Contribution of pannexin 1 and connexin 43 hemichannels to extracellular calcium-dependent transport dynamics in human blood-brain barrier endothelial cells
Dysregulation of blood-brain barrier (BBB) transport function is thought to exacerbate neuronal damage in acute ischemic stroke. The purpose of this study was to clarify the characteristics of pannexin (Px) and/or connexin (Cx) hemichannel(s)-mediated transport of organic anions and cations in human...
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Veröffentlicht in: | The Journal of pharmacology and experimental therapeutics 2015-04, Vol.353 (1), p.192-200 |
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creator | Kaneko, Yosuke Tachikawa, Masanori Akaogi, Ryo Fujimoto, Kazuhisa Ishibashi, Megumi Uchida, Yasuo Couraud, Pierre-Olivier Ohtsuki, Sumio Hosoya, Ken-ichi Terasaki, Tetsuya |
description | Dysregulation of blood-brain barrier (BBB) transport function is thought to exacerbate neuronal damage in acute ischemic stroke. The purpose of this study was to clarify the characteristics of pannexin (Px) and/or connexin (Cx) hemichannel(s)-mediated transport of organic anions and cations in human BBB endothelial cell line hCMEC/D3 and to identify inhibitors of hemichannel opening in hCMEC/D3 cells in the absence of extracellular Ca(2+), a condition mimicking acute ischemic stroke. In the absence of extracellular Ca(2+), the cells showed increased uptake and efflux transport of organic ionic fluorescent dyes. Classic hemichannel inhibitors markedly inhibited the enhanced uptake and efflux. Quantitative targeted absolute proteomics confirmed Px1 and Cx43 protein expression in plasma membrane of hCMEC/D3 cells. Knockdown of Px1 and Cx43 with the small interfering RNAs significantly inhibited the enhanced uptake and efflux of organic anionic and cationic fluorescent dyes. Clinically used cilnidipine and progesterone, which have neuroprotective effects in animal ischemia models, were identified as inhibitors of hemichannel opening. These findings suggest that altered transport dynamics at the human BBB in the absence of extracellular Ca(2+) is at least partly attributable to opening of Px1 and Cx43 hemichannels. Therefore, we speculate that Px1 and Cx43 may be potential drug targets to ameliorate BBB transport dysregulation during acute ischemia. |
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The purpose of this study was to clarify the characteristics of pannexin (Px) and/or connexin (Cx) hemichannel(s)-mediated transport of organic anions and cations in human BBB endothelial cell line hCMEC/D3 and to identify inhibitors of hemichannel opening in hCMEC/D3 cells in the absence of extracellular Ca(2+), a condition mimicking acute ischemic stroke. In the absence of extracellular Ca(2+), the cells showed increased uptake and efflux transport of organic ionic fluorescent dyes. Classic hemichannel inhibitors markedly inhibited the enhanced uptake and efflux. Quantitative targeted absolute proteomics confirmed Px1 and Cx43 protein expression in plasma membrane of hCMEC/D3 cells. Knockdown of Px1 and Cx43 with the small interfering RNAs significantly inhibited the enhanced uptake and efflux of organic anionic and cationic fluorescent dyes. Clinically used cilnidipine and progesterone, which have neuroprotective effects in animal ischemia models, were identified as inhibitors of hemichannel opening. These findings suggest that altered transport dynamics at the human BBB in the absence of extracellular Ca(2+) is at least partly attributable to opening of Px1 and Cx43 hemichannels. Therefore, we speculate that Px1 and Cx43 may be potential drug targets to ameliorate BBB transport dysregulation during acute ischemia.