Zinc Enhancement of 17β-Estradiol's Anabolic Effect in Osteoblastic MC3T3-E1 Cells

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Abstract
The anabolic effect of 17β-estradiol in osteoblastic MC3T3-E1 cells was investigated. The cells were cultured for 3 days in the medium containing either vehicle or 17β-estradiol (10−11–10−9 M). 17β-Estradiol significantly increased alkaline phosphatase activity and protein concentration in the cells. The steroid (10−9 M) also significantly elevated the cell numbers and the cellular DNA content. The anabolic effect by 17β-estradiol was blocked by the presence of dipicolinate (10−3 M), a chelator of zinc ion, suggesting a role of cellular zinc in osteoblastic cell function. The presence of zinc sulfate (10−5 M) or β-alanyl-L-histidinato zinc (AHZ) (10−5 M) significantly enhanced the 17β-estradiol (10−10 or 10−9 M)-induced increase of alkaline phosphatase activity and protein concentration in the cells; the effect of AHZ was greater than that of zinc sulfate. The enhancement by zinc compounds was not based on the augmentation of osteoblastic cell numbers. The co-addition of cycloheximide (10−6 M), an inhibitor of protein synthesis, completely blocked the zinc compound (10−5 M)-induced enhancement of 17β-estradiol's (10−9 M) effect to increase alkaline phosphatase activity and protein concentration in the cells. Moreover, the anabolic effect of 17β-estradiol together with or without zinc compounds was abolished by the presence of staurosporine (10−8 M), an inhibitor of protein kinase C, or of okadaic acid (10−7 M), an inhibitor of protein phosphatase. The present study demonstrates that the anabolic effect of 17β-estradiol is enhanced by zinc-chelating dipeptide in osteoblastic MC3T3-E1 cells, and that the enhancing effect may involve protein synthesis and protein kinase activity.

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