Figure 5: MAN2C1 promotes the formation of DU145 cell-derived xenograft tumours by inactivating PTEN.

(a) EV and MAN2C1 expressing DU145 cells (5×10⁶) were implanted into flanks of NU/NU mice. Five mice were used for each treatment and animals were terminated at 38 days. Tumour volumes are presented as means±standard derivations. (b) DU145 cells were infected with retrovirus expressing shRNA for MAN2C1, PTEN or both, and then cultured in the presence of puromycin (for MAN2C1 shRNA retrovirus), hygromycin (for PTEN shRNA retrovirus) or both for 2–3 days to achieve 100% infection. A total of 2×10⁵ cells were subsequently implanted into NOD/SCID mice and the mice were killed after 8 weeks. Tumour volumes are presented as means±standard derivations. The asterisk indicates statistical significance (two-tailed Student's t-test) for indicated comparisons, P=0.014 for Ctrl shRNA versus PTEN shRNA, P=0.024 for Ctrl shRNA versus MAN2C1 shRNA, P<0.001 for both Ctrl shRNA versus PTEN shRNA/MAN2C1 shRNA and MAN2C1 shRNA versus MAN2C1/PTEN shRNAs. (c) Representative immunohistochemistry to demonstrate MAN2C1, PTEN and AKT-P expression in tumours derived from each of the cell lines in b. Scale bar, 10 μm. (d) DU145 cells were transfected with an empty retrovirus (EV) or retrovirus expressing MAN2C1, C-MAN236 or Cat-MAN, followed by selection of cells with hygromycin for 2 days to ensure 100% transfection. Five nude mice were s.c. injected with 5×10⁶ of the respective cells. Tumour volumes are presented as means±standard derivations. The asterisk represents P<0.05 for all transgene-derived xenograft tumours versus EV xenograft tumours and MAN2C1 tumours versus the tumours of C-MAN236 and Cat-MAN. All statistical analysis was performed using two-tailed Student's t-test.