Tumor-associated Gene Expression And Epigenetic Modification Of Colorectal Polyps In Children

Colorectal polyp is one of the common causes of hematochezia in children, whichthe juvenile polyp is the most common histological type. Its mechanism is not clear. Most scholars believe that the prognosis is good, but there is still some risk of recurrence and malignant potential. In recent years, the role of genetic and epigenetic mechanisms in the development of tumor has been paid more attention. The studies on adult have shown that intestinal stem cells of β-catenin over-accumulation due to mutations and other reasons, leading to the intestinal stem cell self-renewal, clone proliferation and differentiation faster than normal intestinal epithelial cell apoptosis rate, resulting in intestinal epithelial cells continue to amplified and over adjacent normal intestinal crypts. It was considered to be the reasons for the formation of polyps. The cell signal transduction pathway such as Wnt, PTEN and BMP signaling pathways regulate β-catenin levels of intestinal stem cells, leading to the dynamic equilibrium between intestinal epithelial cell renewal and apoptosis, so as to control intestinal epithelial cell regeneration and the normal development process. APC is a tumor suppressor gene, which is the classic Wnt pathway blocker. PTEN is phosphatidylinositol3-kinase/protein kinase (PI3K/AKT) pathway inhibitor, which regulates the rate of cell renewal and apoptosis together with Wnt pathway. However, tumor suppressor gene APC, PTEN mutations in the most common juvenile colon polyps in children has not been reported.Epigenetic modifications such as DNA methylation, histone acetylation is currently a hot research field of tumor. Many gene promoter region CpG island methylations has become a cancer detection, treatment and prognosis of molecular markers in colorectal cancer in adults. The state of histone acetylation was regulated by the histone acetyltransferase (HATs) and histone deacetylases (HDACs). HATs and HDACs control the dynamic balance between chromatin structure and gene expression, and their dysfunction in tumor development is an important molecular mechanism. However, the occurrence of juvenile polyps in children, whether there is development of APC gene promoter methylation changes and histone HATs and HDACs dysfunction is unclear.ObjectiveTumor suppressor genes APC, PTEN and oncogene β-catenin mRNA and protein expression in colorectal polyps, polyp pedicle mucosa and normal colonic mucosa tissue, and polyps APC gene promoter methylation status, acetyltransferase and deacetylase expression of intestinal polyps in children were performed to understand tumor-associated gene APC, PTEN and β-catenin expression and epigenetic changes, so as to attempt to clarify the pathogenesis of intestinal polyp formation, to provide a theoretical basis for the new strategies in the prevention and treatment of colorectal polyps in children.MethodsSpecimens of polyps diagnosed by colonoscopy in our hospital after electronic removing, polyp pedicle mucosa and normal colonoscopy colon mucosa in the same period were collected. The tumor suppressor gene APC, PTEN and oncogene β-catenin, and acetyltransferase (CBP, PCAF, P300), deacetylases (HDAC4, HDAC1) mRNA expression in colorectal polyps, polyp pedicle mucosa and normal mucosa organization in children was detected by RT-PCR. The protein expression levels of APC, PTEN, β-catenin were measured by immunohistochemical method. APC gene promoter methylation status in polyps and normal colon mucosa were performed by applied bisulfite genomic sequencing (bisulfite genomic sequencing).Resultsβ-catenin mRNA expression both in multiple polyps and in single polyps was significantly increased than that in normal colorectal mucosa, while the PTEN and APC mRNA expression were decreased, but the difference was not statistically significant. β-catenin mRNA expression in polyp pedicle mucosa regardless of multiple polyps or single polyps was significantly increased than that in normal colorectal mucosa. PTEN mRNA expression was decreased, but the difference had not reached significant, while the APC had not obvious change. APC and PTEN protein expression detected by immunohistological methods were significantly decreased both in multiple polyps and in single polyps than that in normal colonic mucosa, while β-catenin protein expression was increased, but the difference had no statistically significance. Our study found that P-catenin was fully expressed on the membrane in colorectal polyp, polyp pedicle mucosa or normal mucosa tissue. In addition to10%methylation rate of APC CpG island in individual polyp pedicle mucosa, sequencing indicated that APC gene amplification products in most of colorectal polyp, polyp pedicle mucosa and normal mucosa have become TpG from CpG, that means no methylation. Acetyltransferase CBP both in multiple polyps and single polyps was significantly decreased than that in normal colonic mucosa, while deacetylase HDACl was increased (P=0.051), but the other acetyltransferase was no significant difference. Acetyltransferase CBP both in multiple polyp pedicle mucose and single polyp pedicle mucose was significantly decreased than that in normal colonic mucose, the other acetyltransferase had not reached significant among three groups.ConclusionColorectal polyps in children, whether single, multiple polyps or its pedicle mucosa, the gene transcription level of β-catenin expression was significantly increased, while APC and PTEN protein level expression was significantly decreased. Our study also found that the degree of acetylation was reduced, while the APC gene methylation events occurred less frequently. It was suggested that decreased expression of tumor suppressor genes and increased expression of oncogenes may be involved in the formation of colorectal polyps in children, and decreased acetylation degree may be the characteristic of epigenetic.