The metabolite 1,25-dihydroxyvitamin D (1,25(OH)2D) is synthesized from its precursor 25-hydroxyvitamin D (25(OH)D) by human
osteoblasts leading to stimulation of osteoblast differentiation in an autocrine or paracrine way. Osteoblast differentiation
is also stimulated by mechanical loading through activation of various responses in bone cells such as nitric oxide signaling.
Whether mechanical loading affects osteoblast differentiation through an enhanced synthesis of 1,25(OH)2D by human osteoblasts
is still unknown. We hypothesized that mechanical loading stimulates the synthesis of 1,25(OH)2D from 25(OH)D in primary human
osteoblasts. Since the responsiveness of bone to mechanical stimuli can be altered by various endocrine factors, we also investigated
whether 1,25(OH)2D or 25(OH)D affect the response of primary human osteoblasts to mechanical loading.
Primary human osteoblasts
were pre-incubated in medium with/without 25(OH)D3 (400 nM) or 1,25(OH)2D3 (100 nM) for 24 h and subjected to mechanical loading
by pulsatile fluid flow (PFF). The response of osteoblasts to PFF was quantified by measuring nitric oxide, and by PCR analysis.
The effect of PFF on the synthesis of 1,25(OH)2D3 was determined by subjecting osteoblasts to PFF followed by 24 h post-incubation
in medium with/without 25(OH)D3 (400 nM).
We showed that 1,25(OH)2D3 reduced the PFF-induced NO response in primary human
osteoblasts. 25(OH)D3 did not significantly alter the NO response of primary human osteoblasts to PFF, but 25(OH)D3 increased
osteocalcin and RANKL mRNA levels, similar to 1,25(OH)2D3. PFF did not increase 1,25(OH)2D3 amounts in our model, even though
PFF did increase CYP27B1 mRNA levels and reduced VDR mRNA levels. CYP24 mRNA levels were not affected by PFF, but were strongly
increased by both 25(OH)D3 and 1,25(OH)2D3.
In conclusion, 1,25(OH)2D3 may affect the response of primary human osteoblasts
to mechanical stimuli, at least with respect to NO production. Mechanical stimuli may affect local vitamin D metabolism in
primary human osteoblasts. Our results suggest that 1,25(OH)2D3 and mechanical loading, both stimuli of the differentiation
of osteoblasts, interact at the cellular level.