Inhibitory effect of beta-cryptoxanthin on osteoclast-like cell formation in mouse marrow cultures

The carotenoid beta-cryptoxanthin has been shown to have an inhibitory effect on bone-resorping factor-stimulated bone resorption in rat bone tissues in vitro. The effect of beta-cryptoxanthin on osteoclast-like cell formation in mouse marrow culture in vitro was investigated. The bone marrow cells were cultured for 7 days in alpha-minimal essential medium containing a bone-resorbing agent [parathyroid hormone (1-34) (PTH), prostaglandin E(2), 1,25-dihydroxyvitamin D(3), lipopolysaccharide, or tumor necrosis factor-alpha (TNFalpha)] with an effective concentration. Osteoclast-like cell formation was estimated by staining for tartrate-resistant acid phosphatase, a marker enzyme of osteoclasts. The presence of PTH (10(-7)M), prostaglandin E(2) (10(-5)M), 1,25-dihydroxyvitamin D(3) (10(-7)M), lipopolysaccharide (10 microg/mL), or TNFalpha (10 ng/mL) induced a remarkable increase in osteoclast-like multinucleated cells. These increases were significantly inhibited in the presence of beta-cryptoxanthin (10(-8) to 10(-6)M). beta-Cryptoxanthin (10(-7) and 10(-6)M) significantly inhibited dibutyryl cyclic adenosine monophosphate (DcAMP) (10(-5)M) or phorbol 12-myristate 13-acetate (PMA) (10(-5)M), an activator of protein kinase C, induced osteoclast-like cell formation. Also, beta-cryptoxanthin (10(-7) and 10(-6)M) had a significant inhibitory effect on osteoclast-like formation induced by receptor activator of NF-kappaB ligand (RANKL) (10 and 20 ng/mL) in the presence of macrophage colony-stimulating factor (M-CSF) (10 and 20 ng/mL). The stimulatory effect of RANKL and M-CSF on osteoclast-like cell formation was significantly enhanced in the presence of PMA, while such an effect was not seen by DcAMP. beta-Cryptoxanthin (10(-6)M) significantly inhibited osteoclast-like cell formation induced by RANKL and M-CSF in the presence of PMA or DcAMP. Moreover, the inhibitory effect of beta-cryptoxanthin on RANKL plus M-CSF-, PTH-, or TNFalpha-induced osteoclast-like cell formation was not observed in the presence of cycloheximide (10(-7)M), an inhibitor of protein synthesis at translational process, or 5,6-dichloro-1-beta-d-ribofuranosylbenzimidazole (10(-6)M), an inhibitor of transcription. This study demonstrates that beta-cryptoxanthin has a potent inhibitory effect on osteoclast-like cell formation in mouse marrow culture. The inhibitory action of beta-cryptoxanthin may partly involve in a newly synthesized protein component which is related to RANKL stimulation in osteoclastogenesis.