PTH Stimulated Growth and Decreased Col‐X Deposition are Phosphotidylinositol‐3,4,5 Triphosphate Kinase and Mitogen Activating Protein Kinase Dependent in Avian Sterna

Abstract

Type X collagen (Col-X) deposition is a marker of terminal differentiation during chondrogenesis, in addition to appositional growth and apoptosis. The parathyroid hormone/parathyroid hormone related peptide (PTH/PTHrP) receptor, or PPR, is a G-Protein coupled receptor (GPCR), which activates several downstream pathways, moderating chondrocyte differentiation, including suppression of Col-X deposition. An Avian sterna model was used to analyze the PPR GPCR downstream kinase role in growth rate and extracellular matrix (ECM) including Col-II, IX, and X. Phosphatidylinositol kinase (PI3K), mitogen activating protein kinase (MAPK) and protein kinase A (PKA) were inhibited with specific established inhibitors LY294002, PD98059, and H89, respectively to test the hypothesis that they could reverse/inhibit the PTH/PTHrP pathway. Excised E14 chick sterna were PTH treated with or without an inhibitor and compared to controls. Sternal length was measured every 24 hr. Cultured sterna were immuno-stained using specific antibodies for Col-II, IX, or X and examined via confocal microscopy. Increased growth in PTHtreated sterna was MAPK, PI3K, and PKA dose dependent, suggesting growth was regulated through multiple pathways. Col-X deposition was rescued in PTH-treated sterna in the presence of PI3K or MAPK inhibitors, but not with the PKA inhibitor. All three inhibitors moderately disrupted Col-II and Col-IX deposition. These results suggest that PTH can activate multiple pathways during chondrocyte differentiation. Anat Rec,

Mechanism of parathyroid hormone-mediated suppression of calcification markers in human intervertebral disc cells

“…This strongly implicated that at least in NP cells, MAPK signalling is instrumental in the PTH-mediated inverse regulation of COL II and COL X expression. This is in agreement with a recent study (Harrington et al, 2010), showing a rescue of COL X expression by MAPK inhibitors in a model of avian sterna.…”

“…This is in agreement with earlier reports showing that PKA is not involved in PTH mediated reduction in COL X expression in avian sterna (Harrington et al, 2010). The lack of cumulative effect of forskolin and PTH suggested that these two agents share the same pathway to exert their effects.…”

Multiple Roles of the SO42−/Cl−/OH− Exchanger Protein Slc26a2 in Chondrocyte Functions

“…Considering the central role of Slc26a2 in chondrocyte biology, our findings indicate a major role for PI3K in regulation of chondrocytes (29). Interestingly, inhibition of basal PI3K activity inhibits basal growth and the effects of parathyroid hormone, C-type natriuretic peptide, and IGF-1 in explant cultures (32)(33)(34). In addition, expression of active Akt in murine cartilage promotes chondrocyte differentiation, whereas dominantnegative Akt delays chondrocyte differentiation (35).…”

Suppression of Nkx3.2 by phosphatidylinositol-3-kinase signaling regulates cartilage development by modulating chondrocyte hypertrophy

“…In chondrocyte regulation, PI3K and Rac1 signaling pathways have been implicated in chondrogenic differentiation and hypertrophic maturation [40], as well as in proteoglycan biosynthesis [40,41]. Most relevant to this study, LY294002, a pharmacological inhibitor of PI3K, has been shown to suppress cartilage hypertrophy in ex vivo long bone cultures [42].…”