The effect of PTH on osteoblast and osteocyte marker gene and protein expression in IDG-SW3 cells and primary osteocytes.

<p><b>(A)</b> Real-time PCR analysis of IDG-SW3 cells cultured over a 29 day time course and treated with 50nM PTH, or PBS control, for 24 hours at days 0, 7, 14, 21 and 28. Expression was normalized to <i>Actb</i> and is relative to day 1 control samples. <i>Kera</i> and <i>E11</i> were upregulated by PTH treatment, whereas <i>Dmp1</i>, <i>Phex</i>, <i>Mepe</i> and <i>Sost</i> expression was decreased, particularly at the later time points (n = 3±SD, *p<0.05). <b>(B)</b> Real-time PCR analysis of osteocyte-enriched bone fragments cultured in the presence of 50nM PTH, or PBS, for 24 hours. Expression was normalized to <i>Actb</i> and is relative to control samples. <i>Kera</i> and <i>E11</i> were upregulated by PTH treatment. <i>Dmp1</i>, <i>Phex</i>, <i>Mepe</i> and <i>Sost</i> expression was decreased in the primary osteocytes (n = 4±SD, *p<0.05). <b>(C)</b> IDG-SW3 cells were cultured over a 30 day time course and treated with 50nM PTH, or PBS, for 48 hours at days 0, 7, 14, 21 and 28. E11 and sclerostin expression was assessed by western blotting. <b>(D)</b> Quantitation of E11 protein expression with and without PTH treatment, relative to day 2 control samples. <b>(E)</b> GFP expression in mature (day 28) IDG-SW3 cells treated with 50nM PTH for 48 hours. Scale bar = 20μm. <b>(F)</b> Quantification of GFP in IDG-SW3 cells cultured over a 30 day time course and treated with 50nM PTH for 48 hours at days 0, 7, 14, 21 and 28 (n = 4±SD, *p<0.05).