Abstract 16253: Cyclosporine-a Mediated Impairment of Human Coronary Artery Smooth Muscle Cell Phospho-nrf2 and Vascular Injury Function Are Rescued by Tert-Butylhydroquinone
Background: The association of Cyclosporine (CsA) with cardiac allograft vasculopathy (CAV) may, in part, be via oxidative stress in coronary arteries. We previously demonstrated that CsA-mediated impairement of the antioxidant Nrf2 pathway was rescued by Tert-butylhydroquinone (tBHQ), an Nrf2 agonist. Nrf2 phosphorylation is a key step upstream of its nuclear localization and subsequent upregulation of antioxidant enzymes. We hypothesized that CsA mediated vasomotor dysfunction is characterized by a reduction in phospho Nrf2 (pN) levels and rescued by tBHQ.
Methods and Results: Lewis rats (n=8) received tBHQ (50mg/kg), CsA (5mg/kg) ± tBHQ, or Saline (CON) daily for 2 weeks. Thoracic aortic segments were excised and exposed to phenylephrine (Phe, 100nM) followed by acetylcholine (ACh) and sodium nitroprusside (SNP) to assess for endothelial dependent (Edep) and independent (Eind) relaxation respectively. We measured ACh and SNP half-maximum vasodilation (ED50) as well as sensitivity to Endothelin-1 (ET-1, 15nM) induced vasospasm.
Human coronary artery smooth muscle cells (HCASMC, n=6-8) were incubated with tBHQ (80uM), CsA (50ug/ml) ± tBHQ, or DMSO (control) for 24hrs. Using Western Blotting, we measured pN/whole cell Nrf2 (pN/N) ratio.
CsA resulted in lower Edep vasorelaxation compared to CON and tBHQ (ED50: CsA. 90±7%, CON, 75±8%, tBHQ, 80±3%, p<0.01), but not compared to CsA + tBHQ (ED50: 92±9%). However, CsA resulted in lower Eind vasorelaxation compared to all groups (ED50: CsA, 99±4%, CON, 41±8%, tBHQ, 62±3%, and CsA + tBHQ, 78%±10, p<0.05). Compared to CON, CsA demonstrated increased sensitivity to vasospasm which was normalized in tBHQ+CsA (CON, 100±5%, CsA, 121±13%, tBHQ+CsA, 108±9%, p<0.05).
In HCASMCs, CsA significantly reduced pN/N ratio compared to CON and tBHQ (CsA, 39.7±1.2%, CON, 100±5.6%, tBHQ, 416±40%, p<0.05). CsA + tBHQ pN/N ratio (135%±8.9%, p=0.01) was significantly higher than CsA alone.
Conclusion: Previously we showed that augmenting nitric oxide (NO) levels rescued CsA mediated Edep vascular dysfunction. Here we show the novel observation that tBHQ augments vasomotor function, via an Eind pathway which may be associated with normalizing phosphorylation of Nrf2 in smooth muscle cells.
- © 2013 by American Heart Association, Inc.