title: Profiling of kidney vascular endothelial cell plasma membrane proteins by liquid chromatography-tandem mass spectrometry.
abstract: BACKGROUND
  Vascular endothelial cells (VECs) play crucial roles in physiological and pathologic conditions in tissues and organs. Most of these roles are related to VEC plasma membrane proteins. In the kidney, VECs are closely associated with structures and functions; however, plasma membrane proteins in kidney VECs remain to be fully elucidated.
  
  
  METHODS
  Rat kidneys were perfused with cationic colloidal silica nanoparticles (CCSN) to label the VEC plasma membrane. The CCSN-labeled plasma membrane fraction was collected by gradient ultracentrifugation. The VEC plasma membrane or whole-kidney lysate proteins were separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis and digested with trypsin in gels for liquid chromatography-tandem mass spectrometry. Enrichment analysis was then performed.
  
  
  RESULTS
  The VEC plasma membrane proteins were purified by the CCSN method with high yield (approximately 20 μg from 1 g of rat kidney). By Mascot search, 582 proteins were identified in the VEC plasma membrane fraction, and 1,205 proteins were identified in the kidney lysate. In addition to 16 VEC marker proteins such as integrin beta-1 and intercellular adhesion molecule-2 (ICAM-2), 8 novel proteins such as Deltex 3-like protein and phosphatidylinositol binding clathrin assembly protein (PICALM) were identified. As expected, many key functions of plasma membranes in general and of endothelial cells in particular (i.e., leukocyte adhesion) were significantly overrepresented in the proteome of CCSN-labeled kidney VEC fraction.
  
  
  CONCLUSIONS
  The CCSN method is a reliable technique for isolation of VEC plasma membrane from the kidney, and proteomic analysis followed by bioinformatics revealed the characteristics of in vivo VECs in the kidney.
authors.affiliation: Department of Structural Pathology, Institute of Nephrology, Graduate School of Medical and Dental Sciences, Niigata University, Niigata, Japan.
authors.forename: Zan
authors.initials: Z
authors.lastname: Liu
authors.forename: Bo
authors.initials: B
authors.lastname: Xu
authors.forename: Masaaki
authors.initials: M
authors.lastname: Nameta
authors.forename: Ying
authors.initials: Y
authors.lastname: Zhang
authors.forename: Sameh
authors.initials: S
authors.lastname: Magdeldin
authors.forename: Yutaka
authors.initials: Y
authors.lastname: Yoshida
authors.forename: Keiko
authors.initials: K
authors.lastname: Yamamoto
authors.forename: Hidehiko
authors.initials: H
authors.lastname: Fujinaka
authors.forename: Eishin
authors.initials: E
authors.lastname: Yaoita
authors.forename: Masayuki
authors.initials: M
authors.lastname: Tasaki
authors.forename: Yuki
authors.initials: Y
authors.lastname: Nakagawa
authors.forename: Kazuhide
authors.initials: K
authors.lastname: Saito
authors.forename: Kota
authors.initials: K
authors.lastname: Takahashi
authors.forename: Tadashi
authors.initials: T
authors.lastname: Yamamoto
mesh_terms: D000818:Animals; D002462:Cell Membrane / Q000737:chemistry*; D002853:Chromatography, Liquid / Q000379:methods; D042783:Endothelial Cells / Q000737:chemistry*; D007150:Immunohistochemistry; D007668:Kidney / Q000166:cytology*; D008565:Membrane Proteins / Q000737:chemistry* / Q000302:isolation & purification; D053758:Nanoparticles; D040901:Proteomics / Q000379:methods; D051381:Rats; D017208:Rats, Wistar; D012822:Silicon Dioxide; D053719:Tandem Mass Spectrometry / Q000379:methods; D059467:Transcriptome
chemical_list: D008565:Membrane Proteins; D012822:Silicon Dioxide