MicroRNA Profile In Distal Gastric Adenocarcinoma And Preliminary Exploration On The Mechanism Of MiR-204

MicroRNAs (miRNAs) are a class of endogenous small non-coding RNAs of 21-25 nt that are found in diverse organisms including plants and animals and involve in negatively post-transcriptional gene expression regulation, mainly through partial base-pairing to the complementary sites within the 3'-UTR (3'-untranslated region,3'-UTR) of their target messenger RNAs (mRNAs). Through years of researches, it was found that miRNAs play a very important role in controlling various cellular processes such as proliferation, differentiation, apoptosis and tumorigenesis. Recently, some miRNAs were observed to function as oncogene or tumor suppressor in the initiation and development of gastric cancer, which provides a novel tool for the diagnosis, treatment and prognostic evaluation of gastric cancer. However, the biological functions of most miRNAs are still largely illusive until now and their functional mechanisms need to be further investigated. Additionally, Several stydies have reported that cardia adenocarcinoma has its specific pathogenesis.To screen and identify the differentially expressed miRNAs in distal (corpus and antrum) gastric adenocarcinoma and then evaluate the relationship of miRNAs' expression and clinicopathological factors in patients with distal gastric adenocarcinoma. Furthermore, the functional mechanism of corresponding miRNAs in the course of development and progression of gastric carcinoma were preliminarily investigated.1. Three distal gastric adenocarcinoma tissues and their paired normal tissues conserved in liquid nitrogen were selected from the tissues archives in the Labs of our department and their pathological features were confirmed through HE staining.2. Agilent human oligonucleotide miRNA microarray were used to profile miRNAs expression in aforementioned matched tissues. Furthermore, five miRNAs with differential expressions in distal gastric adenocarcinoma were selected and validated in an independent set of 20 distal gastric adenocarcinoma and paired normal tissues by using real-time RT-PCR. Then, the relationships between miRNAs expression levels and age, tumor size, staging, grading and lymphonode metastasis were investigated. Additionally, the differential expression levels between 17 cardia adenocarcinoma and distal gastric adenocarcinoma, the expression conditions in serum and several gastric tumor cell lines were observed by using the same method.3. MTS method and flow cytometry were employed to observe the proliferation and apoptosis of tumor cells that transfected with specific miRNA.4,To predict the possible targets of miRNA through bioinformatics.1. Pathological conditions are validated through HE staining for distal gastric adenocarcinoma and paired normal tissues. By using miRNAs microarray,47 miRNAs were found to be differentially expressed between tumor tissues and normal tissues, with 24 miRNAs up-regulated while 23 down-regulated respectively. Among these miRNAs, miR-204, miR-196b, miR-101, miR-107 and other 23 miRNAs are reported here in distal gastric adenocarcinoma for the first time.2. Differential expression levels of miR-196b, miR-204 and miR-375 found with miRNA microarray are validated by using real time RT-PCR. However, no relationship between the expression levels of these three miRNAs and clinopathological factors is observed. In addition, level of miR-204 is found to increased in serum of gastric adenocarcinoma comparing normal controls. It yields an AUC (the area under the ROC curve) of 0.844 (95% CI:0.579-0.973; p=0.002) with 100% sensitivity and 75% specificity in discriminating gastric adenocarcinoma from healthy controls, using the cut-off value 0.0343 (normalized). The expressions of miR-375 in distal gastric adenocarcinma and cardia adenocarcinoma are both decreased, while the latter is more significant.3. There are no significant impact on the proliferation and apoptosis of gastric tumor cell lines after transfecting mature miR-204 mimics, independent of the transfectional concentration or time points.4. Bioinformatics indicates that BCL-2, NR3C1 and SOCS6 may be the targets of miR-204.1.There is a specific miRNA profile in distal gastric adenocarcinoma.2.There are no significant correlation between levels of specific miRNAs and clinopathological factors in distal gastric adenocarcinoma. The levels of specific miRNAs in distal gastric adenocarcinoma and cardia adenocarcinoma are different. The abundance of miR-204 is significant higher in patients with gastric adenocarcinoma comparing healthy controls. And it may be a potential biomarker for gastric adenocarcinoma.3. There are no significant impact on the proliferation and apoptosis of gastric tumor cell lines after over expression of mature miR-204.4. BCL-2, NR3C1 and SOCS6 may be targets of miR-204.