Abstract 1486: Preconditioning with S-nitrosoglutathione (GSNO) Results in S-nitrosylation and Altered Activity of the Sarcoplasmic Reticulum Ca2+-ATPase (SERCA) and the Mitochondrial F1-ATPase
Recent studies suggest that protein S-nitrosylation might play an important role in nitric oxide (NO)-related redox signaling to protect against oxidative stress. In this study, we found that S-nitrosoglutathione (GSNO), a biological S-nitrosothiol, elicited a cardioprotective effect similar to ischemic preconditioning (IPC). Hearts from C57BL/6J mice were perfused in the Langendorff mode. Hearts were subjected to:
GSNO preconditioning (GSNO-PC),
followed by 20 min of no-flow ischemia and reperfusion. Comparable to the effect of IPC, GSNO-PC significantly improved post-ischemic recovery of left ventricular developed pressure (LVDP) (54.8 ± 7.9% in GSNO-PC, 62.1 ± 6.7% in IPC, vs 36.8 ± 4.8% in control) and reduced infarct size (8.2 ± 2.4% of total area in GSNO-PC, 6.4 ± 1.1% in IPC, vs 28.2 ± 5.6% in control). GSNO-PC and IPC both significantly increased the S-nitrosothiol (SNO) contents in heart membrane fractions (9.2 ± 0.5 in GSNO-PC, 9.9 ± 0.4 in IPC, vs 3.4 ± 0.6 pmol SNO/mg protein in control). Using a combination of biotin switch method, streptavidin chromatography, and mass spectrometry proteomic analysis, we identified several candidate S-nitrosylated proteins in GSNO-PC hearts, including the L-type Ca2+ channel α1 subunit, the cardiac sarcoplasmic reticulum (SR) Ca2+-ATPase (SERCA), the α1 and β subunits of the mitochondrial F1-ATPase, cardiac muscle α myosin heavy chain, and myosin light chain 1. We examined the functional effect of S-nitrosylation on the activity of SERCA and the F1-ATPase by measuring the effects of GSNO treatment and subsequent S-nitrosylation on Ca2+ uptake into SR and oligomycin sensitive F1-ATPase activity. GSNO (0–1 mM) dose-dependently increased SERCA activity in isolated SR and decreased the mitochondrial F1-ATPase activity in the mitochondrial fraction, respectively. Because the F1-ATPase has been shown to run in reverse and consume ATP during ischemia, a decrease in activity of the F1-ATPase during ischemia would reduce ischemic ATP breakdown and promote cardioprotection. Thus, GSNO can elicit a cardioprotective role by S-nitrosylation and alterations in the activity of proteins such as SERCA2a and the mitochondrial F1-ATPase.