Middle ear cholesteatoma consists of a layer of keratinized stratified squamous epithelium overlying subepithelial granuloma tissue with inflammatory cell infiltration, and is feared for its destructive properties. The bone resorption mechanism in cholesteatoma is thought to be a cellular and biochemical reaction associated with the chronic inflammatory component of cholesteatoma. The purposes of this study are, on one hand, to analyze immunohistochemically the interaction between subepithelial granuloma tissue and eroded ossicles in cholesteatoma using a new decalcifying technique, and on the other hand, to analyze biochemically the bone resorbing activity produced by cholesteatoma tissue in culture. It was found that numerous number of T-lymphocytes, which predominantly belonged to helper/inducer T-cell subset, and HLA-DR positive cells were accumulated beneath the epithelial layer and near the eroded ossicles. The bone resorbing activity produced by cholesteatoma tissue was suggested to be attributable to interleukin-1α and prostaglandin E2, whereas the granulation tissue in cases of simple chronic otitis media without cholesteatoma produced only prostaglandin E2 but not interleukin-1α.