Middle ear cholesteatoma is characterized by the presence of a keratinizing squamous epithelium with hyperproliferative features. Such growth can only be supported by abundant blood vessels. The presence and distribution of blood vessels in cholesteatoma was studied to determine the mechanisms responsible for its origin and maintenance. Cholesteatoma (n = 30) and retroauricular skin samples (n = 30) were studied with indirect immunoperoxidase and alkaline phosphatase anti-alkaline phosphatase methods. Antibodies were used to recognize endothelial cells (von Willebrand Factor VII), vascular basal membrane components (type VI collagen), intercellular adhesion molecules (ICAM-1, 1CAM-2), human histocompatibility antigen (HLA-II) as a marker for cellular activation, angiogenetic growth factors and their receptors (TGF-alpha and VEGF), lymphocytes (CD3), and macrophages (KiM8). The cholesteatoma stroma had numerous vessels with intact basal membrane. The vessel concentration was higher in regions with abundant macrophage infiltration. Perivascular cell infiltrates were positive for antibodies against angiogenetic factors and HLA-II. Endothelial cells had increased expression of intercellular adhesion molecules and angiogenetic growth factor receptors. These results confirm the presence of increased vascularization in cholesteatoma, which may play an important role in sustaining continuous abnormal growth.