<p>Femur-derived BMSCs were harvested from 3 male <i>Per1</i>::<i>luc</i> rats (5 months old). BMSCs from passages 3 to 5 were used for the experiments. <b>A</b>. Time-lapse microscopy depicted the periodic activation of <i>Per1</i>::<i>luc</i> reporter gene expression in the BMSCs. <b>B</b>. <i>Per1</i>::<i>luc</i> reporter gene expression was measured by luminometry. The experiment was performed with three dishes in each group. The baseline-subtracted luminometry data demonstrated the circadian activation of <i>Per1</i>::<i>luc</i> expression when BMSCs were cultured on polypropylene dishes with 1 nM vitamin D supplementation; this was reduced on the Ti discs. Strikingly, the circadian rhythm of <i>Per1</i>::<i>luc</i> expression was nearly completely lost on the B-DAE-DCD discs. <b>C</b>. The loss of <i>Per1</i>::<i>luc</i> circadian expression on the B-DAE-DCD discs was depicted by the decreased peak-to-trough amplitude and peak-to-peak period. **: p<0.01 by Wilks Lambda for the amplitude and period. <b>D</b>. The raw amplitude measurements depicted the stable expression of the <i>Per1</i>::<i>luc</i> reporter gene after the initial forskolin shock when the BMSCs were cultured on the polypropylene dishes or machined-polished Ti discs. In contrast, <i>Per1</i>::<i>luc</i> reporter gene expression remained low when the BMSCs were cultured on the B-DAE-DCD discs. <b>E</b>. The average raw amplitude of the <i>Per1</i>::<i>luc</i> reporter gene was significantly lower in the B-DAE-DCD group than the polypropylene and machine-polished groups; however, it was above the background level of the no cell negative control. **: p<0.01 by ANOVA. <b>F</b>. When BMSCs were cultured on B-DAE-DCD Ti discs without vitamin D supplementation, the lost <i>Per1</i>::<i>luc</i> circadian expression was partially recovered. <b>G</b>. The raw amplitudes from the luminometry data indicated that vitamin D supplementation did not affect Per1 expression levels in BMSCs cultured on the machined-polished Ti discs or B-DAE-DCD discs.</p
