Publications by authors named "Kazuo Kitamura"

256 Publications

A likely unavoidable clinical scenario during treatment for venous thromboembolism complicated with severe immune thrombocytopenia: A case report.

Clin Case Rep 2021 09 18;9(9):e04805. Epub 2021 Sep 18.

Department of Internal Medicine, Circulatory and Body Fluid Regulation University of Miyazaki Miyazaki Japan.

Patients with immune thrombocytopenia have increased risks of bleeding and thrombosis. The acute-phase treatment for venous thromboembolism complicated with severe immune thrombocytopenia involves a "platelet dilemma" in therapeutic decision-making.
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http://dx.doi.org/10.1002/ccr3.4805DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8449094PMC
September 2021

Adrenomedullin: A Novel Therapeutic for the Treatment of Inflammatory Bowel Disease.

Biomedicines 2021 Aug 23;9(8). Epub 2021 Aug 23.

Department of Projects Research, Frontier Science Research Center, University of Miyazaki, Miyazaki 889-1692, Japan.

Adrenomedullin (AM) is a bioactive peptide with various physiological functions, including vasodilation, angiogenesis, anti-inflammation, organ protection, and tissue repair. AM suppresses inflammatory cytokine production in the intestinal mucosa, improves vascular and lymphatic regeneration and function, mucosal epithelial repair, and immune function in the intestinal bacteria of animal models with intestinal inflammation. We have been promoting translational research to develop novel therapeutic agents for inflammatory bowel disease (IBD) using AM and have started clinical research for IBD patients since 2010. A multicenter clinical trial is currently underway in Japan for patients with refractory ulcerative colitis and Crohn's disease. Moreover, since current AM administration is limited to continuous intravenous infusion, the development of a subcutaneous formulation using long-acting AM is underway for outpatient treatment.
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http://dx.doi.org/10.3390/biomedicines9081068DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8391925PMC
August 2021

A database and deep learning toolbox for noise-optimized, generalized spike inference from calcium imaging.

Nat Neurosci 2021 09 2;24(9):1324-1337. Epub 2021 Aug 2.

Friedrich Miescher Institute for Biomedical Research, Basel, Switzerland.

Inference of action potentials ('spikes') from neuronal calcium signals is complicated by the scarcity of simultaneous measurements of action potentials and calcium signals ('ground truth'). In this study, we compiled a large, diverse ground truth database from publicly available and newly performed recordings in zebrafish and mice covering a broad range of calcium indicators, cell types and signal-to-noise ratios, comprising a total of more than 35 recording hours from 298 neurons. We developed an algorithm for spike inference (termed CASCADE) that is based on supervised deep networks, takes advantage of the ground truth database, infers absolute spike rates and outperforms existing model-based algorithms. To optimize performance for unseen imaging data, CASCADE retrains itself by resampling ground truth data to match the respective sampling rate and noise level; therefore, no parameters need to be adjusted by the user. In addition, we developed systematic performance assessments for unseen data, openly released a resource toolbox and provide a user-friendly cloud-based implementation.
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http://dx.doi.org/10.1038/s41593-021-00895-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7611618PMC
September 2021

Combined evaluation of plasma B-type natriuretic peptide and urinary liver-type fatty acid-binding protein/creatinine ratio is related to worsening renal function in patients undergoing elective percutaneous coronary intervention.

Clin Exp Nephrol 2021 Dec 13;25(12):1319-1328. Epub 2021 Jul 13.

Department of Cardiovascular Medicine, National Cerebral and Cardiovascular Center, 6-1 Kishibe-Shimmachi, Suita, Osaka, 564-8565, Japan.

Background: There are few reports on the significance for the combined evaluation of blood humoral factors and urinary biomarkers in terms of worsening renal function (WRF) after coronary angiography (CAG)/percutaneous coronary arterial intervention (PCI).

Method And Results: Urinary liver type-fatty acid-binding protein (L-FABP), neutrophil gelatinase associated lipocalin (NGAL), and adrenomedullin (AM) were measured less than 24 h before and 3 h, 6 h, 1 day, and 2 days after CAG/PCI. WRF was defined as a > 20% decrease in the estimated GFR. WRF occurred in seven of 100 patients and the increase in L-FABP/creatinine (Cr) at 1 day after CAG/PCI was significantly higher in the WRF group than in the non-WRF group. Plasma B-type natriuretic peptide (BNP) before CAG/PCI and L-FABP/Cr at 1 day after CAG/PCI were independent predictors for WRF. The areas under the receiver-operating characteristic curves were as follows: 0.760 for BNP before CAG/PCI, 0.731 for L-FABP/Cr at 1 day after CAG/PCI, and 0.892 for BNP and L-FABP/Cr. Urinary AM levels after PCI/CAG were negatively correlated only to serum potassium levels. Gene expressions of AM and AM-receptor were detectable in renal tubule epithelial cells. AM increased intracellular second messenger levels in a dose-dependent manner.

Conclusions: Our results suggest that combined evaluation of plasma BNP and urinary L-FABP/Cr is useful as a predictor of renal dysfunction in CAG/PCI patients.
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http://dx.doi.org/10.1007/s10157-021-02113-9DOI Listing
December 2021

The usefulness of plasma levels of mature and total adrenomedullin as biomarkers indicating the magnitude of surgical stress responses: A single-center, prospective, observational study.

J Clin Transl Res 2021 Jun 14;7(3):302-310. Epub 2021 May 14.

Department of Anesthesiology, Faculty of Medicine, University of Miyazaki, Miyazaki 889-1692, Japan.

Background And Aim: Adrenomedullin (AM), a vasodilatory peptide, is known for its pleiotropic actions. AM levels are increased under inflammatory conditions such as sepsis and can be useful as a prognostic biomarker. However, there are only a few reports on the physiological actions of AM in the perioperative period. The aim of this single-center, prospective, and observational study was to investigate the changes in the plasma levels of mature AM (mAM) and total AM (tAM) observed during the perioperative period. In addition, we aimed to determine the association between each AM level and immune-inflammatory parameters to explore the usefulness of AM as a biomarker of the magnitude of surgical stress responses.

