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Background Vasospasm occurs in conduits used for vascular reconstructions. The small heat shock proteins, HSP20 and HSP27, coordinately regulate vascular smooth muscle tone. Phosphorylated HSP20 is associated with vasorelaxation, and phosphorylated HSP27 inhibits the phosphorylation of HSP20 and relaxation. We hypothesized that the relationship between the phosphorylated states of these two proteins might dictate the tone of a vessel and may contribute to vasospasm. Study design Sodium nitroprusside relaxation of vascular smooth muscle was recorded using pig coronary artery and human saphenous vein. Segments were frozen and homogenized, and extracted proteins were separated by one- and two-dimensional gel electrophoresis, transferred to Immobilon (Millipore), and probed with anti-cGMP-dependent protein kinase (anti-PKG), -HSP20, -HSP27, and -phosphoHSP27 antibodies. Band intensity was estimated using densitometry. Results Pig coronary artery completely relaxed (100%) with SNP (10 −7 M), but human saphenous vein only partially relaxed (20%). The levels of cGMP-dependent protein kinase and HSP20 were similar in the two tissue types. Human saphenous vein had significantly higher levels of HSP27 versus pig coronary artery (30.14 ± 0.8 versus 6.62 ± 0.2 pixels/mg; p ≤ 0.001) and phosphoHSP27 (8.29 ± 3.43 versus 0.012 ± 0.008 pixels/mg; p ≤ 0.001). Conclusions Human saphenous vein contained significantly higher levels of HSP27 and pHSP27. Increased levels of phosphorylated HSP27 might contribute to vasospasm in human saphenous vein.

Introduction: HSP27 and HSP20, small heat shock proteins, have been implicated in the regulation of vascular smooth muscle (VSM) tone. Stress induced hypertension leads to increased expression of HSP27 in VSM. Transduction of VSM with HSP20 leads to vasorelaxation. Phosphorylated HSP27 (pHSP27) inhibits the activation of HSP20. We hypothesize that expression of HSP27 is greater in VSM refractory to cyclic nucleotide vasorelaxation. Methods: Sodium nitroprusside (SNP 10-7M) induced relaxation of VSM was measured using segments of pig coronary artery (PCA) and human saphenous vein (HSV) in a muscle bath. The levels of PKG, HSP20, phosphorylated HPS20 (pHSP20), HSP27 and pHSP27 in tissue extracts were determined by one- and two-dimensional gel electrophoresis and western blotting. Band intensity was estimated by densitometry. Results: PCA completely relaxed with SNP (100%), however, HSV did not (60%). Both tissue types had similar levels of expression of PKG and HSP20. The levels of HSP27 were 5 fold higher in the HSV (6.62 + 0.15 vs. 30.14 + 0.80) and pHSP27 was undetectable in PCA (0 vs.2.25 + 0.1). The levels of pHSP20 were undetectable in HSV. Conclusions: HSV segments refractory to relaxation with SNP had similar levels of expression of PKG and HSP20 as PCA segments demonstrating that the mediators of cyclic nucleotide vasorelaxation were present in both tissues. Increased levels of pHSP27 inhibit the phosphorylation of HSP20, and therefore, vasorelaxation of the HSV segments. Intracellular pHSP20 may be necessary and sufficient for VSM relaxation.