC
February 2019

RGB-Color Intensiometric Indicators to Visualize Spatiotemporal Dynamics of ATP in Single Cells.

Angew Chem Int Ed Engl 2018 08 18;57(34):10873-10878. Epub 2018 Jul 18.

Cell Signaling Group, Waseda Bioscience Research Institute in Singapore (WABIOS), Singapore, 138667, Republic of Singapore.

Adenosine triphosphate (ATP) provides energy for the regulation of multiple cellular processes in living organisms. Capturing the spatiotemporal dynamics of ATP in single cells is fundamental to our understanding of the mechanisms underlying cellular energy metabolism. However, it has remained challenging to visualize the dynamics of ATP in and between distinct intracellular organelles and its interplay with other signaling molecules. Using single fluorescent proteins, multicolor ATP indicators were developed, enabling the simultaneous visualization of subcellular ATP dynamics in the cytoplasm and mitochondria of cells derived from mammals, plants, and worms. Furthermore, in combination with additional fluorescent indicators, the dynamic interplay of ATP, cAMP, and Ca could be visualized in activated brown adipocyte. This set of indicator tools will facilitate future research into energy metabolism.
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http://dx.doi.org/10.1002/anie.201804304DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6456769PMC
August 2018

Bacterial metabolite S-equol modulates glucagon-like peptide-1 secretion from enteroendocrine L cell line GLUTag cells via actin polymerization.

Biochem Biophys Res Commun 2018 07 21;501(4):1009-1015. Epub 2018 May 21.

Department of Life Sciences, Graduate School of Arts and Sciences, The University of Tokyo, 3-8-1 Komaba, Meguro, Tokyo 153-8902, Japan; Department of Biological Sciences, Graduate School of Science, The University of Tokyo, 7-3-1 Hongo, Bunkyo, Tokyo 113-0033, Japan. Electronic address:

S-equol is one of gut bacterial metabolites produced from soybean isoflavone daizein. While S-equol is known to promote glucose-induced insulin secretion from pancreatic β cells, whether S-equol affects glucagon-like peptide-1 (GLP-1) secretion from enteroendoceine L cells remains unclear. Here we assessed the effect of S-equol on GLP-1 secretion from mouse enteroendocrine L cell line GLUTag cells. GLUTag cells expressed GPR30 and estrogen receptors, which are putative S-equol receptors. Application of S-equol induced an increase in intracellular Ca levels via GPR30. However, S-equol did not enhance GLP-1 exocytosis, and long-term treatment of S-equol suppressed GLP-1 secretion. Moreover, immunocytochemistry revealed that S-equol increased the density of cortical actin filaments via G signaling under GPR30. These data suggest that S-equol prevents GLP-1 secretion as a result of competing regulation between Ca mobilization and actin reorganization.
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http://dx.doi.org/10.1016/j.bbrc.2018.05.100DOI Listing
July 2018

Transient receptor potential ankyrin 1 channels are involved in spontaneous peptide hormone release from astrocytes.

Biochem Biophys Res Commun 2018 07 24;501(4):988-995. Epub 2018 May 24.

Department of Life Sciences, Graduate School of Arts and Sciences, The University of Tokyo, 3-8-1 Komaba, Meguro, Tokyo, 153-8902, Japan. Electronic address:

Astrocytes, a large population of glial cells, detect neurotransmitters and respond by increasing intracellular Ca concentration ([Ca]) and releasing chemical molecules called gliotransmitters. Recently discovered Ca influx through transient receptor potential ankyrin 1 (TRPA1) channels is reported to cause spontaneous [Ca] increase in astrocytes. While several physiological functions of TRPA1-mediated spontaneous Ca signal have been revealed, relation with gliotransmitter release, especially peptide hormone exocytosis is largely unknown. We therefore explored the [Ca] and exocytosis dynamics in rat astrocyte cell line C6 cells and primary astrocytes. TRPA1-mediated spontaneous [Ca] transients were observed in both C6 cells and primary astrocytes. Total internal reflection fluorescence microscopy revealed that Venus-tagged brain-derived neurotrophic factor and neuropeptide Y were released spontaneously from astrocytes. Activation of TRPA1 channels enhanced the frequency of peptide hormone exocytosis, and inhibition of TRPA1 channels decreased the number of peptide hormone exocytosis. These results suggest that TRPA1-mediated spontaneous [Ca] increase modulates the spontaneous release of peptide hormones from astrocytes.
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http://dx.doi.org/10.1016/j.bbrc.2018.05.097DOI Listing
July 2018

Concordance between Aspergillus-specific precipitating antibody and IgG in allergic bronchopulmonary aspergillosis.