Methods: The levels of both mAM and tAM, in addition to the levels of presepsin, interleukin-6, procalcitonin, white blood cell, and C-reactive protein, were measured in blood samples obtained during the perioperative period. Other laboratory data, including sequential organ failure assessment (SOFA) and acute physiology and chronic health evaluation (APACHE) II scores, were obtained from individual clinical records. Correlations between each AM and clinical parameters were determined using Spearman's rank correlation. <0.05 were considered statistically significant.

Results: One hundred and twenty-three perioperative patients scheduled for three types of surgical procedures, including cardiopulmonary bypass surgery, abdominal surgery, and cervical laminoplasty, were included in this study. There was a moderate to strong correlation between each AM and immune-inflammatory parameters, SOFA score, and APACHE II score, as related to surgical trauma. Specifically, the strongest correlation was observed between each AM and SOFA score.

Conclusions: These findings suggest that plasma AM levels may represent the most important inflammatory mediators that are evident in surgical stress responses.

Relevance For Patients: Since the levels of both tAM and mAM show the same trend, mAM and tAM may be equally used as biomarkers for the evaluation of the physiological status of surgical patients.

Trial Registration: This observational study was retrospectively registered with Japanese Clinical Trial Registry "UMIN-CTR" on March 19, 2018, and was given a trial ID number UMIN000031792.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8221752PMC
June 2021

Intracellular glutamine level determines vascular smooth muscle cell-derived thrombogenicity.

Atherosclerosis 2021 07 26;328:62-73. Epub 2021 May 26.

Department of Pathology, Faculty of Medicine, University of Miyazaki, Japan. Electronic address:

Background And Aims: The everolimus-eluting stent (EES), one of the effective stents for in-stent restenosis (ISR), has a lower incidence of stent thrombosis; however, the underlying mechanism remains unknown. This study aimed to identify the effects of everolimus on vascular metabolism and thrombogenicity and examine their mechanistic link.

Methods: EESs and bare-metal stents were implanted in rabbit iliac arteries with smooth muscle cell (SMC)-rich neointima induced by endothelial denudation. Four weeks after stent implantation, the stented arteries were examined for histological analysis and metabolomics. Additionally, everolimus effects in coronary artery SMCs metabolism, tissue factor (TF) expression, and procoagulant activity were assessed in vitro.

Results: EES-implanted arteries showed decreased neointima formation, less SMCs infiltration, and reduced TF expression. Concomitantly, they were metabolically characterized by increased levels of metabolites in amino acids, such as glutamine. Similarly, everolimus increased intracellular glutamine levels, decreased TF expression, and reduced procoagulant activity in SMCs in vitro. On the contrary, exogenous glutamine administration also increased intracellular glutamine level, decreased TF expression, and reduced procoagulant activity despite enhanced mammalian target of rapamycin (mTOR) activity.

Conclusions: Intracellular glutamine level is likely to determine vascular SMC-related thrombogenicity regardless of mTOR pathway activity. Therefore, increased intracellular glutamine level might contribute partially to the beneficial effect of EES use on stent thrombosis.
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http://dx.doi.org/10.1016/j.atherosclerosis.2021.05.012DOI Listing
July 2021

Differential effects of the formin inhibitor SMIFH2 on contractility and Ca handling in frog and mouse cardiomyocytes.

Genes Cells 2021 Aug 17;26(8):583-595. Epub 2021 Jun 17.

Department of Pharmacology, Faculty of Medicine, University of Miyazaki, Miyazaki, Japan.

Genetic mutations in actin regulators have been emerging as a cause of cardiomyopathy, although the functional link between actin dynamics and cardiac contraction remains largely unknown. To obtain insight into this issue, we examined the effects of pharmacological inhibition of formins, a major class of actin-assembling proteins. The formin inhibitor SMIFH2 significantly enhanced the cardiac contractility of isolated frog hearts, thereby augmenting cardiac performance. SMIFH2 treatment had no significant effects on the Ca sensitivity of frog muscle fibers. Instead, it unexpectedly increased Ca concentrations of isolated frog cardiomyocytes, suggesting that the inotropic effect is due to enhanced Ca transients. In contrast to frog hearts, the contractility of mouse cardiomyocytes was attenuated by SMIFH2 treatment with decreasing Ca transients. Thus, SMIFH2 has opposing effects on the Ca transient and contractility between frog and mouse cardiomyocytes. We further found that SMIFH2 suppressed Ca -release via type 2 ryanodine receptor (RyR2); this inhibitory effect may explain the species differences, since RyR2 is critical for Ca transients in mouse myocardium but absent in frog myocardium. Although the mechanisms underlying the enhancement of Ca transients in frog cardiomyocytes remain unclear, SMIFH2 differentially affects the cardiac contraction of amphibian and mammalian by differentially modulating their Ca handling.
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http://dx.doi.org/10.1111/gtc.12873DOI Listing
August 2021

Plasma adrenomedullin level and year-by-year variability of body mass index in the general population.

Peptides 2021 08 5;142:170567. Epub 2021 May 5.

Frontier Science Research Center, Faculty of Medicine, University of Miyazaki, Kiyotake, Miyazaki, 889-1692, Japan.

