Boyden Chamber Assay to Study of Cell Migration Induced by Metalloprotease Cleaved-CD95L.

Abstract

CD95 receptor, also called Fas or Apo-1, is a member of the tumor necrosis factor receptors (TNF-R) superfamily (Itoh and Nagata, J Biol Chem 268:10932-10937, 1993). Its cognate ligand, CD95L, is a transmembrane cytokine, which can be cleaved by metalloproteases (Matsuno et al., J Rheumatol 28:22-28, 2001; Vargo-Gogola et al., Arch Biochem Biophys 408:155-161, 2002; Kiaei et al., Exp Neurol 205:74-81, 2007; Schulte et al., Cell Death Differ 14:1040-1049, 2007) releasing a soluble ligand into the bloodstream. Recent work has shown that this metalloprotease-cleaved CD95L (cl-CD95L) is involved in carcinogenesis (Malleter et al., Cancer Res 73:6711-6721, 2013). Cl-CD95L also fuels the inflammatory process in patients affected by systemic lupus erythematosus by promoting the accumulation of activated T lymphocytes in enflamed organs (Tauzin et al., PLoS Biol 9:e1001090, 2011). This chapter aims at describing the methodology used to measure the chemoattractive effect of cl-CD95L on human cancer cells and lymphocytes.