Fig. 6. Production of MMPs in co-cultures. (A) Zymography of conditioned media obtained from co-cultures of DSC and HTR-8/SVneo cells. HTR-8/SVneo cells were added to wells of DSC monolayers. The conditioned media were collected after 24 hours and analysed by zymography. The cells were lysed and loaded for zymography or assayed for MT1-MMP activity. Quantitative determination of total MT1-MMP was performed using the Biotrak MT1-MMP activity assay system (Amersham Biosciences Europe, Orsay, France) according to the manufacturer's instructions. Equal amounts of HTR-8/SVneo and DSC cells were cultivated alone and then processed for zymography and MT1-MMP activity. The increase in MT1-MMP activity in co-cultures was significantly higher (P=0.002) than the summed activity of the enzyme produced by DSC and HTR-8/SVneo cells. (B) Inhibition of haptotaxis. GM6001 (10 µM) was added to block migration of DiI-labeled HTR-8/SVneo cells in the presence of an equal number of DSC. HTS FluoroBlok^TM inserts were previously coated underside with gC1q1 or FN. The migratory process was evaluated at different times (2, 4, 6 and 18 hours). P values were determined versus control samples. Open squares, P>0.05; asterisks, P<0.01.