FIGURE 3

Effects of the exposure to the olfactorin-enriched CM (olfactorin_FLAG) on the chemomigration of GN11 cells. Microchemotaxis experiments were performed in the Boyden's chamber using the CM from COS-7 cells transfected with the empty vector (CM) or with pCMV SPORT6.1 PPT-FLAG-UMODL1 (the olfactorin-enriched CM; olfactorin_FLAG) or pMT21myc-ANOS1 (the anosmin-1-enriched CM; anosmin-1). (A) Comparison of the chemotactic effects exerted by olfactorin- and anosmin-1-enriched CMs on GN11 neurons. Internal controls for the procedure are represented by the chemotaxis to culture medium alone (DMEM) or culture medium with 0.1% Fetal Bovine Serum (FBS) as general chemoattractant. The results are expressed as the number of migrated cells per square mm of the porous membrane in 3 h. Mean ± SD from more than 4–6 independent experiments. *p < 0.05 vs respective CMs; **p < 0.05 vs DMEM. (B) Effect of immunoneutralization with a monoclonal anti-FLAG antibody on GN11 cell chemomigration induced by olfactorin_FLAG. (C) Involvement of HSPG in the chemotaxis of GN11 cells induced by olfactorin_FLAG. The CM was preincubated at 37°C with 30 μg/ml heparin for 30 min, prior the Boyden’s chamber assay. The results (mean ± SD; n. 6) are expressed as the relative chemotactic response of GN11 cells with respect to the stimulus exerted by 0.1% FBS. *p < 0.05 vs respective CM.