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Non‐genomic effects of estrogen on human coronary artery smooth muscle are mediated by nNOS via HSP90 complex

Estrogen ameliorates or exacerbates coronary heart disease. Although estrogen dilates coronary arteries from a variety of species, the molecular basis for this acute, non‐genomic effect is unclear. Moreover, we know very little of how estrogen affects human coronary artery smooth muscle cells (HCASMCs). Heat shock protein 90 (HSP90) facilitates neuronal nitric oxide synthase (nNOS, type 1) activity in vivo. We propose that HSP90 is involved in estrogen signaling in HCASMC by facilitating nNOS activity. We have combined molecular and cellular functional studies with protein analysis to identify novel targets of estrogen action in HCASMCs: nNOS and HSP90. Fluorescence studies demonstrated that 17b‐estradiol (E2) increases NO production, and single‐channel patch‐clamp experiments revealed that activation of the NO/cGMP signaling pathway opens calcium‐activated potassium (BKCa) channels in these cells. Western blot revealed nNOS expression, but failed to detect eNOS or iNOS isoforms. Co‐immunoprecipitation studies revealed that E2 stimulates association of HSP90 with nNOS, whereas HSP90 inhibitors, geldanamycin or radicicol reversed the stimulatory effect of E2 on BKCa channels and on NO production, thereby implicating the nNOS/HSP90 complex as an estrogen transduction component. We conclude that estrogen opens BKCa channels in HCASMC by stimulating nNOS via a transduction sequence involving HSP90 forming a complex with nNOS. These findings provide a molecular mechanism that can explain the clinical observation that estrogen enhances coronary blood flow in patients with diseased or damaged coronary arteries.