</description><identifier>ISSN: 0022-3565</identifier><identifier>EISSN: 1521-0103</identifier><identifier>DOI: 10.1124/jpet.114.220210</identifier><identifier>PMID: 25670633</identifier><language>eng</language><publisher>United States</publisher><subject>Anions ; Biological Transport - drug effects ; Blood-Brain Barrier - cytology ; Blood-Brain Barrier - drug effects ; Blood-Brain Barrier - metabolism ; Calcium - metabolism ; Cations ; Cell Line ; Cell Membrane - metabolism ; Connexin 43 - antagonists & inhibitors ; Connexin 43 - metabolism ; Connexins - antagonists & inhibitors ; Connexins - metabolism ; Endothelial Cells - drug effects ; Endothelial Cells - metabolism ; Extracellular Space - metabolism ; Fluorescent Dyes - metabolism ; Humans ; Ion Channel Gating ; Microvessels - cytology ; Microvessels - metabolism ; Nerve Tissue Proteins - antagonists & inhibitors ; Nerve Tissue Proteins - metabolism</subject><ispartof>The Journal of pharmacology and experimental therapeutics, 2015-04, Vol.353 (1), p.192-200</ispartof><rights>Copyright © 2015 by The American Society for Pharmacology and Experimental Therapeutics.</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c404t-a950a5f6b4b17d0991d2b90498b246aab40ad21fe29a7433ca4e25e0358b471e3</citedby><cites>FETCH-LOGICAL-c404t-a950a5f6b4b17d0991d2b90498b246aab40ad21fe29a7433ca4e25e0358b471e3</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>315,781,785,27929,27930</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/25670633$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Kaneko, Yosuke</creatorcontrib><creatorcontrib>Tachikawa, Masanori</creatorcontrib><creatorcontrib>Akaogi, Ryo</creatorcontrib><creatorcontrib>Fujimoto, Kazuhisa</creatorcontrib><creatorcontrib>Ishibashi, Megumi</creatorcontrib><creatorcontrib>Uchida, Yasuo</creatorcontrib><creatorcontrib>Couraud, Pierre-Olivier</creatorcontrib><creatorcontrib>Ohtsuki, Sumio</creatorcontrib><creatorcontrib>Hosoya, Ken-ichi</creatorcontrib><creatorcontrib>Terasaki, Tetsuya</creatorcontrib><title>Contribution of pannexin 1 and connexin 43 hemichannels to extracellular calcium-dependent transport dynamics in human blood-brain barrier endothelial cells</title><title>The Journal of pharmacology and experimental therapeutics</title><addtitle>J Pharmacol Exp Ther</addtitle><description>Dysregulation of blood-brain barrier (BBB) transport function is thought to exacerbate neuronal damage in acute ischemic stroke. The purpose of this study was to clarify the characteristics of pannexin (Px) and/or connexin (Cx) hemichannel(s)-mediated transport of organic anions and cations in human BBB endothelial cell line hCMEC/D3 and to identify inhibitors of hemichannel opening in hCMEC/D3 cells in the absence of extracellular Ca(2+), a condition mimicking acute ischemic stroke. In the absence of extracellular Ca(2+), the cells showed increased uptake and efflux transport of organic ionic fluorescent dyes. Classic hemichannel inhibitors markedly inhibited the enhanced uptake and efflux. Quantitative targeted absolute proteomics confirmed Px1 and Cx43 protein expression in plasma membrane of hCMEC/D3 cells. Knockdown of Px1 and Cx43 with the small interfering RNAs significantly inhibited the enhanced uptake and efflux of organic anionic and cationic fluorescent dyes. Clinically used cilnidipine and progesterone, which have neuroprotective effects in animal ischemia models, were identified as inhibitors of hemichannel opening. These findings suggest that altered transport dynamics at the human BBB in the absence of extracellular Ca(2+) is at least partly attributable to opening of Px1 and Cx43 hemichannels. Therefore, we speculate that Px1 and Cx43 may be potential drug targets to ameliorate BBB transport dysregulation during acute ischemia.