PROLIFERATING CELL NUCLEAR ANTIGEN IN NORMAL UROTHELIUM AND UROTHELIAL LESIONS OF THE URINARY-BLADDER - A QUANTITATIVE ASSESSMENT USING A TRUE COLOR IMAGE-ANALYSIS SYSTEM

To evaluate proliferating cell nuclear antigen (PCNA) staining for ass essing proliferative activity in routine pathology specimens of urinar y bladder, the bladder carcinoma cell line J82 and a total of 122 spec imens of normal bladder and urothelial lesions were stained with the a ntibody clone PC10 against proliferating cell nuclear antigen. In in v itro plateau cultures the proportion of PCNA-positive cells exceeded t hat of Ki-67-positive cells, and only very few cells were negative. In formalin-fixed tissues, the PCNA staining pattern, which should be co nfined to replicon units in the nucleus, was optimized by 1 h postfixa tion in an organic solvent (methacarn). Sections showed positive nucle ar staining confined to basal and some suprabasal cells in normal urot helium and grade 1 dysplasias, but more generalized nuclear staining i n all other neoplastic lesions. In addition, stromal cells adjacent to invasive tumors showed nuclear positivity in some instances. Using qu antitative true color image analysis of sections counterstained with h emalum, the degree of brown staining of the PCNA reaction product is c ontrasted with the blue staining of the nuclear area. With this method low contrast specific staining not appreciated optically can be relia bly detected. Image analysis data confirmed observations made on nonco unterstained sections and showed significant differences between grade 1 and 2 dysplasias as well as between grade 1 dysplasia and all grade s of papillary tumor. Furthermore, a significant difference in PCNA st aining indices was found between grade 1 and 3 bladder carcinomas. The results indicate that PCNA staining using the PC 10 antibody is not c onfined to the proliferative fraction of neoplastic urothelium. In con trast with data from normal tissue and malignant hematological neoplas ms, the amount of PCNA is regulated differently in urothelial neoplasm s, emphasizing the biological differences between the following two se ts: mild dysplasia and moderate dysplasia; mild dysplasia and papillar y carcinomas. The use of image analysis to standardize the detection p rocess after controlled staining conditions is advisable in order to p rovide reliable data.