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Integration of Droplet Microfluidic Tools for Single-cell Functional Metagenomics:An Engineering Head Start

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Genomics, Proteomics & Bioinformatics 2021年第3期19卷 504-518页

作者： David Conchouso Amani Al-Ma’abadi Hayedeh Behzad Mohammed Alarawi Masahito Hosokawa Yohei nishikawa Haruko Takeyama Katsuhiko Mineta Takashi Gojobori Department of Industrial and Mechanical Engineering Universidad de las Américas PueblaPuebla 72810Mexico Computational Bioscience Research Center King Abdullah University of Science and TechnologyThuwal 23955-6900Saudi Arabia Biological and Environmental Sciences and Engineering Division King Abdullah University of Science and TechnologyThuwal 23955-6900Saudi Arabia Research Organization for Nano&Life Innovation Waseda UniversityTokyo 162-0041Japan Department of Life Science and Medical Bioscience Waseda UniversityTokyo 162-8480Japan Institute for Advanced Research of Biosystem Dynamics Waseda Research Institute for Science and EngineeringWaseda UniversityTokyo 169-8555Japan Computational Bio Big-Data Open Innovation Laboratory AIST-Waseda UniversityTokyo 169–0072Japan Computer Electricaland Mathematical Sciences and Engineering DivisionKing Abdullah University of Science and TechnologyThuwal 23955-6900Saudi Arabia

Droplet microfluidic techniques have shown promising outcome to study single cells at high ***,their adoption in laboratories studying“-omics”sciences is still irrelevant due to the complex and multidisciplinary nature of the *** facilitate their use,here we provide engineering details and organized protocols for integrating three droplet-based microfluidic technologies into the metagenomic pipeline to enable functional screening of bioproducts at high ***,a device encapsulating single cells in droplets at a rate of~250 Hz is described considering droplet size and cell ***,we expand on previously reported fluorescence-activated droplet sorting systems to integrate the use of 4 independent fluorescence-exciting lasers(i.e.,405,488,561,and 637 nm)in a single platform to make it compatible with different fluorescence-emitting *** this sorter,both hardware and software are provided and optimized for effortlessly sorting droplets at 60 ***,a passive droplet merger is also integrated into our pipeline to enable adding new reagents to already-made droplets at a rate of 200 ***,we provide an optimized recipe for manufacturing these chips using silicon dry-etching *** of the overall integration and the technical details presented here,our approach allows biologists to quickly use microfluidic technologies and achieve both single-cell resolution and high-throughput capability(>50,000 cells/day)for mining and bioprospecting metagenomic data.

关键词： Droplet microfluidics Single cell Metagenomics Droplet sorter Biotechnology

Direct Process to Prepare Crystallized Freestanding Membranes of Hydroxyapatite Using Sacrificial Layer of Barium-Compounds

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Journal of Crystallization Process and Technology 2017年第2期7卷 48-53页

作者： Hiroaki nishikawa Takafumi Nishii Faculty of Biology-Oriented Science and Technology Kindai University Kinokawa Japan

Freestanding membrane (FSM) of hydroxyapatite (HA) is a thin sheet of pure HA without any supporting substrates. Our original preparation process of FSM of HA had three steps: The first was the deposition of HA layer on sacrificial layer of solvent-soluble materials, the second was separation of FSM of HA by means of dissolution of sacrificial layer, and the third was post-annealing to crystallize FSM of HA. To date, the post-annealing process was a serious bottleneck of productivity owing to its too long time. In this short report, we proposed a novel sacrificial layer, heatproof and water-soluble Ba-compound, which makes the direct deposition of crystallized HA possible due to its heatproof property because the problem on the original process was that the previous sacrificial layers have no heatproof property and HA layer should be deposited as amorphous. We can deposit the Ba-compound sacrificial layer only in 1 hour followed with the direct deposition of crystallized HA layer, substituting the 20 hours of post-annealing. The FSM of HA was separated successfully from the substrate by means of dissolution of Ba-compound with water. Our novel process can shrink the process time by 19 hours.

