title: Formation of a protein corona on silver nanoparticles mediates cellular toxicity via scavenger receptors.
abstract: Addition of a protein corona (PC) or protein adsorption layer on the surface of nanomaterials following their introduction into physiological environments may modify their activity, bio-distribution, cellular uptake, clearance, and toxicity. We hypothesize that silver nanoparticles (AgNPs) will associate with proteins common to human serum and cell culture media forming a PC that will impact cell activation and cytotoxicity. Furthermore, the role of scavenger receptor BI (SR-BI) in mediating this toxicity was evaluated. Citrate-suspended 20 nm AgNPs were incubated with human serum albumin (HSA), bovine serum albumin (BSA), high-density lipoprotein (HDL), or water (control) to form a PC. AgNPs associated with each protein (HSA, BSA, and HDL) forming PCs as assessed by electron microscopy, hyperspectral analysis, ζ-potential, and hydrodynamic size. Addition of the PC decreased uptake of AgNPs by rat lung epithelial and rat aortic endothelial cells. Hyperspectral analysis demonstrated a loss of the AgNP PC following internalization. Cells demonstrated concentration-dependent cytotoxicity following exposure to AgNPs with or without PCs (0, 6.25, 12.5, 25 or 50 μg/ml). All PC-coated AgNPs were found to activate cells by inducing IL-6 mRNA expression. A small molecule SR-BI inhibitor was utilized to determine the role of SR-BI in the observed effects. Pretreatment with the SR-BI inhibitor decreased internalization of AgNPs with or without PCs, and reduced both cytotoxicity and IL-6 mRNA expression. This study characterizes the formation of a PC on AgNPs and demonstrates its influence on cytotoxicity and cell activation through a cell surface receptor.
authors.affiliation: *Department of Pharmaceutical Sciences, Skaggs School of Pharmacy and Pharmaceutical Sciences, The University of Colorado Anschutz Medical Campus, Aurora, Colorado 80045, Department of Physics and Astronomy, Clemson University, Clemson, South Carolina 29634, Clemson Nanomaterials Center and COMSET, Clemson University, Anderson, South Carolina 29625, and Department of Environmental Engineering and Earth Sciences, Clemson University, Anderson, South Carolina 29625.
authors.forename: Jonathan H
authors.initials: JH
authors.lastname: Shannahan
authors.affiliation: *Department of Pharmaceutical Sciences, Skaggs School of Pharmacy and Pharmaceutical Sciences, The University of Colorado Anschutz Medical Campus, Aurora, Colorado 80045, Department of Physics and Astronomy, Clemson University, Clemson, South Carolina 29634, Clemson Nanomaterials Center and COMSET, Clemson University, Anderson, South Carolina 29625, and Department of Environmental Engineering and Earth Sciences, Clemson University, Anderson, South Carolina 29625 *Department of Pharmaceutical Sciences, Skaggs School of Pharmacy and Pharmaceutical Sciences, The University of Colorado Anschutz Medical Campus, Aurora, Colorado 80045, Department of Physics and Astronomy, Clemson University, Clemson, South Carolina 29634, Clemson Nanomaterials Center and COMSET, Clemson University, Anderson, South Carolina 29625, and Department of Environmental Engineering and Earth Sciences, Clemson University, Anderson, South Carolina 29625.
authors.forename: Ramakrishna
authors.initials: R
authors.lastname: Podila
authors.affiliation: *Department of Pharmaceutical Sciences, Skaggs School of Pharmacy and Pharmaceutical Sciences, The University of Colorado Anschutz Medical Campus, Aurora, Colorado 80045, Department of Physics and Astronomy, Clemson University, Clemson, South Carolina 29634, Clemson Nanomaterials Center and COMSET, Clemson University, Anderson, South Carolina 29625, and Department of Environmental Engineering and Earth Sciences, Clemson University, Anderson, South Carolina 29625.
authors.forename: Abdullah A
authors.initials: AA
authors.lastname: Aldossari
authors.affiliation: *Department of Pharmaceutical Sciences, Skaggs School of Pharmacy and Pharmaceutical Sciences, The University of Colorado Anschutz Medical Campus, Aurora, Colorado 80045, Department of Physics and Astronomy, Clemson University, Clemson, South Carolina 29634, Clemson Nanomaterials Center and COMSET, Clemson University, Anderson, South Carolina 29625, and Department of Environmental Engineering and Earth Sciences, Clemson University, Anderson, South Carolina 29625.
