The Expression Of Beclin1, Lin28b In Colorectal Adenoma

Objective: Present study showed that the morbidity and mortality of colorectal cancer is growing. It is one of the most common digestive carcinomas. Colorectal cancer is the third most common cause of cancer in men and the second in women worldwide, with an estimated 1.36 million total number of new cases in 2012. It remains as the fourth most common cause of cancer death in the world(2008 and 2012), accounting for 8.5% of all cancer deaths and approximating to 694,000 deaths annually. The pathogenesis of colorectal cancer is still not clear, and most people think that most of the colorectal carcinomas result from colorectal adenoma. Colorectal adenoma can be invided into tubular adenoma, tubulovillous adenoma, villous adenoma, it is crucial to strengthen the understanding of colorectal adenoma.The occurrence and development of the colorectal cancer is related to a variety of factors, and the carcinogenesis of intestinal epithelial cells is a multi-step, multi-stage process and involved in the multi-gene. This process not only involves excessive proliferation, but also has close relationship with cell apoptosis.Beclin1 is the mammalian orthologue of the yeast Apg6/Vps30 gene. In 1998, Liang et al found that this molecule was shown to be a novel 60-k Da coiled protein,when the bcl2-interacting gene was used in a study of lethal encephalitis due to central nervous system infection with Sindbis virus. They named it beclin1. It is located on chromosome 17q21 and codes for a protein that binds to the apoptosis-controlling protein bcl-2. It is the key regulator in autophagy. Furthermore, beclin1 is involved in many biological processes, including development, differentiation, stress adaptation, inflammation, tumorigenesis, aging, and cell death. Multiple diseases are associated with deficiency or malfunction of beclin1, which makes it a potentially valuable target for various therapies, including anticancer treatment. There are indications that the Beclin 1 gene plays an important role in tumor growth. Although the mechanism is not known,Some studies suggest that Beclin1 was to inhibit tumor cell proliferation by inducing tumor cell autophagic cell death or apoptosis, or to inhibit the occurrence and development of tumor through the prevention of tumor necrosis reaction and inflammatory reaction. Its expression is down regulated in cervical cancer, endometrial cancer, breast cancer, prostate cancer, and in these cancer tissues exsisted gene mutations. But in colon cancer Beclin1 was high expression, the mechanism is still not very clear.Lin28 is a RNA-binding protein. Lin28 b is a homolog of Lin28, which induces pluripotency in somatic cells when expressed in concert with KLF4, SOX2, and NANOG. Similar to Lin28, Lin28 b contains a cold shock domain and retro-viral-type CCHC zinc fingers that confer RNA-binding ability, and inhibit biogenesis of tumor-suppressive micro RNAs of the let-7 family. Evidences demonstrate that Lin28 b is implicated in multiple developmental processes, largely as a consequence of its ability to repress let-7 biogenesis. Induction of expression with exogenous Lin28 b promotes cancer cell proliferation.The aim of the study is to detect the expression levels of Beclin1 and Lin28 b in colorectal carcinoma, colorectal adenomas(tubular adenoma, mixed pattern of adenoma,villous adenoma), normal colorectal mucosa, and analyze the relationships among different histological types of colorectal adenomas and colorectal carcinoma,also to explore their effect and significance in the development of the colorectal adenoma, and to provide the new clue for the early diagnosis and therapy for the tumors.Method: We collected 100 colorectal carcinoma tissues after surgical resection and 60 normal colorectal mucosa tissues(adjacent to carcinoma, at least 5cm away from the cancer tissue from the stump of the surgery) during January 2012 to August 2014. None of the 100 patients underwent preoperative chemotherapy, radiation or targeted therapy before surgical resection. 