Pericyte abundance affects sucrose permeability in cultures of rat brain microvascular endothelial cells.
Identifieur interne : 002966 ( Main/Exploration ); précédent : 002965; suivant : 002967Pericyte abundance affects sucrose permeability in cultures of rat brain microvascular endothelial cells.
Auteurs : Fiona E. Parkinson [Canada] ; Cindy HackingSource :
- Brain research [ 0006-8993 ] ; 2005.
English descriptors
- KwdEn :
- MESH :
- chemical , metabolism : Sucrose.
- cytology : Endothelial Cells, Pericytes.
- metabolism : Blood-Brain Barrier, Endothelial Cells, Pericytes.
- methods : Cell Separation, Coculture Techniques.
- physiology : Capillary Permeability.
- Animals, Cells, Cultured, Rats.
Abstract
The blood-brain barrier is a physical and metabolic barrier that restricts diffusion of blood-borne substances into brain. In vitro models of the blood-brain barrier are used to characterize this structure, examine mechanisms of damage and repair and measure permeability of test substances. The core component of in vitro models of the blood-brain barrier is brain microvascular endothelial cells. We cultured rat brain microvascular endothelial cells (RBMEC) from isolated rat cortex microvessels. After 2-14 days in vitro (DIV), immunohistochemistry of these cells showed strong labeling for zona occludens 1 (ZO-1), a tight junction protein expressed in endothelial cells. Pericytes were also present in these cultures, as determined by expression of alpha-actin. The present study was performed to test different cell isolation methods and to compare the resulting cell cultures for abundance of pericytes and for blood-brain barrier function, as assessed by 14C-sucrose flux. Two purification strategies were used. First, microvessels were preabsorbed onto uncoated plastic for 4 h, then unattached microvessels were transferred to coated culture ware. Second, microvessels were incubated with an antibody to platelet-endothelial cell adhesion molecule 1 (PECAM-1; CD31) precoupled to magnetic beads, and a magnetic separation procedure was performed. Our results indicate that immunopurification, but not preadsorption, was an effective method to purify microvessels and reduce pericyte abundance in the resulting cultures. This purification significantly reduced 14C-sucrose fluxes across cell monolayers. These data indicate that pericytes can interfere with the development of blood-brain barrier properties in in vitro models that utilize primary cultures of RBMECs.
DOI: 10.1016/j.brainres.2005.04.054
PubMed: 15935996
Affiliations:
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Parkinson</name><affiliation wicri:level="4"><nlm:affiliation>Department of Pharmacology and Therapeutics, University of Manitoba, A203-753 McDermot Avenue, Winnipeg, MB, Canada R3E 0T6. parkins@ms.umanitoba.ca</nlm:affiliation><country>Canada</country><wicri:regionArea>Department of Pharmacology and Therapeutics, University of Manitoba, A203-753 McDermot Avenue, Winnipeg, MB</wicri:regionArea><orgName type="university">Université du Manitoba</orgName><placeName><settlement type="city">Winnipeg</settlement><region type="state">Manitoba</region></placeName></affiliation></author><author><name sortKey="Hacking, Cindy" sort="Hacking, Cindy" uniqKey="Hacking C" first="Cindy" last="Hacking">Cindy Hacking</name></author></titleStmt><publicationStmt><idno type="wicri:source">PubMed</idno><date when="2005">2005</date><idno type="RBID">pubmed:15935996</idno><idno type="pmid">15935996</idno><idno type="doi">10.1016/j.brainres.2005.04.054</idno><idno type="wicri:Area/PubMed/Corpus">001250</idno><idno type="wicri:explorRef" wicri:stream="PubMed" wicri:step="Corpus" wicri:corpus="PubMed">001250</idno><idno type="wicri:Area/PubMed/Curation">001250</idno><idno type="wicri:explorRef" wicri:stream="PubMed" wicri:step="Curation">001250</idno><idno type="wicri:Area/PubMed/Checkpoint">001250</idno><idno type="wicri:explorRef" wicri:stream="Checkpoint" wicri:step="PubMed">001250</idno><idno type="wicri:Area/Ncbi/Merge">000546</idno><idno type="wicri:Area/Ncbi/Curation">000546</idno><idno type="wicri:Area/Ncbi/Checkpoint">000546</idno><idno type="wicri:doubleKey">0006-8993:2005:Parkinson F:pericyte:abundance:affects</idno><idno type="wicri:Area/Main/Merge">002C79</idno><idno type="wicri:Area/Main/Curation">002966</idno><idno type="wicri:Area/Main/Exploration">002966</idno></publicationStmt><sourceDesc><biblStruct><analytic><title xml:lang="en">Pericyte abundance affects sucrose permeability in cultures of rat brain microvascular endothelial cells.