Plasma levels of the hypotensive peptides of adrenomedullin and atrial and B-type natriuretic peptides (AM, ANP, BNP) are possible biomarkers for cardiovascular diseases. Increased variability of body mass index (BMI) over a certain period of time has been reported to be associated with cardiovascular morbidity or mortality. The aim of this study is to examine clinical significance of those hypotensive peptides as biomarkers by analyzing the relationship between plasma levels of the peptides and year-by-year variability of BMI in the general population without overt cardiovascular diseases. The subjects were 427 local residents (141 males and 286 females) attending their annual health check-up, who had been examined at least 5 times over the preceding period of 10 years. They were divided into two groups of low or high variability by the median of coefficient of variation (CV) of BMI values for each gender. Plasma AM levels of those with high year-by-year variability of BMI were significantly increased, as compared to the group with low variability, in both genders; meanwhile, such a difference was not noted in plasma levels of the natriuretic peptides. No significant differences were found in the basal parameters, which could affect plasma AM level, such as age, BMI, blood pressure or serum creatinine, between two groups. In conclusion, increase in plasma AM was associated with high year-by-year variability of BMI in the general population without overt heart disease. This relationship between the two suggests that increased plasma AM level is a cardiovascular risk marker.
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http://dx.doi.org/10.1016/j.peptides.2021.170567DOI Listing
August 2021

Development of a novel human adrenomedullin derivative: human serum albumin-conjugated adrenomedullin.

J Biochem 2021 May 8. Epub 2021 May 8.

Circulatory and Body Fluid Regulation, Faculty of Medicine, University of Miyazaki, 5200 Kihara, Kiyotake, Miyazaki 889-1692, Japan.

Adrenomedullin is a biologically active peptide with multiple functions. Here, we have developed a novel human serum albumin-adrenomedullin (HSA-AM) conjugate, which was synthesized by the covalent attachment of a maleimide derivative of adrenomedullin to the 34th cysteine residue of human serum albumin via a linker. Denaturing gel electrophoresis and Western blotting for HSA-AM yielded a single band with adrenomedullin immunoreactivity at the position corresponding to a molecular weight (MW) of 73 kDa. Following gel-filtration chromatography, the purified HSA-AM showed a single main peak corresponding with a MW of 73 kDa, indicating that HSA-AM is a monomer. Both adrenomedullin and HSA-AM stimulated the intracellular accumulation of cyclic AMP in HEK-293 cells stably expressing the adrenomedullin 1 receptor. The pEC50 values for adrenomedullin and HSA-AM were 8.660 and 7.208 (equivalent to 2.19 nM and 61.9 nM as EC50), respectively. The bioavailability of HSA-AM compared with that of adrenomedullin was much improved after subcutaneous administration in the rat, which was probably due to the superior resistance of HSA-AM toward endogenous proteases and its reduced clearance from the blood. HSA-AM may be a promising drug candidate for clinical application.
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http://dx.doi.org/10.1093/jb/mvab057DOI Listing
May 2021

Study Protocol for a Randomized, Double-Blind, Placebo-Controlled, Phase-II Trial: AdrenoMedullin for Ischemic Stroke Study.

J Stroke Cerebrovasc Dis 2021 Jun 1;30(6):105761. Epub 2021 Apr 1.

Department of Neurology, National Cerebral and Cardiovascular Center, 6-1 Kishibe-Shimmachi, Suita, Osaka 564-8565, Japan. Electronic address:

Objectives: Adrenomedullin (AM), a vasoactive peptide, has strong anti-inflammatory and angiogenic properties, which have been reported to ameliorate the consequences of ischemic stroke in several animal models. After a phase I study in healthy volunteers, two phase II trials of AM for inflammatory bowel diseases have been recently completed. The current AdrenoMedullin For Ischemic Stroke (AMFIS) study aims to assess the safety and efficacy of AM in patients with acute ischemic stroke.

Materials And Methods: The AMFIS study is an investigator-initiated, randomized, double-blind, phase-II trial. AM or placebo will be administered to patients with non-cardioembolic ischemic stroke within 24 h after stroke onset. In the first cohort of the AMFIS study, patients will be randomly allocated to the investigation treatment A (30 μg/kg of AM in total for 7 days, n = 20) or placebo group (n = 10). In the second cohort, patients will be assigned to the investigation treatment B (56 μg/kg of AM in total for 7 days, n = 20) or placebo group (n = 10).

Results: Serious adverse events related to the protocol treatment will be evaluated as the primary outcome. All adverse events will be analyzed as the secondary outcome. Regarding efficacy endpoints, the change in National Institutes of Health Stroke Scale and modified Rankin Scale scores will be compared between investigation treatment and placebo groups.

Conclusions: AM is expected to be a safe and effective treatment for ischemic stroke.
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http://dx.doi.org/10.1016/j.jstrokecerebrovasdis.2021.105761DOI Listing
June 2021

Development of a novel AlphaLISA ImmunoAssay for Big angiotensin-25.

Nephrology (Carlton) 2021 May 30;26(5):479-484. Epub 2020 Dec 30.

Circulatory and Body Fluid Regulation, Faculty of Medicine, University of Miyazaki, Miyazaki, Japan.

We previously described the discovery of Big angiotensin-25 (Bang-25), an angiotensin-related peptide isolated from human urine. Bang-25 consists of the first 25 amino acids of the N-terminus of angiotensinogen (Aogen), with N-linked glycosylation on the 14th amino acid and a cysteine conjugated to the 18th amino acid. Bang-25 is rapidly converted into angiotensin II (Ang II) by chymase. Because Bang-25 is widely distributed in human tissues, including islet cells in the pancreas and podocytes in the kidney, we hypothesized that it may participate in the Ang II production system in these tissues. To test this hypothesis, we developed a specific assay for Bang-25 and used it to examine the urinary concentrations of Bang-25 in patients with diabetes mellitus (DM). The assay used the Amplified Luminescent Proximity Homogeneous Assay (Alpha)-based ELISA method (AlphaLISA) of PerkinElmer Japan and included antibodies specific to the N-terminus of Ang II and the C-terminus of Bang-25. The AlphaLISA ImmunoAssay specifically recognized Bang-25 and had no cross-reactivity with Aogen or Ang I. Bang-25 was detected in healthy volunteers' urine samples but not in their plasma samples. In patients with DM, the urinary Bang-25 concentration was significantly higher than in healthy volunteers. Moreover, the results indicated that the Bang-25 concentration in the urine may offer a different perspective on disease status from that provided by urinary albumin. This assay could provide a useful tool for determining urinary Bang-25, which may prove an important biomarker for diabetic kidney disease.
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http://dx.doi.org/10.1111/nep.13845DOI Listing
May 2021

Upregulated Kynurenine Pathway Enzymes in Aortic Atherosclerotic Aneurysm: Macrophage Kynureninase Downregulates Inflammation.

