[Effects of extracellular signal-regulated kinase/mitogen-activated protein kinase signaling pathway activation on proliferation and cell cycle associated genes in human colon cancer cells]

Abstract

Objective: To investigate the effects of extracellular signal-regulated kinase/mitogen-activated protein kinase (ERK/MAPK) signaling pathway activation on the proliferation and cell cycle associated genes in human colon cancer cells.

Methods: Human colon cells of the line SW1116 were cultured, transfected with the recombinant plasmids pCMV-RAF containing the upstream molecular target RAF, pCMV-MEK containing the upstream molecular target MEK, or blank plasmid pCMV respectively, and then screened in the culture fluid with G418, thus obtaining a stable transfected cell clone. MTT method was used to detect the cell viability. The cell growth curve was drawn by using soft agar colony formation test. Flow cytometry was used to analyze the cell cycle. Real-time PCR was performed to detect the mRNA expression of the tumor-associated genes p21(WAF1), p16(INK4A), and c-myc.

Results: MTT method and cell growth curve showed that the viability and proliferation speed of the SW1116 cells transfected with pCMV-RAF and pCMV-MEK were all significantly greater compared with those of the cells transfected with blank vector (all P < 0.01). The numbers of colony of the SW1116 cells transfected with pCMV-RAF and pCMV-MEK were significantly greater and the sizes of the colonies were ignorantly bigger than those of the cells transfected with blank vector. Light microscopy showed that the SW1116 cells transfected with RAF and MEK were polyploidy with mitochysis. The cells at the G0/G1 phase decreased and the cells at the S phase increased (P < 0.01), and the cells at the G2 phase did not change significantly in number. The expression of p21(WAF1) and that of p16(INK4A) were both down-regulated and the expression of c-myc was up-regulated in the SW1116 cells transfected with pCMV-RAF and pCMV-MEK.

Conclusion: Reducing the G1phase and accelerating G1/S transition, up-regulating the expression of c-myc and down-regulating the expression of p21(WAF1) and p16(INK4A), ERK/MAPK signaling pathway activation promotes proliferation of cancer cells.