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Frequency‐Dependent Effect of Nitric Oxide Donor Nitroglycerin on Bone

Recently, we showed that supplementation with nitric oxide (NO) via donor nitroglycerin (NG) alleviated the ovariectomy and corticosteroid‐induced bone loss in rats. In humans, high doses or frequent applications of NG (i.e., for angina) lead to rapid loss of its efficacy in relieving angina. To examine whether there is a similar effect on the loss of efficacy of NG on bone, we examined the frequency‐dependent effects of NG on bone mineral density (BMD), bone mass, trabecular bone volumes (BV/TV), and blood pressure in rats. Thirty 7‐month‐old female Brown Norway rats underwent ovariectomy, and an additional six rats were sham‐operated. The ovariectomized rats were treated either with vehicle (ovariectomized control), 17β‐estradiol (E2; positive control), or 0.2 mg NG (via dermal application) once, twice, or three times a day. Before and at the end of the 10‐week treatment period, BMD of the lumbar spine was measured by dual‐energy X‐ray absorptiometric (DXA) scanning and expressed as a percentage change. BMD in ovariectomized rats was significantly lower (−2.5 ± 2.0%) compared with the sham‐operated rats (+6.3 ± 5.3%; p < 0.01). Estrogen therapy completely abolished the ovariectomy‐induced potential bone loss (+5.9 ± 3.4%). Application of NG once daily also completely prevented (+6.2 ± 2.8%; p < 0.01) the ovariectomy‐induced bone loss (i.e., it was as effective as estrogen). However, the beneficial effects of NG on BMD were significantly reduced with increased frequency of application of NG (+1.9 ± 2.1%, twice a day and −0.2 ± 3.3% three times a day). Estrogen or once daily administration of NG preserved femur weights, BV/TV, and decreased urinary deoxypyridinoline levels as expected. However, a higher level of serum osteocalcin and bone‐specific alkaline phosphatase levels were maintained only with once daily administration of NG. There were no adverse effects of these doses of NG on blood pressure, but a tendency to lower blood pressure was noticed with increased frequency of NG. These results confirmed our previous findings that NO donors counteract the bone loss associated with estrogen deficiency. However, these beneficial effects of maintaining BMD are lost with increased frequency of NG application.