Characterization of Epithelial Apoptosis in Biopsies of Small-Bowel Allografts Using Cleaved Caspase 3 Immunostaining

Background. The diagnosis of acute cellular rejection (ACR) in small-bowel allograft biopsies depends in part on quantification of crypt apoptotic bodies (ABs). The definition of ABs varies between authors. Recently, immunoperoxidase stains specific for apoptotic material have been used in paraffin-embedded tissue. The aim was to characterize AB morphology and quantify ABs using immunoperoxidase and hematoxylin and eosin (H&E) staining. Methods. Allograft biopsies with diagnoses of negative for ACR, indeterminate for ACR, and mild ACR were selected. Sections were stained for cleaved caspase 3 (CC3). The number of ABs per 10 crypt epithelial cells was compared by H&E and CC3 staining. Results. A total of 39 cases (15 negative, 12 indeterminate, 12 mild ACR) were obtained. CC3 staining revealed that ABs varied from well-developed “classical” exploding crypt cells to intraepithelial clusters of basophilic material. In 61%, the number detected by CC3 was higher than on H&E stains (mean 4.30 and 3.56; P < .002), with good overall correlation. With the latter definition, AB numbers by H&E were significantly higher in most cases (72%), with the diagnosis of mild ACR being downgraded in 50% if only classical ABs were counted. Conclusions. ABs have a range of morphology in biopsies of intestinal allografts, and H&E identifies the majority. The results suggest that AB counts should include basophilic clusters.