Vascular oxygen sensing: detection of novel candidates by proteomics and organ culture

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Abstract
We have shown that the specific inhibition of hypoxia-induced relaxation by organ culture in porcine coronary arteries can be mimicked by treatment of control vessels with the protein synthesis inhibitor, cycloheximide. We hypothesize that organ culture of vascular smooth muscle results in the decreased expression of proteins that are critical for vascular oxygen sensing. Using two-dimensional gel electrophoresis and mass spectroscopy, we identified such candidate proteins. The expressions of the smooth muscle-specific protein, SM22, and tropomyosin are decreased after 24 h in organ culture. These results were confirmed by Western blot analysis. Other smooth muscle proteins (actin and calponin) exhibited little change. We also demonstrate a 50% downregulation in the small G protein, Rho, a potent modulator of Ca2+-independent force. These results indicate that organ culture preferentially inhibits the expression of certain smooth muscle proteins. This change in protein expression after organ culture correlates with the specific inhibition of hypoxic vasorelaxation. These results provide novel target pathways for investigation that are potentially important for vascular oxygen sensing.