Authors: Matthew H Crouthamel Edward J Kelly Rodney J Y Ho
Publish Date: 2011/05/03
Volume: 21, Issue: 1, Pages: 113-130
Abstract
For many CNS acting drugs penetration into the central nervous system CNS is limited by the bloodCNSbarriers In an effort to quantitate the role of the protein components that make up the bloodCNSbarriers we created transgenic mice that allow conditional gene knockout using Cre/loxP technology We targeted the expression of Crerecombinase to the choroid plexus the bloodcerebral spinal fluid barrier using the lymphotropic papovavirus control region LPVcr and to brain endothelium the blood–brainbarrier using the proximal promoter region of the human von Willebrand Factor gene hVWFf We verified that LPVcr restricts expression to the choroid plexus in adult mice by using the LPVcr to drive nLacZ expression in transgenic mice The LPVCre and hVWFCre plasmids were then constructed and tested for Crerecombinase function in vitro and subsequently used to create transgenic mice The resulting transgenic mice were characterized for celltype specific Cremediated endonuclease activity by crossing them with transgenic mice containing a loxPflankedLacZ/EGFP dual reporter gene Z/EG The dual CreZ/EG transgenic offspring were evaluated for the location of EGFP mRNA expression by reverse transcriptase PCR and for protein expression by immunohistochemistry Immunohistochemistry for EGFP verified expression in the target cells and no ectopic expression outside of the expected cell types The LPVCre0607 transgenic line expressed functional Cre only in the choroid plexus and hVWFCre1304 line in brain endotheliumWe thank Terry Kavanagh for the use of the Cryocut cryomicrotome Richard Palmiter for the pLacF plasmid Nancy Jahroudi for the pHGHK plasmid Terry Van Dyke for the pLST plasmid Corrinne Lobe for the Z/EG plasmid Washington University ES Cell Core for the pTurboCre plasmid and Kerstin Verdina for assistance in editing This work was supported by the National Institute of Health grants NS39178 AI077390 and AI52663 Rodney J Y Ho is also supported by the Milo Gibaldi Endowment
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