100 colorectal adenomas including 30 tubular adenoma, 35 mixed pattern of adenoma, 35 villous adenoma were collected from endoscopy center of 252 hospital. All the specimens were reviewed and filtrated by senior pathologist. Some of the fresh tissues were obtained and put into the RNA preservation solution, maintained at-20℃, which were used for Fluorescence quantitative PCR. The other part were placed into 10% formalin fixed, embedded by paraffin, then performing continuous sections, which were used for Immunohistochemistry.1 HE staining was used to observe pathological types and the degree of histological differentiation.2 Immunohistochemistry was used to detect the location and expression of Beclin1 and Lin28 b in the tissues.3 Using fluorescence quantitative polymerase chain(PCR) to detect Beclin1 m RNA and Lin28 b m RNA expression.The results were analyzed by chi-square criterion analysis and One-way analysis of variance with SPSS 18.0. The test was considered to be statistically significant when P<0.05.Results:1 Immunohistochemistry: The positive substance of Beclin1 and Lin28 b which stained yellow, were located in the cytoplasm.The results of the immunohistochemistry were that the positive expression rates of Beclin1 protein in colorectal carcinoma tissues, colorectal adenoma and normal colorectal mucosa were 85%, 73%, 35%, respectively, with significant differences among them(P<0.05). The positive expression rates of beclin1 in tubular adenoma, mixed pattern of adenoma, villous adenoma were 43.3%, 60%, 82.9%, respectively, also with significant differences among them(P<0.05). There was a significant correlation between raised expression of Beclin1 and unfavorable variables, including nodal metastasis, Dukes stage, differentiation degree. There was no obvious difference in age and sex. The positive expression rates of Lin28 b in colorectal carcinoma tissues, colorectal adenoma and normal colorectal mucosa were 80%, 65%, 30%, respectively, with significant differences among them(P<0.05). The positive expression rates of Lin28 b in tubular adenoma, mixed pattern of adenoma, villous adenoma were 33.3%, 65.7%, 77.1%, respectively, also with significant differences among them(P < 0.05). There was a significant correlation between the expression of Lin28 b and unfavorable variables, including nodal metastasis, Dukes stage, differentiation degree. There was no obvious difference in age and sex.2 Fluorescence quantitative PCR: Semi-quantitative Taq Man PCR were used to detect the expression of beclin1 and Lin28 b of colorectal cancer and colorectal villous adenoma, mixed colorectal adenoma, colorectal tubular adenoma, colorectal normal mucosa, and the beclin1 m RNA expression of the colorectal carcinoma, villous adenoma, the mixed pattern of adenoma, tubular adenoma, normal colorectal mucosa were 8.196±0.760, 5.680±1.178, 3.898±0.520, 3.566±0.625, 0.312±0.419, respectively. The expression of beclin1 gradually decreased in he colorectal carcinoma, villous adenoma, the mixed pattern of adenoma, tubular adenoma, normal colorectal mucosa. F=75.11074, P<0.05. SNK multiple comparisons displayed that the differences about any two groups of the total mean showed statistical significance. The Lin28 b m RNA expression of the colorectal carcinoma, villous adenoma, the mixed pattern of adenoma, tubular adenoma, normal colorectal mucosa were 17.994± 2.044, 11.080 ± 0.947, 7.232 ± 1.580, 3.966 ± 0.833, 1.656 ± 0.740, respectively. The expression of Lin28 b gradually decreased in the colorectal carcinoma, villous adenoma, the mixed pattern of adenoma, tubular adenoma, normal colorectal mucosa. F=117.5025,P<0.05. SNK multiple comparisons displayed that the differences about any two groups of the total mean showed statistical significance.Conclusion:Both the Beclin1 and Lin28 b expressed highly in colorectal cancer tissue, lowly in normal mucosa. As the villus grows higher in adenoma, the expression of Beclin1 and Lin28 b presented a gradual upward trend. Beclin1 and Lin28 b both showed highest expression level in colorectal carcinoma. Beclin1 and Lin28 b may participate in the transformation process of colorectal adenoma to colorectal cancer. They may be the molecular biological indicators for predicting malignant transformation of colorectal adenoma. Fourthmore, they may also provide the molecular biology basis for colorectal cancer targeted therapy.