Up-regulation of MT1-MMP expression in monocytes bound to activated endothelium or VCAM-1 or ICAM-1 ligands. (A) MT1-MMP cell-surface expression was analyzed by flow cytometry of fresh isolated monocytes (thin solid line) and of monocytes attached for 16 hours to endothelium stimulated with 20 ng/mL TNF-α for 12 hours. MT1-MMP cell-surface expression was also analyzed in monocytes attached to plates coated with 10 μg/mL recombinant forms of VCAM-1 or ICAM-1 for 16 hours. X63 or labeled avidin was used as negative controls of staining of monocytes on endothelium or on VCAM-1 or ICAM-1, respectively (dotted line). One independent experiment of 3 is represented. (B) The arithmetic means and SD of the percentage of MT1-MMP–positive monocytes (subtracting X63 or labeled-avidin negative control values) under different conditions are shown. n = 3. ^*P < .05 (versus fresh) and ^**P < .04 and ^***P < .01 (versus BSA). (C) MT1-MMP cell-surface expression was analyzed as described in panel A in monocytes adhered for 16 hours to different doses of BSA, VCAM-1, or ICAM-1. The arithmetic means of the percentage of MT1-MMP–positive monocytes (subtracting labeled-avidin negative control values) are represented for each condition. n = 3. (D) MT1-MMP protein content was estimated by Western blot analysis of total cellular extracts from fresh monocytes or monocytes attached to plates coated as described for panel A. LPS stimulation for 16 hours was included as a positive control. Anti–MT1-MMP LEM-2/15 mAb recognizes immature and mature forms of MT1-MMP of 63 and 60 kDa, respectively. The arithmetic means and SD of the fold induction of the ratio of MT1-MMP to β-actin levels estimated by densitometry are represented (an arbitrary value of 1 was assigned to the MT1-MMP/β-actin ratio in freshly isolated monocytes). n = 4. ^***P < .01.