Many otologists were interested in the cholesterol level in the cholesteatoma matrixs and discussed the origin of this cholesterol. At present, the origin of the cholesterol is uncertain. In this study, we cultured the skin of the external auditory canal and ear drum, in-vitro and measured the synthesis of cholesterol in these cultures by using radioactive material. The results showed that cholesterol, especially esterified cholesterol is actively synthesized in these skins. This study appears to indicate that the cholesterol of the cholesteatoma matrix is actively produced in the cholesteatoma matrix.