Density gradient centrifugation of colonic fluid after segmental lavage: Amethod of purification of exfoliative epithelial colonic cells for cytological interpretation and image cytometry in patients with long-standing ulcerative colitis

Objectives: Patients with extensive, long-standing ulcerative colitis (UC) have an increased risk for developing colorectal cancer. In this study, we wanted to establish a method for retrieving cytological material after segm ental colonic lavage for further cytopathological investigations and for pe rforming DNA image cytometry. Methods: Ten patients with long-standing and extensive ulcerative colitis and 10 patients without macroscopic abnormalit ies were investigated. After segmental colonic lavage during routine colono scopy a three-layer (1.146, 1.075, and 1.046 g/ml, respectively) density gr adient centrifugation of the retrieved colonic fluid was performed for isol ation and purification of the epithelial cells. For identification of the e pithelial cells flow cytometry with monoclonal antibody against cytokeratin and counterstaining with propidium iodine was performed. The smears obtain ed were stained for routine cytopathological interpretation and for DNA ima ge cytometry. Results: In eight of 10 UC patients and in nine of 10 control group patients adequate cytological material could be obtained. The band o n top of the density gradient at 1.046 g/ml could be identified as the epit helial cells. Atypical cells were found in smears of three UC patients. In these patients and in one additional patient aneuploid stemlines could be d etected. In smears of control group patients neither atypical cells nor ane uploidy were present. Conclusions: Isolation and purification of epithelial cells after segmental colonic lavage by using density gradient centrifugat ion was performed. This cytological material is adequate for cytopathologic al interpretation and for DNA image cytometry. Information about atypical c ells and DNA aneuploidy as an additional marker of malignant transformation in UC patients was obtained. The combination of cytological examination an d DNA image cytometry might improve the detection of UC patients with high risk for colorectal cancer. (Am J Gastroenterol 1999;94:404-409. (C) 1999 b y Am. Cell. of Gastroenterology).