Study Of MiRNAs Expression Profile On Colorectal Cancers

Colorectal cancer (CRC) remains a common malignant tumor of digestive tract in the worldwide. Several dietary and other lifestyle factors have been implicated in the development of colorectal cancer. As our country’s economy developed at high-speed, people’s living level improved greatly and lifestyle has been Western, the incidence of CRC has improved significantly year by year. In recent years, the incidence and death rate of CRC has risen to No.5and No.5in our country. According to the changes of initiation and development of CRC in other countries and the dynamic of incidence rate of cancer in all regions of China in recent years, we can see that CRC will be one of the fastest rising malignancy cancers in China of this century.MicroRNAs (miRNAs) are small nucleotide RNA species that are expressed from specialized genes. The first miRNA was characterized in the nematode by Lee et al, and was termed as lin-14. It has been reported that miRNAs plays an extremely important role in cell growth, cell death, apoptosis, and metastasis during the carcinogenesis and tumor progression.MiRNAs have been found to have links with multiple types of cancer. It acts as oncogene or cancer suppressor gene and plays an important part in regulation function. In recent years, a number of informative profiling studies have determined the levels of miRNAs in various types of cancer cell lines and tumors. Multiple functional studies on tumor suppressive or oncogenic miRNAs were identified from profiling, screens or due to their cancer-associated genomic loci. Numerous miRNAs and target genes formed a complex network. Many studies showed that miRNA-31has different expression levels in cancers. Previous studies found that miR-31over-expressed in colon cancer, head and neck cancer, hepatocellular cancer, oral cavity squamous cell cancer and lung cancer; low-expressed in bladder cancer, prostate cancer, gastric cancer, breast cancer, serous ovarian cancer and so on. Many evidence demonstrated the involvement of miR-31in tumor metastasis. For example.the high expression of miR-31could prevent the metastasis of breast cancer, but promote the invasion of CRC. This discrepancy could have resulted from the different role of miR-31in these cancers. Ezrin (also known as P81or villin-2) is a member of the ERM protein (Ezrin, Radixm, Moesin family). It is connexin between cell membrane and cytoskeleton and involved in the motility of cancer cells.The research of miRNA is still in the initial stage. But we still don’t know the relationship between the expression of miRNA and the development, invasion and metastasis of CRC. In this study, we focused on the expression of miRNAs in CRC and the adjacent tissues by miRNA gene microarray, determined some cellular functions and molecular pathways targeted by these differentially expressed miRNAs, and discussed differential expression profile and the judgement of clinical biology characteristics in the CRC tissue.11CRC and adjacent tissue treated with surgery were collected and frozen immediately. For microarray analysis, the AFFX microRNA biochip was used. Total RNA was isolated from samples and more than thousand miRNAs were analyzed. The interested miRNAs were conformed by a real-time PCR analysis. This approach enabled the identification of miRNAs whose expression is significantly altered in CRC compared with the normal tissues from the same patient. In result,25miRANs were found to be differentially expressed in tumors compared with the adjacent tissues (P<0.001), including15over-expressed miRNAs (has-miR-31,has-miR-767-5. has-miR-105, has-miR-196b-star, has-miR-224, has-miR-483-5p, has-miR-1246, has-miR-196b, has-miR-663b, has-miR-584, has-miR-141, has-miR-18a-star, has-miR-19a, has-miR-452, has-miR-622) and10low-expressed miRNAs (miR-215, miR-4298, mi-R-139-5p, has-miR-490-5p. miR-3201, miR-363. miR-187. miR-133a. miR-1825. miR-30). The further result of qRT-PCR(Real-Time quantitative fluorescence PCR) showed that the expression of miR-31. miR-105. miR-196b. miR-224, miR-483-5p and miR-767-5p in tumor has up-regulated compared with carcinoma side normal tissue(P<0.05). While miR-139-5p, miR-215. miR-490-5p and miR-4298are obviously down-regulated which consist with chip result(P<0.05).On that basis, including miR-31. miR-105and miR-196b. strongly up-regulated in CRC were selected to the further Real time PCR study with more samples of CRC. The samples are from53of CRC and15of adjacent tissue. And this study used qRT-PCR to comparatively analyse its differential expression and the judgement of clinical biology characteristics. Meanwhile, we analysed gene Ezrin-closely related to miR-31via Targets Scan search method. And we determined its expression in CRC tissue by immunohistochemistry and RT-PCR analysis. We found miR-31was correlated with histological type (P <0.01) and the clinical stage (P<0.05). And no statistically difference was found between miR-31expression and lymph node metastasis on differentiation (P>0.05); miR-105was positive correlated with histological type, clinical stage and lymph node metastasis (P<0.05); while miR-196-5p was only correlated with the histological type of CRC. Additionally, the over-expression of Ezrin protein and mRNA were indicated by western blotting and RT-PCR in CRC, respectively. Analysis of IHC result demonstrated Ezrin was correlated with the metastasis of CRC.Conclusion:Colorectal cancer exists obvious differences of miRNAs expression, miRNAs specifically expressed may become colorectal cancer potential diagnosis and treatment of molecular targets. miR-31, miR-105and miR-196-b high expression indicates poor outcome of colorectal cancer. miR-31related gene Ezrin gene mRNA and protein levels high expression predicts colorectal cancer adverse outcomes, namely the lymph node metastasis.