J Atheroscler Thromb 2021 Nov 10;28(11):1214-1240. Epub 2020 Dec 10.

Department of Pathology, Faculty of Medicine, University of Miyazaki.

Aims: Inflammation and hypertension contribute to the progression of atherosclerotic aneurysm in the aorta. Vascular cell metabolism is regarded to modulate atherogenesis, but the metabolic alterations that occur in atherosclerotic aneurysm remain unknown. The present study aimed to identify metabolic pathways and metabolites in aneurysmal walls and examine their roles in atherogenesis.

Methods: Gene expression using microarray and metabolite levels in the early atherosclerotic lesions and aneurysmal walls obtained from 42 patients undergoing aortic surgery were investigated (early lesion n=11, aneurysm n=35) and capillary electrophoresis-time-of-flight mass spectrometry (early lesion n=14, aneurysm n=38). Using immunohistochemistry, the protein expression and localization of the identified factors were examined (early lesion n=11, non-aneurysmal advanced lesion n=8, aneurysm n=11). The roles of the factors in atherogenesis were analyzed in macrophages derived from human peripheral blood mononuclear cells.

Results: Enrichment analysis using 35 significantly upregulated genes (log2 ratio, >3) revealed the alteration of the kynurenine pathway. Metabolite levels of tryptophan, kynurenine, and quinolinic acid and the kynurenine-to-tryptophan ratio were increased in the aneurysmal walls. Gene and protein expression of kynureninase and kynurenine 3-monooxygenase were upregulated and localized in macrophages in the aneurysmal walls. The silencing of kynureninase in the cultured macrophages enhanced the expression of interleukin-6 and indoleamine 2,3-dioxygenase 1.

Conclusion: Our study suggests the upregulation of the kynurenine pathway in macrophages in aortic atherosclerotic aneurysm. Kynureninase may negatively regulate inflammation via the kynurenine pathway itself in macrophages.
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http://dx.doi.org/10.5551/jat.58248DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8592691PMC
November 2021

Adrenomedullin for steroid-resistant ulcerative colitis: a randomized, double-blind, placebo-controlled phase-2a clinical trial.

J Gastroenterol 2021 02 2;56(2):147-157. Epub 2020 Nov 2.

Division of Circulatory and Body Fluid Regulation, Department of Internal Medicine, Faculty of Medicine, University of Miyazaki, 5200 Kihara, Miyazaki, Miyazaki, 889-1692, Japan.

Background: Adrenomedullin (AM) is a bioactive peptide having many pleiotropic effects, including mucosal healing and immunomodulation. AM has shown beneficial effects in rodent models and in preliminary study for patients with ulcerative colitis (UC). We performed a clinical trial to investigate the efficacy and safety of AM in patients with UC.

Methods: This was a multi-center, double-blind, placebo-controlled phase-2a trial evaluating 28 patients in Japan with steroid-resistant UC. Patients were randomly assigned to four groups and given an infusion of 5, 10, 15 ng/kg/min of AM or placebo for 8 h per day for 14 days. The primary endpoint was the change in Mayo scores at 2 weeks. Main secondary endpoints included the change in Mayo scores and the rate of clinical remission at 8 weeks, defined as a Mayo score 0.

Results: No differences in the primary or secondary endpoints were observed among the four groups at 2 weeks. Despite the insufficient tracking rate, the Mayo score at 8 weeks was only significantly decreased in the high-dose AM group (15 ng/kg/min) compared with the placebo group (- 9.3 ± 1.2 vs. - 3.0 ± 2.8, P = 0.035), with its rate of clinical remission at 8 weeks being significantly higher (3/3, 100% vs. 0/2, 0%, P = 0.025). We noted mild but no serious adverse events caused by the vasodilatory effect of AM.

Conclusions: In this double-blind randomized trial, we observed the complete remission at 8 weeks in patients with steroid-resistant UC receiving a high dose of AM.

Clinical Trial Registry: JAPIC clinical trials information; Japic CTI-205255 (200410115290). https://www.clinicaltrials.jp/cti-user/trial/Search.jsp .
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http://dx.doi.org/10.1007/s00535-020-01741-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7862507PMC
February 2021

Activation of Calcitonin Gene-Related Peptide and Adrenomedullin Receptors by PEGylated Adrenomedullin.

Biol Pharm Bull 2020 ;43(11):1799-1803

Circulatory and Body Fluid Regulation, Faculty of Medicine, University of Miyazaki.

Adrenomedullin (AM) improves colitis in animal models and patients with inflammatory bowel disease. We have developed a PEGylated AM derivative (PEG-AM) for clinical application because AM has a short half-life in the blood. However, modification by addition of polyethylene glycol (PEG) may compromise the function of the original peptide. In this paper, we examined the time course of cAMP accumulation induced by 5 and 60 kDa PEG-AM and compared the activation of calcitonin gene-related peptide (CGRP), AM1 and AM2 receptors by AM, 5 and 60 kDa PEG-AM. We also evaluated the effects of antagonists on the action of 5 and 60 kDa PEG-AM. PEG-AM stimulated cAMP production induced by these receptors; the increase in cAMP levels resulting from application of PEG-AM peaked at 15 min. Moreover, PEG-AM activity was antagonized by CGRP (8-37) or AM (22-52) (antagonists of CGRP and AM receptors, respectively) and the maximal response was not suppressed. These findings indicate that the effects of PEG-AM are similar to those of native AM.
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http://dx.doi.org/10.1248/bpb.b20-00373DOI Listing
July 2021

Improved hyperacuity estimation of spike timing from calcium imaging.

