| ObjectiveMolecular changes occurring with tumor formation and metastasis need to be identified in order to define novel diagnostic markers and target genes for therapeutic intervention, which can be revealed by u-sing subtractive hybridization strategy among the profiling of differently expressed genes from primary tumor samples. Ribosomal protein S24 was identified as a gene up - regulated in human gastric cancer in a suppression subtractive hybridization screen ( SSH) of tumor minus normal tissue from the previous work of Sun Xiuju of China Medical U-niversity. Establishing a greater protein synthesis output may be a necessary step for tumor cells to sustain their rapid proliferation. At present, a large number of researches show that misregulation of protein synthesis is emerging as a major contributory factor in cancer development and several components of the eukaryotic protein synthesis apparatus, such as ribosomal protein, have been associated with onco-genic transformation and malignant progression. Alternative splicing is a versatile form of genetic control whereby a common pre - mRNA is processed into multiple mRNA isoforms differing in their precise combination of exon sciences. The alternatively splicing variants of the same gene product can function in more than one distinct capacity. Expression of alternatively spliced variants mRNA at specific stages of development or in specific cells and tissues contributes to the function-al diversity of the human genome. Certain splice variants that are associated with induction of cell death, regulation of cellular proliferation and differentiation, cell signaling, and angiogenesis are present in a variety of tumors. The essential nature of this process is underscored by the finding that its misregulation is a common characteristic of human disease. There are a few literatures on the expression of ribosomal protein or alternatively splicing transcript in gastric cancer. It has been found that rpS24 mRNAs from multiple human tissues and cell lines revealed two mRNA isoforms which differ from each other. In this study, we employ RT - PCR with splice - specific primer to assess the expression of alternatively splicing transcripts of rpS24 mRNA in gastric cancer, and explore their clinical significance.Materials and Methods1. Subjects The tissues of sixty patients with gastric cancer and seven patients with gastric benign ulcer were collected from oncology department and endoscopy center, the first affiliated hospital, China Medical University from Jan, 2002 to Jan, 2003.2. Extraction of total RNA Total RNAs of the cells in their respective tissue samples were isolated by TRIZOL reagent.3. RT-PCR RNA was changed into cDNA by the process of reverse transcription. The cDNAs were amplified by PCR reaction with the primers of rpS24 splice - specific locus and (3 - actin intrinsic control.4. Detection of PCR products and semi - quantitation The expression of rpS24 spliced variants was detected by 10% PAGE electro-phoresis and analyzed in a form of the band density of their cDNAfragments by Gel work - ID software, lastly the values which were calculated and represented the expression level of mRNA.5. Statistical Analysis Statistical analysis was performed by t test.Results1. Expression of rpS24 mRNA in the patients with gastric cancer and benign ulcerrpS24 mRNA is overexpressed in gastric cancer compared with paired normal mucosa (tumor/normal ratio range, 0. 892 to 3. 568; x s, 1.815 0.4971). There is a significant increase of the expression of rpS24 mRNAs in gastric cancer compared with adj'acent normal mucosa (x s, 0. 6570 0.1314 versus 0. 3713 0. 0582, respective; n =60, P <0. 01). No significant difference is found between gastric benign ulcer and its paired adjacent mucosa (n = 7).2. Expression of rpS24 mRNA splicing variants in the patients with gastric cancer and benign ulcerTwo gastric cancers show rpS24a alone, whereas in all other 58 (96.7%) specimens rpS24c is present wi... |
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