Effects of endogenous TNC on cell proliferation and migration.

<p>PANC-1 cells were stably transfected with a vector driving the expression of <i>large</i> TNC (PANC-T2, PANC-T24 and PANC-T27 cells) and with the empty vector (PANC-C21, PANC-C23 and PANC-C27 cells). (A) Immunoblotting of precipitated cell culture medium of the transfected PANC-1 cells grown up to 80% confluence. To ensure that a comparable number of transfected cells was the source of secreted TNC, GAPDH expression in whole cell extracts was tested. PANC-T2 cells show the highest levels of secreted TNC, while lower levels are observed in PANC-T24 and PANC-T27. The control clones in comparison do not show any TNC secretion. (B) Cells were grown in complete medium. Growth was determined with MTT assay at different time points (24, 48 and 72 hours). Data are calculated as mean +/− s.e.m. of three experiments and are expressed as percentage compared to the PANC-1 cells (°p<0.05, *p<0.01, **p<0.001, Students two-tailed t test). The proliferation rate is significantly higher for PANC-T2 at all time points (p<0.001), for PANC-T24 at 24 hours (p<0.001) and at 48 hours (p = 0.022) and for PANC-T27 at 24 (p<0.001) and 48 hours (p = 0.009). (C) The proliferation of all the PANC-1 positive clones (PT) compared to the mock transfected cells (PC) is significantly higher for each tested time point (24 hours: p = 0.004, 48 hours: p = 0.012, 72 hours: p = 0.026) (D) Transfected PANC-1 cells were plated, after 24 hours medium was changed with medium containing 0.1% FBS and, after overnight incubation, the monolayer was scraped with a 10 µl pipette tip. Data are calculated as described in B. In comparison to the non transfected PANC-1 cells, migration is significantly faster for PANC-T2 (p<0.001 at both time points) and for PANC-T27 (p = 0.042 at 24 hours; p = 0.009 at 48 hours). (E) All together, the PANC-1 positive clones (PT) migrate significantly faster compared to the mock transfected cells (PC) at both time points (p<0.001).