Two distinct populations of tumor necrosis factor-stimulated gene-6 protein in the extracellular matrix of expanded mouse cumulus cell-oocyte complexes

After the luteinizing hormone surge, the cumulus cell-oocyte complexes (COC s) in the preovulatory follicles produce a viscoelastic extracellular matri x, a process that requires the synthesis of hyaluronan as well as the incor poration of some components of the inter-a-trypsin inhibitor (I alphaI) fam ily. In this study we report, that a hyaluronan-binding protein, the transl ated product of tumor necrosis factor-stimulated gene-6 (TSG-6), is also sp ecifically accumulated in this matrix. TSG-6 mRNA expression is quickly upr egulated and peaks at similar to 1500 copies/cell 4 h after the ovulatory s timuli as assessed by quantitative reverse transcription-polymerase chain r eaction. Immunohistochemistry reveals the colocalization of the TSG-6 prote in and hyaluronan around the cumulus and granulosa cells. The TSG-6 protein exists in two distinct populations in the COC matrix as demonstrated by We stern-blot analysis. One population is a monomer that is anchored to the ma trix by a noncovalent interaction. The second population is a covalent comp lex with either of the heavy chains of I alphaI and is bound to hyaluronan through a strong interaction that is resistant to denaturing conditions. Th e specific incorporation of the TSG-6 protein into the COC matrix suggests a structural role for this molecule. (C) 2001 Academic Press.