Matrix metalloproteinase-dependent regulation of extracellular matrix shapes the structure of sexually differentiating mouse gonads

The extracellular matrix (ECM) proteins play an important role in the establishment of the sex-dependent structure of developing gonads. The matrix metalloproteinases (MMPs) are the major players in the regulation of ECM. Our hypothesis was that the MMPs-dependent regulation of EMC is crucial for the establishment of the correct, either testis or ovary, structure of developing gonad. We cultured developing mouse gonads in vitro in the presence of the MMPs inhibitors (alpha-2-macroglobulin, leupeptin, phosphoramidon) or the MMPs activator, APMA (4-aminophenylmercuric acetate). These inhibitors and activator inhibit/activate, to a different degree, matrix metalloproteinases, but the exact mechanism of inhibition/activation remains unknown. We found that the MMP inhibitors increased accumulation of ECM in the developing gonads. The alpha-2-macroglobulin had the weakest, and the phosphoramidon the strongest effect on the ECM and the structure of the gonads. The alpha-2-macroglobulin caused a slight increase of ECM and did not disrupt the gonad structure. Leupeptin led to the strong accumulation of ECM, resulted in the formation of the structures resembling testis cords in both testes and ovaries, and caused increase of apoptosis and complete loss of germ cells. Phosphoramidon caused the strongest accumulation of ECM, which separated individual cells and completely prevented intercellular adhesion both in the testes and in the ovaries. As a result of aberrant morphology, the sex of the phosphoramidon-treated gonads was morphologically unrecognizable. The APMA - the activator of MMP caused ECM loss, which led to the loss of cell adhesion, cell dispersion and an aberrant morphology of the gonads. These results indicate that the ECM accumulation is MMPs-dependent and that the correct amount and distribution of ECM during gonad development plays a key role in the formation of the gonad structure.