Figure 6

Association of PEc with metastases. (A) qRT-PCR analysis indicated a significant increase of PEc expression in HPrEC-PEc cells compared with mHPrEC cells, (reference gene: GAPDH) (Student t test, P < 0.001, n = 3). (B) Similar results were observed by immunofluorescence analysis. (C) The growth rate was significantly induced as assessed by MTT assay, in HPrEC-PEc cells and in mHPrEc cells after the exogenous administration of PEc (17 nmol/L) at 24 and 48 h, as compared with the untreated immortalized mHPrEC cells (p < 0.001 in both cases, Student t test, n = 3. Error bars refers to SD). (D) PEc administration induced ERK1/2 phosphorylation in immortalized mHPrEC cells, while the immortalized HPrEC-PEc cells possessed constantly activated ERK1/2 (Western blot). (E) Kaplan Meier survival analysis of orthotopically injected SCID mice with mHPrEC or HPrEC-PEc cells. The mice injected with HPrEC-PEc cells (seven) presented a statistically significant decrease in survival at 12 wks compared with the mice injected with immortalized HPrEC cells (nine) (P < 0.002) (Cum. Survival: cumulative survival). (F,G,H) Orthotopic injections of HPrEC-PEc cells in SCID mice were associated with metastases in proximal tissues (seminal cord, colon, peritoneum).