title: Roundabout 4 regulates blood-tumor barrier permeability through the modulation of ZO-1, Occludin, and Claudin-5 expression.
abstract: The blood-tumor barrier (BTB) restricts the delivery of chemotherapeutic drug molecules to tumor tissues. We found that the endothelial cell (EC) receptor molecule Roundabout 4 (Robo4) is endogenously expressed in human brain microvascular ECs and that it is upregulated in a BTB model of glioma cocultured ECs. Knockdown of Robo4 in this BTB model increased permeability; short hairpin RNA targeting Robo4 (shRobo4) led to decreased transendothelial electric resistance values, increased BTB permeability, and downregulated expression of the EC tight junction proteins ZO-1, occludin, and claudin-5. Roundabout 4 influenced BTB permeability via binding with its ligand, Slit2. Short hairpin RNA targeting Robo4 also increased matrix metalloproteinase-9 (MMP-9) activity and expression in glioma cocultured ECs; pretreatment with the MMP inhibitor GM6001 partially blocked the effects of shRobo4 on the transendothelial electric resistance values and ZO-1 and occludin expression. Short hairpin RNA targeting Robo4 also upregulated the phosphorylation of Src and Erk1/2; the Src inhibitor PP2 and the Erk1/2 inhibitor PD98059 blocked shRobo4-mediated alteration in ZO-1 and occludin expression. Together, our results indicate that knockdown of Robo4 increased BTB permeability by reducing EC tight junction protein expression, and that the Src-Erk1/2-MMP-9 signal pathways are involved in this process. Thus, Robo4 may represent a useful future therapeutic target for enhancing BTB permeability.
authors.affiliation: From the Department of Neurosurgery, Shengjing Hospital of China Medical University (HC, JL, ZL, YH, YL); and Department of Neurology, The First Affiliated Hospital (WL, XS); Department of Neurobiology, College of Basic Medicine (YX, PW, LL); and Institute of Pathology and Pathophysiology (YX, PW, LL), China Medical University, Shenyang, People's Republic of China.
authors.forename: Heng
authors.initials: H
authors.lastname: Cai
authors.forename: Wenjing
authors.initials: W
authors.lastname: Liu
authors.forename: Yixue
authors.initials: Y
authors.lastname: Xue
authors.forename: Xiuli
authors.initials: X
authors.lastname: Shang
authors.forename: Jing
authors.initials: J
authors.lastname: Liu
authors.forename: Zhen
authors.initials: Z
authors.lastname: Li
authors.forename: Ping
authors.initials: P
authors.lastname: Wang
authors.forename: Libo
authors.initials: L
authors.lastname: Liu
authors.forename: Yi
authors.initials: Y
authors.lastname: Hu
authors.forename: Yunhui
authors.initials: Y
authors.lastname: Liu
mesh_terms: D001812:Blood-Brain Barrier / Q000378:metabolism* / Q000473:pathology; D001932:Brain Neoplasms / Q000473:pathology*; D002199:Capillary Permeability / Q000235:genetics; D002478:Cells, Cultured; D062507:Claudin-5 / Q000378:metabolism; D017097:Electric Impedance; D042783:Endothelial Cells / Q000378:metabolism / Q000473:pathology; D015972:Gene Expression Regulation, Neoplastic / Q000235:genetics* / Q000502:physiology; D005910:Glioma / Q000473:pathology*; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins / Q000378:metabolism; D020780:Matrix Metalloproteinase 9 / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000378:metabolism; D062793:Occludin / Q000378:metabolism; D011956:Receptors, Cell Surface / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction / Q000235:genetics; D062725:Tight Junction Proteins / Q000235:genetics / Q000378:metabolism*; D057705:Transendothelial and Transepithelial Migration / Q000502:physiology; D062826:Zonula Occludens-1 Protein / Q000378:metabolism; D019061:src-Family Kinases / Q000378:metabolism; D000098552:Slit Homolog 2 Protein; D000097905:Roundabout Proteins
chemical_list: D062507:Claudin-5; D049452:Green Fluorescent Proteins; D036341:Intercellular Signaling Peptides and Proteins; D020780:Matrix Metalloproteinase 9; D009419:Nerve Tissue Proteins; D062793:Occludin; D011956:Receptors, Cell Surface; D062725:Tight Junction Proteins; D062826:Zonula Occludens-1 Protein; D019061:src-Family Kinases; D000098552:Slit Homolog 2 Protein; C462951:ROBO4 protein, human; D000097905:Roundabout Proteins; C485184:enhanced green fluorescent protein