Figure 3. MMP-13 acts as a downstream effector of JAK2/STAT3 activation and cell invasion stimulated by leptin.
A. PANC-1 and AsPC-1 cells were left untreated or were treated with leptin for 24 h, followed by a quantitative reverse transcription-PCR analysis of the expression of MMP-2, MMP-7, MMP-9, and MMP-13 mRNA. The leptin induced the transcriptional activation of MMP-13. B-D. PANC-1 or AsPC-1 cells were treated with leptin alone or in combination with the JAK2 inhibitor AG490 for 24 h. Untreated cells were used as controls. B. The cell lysates were subjected to a western blot analysis of the MMP-13 protein. The histogram shows the densitometric analysis of the bands; the data are expressed as the fold changes in the levels of MMP-13 with respect to β-actin. C. The MMP-13 enzyme activity in the culture supernatants was analyzed using collagen zymography. D. The MMP-13 mRNA level of the treated cells was also analyzed using quantitative reverse transcription-PCR. The leptin enhanced the MMP-13 expression and activity via the JAK2/STAT3 pathway. E, F. PANC-1 cells were infected with a recombinant lentivirus carrying the human MMP-13 gene (MMP-13-overexpressing cells, PANC-1-MMP13) or an empty virus (control cells, PANC-1-Lv). E. A western blot analysis of the expression of MMP-13 in the PANC-1-MMP13 and PANC-1-Lv cells. F. Both of the infected cells were subjected to an in vitro scratch assay (left panel) and a Matrigel-based invasion assay (right panel). The MMP-13 overexpression significantly enhanced the invasion, but not the migration, of the pancreatic cancer cells. G, H. PANC-1 or AsPC-1 cells were treated with leptin alone or in combination with the MMP-13 inhibitor CL82198. Untreated cells were used as controls. The treated cells were subjected to an in vitro scratch assay G. and a Matrigel-based invasion assay H.. The inhibition of MMP-13 attenuated the leptin-stimulated invasion of the pancreatic cancer cells. The data are presented as the mean ± SEM of 3-4 independent experiments. * P < 0.05, ** P < 0.01, compared with the control cells; NS P > 0.05, # P < 0.05, ## P < 0.01, compared with the leptin-treated or MMP-13-overexpressing cells.