Estrogen plays an important role in skeletal physiology by maintaining a remodeling balance between the activity of osteoblasts and osteoclasts. In an attempt to decipher the mechanism through which estrogen elicits its action on osteoblasts, experimentation necessitated the development of a culturing environment reduced in estrogenic compounds. The selected medium (OPTI-MEM) is enriched to sustain cultures under reduced fetal bovine serum (FBS) conditions and is devoid of the pH indicator phenol red, a suspected estrogenic agent. This protocol reduced the concentration of FBS supplementation to 0% through successive 24 h incubations with diminishing amounts of total FBS (1%, 0.1%, and 0%). The protocol does not appear to alter the viability, cell morphology, or osteoblast-like phenotype of 7F2 and UMR-106 cell lines when compared with control cells grown in various concentrations of FBS. Although the rate of mitotic divisions declined, the 7F2 and UMR-106 cultures continued to express osteoblast-specific markers and exhibited estrogen responsiveness. These experimental findings demonstrate that the culture protocol developed did not alter the osteoblast nature of the cell lines and provides a model system to study estrogen's antiresorptive role on skeletal turnover.

Doxorubicin is the most commonly prescribed chemotherapeutic agent. Doxorubicin causes an increase in free radical production in the myocardium and cardiac muscle dysfunction. Patients receiving chemotherapy also experience weakness and fatigue leading us to hypothesize doxorubicin (1) increases muscle‐derived oxidants and (2) depresses diaphragm and limb muscle force in mice. Doxorubicin treatment (20μM, 1hr) in vitro increased oxidant activity in diaphragm muscle as measured using DCF fluorescence (n=3, p<0.04). We injected C57B6 male mice intraperitoneally with doxorubicin (20 mg/kg), a cumulative dose commonly used in cardiotoxicity studies. Diaphragm and extensor digitorum longus (EDL) were excised three days post injection, compared to controls, doxorubicin lowered maximal force in diaphragm by 62.2 ± 7.1% (mean ± SE; n=5, p<0.01) and EDL by 28.7 ± 7.9% (n=3, p<0.03). Doxorubicin depressed diaphragm twitch characteristics, including peak force (−37.7 ± 12.5%; n=5, p<0.01) and time to peak force (Doxorubicin 0.15 ± 0.02 N s−1, Control 0.29 ± 0.02 N s−1). At the end of the fatigue trial, force was lower in doxorubicin treated mice 26.9 ± 12.2% of control values. The stability of the muscle in vitro was not altered. Our data shows that doxorubicin causes skeletal muscle weakness and predisposes muscle to fatigue, which may be due to an increase in muscle‐derived oxidants.Supported by: HL 59878 (MBR)

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