JARARHAGIN STIMULATES EPITHELIAL CELL MIGRATION, PROMOTES ACTIN POLYMERIZATION AND THE RECRUITMENT OF SPECIFIC INTEGRINS TO FOCAL CONTACTS

Costa, E.P; Santos, M.F.

Department of Histology and Embriology, Institute of Biomedical Sciences, USP

Jararhagin(JG), a snake venom metalloprotease, induces the influx of leukocytes in vivo in an air pouch model, without being chemoctatic(Costa et al., 2002). In this study we analysed the direct effects of JG on epithelial cell adhesion and migration, F-actin arrangement, laminin and fibronectin distribution, as well as the expression of some laminin and fibronectin-binding integrins.

Methods and Results: IEC-6 cells were grown to confluence, when 1/3 of the monolayer was removed in order to stimulate spontaneous migration. Migrating cells were treated with several doses of JG, and maximum stimulation was obtained with 1,25mg/ml (Matrigel) or 5 mg/ml (treated plates). Cell adhesion was also studied in the presence of JG during 20 min. An inhibition of 45% and 40% was observed with the doses of 1,25 and 5 mg/ml JG, rescpectively. The distribution of some integrins (a2 and av subunits, a6b1), fibronectin and laminin was studied using immunofluorescence with comercial antibodies (Chemicon) and confocal analysis (Nikon). F-actin was stained with rodhamine-phalloidin (Molecular Probes). JG stimulated actin polymerization in stress fibers, filopodia and membrane ruffles after 15 min. The pericellular fibronectin matrix was lost in migrating cells, while laminin distribution was less affected. The toxin also stimulated the recruitment of a6b1 and av integrin subunit to focal contacts. The distribution of the a2 subunit, also present in focal contacts, was not altered.

Conclusion: Jararhagin stimulates the migration of epithelial cells in vitro through a mechanism the involves both quantitative and qualitative changes in cellular adhesion to the substrate and the formation of actin-rich cellular processes.

Financial support: FAPESP

CORRESPONDENCE TO:

Érica Pereira da Costa, Avenida Inocencio Pires de Oliveira, 441, Cotia, SP, CEP: 06705-125, Brasil, Email: ericacosta22@hotmail.com