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The bone remodeling process is influenced by various factors, including estrogens and transmitters of the endocannabinoid system. In osteoblasts, cannabinoid receptors 2 (CB-2) are expressed at a much higher level compared to CB-1 receptors. Previous studies have shown that estrogens could influence CB-2 receptor expression. In the present study, the possible interactions of a specific CB-2 agonist and a specific CB-2 antagonist/inverse agonist with 17-β-estradiol were investigated in primary human osteoblasts (HOB). HOB cells were cultured in phenol red-free osteoblast growth medium (37°C, 5% CO 2 ). In their 5th passage, HOB were exposed to different concentrations of i) 17-β-estradiol (1, 10 and 100 nM); ii) a specific CB-2 agonist (R,S)-AM1241 (1 and 7.5 µM); and iii) a specific CB-2 antagonist/inverse agonist AM630 (10 µM) and to selected combinations of the substances. After 24 and 48 h of incubation, HOB proliferation activity was measured using a WST-8 assay. Alkaline phosphatase activity was also evaluated using spectrophotometry. Concomitant exposure of HOB to 17-β-estradiol (10 nM) and to specific CB-2 antagonist/inverse agonist (10 µM) showed similar HOB proliferation activity to HOB incubated with 17-β-estradiol only at a 100 nM concentration. By contrast, concomitant incubation of HOB with 17-β-estradiol (10 nM) and specific CB-2 agonist (7.5 µM) resulted in decreased HOB proliferation activity as compared to HOB incubated with 17-β-estradiol only (10 nM). Similar findings were observed after 24 and 48 h of incubation. In all the experiments, HOB successfully passed the alkaline phosphatase differentiation test. In conclusion, for the first time a synergistic interaction between 17-β-estradiol and specific CB-2 antagonist/inverse agonist was observed in HOB. Understanding the molecular pathways of this interaction would be of great importance in developing more efficient and safer drugs for treating or preventing bone diseases.

biomedical reports

A synergistic interaction of 17-β-estradiol with specific cannabinoid receptor type 2 antagonist/inverse agonist on proliferation activity in primary human osteoblasts