Expression of the receiver for mannose in cells of Schwann and Schwannoma ST88-14.
The mannose receptor (MR) is a transmembrane glycoprotein that is expressed in several cell types but little or no information is available on Schwann cells (SC). We show that rodent SC in primary cultures of dissociated cells or explantnerve cultures and a human Schwannoma cell line (ST88-14) bind the exogenous MR ligand neoglycoprotein mannosyl/bovine serum albumin-fluorescein isothiocyanate (man/BSA-FITC) in a highly specific manner. After incubation with man/BSA-FITC, flow cytometry demonstrates 62% positive ST88-14 cells and 90% positive SC cells, a dose-dependent increase in tagged ligands and near total inhibition of binding by 250 mM D-mannose or ~ 1.1 amp;#956;M of the highly mannosylatedprotein horseradish peroxidase (HRP). Treatment of cultured SC with dexamethasone (0.1 amp;#956;g/ml) or interferon amp;#61543; (IFN-amp;#61472;amp;#61543; ? 100 U/ml) followed by tagging with man/BSA-FITC and analysis by flow cytometry shows an increase and a decrease, respectively, of the ligand uptake by the putative receptor. Sciatic nerve explants cultured in the presence of IFN-amp;#61472;amp;#61543; show colocalization of man/BSA-FITC and MHC class II immunoreactivity. Ultrastructural analysis of ST88-14 cells afterincubation with HRP-colloidal gold without or with subsequent chasing at 37ºC mostra localização inicial na superfície cellular e internalização ation on the cell surface and temperature- and time dependent internalization of the probe. ST88-14cells and dissociated SC and recently teased nerves exhibit reactivity to a polyclonal antibody against the C-terminal domain of the mouse macrophage MR (anti-cMR). Inthe case of teased nerves, immunoreactivity was particularly abundant in regions topographically homologous to paranodal loops. Furthermore, ultrastructural immunohistochemistry of Lowicryl-embedded sciatic nerve from normal animalsshowed reactivity to anti-cMR with maximal deposition of Au-tagged secondary antibody in myelinating or non-myelinating SC somata. SDS-PAGE separated proteins studied by endogenous lectin binding of SC and ST88-14 extracts show that both share a unique HRP-binding protein of about 180 kDa with peritoneal macrophages. A single band of 180 kDa was also obtained in immunoblots with anticMR. Our results suggest that Schwannoma cells and SC in vitro and in situ expressa MR-like protein in a prospectively functional state and that both types may be useful models for studies regarding the role of this receptor in different infectious/inflammatory states or in homeostasis of the peripheral nervous system.
Advisor:Leny Alves Cavalcante; Suzana Corte Real Faria
School:Faculdades Oswaldo Cruz
Source Type:Master's Thesis
Date of Publication:09/26/2006