Rho GTPase protein expression and activation in murine monocytes/macrophages are not modulated by model biomaterial surfaces in serum-containing in vitro cultures

摘要

The Rho GTPase cellular signaling cascade was investigated in pro-monocyte and (monocyte-)macrophage cells by examining GTPase expression and activation in serum-containing cultures on model biomaterials. Abundance of Rho GDI and the Rho GTPase proteins RhoA, Cdc42 and Rac1 was determined in cells grown on tissue-culture polystyrene, polystyrene, poly-L-lactide and Teflon (R) AF surfaces. Protein expression was compared based on cell maturity (promonocyte to monocyte to macrophage lineages) and by model surface chemistry: Rho proteins were present in the majority of macrophage cells tested on model surfaces suggesting that a pool of Rho proteins is readily available for signaling events in response to numerous activating cues, including biomaterials surface encounter. Rho GTPase activation profiles in these cell lines indicate active Cdc42 and Rho proteins in RAW 264.7, Rac1 and Rho in J774A.1, and Cdc42 and Rac1 in IC-21 cell lines, respectively. Collectively, these proteins are known to play critical roles in all actin-based cytoskeletal rearrangement necessary for cell adhesion, spreading and motility, and remain important to establishing cellular responses required for foreign body reactions in vivo. Differences in Rho GTPase protein expression levels based on cell sourcing (primary versus secondary-derived cell source), or as a function of surface chemistry were insignificant. Rho GTPase expression profiles varied between pro-monocytic non-adherent precursor cells and mature adherent monocyte/macrophage cells. The active GTP-bound forms of the Rho GTPase proteins were detected from monocyte-macrophage cell lines RAW 264.7 and J774A.1 on all polymer surfaces, suggesting that while these proteins are central to cell adhesive behavior, differences in surface chemistry are insufficient to differentially regulate GTPase activation in these cell types. Active Cdc42 was detected from cells cultured on the more-polar tissue-culture polystyrene and poly-L-lactide surfaces after several days, but absent from those grown on apolar polystyrene and Teflon (R) AF, indicating some surface influence on this GTPase in serum-containing cultures.