Figure 2. Effect of endogenous PTHrP on LoVo cell proliferation, migration and invasion.

(A) Proliferation. Cells were plated in 96-well dishes at 10⁴ cells/well. Cell number was determined at the indicated time intervals. For si and sc, each point is the mean ± SEM of three independent experiments each for si1 and si2 or the respective scrambled (sc) siRNA control. (B and C) Migration and invasion. Cells (0.5 × 10⁶) were loaded with Calcein-AM and plated onto FluoroBlok inserts in the absence of FBS. To measure invasion, the inserts were overlaid with Matrigel. FBS (10%) was used as the chemoattractant. Migration and invasion were measured after 4 h. In (B and C), 1 and 2 refer to two independent PTHrP siRNA-transfected cell lines. Each bar is the mean ± SEM of three independent experiments. (D) Migration was analyzed using a monolayer scratch assay. Images at 0 and 5 days after wounding are shown. The arrowed lines mark the edges of the monolayer. Magnification, ×10. (E) Quantitation of cell migration 3, 4, and 5 days after wounding the cell monolayer. si is the mean ± SEM of three independent experiments each for si1 and si2. * = Significantly different from the control value (P < 0.001).