A hallmark of mucoid otitis media (MOM, i.e., chronic otitis media with mucoid effusion) is mucus accumulation in the middle ear cavity, a condition that impairs transduction of sounds in the ear and causes hearing loss. The mucin identities of mucus and the underlying mechanism for the production of mucins in MOM are poorly understood. In this study, we demonstrated that the MUC5B and MUC4 were major mucins in MOM that formed distinct treelike polymers (mucus strands). The MUC5B and MUC4 mRNAs in the middle ear mucosa with MOM were up regulated 5-fold and 6-fold, compared with the controls. This upregulation was accompanied by the extensive proliferation of the MUC5B- and MUC4-producing cells in the middle ear epithelium. Further study indicated that the mucin hyperproduction was significantly linked to CD4+ and CD8+ T cells and/or CD68+ monocyte macrophages. It suggests that MUC5B and MUC4 expression may be regulated by the products of these cells.