Sci Rep 2020 10 20;10(1):17844. Epub 2020 Oct 20.

ATR Brain Information Communication Research Laboratory Group, Advanced Telecommunications Research Institute International, Kyoto, Japan.

Two-photon imaging is a major recording technique used in neuroscience. However, it suffers from several limitations, including a low sampling rate, the nonlinearity of calcium responses, the slow dynamics of calcium dyes and a low SNR, all of which severely limit the potential of two-photon imaging to elucidate neuronal dynamics with high temporal resolution. We developed a hyperacuity algorithm (HA_time) based on an approach that combines a generative model and machine learning to improve spike detection and the precision of spike time inference. Bayesian inference was performed to estimate the calcium spike model, assuming constant spike shape and size. A support vector machine using this information and a jittering method maximizing the likelihood of estimated spike times enhanced spike time estimation precision approximately fourfold (range, 2-7; mean, 3.5-4.0; 2SEM, 0.1-0.25) compared to the sampling interval. Benchmark scores of HA_time for biological data from three different brain regions were among the best of the benchmark algorithms. Simulation of broader data conditions indicated that our algorithm performed better than others with high firing rate conditions. Furthermore, HA_time exhibited comparable performance for conditions with and without ground truths. Thus HA_time is a useful tool for spike reconstruction from two-photon imaging.
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http://dx.doi.org/10.1038/s41598-020-74672-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7576127PMC
October 2020

The Cytokine Expression in Patients with Cardiac Complication after Immune Checkpoint Inhibitor Therapy.

Intern Med 2021 Feb 19;60(3):423-429. Epub 2020 Sep 19.

Department of Pharmacy, University of Miyazaki Hospital, Japan.

We herein report the cytokine expression at different stages for three patients who developed cardiac complications after immune checkpoint inhibitor (ICI) therapy. Case 1 with biopsy-proven myocarditis showed increased levels of interleukin (IL)-8, monocyte chemotactic and activating factor, and granulocyte macrophage colony-stimulating factor (GM-CSF) when he developed Takotsubo cardiomyopathy. Case 2 with subclinical myocarditis showed predominant activation of IL-8 during the progressive clinical course. Case 3 with cytokine-releasing syndrome showed substantial activations of IL-6, IL-8, GM-CSF, and interferon-γ. Our data suggest the development of unique cytokine activation in individual patients with cardiac complications after ICI therapy.
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http://dx.doi.org/10.2169/internalmedicine.5317-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7925267PMC
February 2021

Thrombin rapidly digests adrenomedullin: Synthesis of adrenomedullin analogs resistant to thrombin.

Biochem Biophys Res Commun 2020 08 19;529(3):778-783. Epub 2020 Jul 19.

Circulatory and Body Fluid Regulation, Faculty of Medicine, University of Miyazaki, Kiyotake, Miyazaki, 889-1692, Japan. Electronic address:

Human adrenomedullin (AM) functions as a circulating hormone and as a local paracrine mediator with multiple biological activities. We investigated the metabolism of AM by examining its fragmentation in human serum. Adrenomedullin was rapidly cleaved in human serum, but was relatively stable in plasma. We showed that AM was rapidly digested by thrombin in serum, with AM(13-44) as the main product. On the basis of these data, we prepared AM analogs in which Arg-44 was replaced by Ala, Lys, and D-Arg, respectively. These analogs were resistant to thrombin and showed comparable biological activity to native AM. Furthermore, the bioavailabilities of these peptides were improved after subcutaneous administration in rats. These AM analogs may be promising drug candidates for clinical applications.
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http://dx.doi.org/10.1016/j.bbrc.2020.06.057DOI Listing
August 2020

Activation of the reward system ameliorates passive cutaneous anaphylactic reaction in mice.

Allergy 2020 12 29;75(12):3275-3279. Epub 2020 Jun 29.

Department of Immunology, Faculty of Medicine, University of Yamanashi, Yamanashi, Japan.

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http://dx.doi.org/10.1111/all.14442DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7754286PMC
December 2020

Questionnaire in patients with aborted sudden cardiac death due to coronary spasm in Japan.

Heart Vessels 2020 Dec 12;35(12):1640-1649. Epub 2020 Jun 12.

Toyama Prefectural Central Hospital, Toyama, Japan.

Objectives: We investigated the medical or mechanical therapy, and the present knowledge of Japanese cardiologists about aborted sudden cardiac death (ASCD) due to coronary spasm.

Methods: A questionnaire was developed regarding the number of cases of ASCD, implantable cardioverter-defibrillator (ICD), and medical therapy in ASCD patients due to coronary spasm. The questionnaire was sent to the Japanese general institutions at random in 204 cardiology hospitals.

Results: The completed surveys were returned from 34 hospitals, giving a response rate of 16.7%. All SCD during the 5 years was observed in 5726 patients. SCD possibly due to coronary spasm was found in 808 patients (14.0%) and ASCD due to coronary spasm was observed in 169 patients (20.9%). In 169 patients with ASCD due to coronary spasm, one or two coronary vasodilators was administered in two-thirds of patients [113 patients (66.9%)], while more than 3 coronary vasodilators were found in 56 patients (33.1%). ICD was implanted in 117 patients with ASCD due to coronary spasm among these periods including 35 cases with subcutaneous ICD. Majority of cause of ASCD was ventricular fibrillation, whereas pulseless electrical activity was observed in 18 patients and complete atrioventricular block was recognized in 7 patients. Mean coronary vasodilator number in ASCD patients with ICD was significantly lower than that in those without ICD (2.1 ± 0.9 vs. 2.6 ± 1.0, p < 0.001). Although 16 institutions thought that the spasm provocation tests under the medications had some clinical usefulness of suppressing the next fatal arrhythmias, spasm provocation tests under the medication were performed in just 4 institutions.