authors.forename: Hilary
authors.initials: H
authors.lastname: Emerson
authors.affiliation: *Department of Pharmaceutical Sciences, Skaggs School of Pharmacy and Pharmaceutical Sciences, The University of Colorado Anschutz Medical Campus, Aurora, Colorado 80045, Department of Physics and Astronomy, Clemson University, Clemson, South Carolina 29634, Clemson Nanomaterials Center and COMSET, Clemson University, Anderson, South Carolina 29625, and Department of Environmental Engineering and Earth Sciences, Clemson University, Anderson, South Carolina 29625.
authors.forename: Brian A
authors.initials: BA
authors.lastname: Powell
authors.affiliation: *Department of Pharmaceutical Sciences, Skaggs School of Pharmacy and Pharmaceutical Sciences, The University of Colorado Anschutz Medical Campus, Aurora, Colorado 80045, Department of Physics and Astronomy, Clemson University, Clemson, South Carolina 29634, Clemson Nanomaterials Center and COMSET, Clemson University, Anderson, South Carolina 29625, and Department of Environmental Engineering and Earth Sciences, Clemson University, Anderson, South Carolina 29625.
authors.forename: Pu Chun
authors.initials: PC
authors.lastname: Ke
authors.affiliation: *Department of Pharmaceutical Sciences, Skaggs School of Pharmacy and Pharmaceutical Sciences, The University of Colorado Anschutz Medical Campus, Aurora, Colorado 80045, Department of Physics and Astronomy, Clemson University, Clemson, South Carolina 29634, Clemson Nanomaterials Center and COMSET, Clemson University, Anderson, South Carolina 29625, and Department of Environmental Engineering and Earth Sciences, Clemson University, Anderson, South Carolina 29625 *Department of Pharmaceutical Sciences, Skaggs School of Pharmacy and Pharmaceutical Sciences, The University of Colorado Anschutz Medical Campus, Aurora, Colorado 80045, Department of Physics and Astronomy, Clemson University, Clemson, South Carolina 29634, Clemson Nanomaterials Center and COMSET, Clemson University, Anderson, South Carolina 29625, and Department of Environmental Engineering and Earth Sciences, Clemson University, Anderson, South Carolina 29625.
authors.forename: Apparao M
authors.initials: AM
authors.lastname: Rao
authors.affiliation: *Department of Pharmaceutical Sciences, Skaggs School of Pharmacy and Pharmaceutical Sciences, The University of Colorado Anschutz Medical Campus, Aurora, Colorado 80045, Department of Physics and Astronomy, Clemson University, Clemson, South Carolina 29634, Clemson Nanomaterials Center and COMSET, Clemson University, Anderson, South Carolina 29625, and Department of Environmental Engineering and Earth Sciences, Clemson University, Anderson, South Carolina 29625 jared.brown@ucdenver.edu.
authors.forename: Jared M
authors.initials: JM
authors.lastname: Brown
mesh_terms: D000327:Adsorption; D000818:Animals; D001011:Aorta / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D042783:Endothelial Cells / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D015850:Interleukin-6 / Q000235:genetics / Q000378:metabolism; D008075:Lipoproteins, HDL / Q000737:chemistry / Q000378:metabolism*; D008168:Lung / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D053768:Metal Nanoparticles / Q000737:chemistry / Q000633:toxicity*; D010316:Particle Size; D011487:Protein Conformation; D000066970:Protein Corona / Q000737:chemistry / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D051122:Scavenger Receptors, Class B / Q000037:antagonists & inhibitors / Q000378:metabolism*; D012709:Serum Albumin / Q000737:chemistry / Q000378:metabolism*; D012710:Serum Albumin, Bovine / Q000737:chemistry / Q000378:metabolism*; D000075462:Serum Albumin, Human; D012834:Silver / Q000737:chemistry / Q000633:toxicity*; D013499:Surface Properties; D015854:Up-Regulation
chemical_list: C422455:ALB protein, human; D015850:Interleukin-6; D008075:Lipoproteins, HDL; D000066970:Protein Corona; D012333:RNA, Messenger; C495125:Scarb1 protein, rat; D051122:Scavenger Receptors, Class B; D012709:Serum Albumin; D012710:Serum Albumin, Bovine; D012834:Silver; D000075462:Serum Albumin, Human
keywords: darkfield microscopy; endothelial cells; epithelial cells; hyperspectral microscopy; in vitro toxicity; nanotoxicology