Differential Expressions Of MicroRNAs In Gastric Carcinoma Tissue

BackgroundThe non-coding RNA (ncRNA) plays important roles in the process of life, which has been identified by many investigations. NcRNA exercises various functions in the living cells, including the process of storage and transport of the genetic information, as well as catalysis biochemistry reactions.microRNAs (miRNAs) are small NcRNAs of 22 nucleotides in length, and post-transcriptionally regulate gene expression by the base pairing between positions 2 to 8 from the 5'miRNA with the 3'untranslated region of their taget mRNAs. A kind of living organism has hundreds to thousand different miRNAs. In addition, they have different expressing patterns in different stages of biological development, different biological tissues and different kind of cells.Some miRNAs are expressed broadly in the living organisms. However, some are expressed in the distinct growth stage, or expressed with the specific model of tissues.Many experimental studies and clinical investigations have found that miRNAs acts as oncogene and/or anti-oncogene, thereby promoting or restraining the development of cancers. Over-expressed miRNAs in the tumors are looked upon as oncogene, and these miRNAs are called"oncomiRNA". The initial reports of this effects were come from the study of the chronic lymphocytic leukemia(CLL). Accumulating evidence provided by worldwide scientis suggests miRNA expression is disregulated in many kinds of of diverse cancer, which involved with cancer initiation, development, and varied phneotypies. At present, however, few previous reports on the the miRNAs expressing in gastric have been published in the literature. This study applied with miRNA gene chip analysis and quantitative real-time PCR technology to compared the differences of miRNA's expressing between gastric cancer tissues and normal tissues which were away from the focus of tumor >= 5 cm, and bioinformatics were used to predict target genes and KEGG pathway associated with target gene.Purpose:Applying with miRNA gene chip analysis technology to compared the differences of miRNA's expressing between gastric cancer tissues and normal tissues. Quantitative real-time PCR method was used to verify these miRNAs that demonstrated significant differences in more samples.Method:1,Snap frozen tissues from 7 gastric adenocarcinoma confirmed histologically and 7 normal gastric tissues which were away from the tumor's margine >= 5 cm and without cancer cell histologically were collected duing surgical intervention for patients. The miRNAs expression profiling was performend on Illumina microarray platform for miRNAm, which include following steps: purified total RNA from the tissues, RNA polyadenylation, cDNA synthesis, miRNA-specific primer extension, PCR with fluoresenctly labeled universal primers, hybridization, array readout, and normalization of data. Differential Analysis statistics the different expression of miRNAs.2,Tissues from 25 cases of gastric adenocarcinoma diagnosed pathologically and 25 samples of normal gastric tissue which were away from the tumors >=10cm were collected during operations, and purified total RNA from the tissues.According to the nucleotide sequences deposited in miRNA database, primers were designed for has-miR-181a and has-miR-584. Quantitative real-time PCR analysis were performed using locked nucleic acids(LNAs) linear primers and SYBR GreenⅠto detect the expressions of miR-181a and miR-584 between cancer tissues and normal gastric tissues.3,TargetscanS and miRanda computational approaches were used to predicts miR-181a target genes. The biological processes associated with predict targets were evaluated using KEGG pathway online resource.Result:1,There were 9 miRNAs differential expression in the cancer group(p<0.01), 3 miRNAs including HS-138, HS-153, and HS-157) were down-regulated, the another 6 such as hsa-miR-181a, hsa-miR-21, hsa-miR-21*, hsa-miR-27a, hsa-miR-584 and hsa-miR-93) were up-regulated.2,Use quantitative real-time PCR method, overexpressions of miR-181a and miR-584 were observed respectively in 21 and 13 gastric cancer tissues among 25 patients as compared with normal gastric tissue, however, only miR-181a is significantly statistical up-regulation in carcinoma tissues. Mark of type shifts specimen's sex , age , pathological mechanism as well as having a lymph node or not and the expressing condition of miRNA-181a in cancer tissues adopt chi-square test of the discrete variables,we discoveried that high expression of miRNA-181a in cancer tissues is obvious correlative with having a lymph node or not(P=0.009<0.01), as well as the specimen's age(P=0.023<0.05),but the high expression of miRNA-181a in cancer tissues and specimen's sex , pathological mechanism is without correlativity.3,There was 61 predicted target genes using the intersections of both targetscans and MiRanda as a criteria. These predicted target genes were involved 11 KEGG pathway such as human metabolism process; metabolism interaction, genetical signaling pathyway, cell cycle, and so on.Conclusion:1,There are differential expression of miRNAs in the gastric cancer tissues.2,Over-expression of miR-181a in gastric carcinoma tissues was verified using quantitative real-time PCR method, which predicted target genes involved many KEGG pathway.