A synthetic peptide of link protein stimulates the biosynthesis of collagens II,IX and proteoglycan by cells of the intervertebral disc

To date, there have been no reports on the effect on disc cells of the intervertebral disc (IVD) of the amino terminal peptide of link protein (DHLSDNYTLDHDRAIH) (link N) which is generated by the cleavage of human link protein by stromelysins 1 and 2, gelatinase A and B, and collagenase between His(16) and Ile(17). However, link N has been shown to act as a growth factor and stimulate synthesis of proteoglycans and collagen by chondrocytes of human articular cartilage. There are also no studies on the effect of link N on type IX collagen in any tissue. In the studies reported here, a serum-free pellet culture system has been used to examine whether link N can play a role in maintaining the integrity of disc matrix, specifically at the level of matrix assembly by cells of the IVD. Using this culture system, we determined the capacity of link N to stimulate accumulation of these matrix proteins in the annulus fibrosus (AF) and nucleus pulposus (NP). Gross inspection of separate AF and NP pellet cultures in the absence of link N revealed a progressive increase in size and a transition from "spherical" to "polygonal" pellets after centrifugation. Addition of 10 ng/ml link N resulted in increased pellet sizes for both AF and NP pellet cultures. Link N increased proteoglycan, type II and type IX collagen contents with an increase in DNA content over time. This study demonstrates that link N can act directly on disc cells to stimulate matrix production, which involves increased accumulation of proteoglycan, and types II and IX collagens. This study also identifies the value of pellet cultures for studies of the IVD cells in a serum-free chemically defined medium, in which pellets can continue growing in size in response to growth factors with minimal cell loss. Link N may have value in stimulating the growth and regeneration of the damaged IVD.