Figure 3.

TRIB2 enhances prostate cancer cell growth and invasion and confers resistance to enzalutamide.A, immunoblot analysis showing the levels of selected proteins in TRIB2-OE cells. B, colony growth of transfected and parental cells were analyzed after staining with 0.025% crystal violet. C, in vitro invasion was measured using Matrigel-coated Boyden chambers (the scale bar represents 50 μm). D–H, sensitivity of parental and TRIB2-OE cells to enzalutamide was measured by MTS/PES assay. The role of TRIB2 in enzalutamide resistance was verified by treating LNCaP-TRIB2 cells (E) and ENR cells (F–H) with shRNA (1:10) or by Afatinib (6 μM). Positions of the molecular weight markers for the ladder are indicated along with the Western blot data. I–K, impact of TRIB2 overexpression on enzalutamide resistance was measured in soft-agar colony formation assay and in nude mice giving 30 mg/kg/day for 4 weeks (n = 3) (I, the scale bar represents 20 μm). Representative pictures of tumor-bearing mice from different groups as indicated were taken at the end of the study (the scale bar represents 10 mm). Tumor volumes were measured by vernier calipers and presented as mean values ± SE. The data presented as mean values ± SE. ∗∗p < 0.005; ∗∗∗p < 0.0005. For D, E, G, and H, Two-way ANOVA, Tukey’s multiple-comparison test was applied. TRIB2, Tribbles 2; TRIB2-OE, TRIB2-overexpressing.