关键词： Freestanding Membrane Hydroxyapatite Sacrificial Layer Ba-compounds

Evaluation of the effect of pyrrolidine dithiocarbamate in suppressing inflammation in mice with dextran sodium sulfate-induced colitis

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World Journal of Gastroenterology 2007年第11期13卷 1666-1671页

作者： Ichiro Hirata Shingo Yasumoto Ken Toshina Takuya Inoue Takashi nishikawa Naoko Murano Mitsuyuki Murano Fang-Yu Wang Ken-ichi Katsu Department of Gastroenterology Fujita Health University Jaoan Second Department of Internal Medicine Osaka Medical College Japan Department of Gastroenterology Jinling Hospital Nanjing University School of Medicine Nanjing 210002 Jiangsu Province China

AIM: To evaluate the effect of pyrrolidine dithio- carbamate (PDTC; an NF-κB inhibitor) administered at low (50 mg/kg) and high (100 mg/kg) doses in suppressing colitis in mice with dextran sodium sulfate (DSS)-induced colitis. METHODS: Mice were divided into a DSS-untreated group (normal group), DSS-treated control group, DSS+PDTC-treated groupⅠ(low-dose group), and DSS+PDTC-treated groupⅡ (high-dose group). In each group, the disease activity index score (DAI score), intestinal length, histological score, and the levels of activated NF-κB and inflammatory cytokines (IL-1β and TNF-α) in tissue were measured. RESULTS: The DSS+PDTC-treated groupⅡ exhibited suppression of shortening of intestinal length and reduction of DAI score. Activated NF-κB level and IL-1β and TNF-α levels were significantly lower in DSS+PDTC- treated groupⅡ. CONCLUSION: These findings suggest that PDTC is useful for the treatment of ulcerative colitis.

关键词： Ulcerative colitis DSS-induced colitis Pyrrolidine dithiocarbamate NF-κB Mice

Atomic force microscopy analysis of orientation and bending of oligodeoxynucleotides in polypod-like structured DNA

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Nano Research 2015年第12期8卷 3764-3771页

作者： Tomoki Shiomi Mengmeng Tan Natsuki Takahashi Masayuki Endo Tomoko Emura Kumi Hidaka Hiroshi Sugiyama Yuki Takahashi Yoshinobu Takakura Makiya nishikawa Department of Biopharmaceutics and Drug Metabolism Graduate School of Pharmaceutical Sciences Kyoto University Sakyo-ku Kyoto 606-8501 Japan Institute for Integrated Cell-Material Sciences (WPI-iCeMS) Kyoto University Sakyo-ku Kyoto 606-8501 Japan Department of Chemistry Graduate School of Science Kyoto University Sakyo-ku Kyoto 606-8502 Japan

We previously demonstrated that polypod-like structured DNA, or polypodna, constructed with three or more oligodeoxynucleotides （ODNs）, is efficiently taken up by immune cells such as dendritic cells and macrophages, depending on its structural complexity. The ODNs comprising the polypodna should bend to form the polypod-like structure, and may do so by adopting either a bend- type conformation or a cross-type conformation. Here, we tried to elucidate the orientation and bending of ODNs in polypodnas using atomic force microscopy （AFM）. We designed two types of pentapodnas （i.e., a polypodna with five pods） using 60- to 88-base ODNs, which were then immobilized on DNA origami frames. AFM imaging showed that the ODNs in the pentapodna adopted bend-type conformations. Tetrapodna and hexapodna also adopted bend-type conformations when they were immobilized on frames under unconstrained conditions. These findings provide useful information toward the coherent design of, and the structure-activity relationships for, a variety of DNA nanostructures.

关键词： DNA nanostructure atomic force microscopy self-asse nanotechology structure-activity relationship

Reconsideration of Augmentation Strategies in Electroconvulsive Therapy: Effects of the Concurrent Use of a Reduced Dose of Propofol with Divided Supplemental Remifentanil and Moderate Hyperventilation on Electroconvulsive Therapy-Induced Seizure Production and Adverse Events

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Open Journal of Anesthesiology 2015年第10期5卷 219-226页

作者： Kohki nishikawa Michiaki Yamakage Department of Anesthesia Muroran City General Hospital Muroran Japan Department of Anesthesiology Sapporo Medical University School of Medicine Sapporo Japan

