24R,25-Dihydroxyvitamin D3 [24R,25(OH)₂D₃] controls growth plate development by inhibiting apoptosis in the reserve zone and stimulating response to 1α,25(OH)₂D₃ in hypertrophic cells

Previously we showed that costochondral growth plate resting zone (RC) chondrocytes response primarily to 24R,25(OH)₂D₃ whereas prehypertrophic and hypertrophic (GC) cells respond to 1α,25(OH)₂D₃. 24R,25(OH)₂D₃ increases RC cell proliferation and inhibits activity of matrix processing enzymes, suggesting it stabilizes cells in the reserve zone, possibly by inhibiting the matrix degradation characteristic of apoptotic hypertrophic GC cells. To test this, apoptosis was induced in rat RC cells by treatment with exogenous inorganic phosphate (Pi). 24R,25(OH)₂D₃ blocked apoptotic effects in a dose-dependent manner. Similarly, apoptosis was induced in ATDC5 cell cultures and 24R,25(OH)₂D₃ blocked this effect. Further studies indicated that 24R,25(OH)₂D₃ acts via at least two independent pathways. 24R,25(OH)₂D₃ increases LPA receptor-1 (LPA R1) expression and production of lysophosphatidic acid (LPA), and subsequent LPA R1/3-dependent signaling, thereby decreasing p53 abundance. LPA also increases the Bcl-2/Bax ratio. In addition, 24R,25(OH)₂D₃ acts by increasing PKC activity. 24R,25(OH)₂D₃ stimulates 1-hydroxylase activity, resulting in increased levels of 1,25(OH)₂D₃, and it increases levels of phospholipase A2 activating protein, which is required for rapid 1α,25(OH)₂D₃-dependent activation of PKC in GC cells. These results suggest that 24R,25(OH)₂D₃ modulates growth plate development by controlling the rate and extent of RC chondrocyte transition to a GC chondrocyte phenotype.