Extracellular matrix (ECM), secreted by and surrounding cells in most tissues, gives structure and support to the tissues and exerts an influence upon the cells. Collagens and several glycoproteins (fibronectin, laminin, and entactin) have been isolated and shown to be ECM constituents. Studies now suggest that a family of unique glycoproteins containing collagen-like domains are present in ECM. They are extracted from tissues with strong denaturants in the presence of reducing agents. The exact localization and function of these collagen-like glycoproteins is unknown. This study will focus on a collagen-like 130 kilodalton (kd) glycoprotein isolated from skeletal muscle. The role of this glycoprotein in muscle ECM will be examined using monoclonal antibodies (mAbs) and electron immunohistochemistry to localize the protein. Evidence of binding of the 130 kd glycoprotein to other matrix components will be sought. Copurification and coimmunoprecipitation will be considered an indication of an interaction; microELISA and Western blot techniques will be used to confirm any molecular interactions. Tissue culture will be used to determine whether the 130 kd glycoprotein has any effect upon attachment, spreading, and differentiation of muscle cells. mAbs will be used to stain Western blots to find cross-reactive antigens in other tissues. Fetal and nonmuscle tissue homogenates will be used to see if there are alterations in mw of the antigens during development and if there are tissue specific forms of collagen-like glycoproteins. mAbs will be used to determine whether apparently crossreactive low mw serum proteins detected in preliminary studies with polyclonal sera are related to the matrix molecules. If the serum proteins are related, studies will be undertaken to quantitate them and to see if they are changed in diseases affecting ECM. ECM is altered in many diseases. Connective tissue changes are obvious in arthritis. In diabetes, muscular dystrophy and inflammatory myopathies there is thickening of the basement membrane and/or epimysium surrounding individual muscle fibers. The cause of the thickening and which components are increased are unknown. We will use mAbs against the 130 kd glycoprotein to find out whether it contributes to changes in muscle ECM in the above mentioned diseases.
