Proteomic analysis of pancreatic intraepithelial neoplasia and pancreatic carcinoma in rat models

AIM:To detect the proteomic variabilities of pancreatic intraepithelial neoplasia(PanIN)and pancreatic carcinoma(PC)induced by 7,12-dimethylbenzanthracene(DMBA) in rat models and to identify potential biomarkers.METHODS:Sixty adult male Sprague Dawley rats were randomized into three groups.The rats had DMBA implanted into their pancreas for one(n=20)or two months(n=20)or assigned to the normal group(n =20).The rats were killed after one or two months,and were evaluated histopathologically.Three tissue samples from each group of rats with either normal pancreas,PanIN(PanIN-2)or PC were examined by 2D-DIGE.The different expression spot features were analyzed by matrix-assisted laser desorption/ionizationtime of flight/time of flight(MALDI-TOF/TOF)tandem mass spectrometry.The expression of enolase 1,a differentially expressed protein,was identified by immu-nohistochemistry.RESULTS:There was significant difference in the proportions of neoplastic changes between the 1-and 2-mogroups(P=0.0488).There was an increase in the frequency of adenocarcinomas in the 2-mo group compared with the 1-mo group(P=0.0309).No neoplastic changes were observed in any of the animals in the normal group.Enolase 1,pancreatic ELA3B,necdin,Hbp23,CHD3,hnRNP A2/B1,Rap80,and Gnb2l1 were up-regulated in the PanIN and PC tissues,and CEL,TPT1,NME2,PCK2,an unnamed protein product,and glycine C-acetyltransferase were down-regulated in the PanIN and PC tissues.The immunohistochemical results showed that enolase 1 expression was up-regulated in the pancreatic cancer tissues of rats and humans.CONCLUSION:The pancreatic protein expression changes induced by DMBA suggest potential molecular targets for the early diagnosis and treatment of PC.

Expression and localization of Wolfram syndrome 1 gene in the developing rat pancreas

AIM： To investigate the expression and function of Wolfram syndrome 1 gene （WFS1） during the development of normal pancreas. METHODS： Pancreas from Sprague-Dawley rat fetuses, embryos, young and adult animals were used in this study. Expression levels of WFS1 in pancreas of different development stages were detected by reverse transcription-polymerase chain reation （RT-PCR） and Western blotting. To identify the cell location of WFS1 in the developing rat pancreas, double-immunofluorescent staining was performed using antibodies to specific cell markers and WFS1, respectively. RESULTS： Compared to E15.5, the highest level of WFSl mRNA was detected at E18.5, the level of WFSl mRNA in E15.5 and P0 was less, and at a lowest at adult （P 〈 0.05 vs P0 and adult）, respectively. Compare to E15.5, the highest level of WFS1 was at P14 and lowest at P21 （P 〈 0.05 vs P14 and P21）, respectively. The WFSl positive staining is expressed in the normal developing rat pancreas mainly in the islet beta-cells and mesenchyme at each stage tested. CONCLUSION： These results indicate that WFSl may be involved in some aspects of pancreatic development and further research on WFS1 may provide new evidences to prove the interactions between mesenchyma and epithelia at the same time.

Initial Approach to the Cellular Irradiation Injury of Human Pancreatic Carcinoma Cell Line MIA PaCa-2 by High Dose per Fraction

OBJECTIVE To investigate an initial approach of radiotherapy, which produces cellular radiation injury by high dose in one fraction. METHODS Human pancreatic carcinoma cell line MIA PaCa-2, was cultivated and divided into 5 groups： 0, 2, 5, 10, 17 Gy. Cultivated cells were irradiated by 6MV-X ray in one fraction. Analysis were done as follows： comet assay, which assessed the level of DNA damage in the treated cells right after the cell was irradiated, flow cytometry, which was performed at 0, 6, 12, 24, 36 h after the cell line treated to ladder, which quantitatively asses changes of its cell cycle, DNA assessed the degree of DNA injury after 6 and 12 h, and histological examination, which analyzed cellular morphology after 24 h. RESULTS （1） After X-ray irradiated, the morphological change of human pancreatic carcinoma cell line （MIA PaCa-2） was mainly swelling. （2） When the dose of radiation was lower than 10 Gy, increasing the dose could greatly improve cell necrosis, apoptosis and blockage of cell cycle in GJM phase, which was consistent with the theory of radiation biology. （3） When radiation dose was more than the 10 Gy, the peak of apoptotic necrosis appeared strong and early. （4） The degree of DNA injury was also related to the dose of radiation therapy and most obvious in the 10 Gy group and not so obvious in the 17 Gy group. （5） When dose was less than 10 Gy, DNA ladder was a single electrophoretic band; in the 10 Gy group, the electrophoresis showed a multiple ladder band; when dose was more than 10 Gy, a vague and irregular band appeared on the electrophoresis. CONCLUSION Oncotic necrosis may be the main cell death style when dose per fraction is high, which differs from conventional dose fraction radiation therapy.

Expression and localization of paxillin in rat pancreas during development

AIM:To investigate the expression and localization of paxillin in rat pancreas during development. METHODS:Pancreata from Sprague Dawley rat fetuses, embryos, young animals, and adult animals were used in this study. Expression levels of paxillin in pancreata of different development stages were detected by reverse transcription polymerase chain reaction and Western blotting. To identify the cell location of paxillin in the developing rat pancreas, immunohistochemistry and double-immunofluorescent staining were performed using antibodies for specific cell markers and paxillin, respectively. RESULTS:The highest paxillin mRNA level was detected at E15.5 (embryo day 15.5) following a decrease in the later developmental periods (P < 0.05 vs E18.5, P0 and adult, respectively), and a progressively increased paxillin protein expression through the transition from E15.5 to adult was detected. The paxillin positive staining was mainly localized in rat islets of Langerhans at each stage tested during pancreas development. CONCLUSION:The dynamic expression of paxillin in rat pancreas from different stages indicates that paxillin might be involved in some aspects of pancreatic development.