Study On Differential Proteome And Immunohistochemistry In Adenocarcinoma And Normal Mucosal Tissues Of Stomach

Part One:Establishment of 2-DE Profile and Analysis of Differential Proteins in Adenocarcinoma and Normal Mucosal Tissues of StomachBackground and objective:Gastric carcinoma is a disease related to multiple factors. Past studies are mostly concentrated on a single or a few related genes and their protein products. These studies are unable to reflect the overall changes of DNA in gastric carcinoma. Some new techniques have been used to study the genomic DNA or mRNAs, such as comparative genomic hybridization and microarray. But the changes of DNA or mRNAs cannot fully reflect the changes of intra-cellular proteins. In addition, protein functions are affected by serial events including post-translational modification, cellular compartment localization, conformational changes and the interaction between proteins or protein and other biological molecules. Studying gastric carcinoma at the proteome level in combination with genomic DNA and mRNAs information will advance our understanding of the process of carcinogenesis in gastric carcinoma. In the study, we established a 2-DE profile of differential protein expressions of normal gastric mucosal tissues and gastric cancer by using 2-DE and mass-spectrometry techniques. We identified several proteins that are significantly overexpressed or down regulated in gastric cancer and demonstrated their significant role in gastric cancer etiology.Methods:Normal gastric mucosal tissues and poorly-differentiated human adenocarcinomas were obtained directly from the patients with gastric cancer who underwent surgical resection. Two-dimensional gel electrophoresis is used to construct the high-resolution and repetitiveness 2-DE profile of human gastric carcinoma and normal gastric mucosal tissues. Differential protein spots in 2-DE were identified by peptide mass-spectrometry fingerprinting or peptide sequence label.Results:We successfully generated high-resolution 2-DE profiles of human gastric carcinoma tissues and normal gastric mucosal tissues. The profile is highly repeatable. We identified 29 differential protein spots,10 of them were up-regulated and 19 down-regulated. Among these differentially expressed proteins, some were known expressed in other tumors. Using bioinformatic methods and the protein information in ExPASy and NCBI database, we initially analyzed the subcellular localization and functional classification of these differentially expressed proteins.Conclusion:Using high-resolution 2-DE profiling, we identified many proteins differentially expressed between gastric carcinoma tissues and normal gastric mucosal tissues. The study is very important in that it not only attributes to our understanding of,the etiology of gastric carcinoma, but also can potentially be used for clinical diagnosis. Part Two:Expression of Annexin A1 and Galectin-1 in Gastric Cancer and Normal Mucosal Tissues and Their Clinicopathological SignificancesBackground We studied differential expression of proteins using 2-dimensional gel electrophoresis followed by mass spectrometry and found 10 up-regulated and 19 down-regulated proteins in gastric cancer tissues as described in part one. To confirm above experimental results, we selected one up-regulated protein (galectin-1, gal-1) and one down-regulated protein (annexin A1, ANXA1) and examined their levels of expression in gastric cancer and normal mucosal tissues by EnVisionTM immunohistochemical study.Objective Examine the levels of expression of gal-1 and ANXA1 in gastric cancer and normal mucosal tissues and their clinicopathological significancesMethods EnVisionTM immunohistochemistry was used to determine the levels of gal-1 and ANXA1 in conventional paraffin-embedded sections from resected specimens of gastric cancer (n= 45) and peritumoral normal mucosal tissues (n= 45).Results The positive rate and mean level of ANXA1 were significantly lower in gastric cancer tissues than that of normal mucosal tissues (62.2% vs 86.7%, x2= 7.07, P<0.01; 2.7±1.1 vs 4.5±0.8, t= 9.01, P<0.01). The positive rate and mean level of gal-1 were significantly higher in gastric cancer tissues than that of normal mucosal tissues (66.7% vs 6.7%, x= 34.88, P<0.01; 3.4±1.0 vs 0.3±0.9, t= 15.50, P<0.01). The positive rates and scoring means of ANXA1 were significantly higher in well-differentiated adenocarcinoma, invasive carcinomas with a depth of T1 or T2 and without regional lymph node metastasis than those in poorly-differentiated adenocarcinoma, invasive carcinomas of depth T3 or T4 and with regional lymph node metastasis (P<0.05 or P<0.01). The positive rates and scoring means of gal-1 were significantly lower in well-differentiated adenocarcinoma, invasive carcinomas with a depth of T1 or T2 and without regional lymph node metastasis than those in poorly-differentiated adenocarcinoma, invasive carcinomas of depth T3 or T4 and with regional lymph node metastasis (P< 0.05 or P<0.01). The negative correlation was found between gal-1 and ANXA1 in gastric cancer tissues (r=-0.36, P<0.01).Conclusions ANXA1 and/or gal-1 are important biological markers for gastric cancer. Their levels of expression are important markers for carcinogenesis, biological behaviors, progression and prognosis of gastric cancer. Gastric adenocarcinomas with depressed ANXA1 and/or over expression of gal-1 are likely to have poor outcomes.