Hemin activation of an inducible isoform of nitric oxide synthase in vascular smooth-muscle cells

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Abstract
✓ Hemin is a prominent breakdown product of hemoglobin, and high levels of hemin are found in the cerebrospinal fluid during subarachnoid hemorrhage—induced vasospasm. The possible role of hemin in modifying vascular function was examined in the present study by testing its effects on nitric oxide synthase (NOS) activity in cultured rat aortic smooth-muscle cells. Nitric oxide synthase activity was estimated from the amounts of accumulated nitrite and nitrate, which are oxidative products of nitric oxide (NO). Hemin (1–100 µM) increased the levels of nitrite and nitrate in culture medium in a dose- and time-dependent manner. The hemin-induced elevation of nitrite and nitrate was inhibited significantly by the NOS inhibitor, Nω-nitro-l-arginine (300 µM), and by the protein synthesis inhibitor, cycloheximide (5 µg/ml). These results indicate that hemin is capable of stimulating the expression of an inducible isoform of NOS (iNOS) in vascular smooth muscle. Transcriptional expression of iNOS is known to cause injurious effects on the maintenance of cellular homeostasis by generating extremely high levels of NO. The generation of hemin from methemoglobin during hemolysis of a subarachnoid blood clot could therefore stimulate an excessive production of NO in vascular smooth-muscle cells. It is postulated that this series of events contributes to the development of vascular injury associated with cerebral vasospasm after aneurysmal subarachnoid hemorrhage.
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