| PrefaceEvidence from pathology and epidemiology studies has been provided for a human model of gastric tumorigenesis with the following sequential stages: chronic atrophic gastritis (CAG) ; intestinal metaplasia (IM) ; and dysplasia( DYS). An imbalance of cell apoptosis and proliferation was found during the multistep and multifactorial process. It has been reported that Caspase - 3, a cell death protease, is most directly correlated with apoptosis because of its central location in the proteases cascade pathway , and can hydrolyze numerous of protein substrates involving in cell biological and morphologic functions, which directly leads to apoptosis. In this study, we immunolocalized Caspase - 3 in both precancerous conditions and carcinoma of the stomach, and then correlated the findings with cell proliferation studied by Ki - 67 immunostaining and apoptosis detected for DNA fragmentation in situ by TdT - mediated dUTP biotin nick end labeling (TUNEL) method , in order to explore the possible biological significance of Caspase - 3 in cell turnover during tumorigenesis of the gastric mucosa.Materials and Methods1. Clinical PathologyOne hundred and eighty - four gastroscopic mucosal biopsy specimens were derived from the fdes of Cancer Institute of China Medical University, including 6 cases of chronic atrophic gastritis( GAG) ,31 intestinal metaplasia (IM) ,114 dysplasia(DYS) , and 20 gastric carcinomas ( GC, classified as intestinal and diffuse types, 10 cases respectively). All dysplasias were classified as 41 cases of mild, 63 moderate, and 10 severe according to dysplasia degree; and 43 adeno-matous, 22 regenerative, 46 cryptal, 3 globoid dysplasia according to histopathological classifications. The ratio of male and female is 2. 24 with a mean age of 55yrs. Thirteen biopsies without any above lesions were obtained as normal control.2. Immunohistochemistry and TUNELParaffin - embedded biopsy specimens that had been fixed in neutral -buffered formaldehyde solution were sectioned (5um) in series and immunostained using and Caspase - 3 polyclonal antibody ( 1 ;200 dilution, DAKO Company, USA) and Ki ?7 monoclonal antibody ( Ready to use, Maixin Company, Fuzhou, China) by SABC immunohistochemitry. Apoptosis was visualized by TUNEL ( Maixin Company, Fuzhou, China). The staining procedures were done based on the manufacturers recommendations.3. Evaluation of ImmunostainingClearly brown staining restricted to cytoplasm or nuclei was considered as positive for Caspase - 3 or Ki - 67 respectively, and blue to dark purple grains restricted to nuclei were recognized as positive forTUNEL Slides were scored semi - quantitatively according to the percentage of positive cells in all counted cells from 5 randomly selected representative fields based on staining distribution and intensity by two authors (Yang L and Xin Y) as follows; positive rate <5% was defined as negative ( - ) ; weakly positive ( + ) : 5 -25% ; moderately positive ( ++ ) : 25% ~ 50% ; strongly positive ( +++ ): > 50%. Meanwhile, the five chosen images were directly captured through a digital camera (Olympus DP 10, Japan) and subsequently transferred to a personal computer controlled operating system and processed with Metamorph Imaging System v4. 6 (UIC company, USA) by threshold area division. Percentage of positive cells was obtained and designated as Labeling Index (LI). LI = Positive Threshold Area/ Total Counted Threshold Area x 100%.4. Statistical AnalysisStatistical analyses were performed using 2, Pearson, Spearman's rho correlation coefficient, and Student s t - test For all statistical analyses , the SPSS system for personal computer were used, with significance defined as P <0. 05.ResultsCaspase - 3 protein positive staining was restricted in cytoplasm. In normal gastric mucosa, Caspase - 3 positive cells were located predominantly in superficial epitheliam, demonstrated an obvious polar distribution. From CAG, IM, to DYS and GC, the polar distribution gradually disappeared, and LI firstly elevated...
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