Allergol Int 2018 Sep;67S:S12-S17

Division of Pulmonary Medicine, Department of Medicine, Tokai University School of Medicine, Kanagawa, Japan.

Background: Several serological tests for specific precipitin or IgG are available to demonstrate type III hypersensitivity reactions to Aspergillus species and are essential for infectious fungal disease diagnosis. These assays are also important for allergic bronchopulmonary aspergillosis (ABPA) diagnosis; however, their concordance in ABPA was not well studied.

Methods: Fifty-two ABPA patients diagnosed based on ISHAM criteria were enrolled. Precipitins and IgG specific to Aspergillus fumigatus, Aspergillus niger, Aspergillus flavus, or Aspergillus terreus were measured using Ouchterlony double immunodiffusion tests and ImmunoCAP method, respectively. A. fumigatus-specific IgG was also determined using complement-fixation (CF) method.

Results: Forty-eight percent of cases were double-positive for A. fumigatus-specific precipitin and IgG (ImmunoCAP), whereas 3 (6%) and 14 (28%) cases were positive for precipitin or IgG alone, respectively. Kappa coefficient between these measurements was 0.32, suggesting poor concordance. Double-positive cases were more likely to present: Aspergillus sp. in sputum culture, lower pulmonary functions, peripheral blood eosinophilia, higher total IgE and A. fumigatus-specific IgG titer than precipitin-negative cases. A. fumigatus-specific IgG (CF) was positive only in 8 (15%) cases. The presence of A. fumigatus-specific precipitin or IgG was associated with antibodies specific for other Aspergillus spp., suggesting cross-reactivity.

Conclusions: Positive rate of A. fumigatus-specific precipitin or IgG (ImmunoCAP) was superior to IgG (CF), but relatively poor concordance was noted between precipitin and IgG (ImmunoCAP). Positive precipitin for A. fumigatus suggests more active diseases. Cross-reactivity may exist between antibodies to different Aspergillus spp. Therefore, the type III hypersensitivity results in ABPA diagnosis should be carefully evaluated.
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http://dx.doi.org/10.1016/j.alit.2018.04.009DOI Listing
September 2018

Species distribution, virulence factors, and antimicrobial resistance of Acinetobacter spp. isolates from dogs and cats: a preliminary study.

Microbiol Immunol 2018 May 12. Epub 2018 May 12.

The United Graduate School of Veterinary Science, Yamaguchi University, 1677-1 Yoshida, Yamaguchi 753-8515, Japan.

We investigated the prevalence of virulence factors and antimicrobial resistance among 67 Acinetobacter spp. isolates, consisting of 21 Acinetobacter baumannii and 46 non-baumannii Acinetobacter from companion animals. The PCR analysis showed that the most prevalent virulence gene was afa/draBC (29.9%), followed by papC (22.4%) and cvaC (20.9%). Antimicrobial susceptibility testing revealed that resistance to gentamicin (14.9%) and ciprofloxacin (11.9%) was relatively prevalent. Five gentamicin- and/or ciprofloxacin-resistant A. baumannii strains were assigned to ST25, ST149, ST164, ST203, and ST1198. All ciprofloxacin-resistant isolates harbored point mutations in gyrA and/or parC. This is the first preliminary monitoring of animal-origin Acinetobacter spp. in Japan.
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http://dx.doi.org/10.1111/1348-0421.12601DOI Listing
May 2018

Bitter tastant quinine modulates glucagon-like peptide-1 exocytosis from clonal GLUTag enteroendocrine L cells via actin reorganization.

Biochem Biophys Res Commun 2018 06 25;500(3):723-730. Epub 2018 Apr 25.

Department of Life Sciences, Graduate School of Arts and Sciences, The University of Tokyo, 3-8-1 Komaba, Meguro, Tokyo 153-8902, Japan; Department of Biological Sciences, Graduate School of Science, The University of Tokyo, 7-3-1 Hongo, Bunkyo, Tokyo 113-0033, Japan. Electronic address:

