PTH Stimulated Type X Collagen Deposition is MAP Kinase Dependent

Objectives: Type X collagen (Col-X) deposition is a marker of terminal differentiation during chondrogenesis. Col-X is localized at sites of calcification, indicating that it is likely involved in early stage endochondral bone formation. Production of type X collagen relies upon signaling events in proliferative and prehypertophic stage chondrocytes, including the Parathyroid Hormone/ Parathyroid Hormone related Peptide (PTH/PTHrP) receptor, or PPR. The objective of this project was to use an avian sterna model to examine the effects of PTH signaling pathways on Col-X deposition. Methods: Whole day-14 chicken sterna were dissected free of all tissue and perichondrial membranes under sterile conditions and cultured for 8 days in Ham's F-12 medium including 1% Pen-Strep, 1% non-essential amino acids, 10-¹¹M dexamethasone, 60 ng/ml insulin, 10-¹¹M triiodothyronine and 100 µg/ml ascorbic acid. Treated sterna were cultured in the presence 100nM PTH (1-34) bovine fragment with or without a specific kinase inhibitor for mitogen activating protein kinase (MAPK) (100μM PD98059) or protein kinase A (PKA, 1μM H89) and compared to controls (n=10/group). Sternal length was measured every 24 hours, and relative Col-X expression levels were determined with real time PCR. Results: Increased growth in PTH-treated sterna was MAPK and PKA dependent. Control sterna had robust deposition of Col-X in the ECM surrounding the chondrocyte lacunae after eight days of culture while sterna treated with PTH had little Col-X deposition. Col-X deposition was rescued in PTH treated sterna with the MAPK inhibitor but not with the PKA inhibitor. Conclusions: The effects of PTH on cartilage growth were MAPK and PKA dependent. However, Col-X expression was dependent upon only the MAPK pathway. This project was supported by NIH/NIDCR F30 DE05743 and the Baylor Oral Health Foundation.