title: Effect of surface-adsorbed proteins and phosphorylation inhibitor AG18 on intracellular protein expression in adherent macrophages.
abstract: Macrophages are believed to play an important role in the host inflammatory response to implanted biomaterials. However, the mechanism of macrophage adhesion to protein-adsorbed substrates and the subsequent activation and inflammation is unresolved. Previously the effect of various surface-adsorbed proteins and increasing concentrations of phosphorylation inhibitor AG18 on intracellular protein expression levels in adherent human monocytic cell line U937 was identified using SDS-PAGE and densitometry. The protein ligands and AG18 concentrations up or down regulated the expression of a set of proteins ranging from approximately 200 to approximately 23 kDa. In the present work, HPLC coupled tandem mass spectroscopy (LC/MS) was used to identify proteins in these bands. We hypothesized that key proteins in macrophage adhesion and activation could be identified by observing protein expression resulting from various surface-adsorbed ligands and AG18 concentrations. Increasing concentrations of AG18 down or up regulate protein expression in adherent U937 on PBS-adsorbed TCPS at approximately 52, approximately 42 and approximately 23 kDa. AG18 concentrations had no effect on cells on albumin (Alb)-adsorbed surfaces but regulated different protein expression in adherent U937 on fibronectin (FN)-adsorbed TCPS at 40 and 80 microm AG18. Both Alb and FN regulate distinct sets of proteins in adherent cells as surface-adsorbed ligands. Based on the data from LC/MS, both surface associated ligand and increasing concentrations of AG18 modulate shifts in intracellular signaling.
authors.affiliation: Department of Biomedical Engineering, University of Wisconsin-Madison, Madison, WI 53705, USA.
authors.forename: Sean T
authors.initials: ST
authors.lastname: Zuckerman
authors.forename: Weiyuan John
authors.initials: WJ
authors.lastname: Kao
mesh_terms: D000327:Adsorption; D000818:Animals; D002448:Cell Adhesion / Q000502:physiology*; D002460:Cell Line; D006801:Humans; D008024:Ligands; D008264:Macrophages / Q000166:cytology / Q000502:physiology*; D011506:Proteins / Q000737:chemistry / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology; D013499:Surface Properties; D020032:Tyrphostins / Q000378:metabolism*
chemical_list: D008024:Ligands; D011506:Proteins; D020032:Tyrphostins