CHARACTERIZATION OF HUMAN TYPE-II PROCOLLAGEN AND COLLAGEN-SPECIFIC ANTIBODIES AND THEIR APPLICATION TO THE STUDY OF HUMAN TYPE-II COLLAGENPROCESSING AND ULTRASTRUCTURE

Type II collagen is the most abundant collagen in articular cartilage and, together with other tissue-specific collagens and proteoglycans, provides the tissue with its shock-absorbing properties and its resili ency to stress. Specific antibodies which recognize various collagen t ypes have been very useful in the study of collagen biosynthesis, stru cture and metabolism in normal and pathological conditions. Antibodies which recognize epitopes of type II collagen have been described prev iously; however, many of these antibodies display cross-reactivity wit h other collagens or with type II collagen from other species, reflect ing the high degree of homology of the helical domains of fibrillar co llagens. In this study, we prepared antibodies to sequential determina nts of human type II procollagen employing synthetic peptides with seq uences deduced from the nucleotide sequence of the human alpha 1(II) p rocollagen cDNA. The antibodies were highly specific for epitopes in e ither the C-terminal propeptide or the telopeptide of the human type I I collagen and did not cross-react with other human interstitial colla gens or with murine type II collagen. These antibodies were used in co njunction with biosynthetic labeling to study the secretion and proces sing of human type II procollagen and collagen in human chondrocytes i n vitro. The results indicated that a lag period of about 90 min was r equired for the secretion of newly synthesized type II procollagen. Co nversion of the secreted procollagen into fully processed alpha-chains and their deposition in the cell layer were first apparent 240 min fo llowing the intitiation of biosynthetic labeling. The antibodies were also used to examine, by immunoelectron microscopy, the structure of t he extracellular matrix produced by human chondrocytes maintained in l ong-term cultures under conditions which permit the preservation of th e cartilage-specific phenotype. These highly specific antibodies provi de valuable tools to study the metabolism and structure of human type II procollagen and collagen in normal and pathologic conditions.