1,25-Dihydroxyvitamin D3Interaction with Dexamethasone and Retinoic Acid: Effects on Procollagen Messenger Ribonucleic Acid Levels in Rat Osteoblast-Like Cells

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Abstract
We previously have reported that 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3], dexamthasone, and retnoic acid inhibit collagen synthesis in rat osteoblast-like cell primary cultures. We also have found that dexamethasone increases 1,250(OH)2D3 receptor levels in these cells. Furthermore, this increase in 1,25-(OH)2D3 receptor level is paralleled by an enhanced inhibition of collagen synthesis when dexamethasone and 1,25-(OH)2D3 are used in combination. In contrast, retinoic acid at high doses decreases 1,25-(OH)2D3 receptor level in rat osteoblast-like cells and atenuates 1,25-(OH)2D3 inhibition of collagen synthesis. In the present study, we have used a [32P]cDNA probe for rat pro.alpha.1 (I) to determine if these osteotropic agents act by modulating steady state procollagen mRNA levels. Hybridization with a [32P]cDNA probe for human actin was used as a control. We find that the steady state levels of procollagen mRNA are decreased in all cases, while there are negligible changes in actin mRNA levels. Dexamethasone, at the low dose of 13 nM, acts synergistically with 1,25-(OH)2D3 in decreasing procollagen mRNA levels. The effects of retinoic acid and 1,25-(OH)2D3 are additive at low doses (13 and 130 nM); however, at a high dose of retinoic acid (1.3 .mu.M), combined treatment with 1,25-(OH)2D3 does not reduce procollagen mRNA levels beyond the decrease due to retinoic acid alone. The reduction in procollagen mRNA level after each of these treatments falls in the same range as inhibition of collagen synthesis measured at the protein level. These data suggest that the synthesis of collagen under these treatments is controlled primarly through modulation of steady state procollagen mRNA levels. The combined effects of 1,25-(OH)2D3 with dexamthasone and with retinoic aacid appear to be related to modulation of 1,25-(OH)2D3 receptor level in rat osteoblast-like cells. However, the mechanisms of their interaction in the regulation of collagen gene expression are not clear and probably involve multiple pathways aside form the changes at the receptor level.