Abstract
We have examined extracts of articular cartilage from rabbits aged 3β100 weeks for evidence of ageβrelated changes in the structure and content of link protein (LP) in this tissue, with the following findings: (a) Two major molecular weight forms of LP were seen on SDSβPAGE (41 and 48 kDa) and the proportion of these changed markedly with age. The 48 kDa species was predominant in young animals (representing about 78% of the total LP at 5 weeks) whereas the 41 kDa species increased in amount with age (representing 35% of the total LP at 100 weeks). A minor form of about 43 kDa, representing less than 20% of the total, was present only during the growth phase. A small amount of fragmented link protein (less than 5% of the total) of about 25β30 kDa was present in samples from mature and aged rabbits only. (b) The quantitation of LP in guanidinium: HCl extracts of cartilage, by radioimmunoassay with monoclonal antibody 8βAβ4, was markedly influenced by the conditions of preparation and pretreatment of samples. Assays of dialyzed guanidine extracts following treatment at 80Β°C for 15 min in 0.025% (w/v) SDS indicated that immature and mature cartilage contains about 50 and 180 ΞΌg of LP/g of tissue, respectively. On the other hand, assays following treatment at 100Β°C for 20 min in 0.1% (w/v) SDS suggested that rabbit cartilage contains about 300 ΞΌg of LP/g of tissue at all ages; finally, assay of CsCl purified proteoglycan samples under these conditions indicated a content of about 500 ΞΌg of LP/g at all ages. (c) Calculations based on the analysis of proteoglycan preparations for aggregating monomer and link protein suggest that a LP:aggregating monomer molar ratio of about 0.9 is maintained in the articular cartilage throughout maturation and aging in the rabbit.