Relationship between expression and methylation status of p16INK4a and the proliferative activity of different areas’ tumour cells in human colorectal cancer

Abstract

The p16(INK4a) gene is a cell cycle inhibitor and a major tumour suppressor protein, but the regulation and effects on tumour cells' invasion process of p16(INK4a) is poorly known. A role for p16(INK4a) in basal cell carcinoma is suggested by the observation that p16(INK4a) was upregulated at the invasive front of the majority of basal cell carcinomas with infiltrative growth patterns, accompanied by cessation of proliferation. In this paper, we explore whether there is a difference of tumour cells' proliferative activity between the centre and the invasion front tissues of human colorectal cancer and its relationship with the expression and methylation status of p16(INK4a) gene. We obtained the centre and the invasion front tissues of colorectal cancer respectively by the technology of laser mircodissection. The expressions of the proliferating cell nuclear antigen ki67 and p16(INK4a) were assessed by immunohistochemistry, methylation-specific polymerase chain reaction (MS-PCR) and reverse-transcription polymerase chain reaction (RT-PCR) in the different areas. There was a significant difference in the expressions of ki67 between the centre and the invasion front tissues (p < 0.05). The difference did not correlate with age, sex, Dukes stage but did correlate with expression of p16(INK4a) gene (chi(2) = 25.37, p < 0.01). Furthermore, hypermethylation of the promoter was the major mechanism of inactivation of p16(INK4a) in the centre areas. Demethylation of the p16(INK4a) promoter, the elevated expression of p16(INK4a) protein and mRNA level was proved in the invasion front. Our finding suggested that enhanced invasion in tumours was accompanied by ceased proliferation in the invasion fronts of human colorectal cancer. This interesting phenomenon may be due to not only the microenvironment, but also the molecular changes such as p16(INK4a) status.