Enteroendocrine L cells in the gastrointestinal tract secrete glucagon-like peptide-1 (GLP-1), which plays an important role in glucose homeostasis. Here we investigated the effect of bitter tastant quinine on GLP-1 secretion using clonal GLUTag mouse enteroendocrine L cells. We found that GLUTag cells expressed putative quinine receptors at mRNA levels. Although application of quinine resulted in an increase of intracellular Ca levels, which was mediated by Ca release from the endoplasmic reticulum and Ca influx through voltage-sensitive Ca channels, quinine had little effect on GLP-1 secretion. Total internal reflection fluorescence microscopy and immunocytochemistry revealed that GLP-1-containing vesicles remained unfused with the plasma membrane and facilitated actin polymerization beneath the plasma membrane after application of quinine, respectively. Interestingly, application of forskolin together with quinine induced GLP-1 exocytosis from the cells. These results suggest that quinine does not induce GLP-1 secretion because it facilitates Ca increase and actin reorganization but not cAMP increase, and both Ca and cAMP are essential for GLP-1 secretion.
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http://dx.doi.org/10.1016/j.bbrc.2018.04.143DOI Listing
June 2018

Tigecycline Susceptibility of Klebsiella pneumoniae Complex and Escherichia coli Isolates from Companion Animals: The Prevalence of Tigecycline-Nonsusceptible K. pneumoniae Complex, Including Internationally Expanding Human Pathogenic Lineages.

Microb Drug Resist 2018 Jul/Aug;24(6):860-867. Epub 2017 Dec 12.

1 Department of Microbiology, Sapporo Medical University School of Medicine , Sapporo, Japan .

Transmission of tigecycline-nonsusceptible pathogenic Enterobacteriaceae from companion animals to human should be a concern because tigecycline is a last-line drug for treating multidrug-resistant Enterobacteriaceae in human medicine. However, tigecycline susceptibility of Enterobacteriaceae isolated from companion animals has not been investigated. In this study, we investigated the tigecycline susceptibility of Klebsiella pneumoniae complex and Escherichia coli isolates from dogs and cats, and evaluated their human pathogenicity potential. Tigecycline susceptibility of K. pneumoniae, including Klebsiella quasipneumoniae (n = 86) and E. coli (n = 100) strains isolated from dogs and cats was investigated. The antimicrobial susceptibility, capsular serotype, multilocus sequence type, ompK36 group, presence of virulence genes, and serum resistance of tigecycline-nonsusceptible isolates were evaluated. All E. coli isolates were susceptible to tigecycline. Two K. pneumoniae (minimum inhibitory concentration [MIC], 4 mg/L) and one K. quasipneumoniae (MIC, 8 mg/L) isolates were tigecycline resistant. Sixteen K. pneumoniae and one K. quasipneumoniae isolates were tigecycline intermediate (2 mg/L). All tigecycline-nonsusceptible isolates (n = 20) were also ciprofloxacin nonsusceptible. These isolates harbored five to nine virulence genes; 16 isolates were resistant to the human serum. In addition, STs of 13 K. pneumoniae isolates were reported to be found in strains isolated from human; isolates considered high-risk clones in human (ST11, ST15, and ST147) were also identified. In conclusion, the isolation of tigecycline-nonsusceptible K. pneumoniae from companion animals is an impact from the viewpoint of One Health approach to antimicrobial resistance that companion animals are a reservoir of human pathogenic lineages.
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http://dx.doi.org/10.1089/mdr.2017.0184DOI Listing
October 2018

Uterine torsion in a full-term pregnant cat.

JFMS Open Rep 2017 Jul-Dec;3(2):2055116917726228. Epub 2017 Aug 18.

Joint Department of Veterinary Clinical Medicine, Faculty of Agriculture, Tottori University, Tottori, Japan.

Case Summary: A 5-year-old intact female Maine Coon cat presented with a 2 day history of lethargy, anorexia and anaemia. The cat had bred 60 days previously and jumped from a height 3 days earlier, which was followed by a worsening of its condition. Ultrasonography revealed that two fetuses had died and one remained alive. Urgent surgical intervention was deemed necessary, and the cat underwent a blood transfusion and laparotomy. The right uterine horn was dark red in appearance and had rotated 360° in the clockwise direction at its base. Subsequently, an ovariohysterectomy and caesarean section were performed, and the fetus in the left uterine horn initially survived. Although the cat appeared to recover from anaemia and physical injury, the kitten died on postoperative day 1.

Relevance And Novel Information: In cases involving only one twisted uterine horn, the fetuses located in the contralateral horn could potentially survive; however, many such fetuses do not survive, and only a few reports have described fetal survival in a pregnant cat with uterine torsion. In the present case, early surgical intervention and blood transfusion allowed us to save the cat. Our findings demonstrate the life-saving abilities of initial support treatment and early surgical intervention for both the pregnant cat and fetuses in cases of acute abdomen caused by uterine torsion.
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http://dx.doi.org/10.1177/2055116917726228DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5564926PMC
August 2017
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