Previous studies have found a 3-10-fold amplification and overexpression of the cyclin D1 gene in about 32% of human esophageal squamous cell carcinoma. The purpose of this study was to evaluate the prevalence of increased expression of the cyclin D1 protein in Barrett's esophagus. Using 69 formalin-fixed and paraffin-embedded human esophageal specimens, which had been removed endoscopically or obtained at surgery during 1993 and 1994, all immunohistochemical analyses were performed using an avidin-biotin complex immunoperoxidase technique. Increased nuclear expression of the cyclin D1 protein was noted in 32 of 69 samples (46%; 44% of the samples from males and 50% of the samples from females). Positive nuclear staining for the cyclin D1 protein in Barrett's disease with intestinal metaplasia was found in 38% of the male cases and 25% of the female cases, whereas in gastric metaplasia it was positive in 33% of men and 48% of women. Nuclear accumulation of the cyclin D1 protein was also found in both dysplastic and nondysplastic lesions, and it was not associated with sex, age, or cigarette or alcohol consumption. Samples from patients taking proton pump inhibitors tended to be less frequently positive (32%) for cyclin D1 nuclear staining when compared to patients taking H2 antagonists (45%) or antacids (55%). These studies suggest that increased expression of cyclin D1 is an early event in the tumorigenic process of esophageal adenocarcinomas and that the increased expression of this gene might predispose the epithelium to malignant transformation.