Due to our previous findings on colorectal cancer, we hypothesized that loss of RKIP may http://www.selleckchem.com/products/epz-5676.html be associated with the frequent occurrence of tumor budding in pancreatic cancer and may play a role in pancreatic carcinogenesis. We therefore undertook the analysis of RKIP expression on a multi punch TMA from a well characterized Inhibitors,Modulators,Libraries cohort of 120 pancreatic ductal adenocarcinomas, their precursor lesions and matched lymph node metastases. RKIP expression was correlated with clinicopathological data and especially with the presence of tumor budding. The REMARK guidelines were used as a basis for this biomarker study. Material and methods Patients and specimen characteristics 120 non consecutive PDAC patients surgically treated between 2000 and 2010 were randomly selected.
Paraffin embedded tissue blocks of primary tumors were retrieved from the Department of Pathology, Inhibitors,Modulators,Libraries Aretaieion University Hospital, University of Athens Medical School, Greece. All histomorphological data were Inhibitors,Modulators,Libraries reviewed from the corresponding hematoxylin and eosin stained slides, while clinical data were obtained from chart reports. Clinicopathological information for all Inhibitors,Modulators,Libraries patients included age, gender, tumor diameter, number of positive lymph nodes and total number of lymph nodes harvested, TNM stage, perineural, as well as blood vessel and lymphatic invasion and resection margin status. Information on post operative therapy was available for all patients. The use of this material was approved Inhibitors,Modulators,Libraries by the local ethics committees of the University of Athens and University of Bern. Assay methods a.
Construction of tissue microarray For each patient, the hematoxylin and eosin slides of the primary tumor from the corresponding whole tissue sections were evaluated and representative areas of the tissue were marked using a felt tip pen for easy detection. To exclude bias because of possible tumor inhibitor Pfizer heterogeneity, each patient had 4 tumor punches taken from formalin fixed, paraffin embedded blocks using a tissue cylinder with a diameter of 0. 6 mm that were subsequently transferred into 1 recipient paraffin block using a homemade semiautomated tissue arrayer. Tissues were obtained from the tumor center and the invasive tumor front so that each patient had at least 4 tumor punches included on this array. Three additional one punch TMAs were con structed including normal pancreatic tissue, precursor lesions and matched lymph node metastases. b. Immunohistochemistry TMA blocks were cut at 4 um and immunostained for pan cytokeratin AE1AE3, that served to highlight areas of tumor budding and RKIP. Sections were de waxed and re hydrated in dH2O. RKIP staining was performed using a Bond Max Autostainer from Leica Microsystem with antigen retrieval performed in citrate buffer at 100 for 20 min.