Effect of estrogen on bone metabolism in tissue culture: enhancement of the steroid effect by zinc

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Abstract
The present investigation was undertaken to clarity the effect of estrogen (17β-estradiol) on bone metabolism in tissue culture. Calvariae were removed from weanling rats (3-week-old females) and cultured for periods up to 96h in Dulbecco's Modified Eagle Medium (high glucose, 4500 mg/dl) supplemented with antibiotics and bovine serum albumin. The experimental cultures contained 10−10 to 10−8 M estrogen. All cultures were incubated at 37°C in 5% CO2/95% air. Bone calcium content was significantly increased by the presence of 10−10 to 10−8 M estrogen. The steroid (10−10 to 10−8 M) also significantly increased alkaline phosphatase activity in the bone, wheres it did not significantly alter acid phosphatase activity. No appreciable effect on bone alkaline phosphatase activity was produced with 17α-estradiol (10−9 and 10−8 M). Tamoxifen (10−6 M), an anti-estrogen, completely blocked the effect of estrogen (10−9 M) of increasing bone alkaline phosphatase activity. Furthermore, bone DNA content was significantly increased by 10−10 to 10−8 M estrogen. Meanwhile, the presence of 10−4 M zinc, which can stimulate bone protein synthesis, significantly enhanced the effect of 10−9 M estrogen to increase DNA content in rat calvaria, while the metal did not enhance the steroid effect on bone calcium content and alkaline phosphatase activity. The presence of 10−7 M cycloheximide completely prevented the stimulatory effect of estrogen (10−9 M) on calcium content, alkaline phosphatase activity, and DNA content in rat calvaria. The present study demonstrates that estrogen has a direct stimulatory effect on bone metabolism in tissue culture and that zinc can enhance the steroid effect on bone DNA.