Effect of PH on the Metabolism of Proteoglycans in the Intervertebral Disc: Investigation in an in Vitro Model With IL-1 Alpha Stimulation and Confirmation in the Cadaveric Human Intervertebral Disc

Intervertebral disc (IVD) degeneration is associated with the proteolytic degradation of proteoglycans (PGs), mainly aggrecan, the presence of catabolic cytokines, such as IL-1α and the development of a concentration gradient of acidic pH. In the IVD, PG metabolism and the involvement of the two major proteinases in degrading aggrecan, the matrix metalloproteinase (MMPs) and aggrecanases (ADAMTSs), under acidic conditions and the presence of IL-1α is not well understood.

In this dissertation, the scaffold-free tissue-engineered annulus fibrosus (AF) and nucleus pulposus (NP) tissues produced by the alginate-recovered-chondrocyte (ARC) method were cultured with or without IL-1α at normal pH (7.4) or lower acidic pH (6.5) in order to determine the interactive effects of pH and IL-1α on (1) the proteoglycan (PG) metabolism and (2) aggrecan fragmentation pattern. Furthermore, using cadaveric human IVDs aggrecanases and the aggrecan fragmentation profile were determined to facilitate the in vivo confirmation of the involvement of aggrecanases in disc degeneration.

The results demonstrated that while IL-1α treatment significantly inhibited PG synthesis at pH 7.4 in NP and AF ARC tissues, this deleterious effect of IL-1α was not observed at pH 6.5 in both the IVD tissues. The retention of PGs in the ARC was higher at acidic pH, even in the presence of IL-1α. Analysis of the aggrecan fragmentation profile demonstrated that with IL-1α treatment, higher levels of aggrecanase generated aggrecan fragments were obtained in both the ARC tissues at pH 7.4, but not at pH 6.5. Further, the ADAMTS-4 enzyme levels were higher in advanced stage (grade 4) of disc degeneration than in the early disc degenerative stage (grade 2). However, the amounts of ADAMTS-5 and aggrecan fragments did not differ between the two degeneration groups.

It has been concluded that IL-1α induces PG catabolism and aggrecanasesmediated aggrecanolysis at pH 7.4 but not at pH 6.5 in the disc tissue. Furthermore, IVD degeneration involves aggrecanase mediated aggrecanolysis.

Year	Entry
2005	Glasson SS, Askew R, Sheppard B, Carito B, Blanchet T, Ma H-L, Flannery CR, Peluso D, Kanki K, Yang Z, Majumdar MK, Morris EA. Deletion of active ADAMTS5 prevents cartilage degradation in a murine model of osteoarthritis. Nature. March 31, 2005;434(7033):644-648.
1992	Ohshima H, Urban JPG. The effect of lactate and pH on proteoglycan and protein synthesis rates in the intervertebral disc. Spine. September 1992;17(9):1079-1082.
1988	Urban JPG, McMullin JF. Swelling pressure of the lumbar intervertebral discs influence of age, spinal level, composition, and degeneration. Spine. February 1988;13(2):179-187.
1981	Urban JPG, Maroudas A. Swelling of the intervertebral disc in vitro. Connect Tissue Res. 1981;9(1):1-10.
2003	Masuda K, Sah RL, Hejna MJ, Thonar EJ-MA. A novel two-step method for the formation of tissue-engineered cartilage by mature bovine chondrocytes: the alginate-recovered-chondrocyte (ARC) method. J Orthop Res. January 2003;21(1):139-148.
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Year

Entry

2005

Glasson SS, Askew R, Sheppard B, Carito B, Blanchet T, Ma H-L, Flannery CR, Peluso D, Kanki K, Yang Z, Majumdar MK, Morris EA. Deletion of active ADAMTS5 prevents cartilage degradation in a murine model of osteoarthritis. Nature. March 31, 2005;434(7033):644-648.

1992

Ohshima H, Urban JPG. The effect of lactate and pH on proteoglycan and protein synthesis rates in the intervertebral disc. Spine. September 1992;17(9):1079-1082.

1988

Urban JPG, McMullin JF. Swelling pressure of the lumbar intervertebral discs influence of age, spinal level, composition, and degeneration. Spine. February 1988;13(2):179-187.

1981

Urban JPG, Maroudas A. Swelling of the intervertebral disc in vitro. Connect Tissue Res. 1981;9(1):1-10.

2003

Masuda K, Sah RL, Hejna MJ, Thonar EJ-MA. A novel two-step method for the formation of tissue-engineered cartilage by mature bovine chondrocytes: the alginate-recovered-chondrocyte (ARC) method. J Orthop Res. January 2003;21(1):139-148.