Voronov I, Heersche J N M, Casper R F, Tenenbaum H C, Manolson M F
Department of Laboratory Medicine and Pathobiology, Mount Sinai Hospital, Toronto, ON, Canada.
Biochem Pharmacol. 2005 Jul 15;70(2):300-7. doi: 10.1016/j.bcp.2005.04.028.
We investigated the effect of representative polycyclic aryl hydrocarbons (PAHs), benzo[a]pyrene (BaP), and 7,12-dimethylbenz[a]anthracene (DMBA) on osteoclast differentiation and function by using dispersed cancellous bone derived rabbit osteoclasts and the RAW264.7 cells. These cells differentiate into osteoclasts when exposed to receptor activator of NF-kappaB ligand (RANKL). The rabbit osteoclasts were exposed to 10(-6) to 10(-9)M BaP or DMBA and the tartrate-resistant acid phosphatase (TRAP)-positive cells were counted. The effect of PAHs on osteoclast differentiation in dispersed rabbit osteoclast-containing stromal cell populations was cell density dependent, suggesting that the cell density of stromal cells, osteoclast precursors, and/or mature osteoclasts are factors regulating the effect of PAHs. To investigate the direct effect of BaP on osteoclast differentiation, RAW264.7 cells were exposed to 10(-5) to 10(-6) M BaP. Treatment of RAW264.7 cells cultured with 25 ng/ml soluble RANKL and 10(-5)M BaP for 5 days decreased osteoclast differentiation, TRAP activity levels, and resorption of bone-like substrata. The inhibition was prevented by 10(-6) to 10(-7) M resveratrol, an aryl hydrocarbon receptor (AhR) antagonist, and by higher concentrations of RANKL. To investigate the ability of RANKL to reverse BaP-mediated inhibition, gene expression was determined by RT-PCR. Cytochrome P450 1B1 (CYP1B1) mRNA, one of the genes activated by BaP, was present only in the groups exposed to BaP; the levels of CYP1B1 mRNA decreased in the presence of increasing concentrations of RANKL. These results suggest that the inhibitory effects of PAHs on osteoclastogenesis are direct and likely involve interaction of the RANKL and PAH signaling pathways.
我们使用源自兔松质骨的破骨细胞和RAW264.7细胞，研究了代表性多环芳烃（PAHs）、苯并[a]芘（BaP）和7,12-二甲基苯并[a]蒽（DMBA）对破骨细胞分化和功能的影响。当暴露于核因子κB受体活化因子配体（RANKL）时，这些细胞会分化为破骨细胞。将兔破骨细胞暴露于10^(-6)至10^(-9)M的BaP或DMBA中，并对抗酒石酸酸性磷酸酶（TRAP）阳性细胞进行计数。PAHs对含破骨细胞的兔分散基质细胞群体中破骨细胞分化的影响取决于细胞密度，这表明基质细胞、破骨细胞前体和/或成熟破骨细胞的细胞密度是调节PAHs作用的因素。为了研究BaP对破骨细胞分化的直接影响，将RAW264.7细胞暴露于10^(-5)至10^(-6)M的BaP中。用25 ng/ml可溶性RANKL和10^(-5)M BaP培养RAW264.7细胞5天，可降低破骨细胞分化、TRAP活性水平以及类骨基质的吸收。10^(-6)至10^(-7)M的白藜芦醇（一种芳烃受体（AhR）拮抗剂）和更高浓度的RANKL可阻止这种抑制作用。为了研究RANKL逆转BaP介导的抑制作用的能力，通过逆转录-聚合酶链反应（RT-PCR）测定基因表达。细胞色素P450 1B1（CYP1B1）mRNA是由BaP激活的基因之一，仅存在于暴露于BaP的组中；在存在浓度不断增加的RANKL时，CYP1B1 mRNA水平降低。这些结果表明，PAHs对破骨细胞生成的抑制作用是直接的，并且可能涉及RANKL和PAH信号通路的相互作用。