Conclusions: In the real world, there was no fundamental strategy for patients with ASCD due to coronary spasm. Each institution has each strategy for these patients. Cardiologists should have the same strategy and the same knowledge about ASCD patients due to coronary spasm in the future.
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http://dx.doi.org/10.1007/s00380-020-01644-7DOI Listing
December 2020

Nephrotic syndrome and Giant cell arthritis concurrently occurring after percutaneous transluminal angioplasty for renal artery stenosis
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Clin Nephrol 2020 Jun;93(6):300-305

An elderly Japanese woman with bilateral renal artery occlusion who developed massive proteinuria after unilateral percutaneous transluminal renal angioplasty (PTRA) is reported. She had a history of percutaneous coronary intervention and subsequently developed refractory hypertension. She was diagnosed with renovascular hypertension caused by bilateral total occlusion of the renal arteries, and underwent PTRA for the left renal artery. Nephrotic-range proteinuria from the left kidney, confirmed by split urine collection from each kidney under cytoscopic examination, and low-grade fever with positive C-reactive protein became obvious after PTRA. Giant cell arteritis (GCA) was also diagnosed by positive findings on fluorodeoxyglucose-positron emission tomography in the common carotid arteries, subclavian arteries, and aorta, but not in the renal arteries. Administration of corticosteroid and angiotensin-converting enzyme inhibitor decreased the proteinuria (> 9 - 2 g/day). Possible mechanisms for the development of nephrotic-range proteinuria and a hypothesis that GCA became obvious after PTRA are discussed.
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http://dx.doi.org/10.5414/CN110000DOI Listing
June 2020

Safety, Tolerability, and Pharmacokinetics of Adrenomedullin in Healthy Males: A Randomized, Double-Blind, Phase 1 Clinical Trial.

Drug Des Devel Ther 2020 6;14:1-11. Epub 2020 Jan 6.

Division of Circulatory and Body Fluid Regulation, Department of Internal Medicine, Faculty of Medicine, University of Miyazaki, Miyazaki, Japan.

Background: Adrenomedullin (AM), an endogenous vasodilative peptide, has immunomodulative effects and acts as an accelerator of mucosal regeneration in the digestive tract. AM has shown beneficial effects in rodent models of inflammatory bowel disease and patients with ulcerative colitis. The present study aimed to evaluate the pharmacodynamic properties and safety of AM in healthy male adults in a phase 1 clinical trial.

Methods: This phase 1, randomized, double-blind, single-center study was conducted on healthy males aged 20-65 years. Subjects received either a placebo, 3 ng/kg/min AM, 9 ng/kg/min AM, or 15 ng/kg/min AM via continuous 12-h intravenous infusion. Other subjects received either placebo or 15 ng/kg/min AM for 8 h per day for 7 days. Adverse events (AEs), vital signs, physical examinations, laboratory tests, electrocardiograms (ECG), and pharmacokinetics were assessed.

Findings: All 24 subjects in the single-dose test completed the study. Of the 12 subjects in multiple dosing test, one from the AM group withdrew owing to a headache. No serious AEs were reported. Hemodynamic parameters were well maintained in all subjects. Slight ECG abnormalities were observed in the single-dose test. The plasma concentration of AM progressively increased in a dose-dependent manner and reached C at the end of administration. Plasma AM rapidly returned to baseline concentrations after termination, with a T of under 60 min.

Interpretation: This is the first phase 1 trial in healthy men evaluating the safety of AM. Our results demonstrate the safety and tolerability of AM for subsequent Phase 2 trials.
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http://dx.doi.org/10.2147/DDDT.S225220DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6955635PMC
December 2020

Podocyte hypertrophic stress and detachment precedes hyperglycemia or albuminuria in a rat model of obesity and type2 diabetes-associated nephropathy.

Sci Rep 2019 12 6;9(1):18485. Epub 2019 Dec 6.

Division of Nephrology, Department of Internal Medicine, Faculty of Medicine, University of Miyazaki, Miyazaki, Japan.

Type2 diabetes-associated nephropathy is the commonest cause of renal failure. Mechanisms responsible are controversial. Leptin-deficient hyperphagic Zucker (fa/fa) rats were modeled to test the hypothesis that glomerular enlargement drives podocyte hypertrophic stress leading to accelerated podocyte detachment, podocyte depletion, albuminuria and progression. By 6weeks, prior to development of either hyperglycemia or albuminuria, fa/fa rats were hyperinsulinemic with high urinary IGF1/2 excretion, gaining weight rapidly, and had 1.6-fold greater glomerular volume than controls (P < 0.01). At this time the podocyte number per glomerulus was not yet reduced although podocytes were already hypertrophically stressed as shown by high podocyte phosphor-ribosomal S6 (a marker of mTORC1 activation), high urinary pellet podocin:nephrin mRNA ratio and accelerated podocyte detachment (high urinary pellet podocin:aquaporin2 mRNA ratio). Subsequently, fa/fa rats became both hyperglycemic and albuminuric. 24 hr urine albumin excretion correlated highly with decreasing podocyte density (R = 0.86), as a consequence of both increasing glomerular volume (R = 0.70) and decreasing podocyte number (R = 0.63). Glomerular podocyte loss rate was quantitatively related to podocyte detachment rate measured by urine pellet mRNAs. Glomerulosclerosis occurred when podocyte density reached <50/10um. Reducing food intake by 40% to slow growth reduced podocyte hypertrophic stress and "froze" all elements of the progression process in place, but had small effect on hyperglycemia. Glomerular enlargement caused by high growth factor milieu starting in pre-diabetic kidneys appears to be a primary driver of albuminuria in fa/fa rats and thereby an under-recognized target for progression prevention. Progression risk could be identified prior to onset of hyperglycemia or albuminuria, and monitored non-invasively by urinary pellet podocyte mRNA markers.
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http://dx.doi.org/10.1038/s41598-019-54692-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6898392PMC
December 2019

Urinary podocyte mRNA is a potent biomarker of anti-neutrophil cytoplasmic antibody-associated glomerulonephritis.