Background: Although several treatment strategies to enhance the efficacy of electroconvulsive therapy (ECT) have been discussed, there have been no reports on the combined use of these treatments. The purpose of this study was to evaluate the efficacy and safety of concurrent use of moderate hyperventilation and a reduced dose of propofol combined with divided remifentanil in ECT practice. Methods: Sixty patients scheduled to receive a total of 300 ECT treatments were randomly assigned to have the three interventions: a standard dose (1 mg/kg) of propofol (group P/N);a standard dose of propofol and moderate hyperventilation with end-tidal pressure of carbon dioxide (ETCO2) of 30 - 35 mmHg (group P/H);and a reduced dose (0.5 mg/kg) of propofol with divided supplemental remifentanil and moderate hyperventilation (group RP/H). Patients in group RP/H received remifentanil 1 μg/kg followed by propofol 0.5 mg/kg for unconsciousness and thereafter remifentanil 1 μg/kg immediately before the ECT stimulus. Results: Patients in group RP/H showed significantly longer durations of electroencephalographic (EEG) seizures in the early phase of ECT course (P < 0.01 and 0.05) and lower electrical stimulus in the late phase of ECT course (P < 0.05 in each) than those in groups P/N and P/H, respectively. Conclusions: The use of a reduced dose of propofol combined with divided supplemental remifentanil under moderate hyperventilation during ECT can offer advantages over the use of a standard dose of propofol with or without moderate hyperventilation in terms of more seizure augmentation and/or lower electrical stimulus.

关键词： Electroconvulsive Therapy Moderate Hyperventilation Reduced Dose of Propofol Seizure Duration Electrical Stimulus Intensity

Changes in Soil Physicochemical Properties Following Land Use Change from Paddy Fields to Greenhouse and Upland Fields in the Southeastern Basin of Dianchi Lake,Yunnan Province,China

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Pedosphere 2013年第2期23卷 169-176页

作者： N.MORITSUKA T.nishikawa S.YAMAMOTO N.MATSUI H.INOUE LI Kun-Zhi T.INAMURA Laboratory of Plant Production Systems Graduate School of AgricultureKyoto University Kagoshima University KANSO Technos Kunming University of Science and Technology

Paddy fields in the southeastern basin of Dianchi Lake have rapidly changed to greenhouses since 1999. A total of 61 surface soil samples, including 43 greenhouse soils, 12 upland soils, and 6 paddy soils, were collected from a flat lowland area mainly used for agricultural production fields in the southeastern basin of Dianchi Lake. Analyses of the soil samples indicated that the greenhouse soils were characterized by a lower organic matter content, lower pH, and higher soluble nutrients than the paddy soils in the area. The lower organic matter content of the greenhouse soils was ascribed to environmental or management factors rather than the clay content of the soil. Accumulation of soluble nutrients, especially inorganic N, was due to over-application of fertilizers, which also caused soil acidification. The average amount of readily available N, P, and K accumulated in the greenhouse soils was estimated to be equal to or higher than the annual input of these nutrients as a fertilizer, indicating that a reduction in fertilizer application was possible and recommended. In contrast, a very low available Si content was observed in the paddy soils, suggesting the need for Si application for rice production.

关键词： available Si greenhouse soils inorganic N phosphorus salinization

Induction of ischemic tolerance in rat liver via reduced nicotinamide adenine dinucleotide phosphate oxidase in Kupffer cells

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World Journal of Gastroenterology 2007年第38期13卷 5071-5078页

作者： Kazuaki Tejima Masahiro Arai Hitoshi Ikeda Tomoaki Tomiya Mikio Yanase Yukiko Inoue Takako nishikawa Naoko Watanabe Natsuko Ohtomo Masao Omata Kenji Fujiwara Department of Gastroenterology University of Tokyo 7-3-1 Hongo Bunkyo-ku Tokyo 113-8655 Japan Department of Gastroenterology Toshiba General Hospital 6-3-22 Higashiooi Shinagawa-ku Tokyo 140-8522 Japan Yokohama Rosai Hospital 3211 Kozukue-cho Kouhoku-ku Yokohama-shi Kanagawa 222-0036 Japan