</title><author><name sortKey="Parkinson, Fiona E" sort="Parkinson, Fiona E" uniqKey="Parkinson F" first="Fiona E" last="Parkinson">Fiona E. Parkinson</name><affiliation wicri:level="4"><nlm:affiliation>Department of Pharmacology and Therapeutics, University of Manitoba, A203-753 McDermot Avenue, Winnipeg, MB, Canada R3E 0T6. parkins@ms.umanitoba.ca</nlm:affiliation><country>Canada</country><wicri:regionArea>Department of Pharmacology and Therapeutics, University of Manitoba, A203-753 McDermot Avenue, Winnipeg, MB</wicri:regionArea><orgName type="university">Université du Manitoba</orgName><placeName><settlement type="city">Winnipeg</settlement><region type="state">Manitoba</region></placeName></affiliation></author><author><name sortKey="Hacking, Cindy" sort="Hacking, Cindy" uniqKey="Hacking C" first="Cindy" last="Hacking">Cindy Hacking</name></author></analytic><series><title level="j">Brain research</title><idno type="ISSN">0006-8993</idno><imprint><date when="2005" type="published">2005</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Animals</term><term>Blood-Brain Barrier (metabolism)</term><term>Capillary Permeability (physiology)</term><term>Cell Separation (methods)</term><term>Cells, Cultured</term><term>Coculture Techniques (methods)</term><term>Endothelial Cells (cytology)</term><term>Endothelial Cells (metabolism)</term><term>Pericytes (cytology)</term><term>Pericytes (metabolism)</term><term>Rats</term><term>Sucrose (metabolism)</term></keywords><keywords scheme="MESH" type="chemical" qualifier="metabolism" xml:lang="en"><term>Sucrose</term></keywords><keywords scheme="MESH" qualifier="cytology" xml:lang="en"><term>Endothelial Cells</term><term>Pericytes</term></keywords><keywords scheme="MESH" qualifier="metabolism" xml:lang="en"><term>Blood-Brain Barrier</term><term>Endothelial Cells</term><term>Pericytes</term></keywords><keywords scheme="MESH" qualifier="methods" xml:lang="en"><term>Cell Separation</term><term>Coculture Techniques</term></keywords><keywords scheme="MESH" qualifier="physiology" xml:lang="en"><term>Capillary Permeability</term></keywords><keywords scheme="MESH" xml:lang="en"><term>Animals</term><term>Cells, Cultured</term><term>Rats</term></keywords></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">The blood-brain barrier is a physical and metabolic barrier that restricts diffusion of blood-borne substances into brain. In vitro models of the blood-brain barrier are used to characterize this structure, examine mechanisms of damage and repair and measure permeability of test substances. The core component of in vitro models of the blood-brain barrier is brain microvascular endothelial cells. We cultured rat brain microvascular endothelial cells (RBMEC) from isolated rat cortex microvessels. After 2-14 days in vitro (DIV), immunohistochemistry of these cells showed strong labeling for zona occludens 1 (ZO-1), a tight junction protein expressed in endothelial cells. Pericytes were also present in these cultures, as determined by expression of alpha-actin. The present study was performed to test different cell isolation methods and to compare the resulting cell cultures for abundance of pericytes and for blood-brain barrier function, as assessed by 14C-sucrose flux. Two purification strategies were used. First, microvessels were preabsorbed onto uncoated plastic for 4 h, then unattached microvessels were transferred to coated culture ware. Second, microvessels were incubated with an antibody to platelet-endothelial cell adhesion molecule 1 (PECAM-1; CD31) precoupled to magnetic beads, and a magnetic separation procedure was performed. Our results indicate that immunopurification, but not preadsorption, was an effective method to purify microvessels and reduce pericyte abundance in the resulting cultures. This purification significantly reduced 14C-sucrose fluxes across cell monolayers. These data indicate that pericytes can interfere with the development of blood-brain barrier properties in in vitro models that utilize primary cultures of RBMECs.</div></front></TEI><affiliations><list><country><li>Canada</li></country><region><li>Manitoba</li></region><settlement><li>Winnipeg</li></settlement><orgName><li>Université du Manitoba</li></orgName></list><tree><noCountry><name sortKey="Hacking, Cindy" sort="Hacking, Cindy" uniqKey="Hacking C" first="Cindy" last="Hacking">Cindy Hacking</name></noCountry><country name="Canada"><region name="Manitoba"><name sortKey="Parkinson, Fiona E" sort="Parkinson, Fiona E" uniqKey="Parkinson F" first="Fiona E" last="Parkinson">Fiona E. Parkinson</name></region></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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