Clin Exp Nephrol 2020 Mar 25;24(3):242-252. Epub 2019 Nov 25.

Department of Nephrology, University of Miyazaki Hospital, Faculty of Medicine, University of Miyazaki, Miyazaki, Japan.

Background: Anti-neutrophil cytoplasmic antibody-associated glomerulonephritis (ANCA-GN) is a critical kidney disease that sometimes results in an unfavorable renal outcome. Cellular crescent formation is a hallmark of ANCA-GN and is associated with renal prognosis, response to treatment, and it was reportedly associated with podocyte detachment. Because there is a need to explore non-invasive biomarkers for the evaluation of ANCA-GN activity, we tested whether urinary podocyte mRNA might be a potent non-invasive biomarker.

Methods: We measured two different types of urinary podocyte mRNA, including podocin mRNA in relation to urine creatinine concentration (U-PodCR) and urinary podocin mRNA in relation to nephrin mRNA (U-PNR), which were reportedly associated with the activity of various glomerular diseases.

Results: In ANCA-GN patients (n = 19), we discovered that U-PodCR was positively correlated with the percent of crescent formation until 50% crescent was reached because of podocyte depletion; U-PNR was correlated with the percent of crescent formation in all patients. Furthermore, patients with high levels of urinary podocyte mRNA exhibited a favorable renal outcome compared with the outcomes of patients with low levels of urinary podocyte mRNA. The levels of urinary podocyte mRNA were correlated with the rate of improvement in estimated glomerular filtration rate.

Conclusions: U-PodCR, U-PNR or a combination of these parameters might serve as a non-invasive potential biomarker in patients with ANCA-GN to predict the percent of crescent formation and renal prognosis.
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http://dx.doi.org/10.1007/s10157-019-01823-5DOI Listing
March 2020

Modular organization of cerebellar climbing fiber inputs during goal-directed behavior.

Elife 2019 10 9;8. Epub 2019 Oct 9.

Department of Neurophysiology, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan.

The cerebellum has a parasagittal modular architecture characterized by precisely organized climbing fiber (CF) projections that are congruent with alternating aldolase C/zebrin II expression. However, the behavioral relevance of CF inputs into individual modules remains poorly understood. Here, we used two-photon calcium imaging in the cerebellar hemisphere Crus II in mice performing an auditory go/no-go task to investigate the functional differences in CF inputs to modules. CF signals in medial modules show anticipatory decreases, early increases, secondary increases, and reward-related increases or decreases, which represent quick motor initiation, go cues, fast motor behavior, and positive reward outcomes. CF signals in lateral modules show early increases and reward-related decreases, which represent no-go and/or go cues and positive reward outcomes. The boundaries of CF functions broadly correspond to those of aldolase C patterning. These results indicate that spatially segregated CF inputs in different modules play distinct roles in the execution of goal-directed behavior.
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http://dx.doi.org/10.7554/eLife.47021DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6844646PMC
October 2019

Non-canonical Expression of Cardiac Troponin-T in Neuroendocrine Ethmoid Sinus Carcinoma Following Immune Checkpoint Blockade.

Front Cardiovasc Med 2019 23;6:124. Epub 2019 Aug 23.

Department of Internal Medicine, Circulatory and Body Fluid Regulation, Faculty of Medicine, University of Miyazaki, Miyazaki, Japan.

We describe the case of a patient with neuroendocrine ethmoid sinus carcinoma, who exhibited markedly elevated levels of serum cardiac troponin-T and creatine kinase (CK)-MB isoenzyme without any symptom after the administration of nivolumab, immune checkpoint inhibitor. The repeated 12-leads-electrocardiogram did not show any changes in the ST-T segments or arrhythmias. The echocardiogram showed normal ranges of left ventricular contraction in the clinical course. Cardiac magnetic resonance imaging showed minimal myocardial edema and inflammation. Blood clots in the metastatic lesion of bone marrow aspirates exhibited positive staining for cardiac troponin-T and CK-MB in the cytoplasm and nucleoplasm of neoplastic cells. Although we did not perform a second cardiac magnetic resonance imaging and autopsy, we postulate that the attack of the neoplastic cells by the immune checkpoint inhibitor or the secretion from neoplastic cell-derived extracellular vesicles may have exacerbated the increase in concentrations of these molecules in the blood. Our case should warrant consideration a false-positive value of cardiac troponin-T and CK-MB can be obtained in cases with malignancy.
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http://dx.doi.org/10.3389/fcvm.2019.00124DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6716019PMC
August 2019

Augmented blood pressure variability following continuous infusion of noradrenaline in rats.

J Hypertens 2020 02;38(2):314-321

Frontier Science Research Center.

Objective: Augmented blood pressure (BP) variability has been shown to be associated with cardiovascular diseases. Activity of the sympathetic nervous system is an important determinant factor of the 24-h profile of BP variability, although it is unknown whether persistent adrenergic activation causes augmented BP variability or not. Here we report that continuous infusion of noradrenalin augments 24-h BP variability in rats.

Methods: Nine-week-old male Wistar rats were continuously infused with subcutaneous 30 μg/h noradrenalin, 150 μg/h of the α1-adrenergic agonist phenylephrine, or 30 μg/h of the β-agonist isoproterenol, for 14 days. Noradrenalin-infused rats were also administered either oral 5 mg/day prazosin or 50 mg/day atenolol during the infusion period. BP variability was evaluated before and after 7 and 14 days of the infusion, using a coefficient of variation of BP recorded every 15 min under an unrestrained condition via an abdominal aortic catheter by a radiotelemetry system.

Results: Continuous infusion of noradrenalin significantly increased 24-h BP variability at 7 and 14 days, slightly elevating BP levels, while this increase in BP variability was partially attenuated by prazosin, but not by atenolol. Continuous infusion of phenylephrine augmented BP variability, but isoproterenol had no effect on the variability.

