Abstract 8670: Homocysteine Induces Reactive Oxygen Species-dependent Apoptosis of Arterial Smooth Muscle Cells Coinciding with (peri)nuclear Nox4 Translocation
Background: Elevated levels of homocysteine form a vascular risk factor. Although a jeopardizing role of homocysteine on endothelial cells has been shown, its effect on smooth muscle cells is contradicting, varying from induction of proliferation to induction of cell death. Recent studies have shown that NADPH oxidase-mediated reactive oxygen species play a role in homocysteine-induced apoptosis in cardiomyocytes and endothelial cells. However to the best of our knowledge, this has not been studied in smooth muscle cells.
Methods: Human arterial smooth muscle cells were incubated with 100 µM homocysteine during 24 hours and were analyzed for cell viability using FACS analysis and caspase 3 activity. The effects of homocysteine on the expression of different NADPH oxidase proteins and -mediated reactive oxygen species production were studied using digital-imaging microscopy.
Results: Incubation of arterial smooth muscle cells with 100 µM homocysteine during 24 hours induced a significant increase in single annexin V positive cells. This increase coincided with a significant increased caspase 3 activity. Furthermore, homocysteine resulted in a significant shift of NADPH oxidase 4 from the cytoplasm to (peri)nuclear regions and a significant increased reactive oxygen species production in the (peri)nucleus and the cytoplasm. No effects of homocysteine on expression levels, nor location of NADPH oxidase 1 and 2 were found. Inhibition of NADPH oxidase-mediated reactive oxygen species production with the flavoenzyme inhibitor diphenylene iodonium led to significant decreased apoptosis levels as assessed by quantification of caspase 3 activity.
Conclusion: 100 µM homocysteine during 24 hours induced reactive oxygen species-dependent apoptosis of human arterial smooth muscle cells coinciding with (peri)nuclear NADPH oxidase 4 translocation.
- © 2011 by American Heart Association, Inc.