title: Immunohistochemical markers in urinary bladder carcinomas from paraffin-embedded archival tissue after storage for 5-70 years.
abstract: OBJECTIVE
  To evaluate archival tissue specimens from bladder tumours and seek molecular changes in samples collected over seven decades previously, as although the frequencies of some cancer types have remained stable during the last 50 years, the incidence of others, including bladder tumours, has increased significantly, and molecular analyses of bladder cancer over periods with an increasing incidence are of interest as the findings might reflect varying external influences.
  
  
  METHODS
  Immunohistochemical staining with the biological markers p53 protein, p16 protein, epidermal growth factor receptor (EGFR), cytokeratin 7 and high molecular weight 34betaE12 cytokeratin (HMW-cytokeratin, characteristic of basal cells) was used on archival, paraffin wax-embedded autopsy/biopsy tissue material collected from 144 patients with invasive bladder cancer (World Health Organisation grade II and III). The cases were selected from the periods 1932-48, 1950-59, 1960-70 and 1990-2004. Control immunohistochemistry was done on available normal tissue (i.e. connective and fatty tissue, heart, lungs and normal urinary bladder epithelium) obtained from the autopsies.
  
  
  RESULTS
  The normal tissues were all largely negative for EGFR, had <1% positively stained nuclei for p53 and strong positive reactions for p16, and in epithelial tissues the two cytokeratins were detected. The positive scores for HMW-cytokeratin in the tumour tissue decreased significantly from approximately 90% to 30% over the 70 years. For p53 there was a higher fraction of positive scores (borderline significant) with time. The p16-positive tumours showed no significant variation, with the highest frequency of positive scores in recent years. Overexpression of EGFR in the tumours was significantly correlated with the occurrence of HMW-cytokeratin and decreased from approximately 85% to 65% (not significant), with the lowest frequency in the samples from 1990 to 2004. Autolysis after death or long storage periods did not compromise good quality in the histochemical analyses of the autopsy tissue.
  
  
  CONCLUSIONS
  The higher frequency of HMW-cytokeratin, lower p53 accumulation and more EGFR expression in grade II and III urinary bladder carcinomas from the 1930s could indicate different phenotypes in bladder cancer during this 70-year period. The successful detection of these protein markers in old archival material allows larger retrospective studies that might increase the understanding of molecular carcinogenesis in bladder cancer.
authors: The Gade Institute, Section for Pathology, and Department of Surgery, Section of Urology, Haukeland University Hospital, Bergen, Norway. Jorunn.Litlekalsoey@student.uib.no
authors: Jorunn
authors: J
authors: Litlekalsoy
authors: Vendil
authors: V
authors: Vatne
authors: Jens G
authors: JG
authors: Hostmark
authors: Ole Didrik
authors: OD
authors: Laerum
mesh_terms: D000368:Aged; D019941:Cyclin-Dependent Kinase Inhibitor p16 / Q000032:analysis; D066246:ErbB Receptors / Q000032:analysis; D006801:Humans; D007150:Immunohistochemistry / Q000592:standards; D053552:Keratin-7 / Q000032:analysis; D016612:Paraffin Embedding; D013997:Time Factors; D014015:Tissue Banks; D014021:Tissue Preservation; D016159:Tumor Suppressor Protein p53 / Q000032:analysis; D001749:Urinary Bladder Neoplasms / Q000737:chemistry* / Q000473:pathology
chemical_list: D019941:Cyclin-Dependent Kinase Inhibitor p16; D053552:Keratin-7; D016159:Tumor Suppressor Protein p53; D066246:ErbB Receptors