Conclusion: Continuous infusion of noradrenalin augmented 24-h BP variability partly through an α1-adrenergic receptor-mediated mechanism in rats, suggesting that the noradrenalin-infused rat is an animal model of augmented BP variability induced by persistent adrenergic activation.
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http://dx.doi.org/10.1097/HJH.0000000000002239DOI Listing
February 2020

Polyethylene glycol-conjugated human adrenomedullin as a possible treatment for vascular dementia.

Peptides 2019 11 23;121:170133. Epub 2019 Aug 23.

Circulatory and Body Fluid Regulation, Faculty of Medicine, University of Miyazaki, Kiyotake, Miyazaki 889-1692, Japan.

Adrenomedullin (AM) is a multifunctional bioactive peptide. Recent studies have shown that AM has protective effects against ischemic brain damage. We recently prepared a long-acting human AM derivative that was conjugated with a 60 kDa polyethylene glycol (PEG-AM), which had an effect similar to that of native AM. In this study, we examined the effect of PEG-AM on four-vessel occlusion model rats, which exhibit vascular dementia. From day 10 to day 14 after surgery, the learning and memory abilities of the rats were examined using a Morris water maze. The rats were treated with a single subcutaneous injection of 1.0 or 10.0 nmol/kg of PEG-AM. PEG-AM treatment reduced the escape latency in the hidden platform test. Furthermore, the treatment increased the time spent in the platform quadrant in the probe test. The data showed that PEG-AM injection prevented memory loss and learning disorders in dose-dependent manner. On day 14, the immunoreactive AM concentration in plasma was 9.749 ± 2.167 pM in the high-dose group (10.0 nmol/kg) and 0.334 ± 0.073 pM in the low-dose group (1.0 nmol/kg). However, even in the low-dose group, a significant effect was observed in both tests. The present data indicate that PEG-AM is a possible therapeutic agent for the treatment of ischemic brain injury or vascular dementia.
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http://dx.doi.org/10.1016/j.peptides.2019.170133DOI Listing
November 2019

Blockade of the angiotensin II type 1 receptor increases bone mineral density and left ventricular contractility in a mouse model of juvenile Paget disease.

Eur J Pharmacol 2019 Sep 2;859:172519. Epub 2019 Jul 2.

Department of Internal Medicine, Circulatory and Body Fluid Regulation, University of Miyazaki, 889-1692, Japan.

Juvenile Paget disease (JPD1), an autosomal-recessive disorder, is characterized by extremely rapid bone turnover due to osteoprotegerin deficiency. Its extra-skeletal manifestations, such as hypertension and heart failure, suggest a pathogenesis with shared skeletal and cardiovascular system components. In spite of this, the effects of anti-hypertensive drugs on bone morphometry remain unknown. We administered an angiotensin II type 1 receptor blocker, olmesartan (5 mg/kg/day) to 8-week-old male mice lacking the osteoprotegerin gene, with and without 1 μg/kg/min of angiotensin II infusion for 14 days. Olmesartan treatment decreased systolic blood pressure, and echocardiography showed increased left ventricular systolic contractility. Three-dimensional micro-computed tomography scans demonstrated that olmesartan treatment increased trabecular bone volume (sham, +176%; angiotensin II infusion, +335%), mineral density (sham, +150%; angiotensin II infusion, +313%), and trabecular number (sham, +407%; angiotensin II infusion, +622%) in the tibia. Olmesartan increased cortical mineral density (sham, +19%; angiotensin II infusion, +24%), decreased the cortical bone section area (sham, -16%; angiotensin II infusion, -18%), decreased thickness (sham, -18%; angiotensin II infusion, -31%), and decreased the lacunar area (sham, -41%; angiotensin II infusion, -27%) in the tibia. Similar trend was observed in the femur. Moreover, olmesartan decreased angiotensin II-induced increases in tartrate-resistant acid phosphatase concentrations in plasma, but it affected neither type I procollagen N-terminal propeptides, nor the receptor activator of nuclear factor kappa-B ligand. Our data suggest that blockade of the angiotensin II type 1 receptor improves bone vulnerability, and helps to maintain the heart's structural integrity in osteoprotegerin-deficient mice.
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http://dx.doi.org/10.1016/j.ejphar.2019.172519DOI Listing
September 2019

mGluR1 in cerebellar Purkinje cells is essential for the formation but not expression of associative eyeblink memory.

Sci Rep 2019 05 14;9(1):7353. Epub 2019 May 14.

Division of Molecular Genetics, Department of Physiology and Cell Biology, Kobe University Graduate School of Medicine, Kobe, Hyogo, 650-0017, Japan.

Classical eyeblink conditioning is a representative associative motor learning that requires both the cerebellar cortex and the deep cerebellar nucleus (DCN). Metabotropic glutamate receptor subtype 1 (mGluR1) is richly expressed in Purkinje cells (PCs) of the cerebellar cortex. Global mGluR1 knock-out (KO) mice show a significantly lower percentage of conditioned response (CR%) than wild-type mice in eyeblink conditioning, and the impaired CR% is restored by the introduction of mGluR1 in PCs. However, the specific roles of mGluR1 in major memory processes, including formation, storage and expression have not yet been defined. We thus examined the role of mGluR1 in these processes of eyeblink conditioning, using mGluR1 conditional KO (cKO) mice harboring a selective and reversible expression of mGluR1 in PCs. We have found that eyeblink memory is not latently formed in the absence of mGluR1 in adult mouse PCs. However, once acquired, eyeblink memory is expressed even after the depletion of mGluR1 in PCs. We thus conclude that mGluR1 in PCs is indispensable for the formation of eyeblink memory, while it is not required for the expression of CR.
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http://dx.doi.org/10.1038/s41598-019-43744-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6517439PMC
May 2019
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