</description><subject>Anions</subject><subject>Biological Transport - drug effects</subject><subject>Blood-Brain Barrier - cytology</subject><subject>Blood-Brain Barrier - drug effects</subject><subject>Blood-Brain Barrier - metabolism</subject><subject>Calcium - metabolism</subject><subject>Cations</subject><subject>Cell Line</subject><subject>Cell Membrane - metabolism</subject><subject>Connexin 43 - antagonists & inhibitors</subject><subject>Connexin 43 - metabolism</subject><subject>Connexins - antagonists & inhibitors</subject><subject>Connexins - metabolism</subject><subject>Endothelial Cells - drug effects</subject><subject>Endothelial Cells - metabolism</subject><subject>Extracellular Space - metabolism</subject><subject>Fluorescent Dyes - metabolism</subject><subject>Humans</subject><subject>Ion Channel Gating</subject><subject>Microvessels - cytology</subject><subject>Microvessels - metabolism</subject><subject>Nerve Tissue Proteins - antagonists & inhibitors</subject><subject>Nerve Tissue Proteins - metabolism</subject><issn>0022-3565</issn><issn>1521-0103</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2015</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNo9UUtr3DAQFqUh2SY591Z07MXJ6GHv-liWviDQS3s2I2vMKsiSK8mQ_Jf-2Mrstqf5Zr4HAx9j7wU8CCH14_NCpSL9ICVIAW_YTrRSNCBAvWU7ACkb1XbtDXuX8zOA0LpT1-xGtt0eOqV27M8xhpKcWYuLgceJLxgCvbjABcdg-Rgvq1b8RLMbTxvvMy-R00tJOJL3q8fER_SjW-fG0kLBUii8siEvMRVuXwNWb-Y16LTOGLjxMdrGJKwXgyk5SrzaYjmRd-j5Fpvv2NWEPtP9Zd6yX18-_zx-a55-fP1-_PTUjBp0abBvAdupM9qIvYW-F1aaHnR_MFJ3iEYDWikmkj3utVIjapItgWoPRu8FqVv28Zy7pPh7pVyG2eXtAwwU1zyIroNeHoRSVfp4lo4p5pxoGpbkZkyvg4Bhq2TYKqlID-dKquPDJXw1M9n_-n8dqL_dXIr8</recordid><startdate>20150401</startdate><enddate>20150401</enddate><creator>Kaneko, Yosuke</creator><creator>Tachikawa, Masanori</creator><creator>Akaogi, Ryo</creator><creator>Fujimoto, Kazuhisa</creator><creator>Ishibashi, Megumi</creator><creator>Uchida, Yasuo</creator><creator>Couraud, Pierre-Olivier</creator><creator>Ohtsuki, Sumio</creator><creator>Hosoya, Ken-ichi</creator><creator>Terasaki, Tetsuya</creator><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7X8</scope></search><sort><creationdate>20150401</creationdate><title>Contribution of pannexin 1 and connexin 43 hemichannels to extracellular calcium-dependent transport dynamics in human blood-brain barrier endothelial cells</title><author>Kaneko, Yosuke ; Tachikawa, Masanori ; Akaogi, Ryo ; Fujimoto, Kazuhisa ; Ishibashi, Megumi ; Uchida, Yasuo ; Couraud, Pierre-Olivier ; Ohtsuki, Sumio ; Hosoya, Ken-ichi ; Terasaki, Tetsuya</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c404t-a950a5f6b4b17d0991d2b90498b246aab40ad21fe29a7433ca4e25e0358b471e3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2015</creationdate><topic>Anions</topic><topic>Biological Transport - drug effects</topic><topic>Blood-Brain Barrier - cytology</topic><topic>Blood-Brain Barrier - drug effects</topic><topic>Blood-Brain Barrier - metabolism</topic><topic>Calcium - metabolism</topic><topic>Cations</topic><topic>Cell Line</topic><topic>Cell Membrane - metabolism</topic><topic>Connexin 43 - antagonists & inhibitors</topic><topic>Connexin 43 - metabolism</topic><topic>Connexins - antagonists & inhibitors</topic><topic>Connexins - metabolism</topic><topic>Endothelial Cells - drug effects</topic><topic>Endothelial Cells - metabolism</topic><topic>Extracellular Space - metabolism</topic><topic>Fluorescent Dyes - metabolism</topic><topic>Humans</topic><topic>Ion Channel Gating</topic><topic>Microvessels - cytology</topic><topic>Microvessels - metabolism</topic><topic>Nerve Tissue Proteins - antagonists & inhibitors</topic><topic>Nerve Tissue Proteins - metabolism</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Kaneko, Yosuke</creatorcontrib><creatorcontrib>Tachikawa, Masanori</creatorcontrib><creatorcontrib>Akaogi, Ryo</creatorcontrib><creatorcontrib>Fujimoto, Kazuhisa</creatorcontrib><creatorcontrib>Ishibashi, Megumi</creatorcontrib><creatorcontrib>Uchida, Yasuo</creatorcontrib><creatorcontrib>Couraud, Pierre-Olivier</creatorcontrib><creatorcontrib>Ohtsuki, Sumio</creatorcontrib><creatorcontrib>Hosoya, Ken-ichi</creatorcontrib><creatorcontrib>Terasaki, Tetsuya</creatorcontrib><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>MEDLINE - Academic</collection><jtitle>The Journal of pharmacology and experimental therapeutics</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Kaneko, Yosuke</au><au>Tachikawa, Masanori</au><au>Akaogi, Ryo</au><au>Fujimoto, Kazuhisa</au><au>Ishibashi, Megumi</au><au>Uchida, Yasuo</au><au>Couraud, Pierre-Olivier</au><au>Ohtsuki, Sumio</au><au>Hosoya, Ken-ichi</au><au>Terasaki, Tetsuya</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Contribution of pannexin 1 and connexin 43 hemichannels to extracellular calcium-dependent transport dynamics in human blood-brain barrier endothelial cells</atitle><jtitle>The Journal of pharmacology and experimental therapeutics</jtitle><addtitle>J Pharmacol Exp Ther</addtitle><date>2015-04-01</date><risdate>2015</risdate><volume>353</volume><issue>1</issue><spage>192</spage><epage>200</epage><pages>192-200</pages><issn>0022-3565</issn><eissn>1521-0103</eissn><abstract>Dysregulation of blood-brain barrier (BBB) transport function is thought to exacerbate neuronal damage in acute ischemic stroke. The purpose of this study was to clarify the characteristics of pannexin (Px) and/or connexin (Cx) hemichannel(s)-mediated transport of organic anions and cations in human BBB endothelial cell line hCMEC/D3 and to identify inhibitors of hemichannel opening in hCMEC/D3 cells in the absence of extracellular Ca(2+), a condition mimicking acute ischemic stroke. In the absence of extracellular Ca(2+), the cells showed increased uptake and efflux transport of organic ionic fluorescent dyes. Classic hemichannel inhibitors markedly inhibited the enhanced uptake and efflux. Quantitative targeted absolute proteomics confirmed Px1 and Cx43 protein expression in plasma membrane of hCMEC/D3 cells. Knockdown of Px1 and Cx43 with the small interfering RNAs significantly inhibited the enhanced uptake and efflux of organic anionic and cationic fluorescent dyes. Clinically used cilnidipine and progesterone, which have neuroprotective effects in animal ischemia models, were identified as inhibitors of hemichannel opening. These findings suggest that altered transport dynamics at the human BBB in the absence of extracellular Ca(2+) is at least partly attributable to opening of Px1 and Cx43 hemichannels. Therefore, we speculate that Px1 and Cx43 may be potential drug targets to ameliorate BBB transport dysregulation during acute ischemia.</abstract><cop>United States</cop><pmid>25670633</pmid><doi>10.1124/jpet.114.220210</doi><tpages>9</tpages><oa>free_for_read</oa></addata></record> |
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subjects | Anions Biological Transport - drug effects Blood-Brain Barrier - cytology Blood-Brain Barrier - drug effects Blood-Brain Barrier - metabolism Calcium - metabolism Cations Cell Line Cell Membrane - metabolism Connexin 43 - antagonists & inhibitors Connexin 43 - metabolism Connexins - antagonists & inhibitors Connexins - metabolism Endothelial Cells - drug effects Endothelial Cells - metabolism Extracellular Space - metabolism Fluorescent Dyes - metabolism Humans Ion Channel Gating Microvessels - cytology Microvessels - metabolism Nerve Tissue Proteins - antagonists & inhibitors Nerve Tissue Proteins - metabolism |
title | Contribution of pannexin 1 and connexin 43 hemichannels to extracellular calcium-dependent transport dynamics in human blood-brain barrier endothelial cells |
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