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      • Real-time detection of acetylcholine release from the human endocrine pancreas

        Rodriguez-Diaz, Rayner,Dando, Robin,Huang, Y Anthony,Berggren, Per-Olof,Roper, Stephen D,Caicedo, Alejandro Nature Publishing Group, a division of Macmillan P 2012 Nature protocols Vol.7 No.6

        Neurons, sensory cells and endocrine cells secrete neurotransmitters and hormones to communicate with other cells and to coordinate organ and system function. Validation that a substance is used as an extracellular signaling molecule by a given cell requires a direct demonstration of its secretion. In this protocol we describe the use of biosensor cells to detect neurotransmitter release from endocrine cells in real-time. Chinese hamster ovary cells expressing the muscarinic acetylcholine (ACh) receptor M3 were used as ACh biosensors to record ACh release from human pancreatic islets. We show how ACh biosensors loaded with the Ca<SUP>2+</SUP> indicator Fura-2 and pressed against isolated human pancreatic islets allow the detection of ACh release. The biosensor approach is simple; the Ca<SUP>2+</SUP> signal generated in the biosensor cell reflects the presence (release) of a neurotransmitter. The technique is versatile because biosensor cells expressing a variety of receptors can be used in many applications. The protocol takes ?? h.

      • KCI등재

        Quality of Life and Anorectal Malformations: A Single-Center Experience

        Gabriella Scirè,Riccardo Gabaldo,Ilaria Dando,Francesco S. Camoglio,Nicola Zampieri 대한소아소화기영양학회 2022 Pediatric gastroenterology, hepatology & nutrition Vol.25 No.4

        Purpose: The treatment and long term clinical outcomes of anorectal malformations (ARM) in children have always been the focus of pediatric surgeons. This study aimed at reporting our experience as far as long-term follow-up of ARM in children is concern. Methods: We enrolled patients treated between 1999 and 2019, and established selection criteria to choose appropriate subjects. A validated questionnaire was used to determine long-term quality of life outcomes. Results: Out of a total of 48 patients treated within the study period, 28 were enrolled in this study. Among the latter, more than 35% had at least one long-time complication, and more than 90% had a good lifestyle. Urinary and fecal continence was achieved in more than 95% of the patients using medical devices. Conclusion: This study aimed to bring up new concepts; taking into consideration all aspects of life in patients with ARM, from school life to sexuality, while evaluating fecal and urinary continence. This is essential for the improvement of the skills of the different specialists involved in the management of these patients, and for the implementation of strategies that can improve postoperative function. Most especially, it will also help improve communication between doctors to ensure an adequate transition of these children into adult life.

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        Modulation of Tight Junctions does Not Predict Oral Absorption of Hydrophilic Compounds: Use of Caco-2 and Calu-3 Cells

        Amrita V. Kamath,Richard A. Morrison,Neil R. Mathias,Sandra A. Dando,Anthony M. Marino,Saeho Chong 대한약학회 2007 Archives of Pharmacal Research Vol.30 No.8

        Permeability estimates using Caco-2 cells do not accurately predict the absorption of hydrophilic drugs that are primarily absorbed via the paracellular pathway. The objective of this study was to investigate whether modulation of tight junctions would help differentiation of paracellularly absorbed compounds. Tight junctions in Caco-2 cell monolayers were manipulated using calcium depletion approaches to decrease the transepithelial electrical resistance (TEER) of the monolayers, and permeability of hydrophilic compounds were measured under these conditions. Permeability of these compounds were also measured in Calu-3 cells, which have tighter junctions than Caco-2 cells. Calcium depletion loosened the tight junctions of Caco-2 cells to varying levels as measured by the decrease in TEER values of the monolayers. While the absolute permeability of all the model compounds increased as the tight junctions were loosened, the ratios of their permeability relative to mannitol permeability were similar. The permeability of these compounds in the tighter Calu-3 cells were also found to be similar to each other. Altering the tight junctions of Caco-2 cells to obtain leakier cell monolayers, or using a cell line with tighter junctions like Calu-3 cells, did not improve differentiation between well absorbed and poorly absorbed hydrophilic drugs. Mere manipulation of the tight junctions to increase or decrease transepithelial electrical resistance does not appear to be a viable approach to predict human absorption for hydrophilic compounds that are primarily absorbed via the paracellular pathway.

      • SCIESCOPUSKCI등재

        Modulation of Tight Junctions does Not Predict Oral Absorption of Hydrophilic Compounds: Use of Caco-2 and Calu-3 Cells

        Kamath, Amrita V.,Morrison, Richard A.,Mathias, Neil R.,Dando, Sandra A.,Marino, Anthony M.,Chong, Sae-Ho 대한약학회 2007 Archives of Pharmacal Research Vol.30 No.8

        Permeability estimates using Caco-2 cells do not accurately predict the absorption of hydrophilic drugs that are primarily absorbed via the paracellular pathway. The objective of this study was to investigate whether modulation of tight junctions would help differentiation of paracellularly absorbed compounds. Tight junctions in Caco-2 cell monolayers were manipulated using calcium depletion approaches to decrease the transepithelial electrical resistance (TEER) of the monolayers, and permeability of hydrophilic compounds were measured under these conditions. Permeability of these compounds were also measured in Calu-3 cells, which have tighter junctions than Caco-2 cells. Calcium depletion loosened the tight junctions of Caco-2 cells to varying levels as measured by the decrease in TEER values of the monolayers. While the absolute permeability of all the model compounds increased as the tight junctions were loosened, the ratios of their permeability relative to mannitol permeability were similar. The permeability of these compounds in the tighter Calu-3 cells were also found to be similar to each other. Altering the tight junctions of Caco-2 cells to obtain leakier cell monolayers, or using a cell line with tighter junctions like Calu-3 cells, did not improve differentiation between well absorbed and poorly absorbed hydrophilic drugs. Mere manipulation of the tight junctions to increase or decrease transepithelial electrical resistance does not appear to be a viable approach to predict human absorption for hydrophilic compounds that are primarily absorbed via the paracellular pathway.

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