Regardless of its origin, cholesteatoma is characterized by the presence of a keratinizing epithelium with an hyperproliferative behavior leading to a very important bone resorption. Previous studies have demonstrated overexpression of interleukin-1 (IL-1 protein in middle ear cholesteatoma by immunohistochemistry and enzyme-linked immunosorbent assay, suggesting a significant role for IL-1-alpha. In this study, the presence of IL-1-alpha messenger ribonucleic acid (mRNA) was quantified by in situ hybridization on frozen sections (n = 10) and by computer-assisted image analysis. Human skin obtained from the external ear canal (n = 10) was used as the control. A higher percentage of cells hybridized for the antisense probes IL-1-alpha mRNA was found in cholesteatoma epithelium. Furthermore, keratinocytes of the suprabasal cell layers were also found to contain specific hybridizations. Some cells in cholesteatoma stroma also contained IL-1-alpha mRNA transcripts. The results of this study confirm the central role of IL-1-alpha in the epithelium hyperproliferation and bone resorption observed in middle ear cholesteatoma.