title: The P2Y2 receptor mediates uptake of matrix-retained and aggregated low density lipoprotein in primary vascular smooth muscle cells.
abstract: The internalization of aggregated low-density lipoproteins (agLDL) mediated by low-density lipoprotein receptor related protein (LRP1) may involve the actin cytoskeleton in ways that differ from the endocytosis of soluble LDL by the LDL receptor (LDLR). This study aims to define novel mechanisms of agLDL uptake through modulation of the actin cytoskeleton, to identify molecular targets involved in foam cell formation in vascular smooth muscle cells (VSMCs). The critical observation that formed the basis for these studies is that under pathophysiological conditions, nucleotide release from blood-derived and vascular cells activates SMC P2Y2 receptors (P2Y2Rs) leading to rearrangement of the actin cytoskeleton and cell motility. Therefore, we tested the hypothesis that P2Y2R activation mediates agLDL uptake by VSMCs.
  
  
  
  Primary VSMCs were isolated from aortas of wild type (WT) C57BL/6 and.P2Y2R-/- mice to investigate whether P2Y2R activation modulates LRP1 expression. Cells were transiently transfected with cDNA encoding a hemagglutinin-tagged (HA-tagged) WT P2Y2R, or a mutant P2Y2R that unlike the WT P2Y2R does not bind the cytoskeletal actin-binding protein filamin-A (FLN-A).
  
  
  
  P2Y2R activation significantly increased agLDL uptake, and LRP1 mRNA expression decreased in P2Y2R-/- VSMCs versus WT. SMCs, expressing P2Y2R defective in FLN-A binding, exhibit 3-fold lower LDLR expression levels than SMCs expressing WT P2Y2R, while cells transfected with WT P2Y2R show greater agLDL uptake in both WT and P2Y2R-/- VSMCs versus cells transfected with the mutant P2Y2R.
  
  
  
  Together, these results show that both LRP1 and LDLR expression and agLDL uptake are regulated by P2Y2R in VSMCs, and that agLDL uptake due to P2Y2R activation is dependent upon cytoskeletal reorganization mediated by P2Y2R binding to FLN-A.
authors: Human Nutrition, Kansas State University, Manhattan, KS, United States.
authors: Tixieanna
authors: T
authors: Dissmore
authors: Indiana University School of Medicine, Indianapolis, IN, United States.
authors: Cheikh I
authors: CI
authors: Seye
authors: School of Graduate Studies, University of Missouri, Kansas City, MO, United States.
authors: Denis M
authors: DM
authors: Medeiros
authors: Department of Biochemistry and Bond Life Sciences Center, University of Missouri, Columbia, United States.
authors: Gary A
authors: GA
authors: Weisman
authors: Animal Sciences and Industry, Kansas State University, Manhattan, KS, United States.
authors: Barry
authors: B
authors: Bradford
authors: Animal Sciences and Industry, Kansas State University, Manhattan, KS, United States. Electronic address: mamedova@ksu.edu.
authors: Laman
authors: L
authors: Mamedova
mesh_terms: D000199:Actins / Q000378:metabolism; D000818:Animals; D001011:Aorta / Q000378:metabolism; D002465:Cell Movement; D002478:Cells, Cultured; D003599:Cytoskeleton / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004705:Endocytosis; D064448:Filamins / Q000378:metabolism*; D005487:Foam Cells / Q000378:metabolism; D006801:Humans; D008077:Lipoproteins, LDL / Q000097:blood*; D026503:Low Density Lipoprotein Receptor-Related Protein-1; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008840:Microfilament Proteins / Q000378:metabolism; D009131:Muscle, Smooth, Vascular / Q000166:cytology; D009154:Mutation; D032389:Myocytes, Smooth Muscle / Q000378:metabolism*; D011973:Receptors, LDL / Q000378:metabolism*; D058473:Receptors, Purinergic P2Y2 / Q000378:metabolism*; D015398:Signal Transduction; D025521:Tumor Suppressor Proteins / Q000378:metabolism*; D014544:Uridine Triphosphate / Q000737:chemistry
chemical_list: D000199:Actins; D064448:Filamins; D008077:Lipoproteins, LDL; D026503:Low Density Lipoprotein Receptor-Related Protein-1; C486645:Lrp1 protein, mouse; D008840:Microfilament Proteins; D011973:Receptors, LDL; D058473:Receptors, Purinergic P2Y2; D025521:Tumor Suppressor Proteins; D014544:Uridine Triphosphate
keywords: LRP1; P2Y2R