Increase of metabolic activity and disruption of normal contractile protein distribution by bilirubin oxidation products in vascular smooth-muscle cells

OBJECT: Cerebral vasospasm is a common cause of morbidity and death following aneurysmal subarachnoid hemorrhage (SAH). Previous research has shown that bilirubin oxidation products (BOXes) are present in the cerebral spinal fluid in patients with SAH-induced cerebral vasospasm and can contribute to vasoconstriction and vasospasm in vitro and in vivo. The events leading to cerebral vasospasm are not understood; however, one component of the occlusion may be due to vascular remodeling. In this study the authors have investigated the actions of BOXes, okadaic acid ([OA], a phosphatase inhibitor), and phorbol-12 myristate-13 acetate ([PMA], a protein kinase activator) on vascular smooth-muscle cell (VSMC) morphology and metabolism. METHODS: Immunohistochemical analysis was performed to assess VSMC morphology and alpha-smooth-muscle actin (alphaSMA) distribution following the application of BOXes, OA, or PMA. Changes in the level of lactate dehydrogenase (LDH) release and oxidative metabolism were also measured. The BOXes, OA, or PMA caused VSMCs to change their shape and exhibit altered alphaSMA distribution. These treatments increased LDH release (p < 0.05), which is an index of increased cell stress. Oxidative metabolism significantly increased at low and high doses of BOXes, that is, 143 +/- 8.5% and 180 +/- 11.8%, respectively (p < 0.0001). Both PMA and OA also caused a significant increase in metabolism. CONCLUSIONS: The authors concluded that BOXes, OA, and PMA alter VSMC morphology and metabolic activity, events that have been observed during vascular remodeling. Although the mechanism remains unclear, the results indicate that BOXes may play a role in the vascular remodeling that occurs following aneurysmal SAH.