Gene and Protein Expression of Polysulfated Proteoglycan ECM Components in the Equine Digital Lamellae

Article Outline

• Take Home Message
• Introduction
• Materials & Methods
• Results
• Discussion and Clinical Relevance
• Conclusion
• Copyright

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Take Home Message 

Genetic and biochemical evidence for large polysulfated proteoglycans in lamellae, but not liver and lung tissue, show that lamellae are functionally specialized to resist compressive forces.

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Introduction 

Our published and on-going studies show ADAMTS-4 (aggrecanase) gene and protein expression is elevated in the lamellae of laminitic horses. ADAMTS-4 cleaves aggrecan and versican, the core proteins of large polysulfated proteoglycans that in complex with hyaluronan form hydrated gels and provide resistance to compressive forces. Here we investigate ADAMTS-4 substrates in lamellae.

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Materials & Methods 

cDNA-specific probes and relative quantitative real time PCR were used to detect the expression levels of genes encoding polysulfated proteoglycans. Ct values were analyzed using the ΔΔCt method. Protein and glycosaminoglycan moieties in lamellar extracts were detected after SDS-PAGE electrophoresis by Western blotting.

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Results 

Genes encoding aggrecan, versican, chondroitin synthase and hyaluronan synthase were found to be consitutively expressed in normal digital lamellae but not, or to a much lesser extent in lungs and liver. Aggrecan core protein was detected in detergent (0.5%NP-40) lysates of lamellae, whereas keratan- and chondroitin- sulfate aggrecan proteoglycans were detected in guanidine hydrochloride extracts of lamellar proteoglycans.

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Discussion and Clinical Relevance 

Gene expression and product analyses indicate that the equine digital lamellae are functionally specialized to resist compressive forces. Consequently, degradation of this material by elevated ADAMTS-4 may compromise lamellar function.

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Conclusion 

The presence of large polysulfated proteoglycans and elevated ADAMTS-4 in laminitic tissue suggest parallels between the pathophysiology of laminitis and osteoarthritis.