AIM: To elucidate the mechanisms of hepatocyte preconditioning by H2O2 to better understand the pathophysiology of ischemic preconditioning. METHODS: The in vitro effect of H2O2 pretreatment was investigated in rat isolated hepatocytes subjected to anoxia/reoxygenation. Cell viability was assessed with propidium iodide fluorometry. In other experiments, rat livers were excised and subjected to warm ischemia/ reperfusion in an isolated perfused liver system to determine leakage of liver enzymes. Preconditioning was performed by H2O2 perfusion, or by stopping the perfusion for 10 min followed by 10 min of reperfusion. To inhibit Kupffer cell function or reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, gadolinium chloride was injected prior to liver excision, or diphenyleneiodonium, an inhibitor of NADPH oxidase, was added to the perfusate, respectively. Histological detection of oxygen radical formation in Kupffer cells was performed by perfusion with nitro blue tetrazolium. RESULTS: Anoxia/reoxygenation decreased hepatocyte viability compared to the controls. Pretreatment with H2O2 did not improve such hepatocyte injury. In liver perfusion experiments, however, H2O2 preconditioning reduced warm ischemia/reperfusion injury, which wasreversed by inhibition of Kupffer cell function or NADPH oxidase. Histological examination revealed that H2O2 preconditioning induced oxygen radical formation in Kupffer cells. NADPH oxidase inhibition also reversed hepatoprotection by ischemic preconditioning. CONCLUSION: H2O2 preconditioning protects hepato- cytes against warm ischemia/reperfusion injury via NADPH oxidase in Kupffer cells, and not directly. NADPH oxidase also mediates hepatoprotection by ischemic preconditioning.

关键词： Diphenyieneiodonium chioride Ischemia/ reperfusion injury Ischemic preconditioning Liver transplantation Oxygen radicals

Whole-exome sequencing for a more accurate diagnosis of intraductal papillary neoplasms of the bile duct

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Gastroenterology Report 2022年第1期10卷 618-621页

作者： Tomoaki Matsumori Norimitsu Uza Nobuyuki Kakiuchi Toshihiro Morita Yoshihiro nishikawa Masahiro Shiokawa Kojiro Taura Yuzo Kodama Hiroshi Seno Department of Gastroenterology and Hepatology Graduate School of MedicineKyoto UniversityKyotoJapan Department of Phathology and Tumor Biology Kyoto UniversityKyotoJapan Department of Hepato-Biliary-Pancreatic Surgery Graduate school of MedicineKyoto UniversityKyotoJapan Department of Gastroenterology Kobe UniversityKobeJapan

Introduction According to the World Health Organization(WHO,2018),intraductal papillary neoplasms of the bile duct(IPNB)are characterized as polypoid masses in dilated bile ducts;additionally,invasive IPNB occasionally presents with a nodular surface or mass formation[1,2].The IPNB diagnostic criteria are,however,often ambiguous,*** IPNB is considered a type of cancer and whether it must have mucus production are debatable[3].Therefore,establishing new IPNB diagnostic criteria based on an alternative approach,such as whole-exome sequencing,is ***,we present a case of IPNB with atypical features on which we performed a detailed genetic analysis to determine the genetic,morphological,and histological features of IPNB in this case.

关键词： neoplasms bile diagnosis

Angle Control of a Pneumatically Driven Musculoskeletal Model Based on Coordination of Agonist-Antagonist Muscle

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Journal of Mechanics Engineering and Automation 2012年第12期2卷 709-719页

作者： Yuki Honda Fumio Miyazaki Atsushi nishikawa Department of Mechanical Science and Bioengineering Graduate School of Engineering Science Osaka University Toyonaka 560-8531 Japan Bioengineering Course Division of Mechanical Engineering and Robotics Faculty of Textile Science and Technology Shinshu University Ueda 386-8567 Japan

In recent years, researchers have been actively pursuing research into developing robots that can be useful in many fields of industry （e.g., service, medical, and aging care）. Such robots must be safe and flexible so that they can coexist with people. Pneumatic actuators are useful for achieving this goal because they are lightweight units with natural compliance. Our research focuses on joint angle control for a pneumatically driven musculoskeletal model. In such a model, we use a one-degree-of-freedom joint model and a five-fingered robot hand as test beds. These models are driven by low pressure-driven pneumatic actuators, and mimic the mechanism of the human hand and musculoskeletal structure, which has an antagonistic muscle pair for each joint. We demonstrated a biologically inspired control method using the parameters antagonistic muscle ratio and antagonistic muscle activity. The concept of the method is based on coordination of an antagonistic muscle pair using these parameters. We have investigated the validity of the proposed method both theoretically and experimentally, developed a feedback control system, and conducted joint angle control by implementing the test beds.

关键词： Musculoskeletal model pneumatic actuator position control muscle coordination agonist-antagonist muscle.