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PAULINE P.L. SO, FLORENCE W.L. TSUI, REINHOLD VIETH, JINDRA H. TUPY, and KENNETH P.H. PRITZKER

ABSTRACT.

Objective. Calcium pyrophosphate dihydrate (CPPD) crystal deposition disease, a common arthritis affecting the elderly, is characterized by the deposition of CPPD crystals in articular joints. The mechanism underlying disease expression is unknown, but factors contributing to the pathogenesis may involve changes in enzymatic activities involving pyrophosphate and phosphate metabolism. Tissue nonspecific alkaline phosphatase (TNAP) is one of the major enzymes regulating pyrophosphate concentrations in articular joints. We hypothesized that inhibition of TNAP activity at pH = 7.4 by endogenous molecules can lead to CPPD disease pathogenesis.

Methods. We investigated the inhibitory effects of the amino acid cysteine on TNAP's phosphatase, inorganic pyrophosphatase, and CPPD crystal dissolution activities. Kinetic parameters V_max, K_M, concentration for 50% inhibition (I₅₀), inhibitor constant (K_I), and specific activities calculated from Initial Velocity, Eadie-Hofstee, Simple, Dixon, and Secondary plots were used to assess enzyme inhibition.

Results. Cysteine inhibited TNAP's phosphatase activity uncompetitively and its inorganic pyrophosphatase activity mix-competitively. CPPD crystal dissolution activity was also inhibited. I₅₀ values demonstrated that high cysteine concentration is required to inhibit 50% of enzyme activity. K_I values suggested that inorganic pyrophosphatase activity is inhibited more than the phosphatase activity. Ca⁺⁺ and Mg⁺⁺ ion concentrations may regulate this inhibition.

Conclusion. The control of endogenous inhibitors, such as cysteine, that interfere with TNAP's ability to regulate CPPD crystal formation and dissolution in joints could be a potential therapeutic option for CPPD crystal deposition disease. (First Release May 15 2007; J Rheumatol 2007;34:1313-22)

Key Indexing Terms:

ALKALINE PHOSPHATASE
CYSTEINE
AMINO ACIDS
CALCIUM PYROPHOSPHATE
ARTHRITIS
CARTILAGE

From the Department of Pathology and Laboratory Medicine, Mount Sinai Hospital; Department of Laboratory Medicine and Pathobiology, and Department of Immunology, University of Toronto; and Department of Cell and Molecular Biology, Toronto Western Research Institute, Toronto, Ontario, Canada.

P.P.L. So, MSc; R. Vieth, PhD, Professor; J.H. Tupy, MSc; K.P.H. Pritzker, MD, FRCPC, Professor, Department of Pathology and Laboratory Medicine, Mount Sinai Hospital, Department of Laboratory Medicine and Pathobiology, University of Toronto; F.W.L. Tsui, PhD, Associate Professor, Department of Cell and Molecular Biology, Toronto Western Research Institute.

Address reprint requests to Dr. K.P.H. Pritzker, Pathology and Laboratory Medicine, Mount Sinai Hospital, 600 University Avenue, Room 6-500-1, Toronto, Ontario M5G 1X5, Canada. E-mail: kpritzker@mtsinai.on.ca

Accepted for publication February 9, 2007.