Regulation of murine osteoblast macrophage colony-stimulating factor production by 1,25(OH)2D3.
Macrophage colony stimulating factor (MCSF) is important for formation of osteoclasts. We investigated the ability of 1,25(OH)2D3 to regulate osteoblast production of MCSF. Mouse calvarial osteoblasts were cultured for 2 days +/- 1,25(OH)2D3. Since 1, 25(OH)2D3 decreased osteoblast proliferation by 17.6 +/- 1% at 10 nM and 11 +/- 4% at 1 nM, the effect of growth rate on MCSF secretion was examined. Limiting cell proliferation by serum did not affect MCSF production. 1,25(OH)2D3 (1 nM) increased MCSF production (U/10(5) cells) maximally by 68 +/- 33% (n = 3) with an ED50 for 1, 25(OH)2D3 of 5 x 10(-11) M. To investigate effects of 1,25(OH)2D3 on MCSF gene regulation, RT-PCR primers were designed to identify the mRNA coding for the membrane-bound isoform of MCSF. Simultaneous RT-PCR of glyceraldehyde-phosphate dehydrogenase (GAP) allowed semiquantitative assessment of MCSF mRNA between treatment groups expressed as the MCSF/GAP RT-PCR product ratio; both MCSF and GAP (+) primers were labeled with 32P-ATP for phosphorimage quantitation. The membrane-bound MCSF/GAP PCR product ratio was not affected by proliferative rate when growth was limited by [serum]. The MCSF/GAP RT-PCR product ratio was dose dependently increased by 1,25(OH)2D3, maximally at 1 nM at 2.2 +/- 0.2 = fold (n = 10). 1,25 (OH)2D3 also increased the expression of an RT-PCR MCSF/GAP product ratio which represented the secreted isoform of MCSF. The ability of 1,25(OH)2D3 to pretranslationally regulate expression of membrane-bound osteoblast MCSF may be important in osteoblast:osteoclast interactions.
Rubin
J
J
Department of Medicine, Veterans Affairs Medical Center, 1670 Clairmont Road, Decatur, Georgia 30033, USA.
Fan
X
X
Thornton
D
D
Bryant
R
R
Biskobing
D
D
D000818:Animals; D002117:Calcitriol / Q000494:pharmacology*; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D005786:Gene Expression Regulation / Q000187:drug effects; D016173:Macrophage Colony-Stimulating Factor / Q000096:biosynthesis* / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D010006:Osteoblasts / Q000378:metabolism*; D012333:RNA, Messenger / Q000096:biosynthesis*
D012333:RNA, Messenger; D016173:Macrophage Colony-Stimulating Factor; D002117:Calcitriol