Nitrosative stress and myocardial sarcoplasmic endoreticular calcium adenosine triphosphatase subtype 2a activity after lung resection in swine.

BACKGROUND: Chronic, disease-associated oxidative stress induces myocardial peroxynitrite formation that may lead to nitrosative inhibition of the calcium cycling protein sarcoplasmic endoreticular calcium adenosine triphosphatase subtype 2a (SERCA2a). The current study was designed to test the hypothesis that the acute oxidative stress associated with lung resection also induces myocardial nitrosative stress and alters SERCA2a activity. METHODS: Ventricular myocardium from 16 swine was studied; 11 animals had undergone left upper lobectomy (n = 7) or sham thoracotomy (n = 4) 3 days before harvest, and 5 were nonoperated controls. Tissue peroxynitrite was assessed by measurement of 3-nitrotyrosine incorporation into proteins. SERCA2a activity was determined from indo-1 uptake by isolated sarcoplasmic reticular membranes. Expression of SERCA2a and its regulatory protein phospholamban were determined by Western blotting, as was the phospholamban phosphorylation state (when dephosporylated, phospholamban inhibits SERCA2a). Mechanical significance of changes in SERCA2a activity was assessed from the force-frequency relation of isometric myocardial trabeculae. RESULTS: Relative to both the control and sham groups, lobectomy animals exhibited a greater than twofold higher myocardial 3-nitrotyrosine incorporation and an approximately 50% lower SERCA2a activity, but no difference in SERCA2a or phospholamban expression or phospholamban phosphorylation. Concomitantly, whereas the trabecular force-frequency relation of control animals was positive, that of lobectomy animals was negative, consistent with impaired calcium cycling. CONCLUSIONS: These data indicate that oxidative/nitrosative stress associated with lung resection influences SERCA2a activity independent of any influence on protein expression or phospholamban phosphorylation. The findings link an acute event with a subcellular process primarily described for chronic illness and suggest a biochemical basis for perioperative changes in myocardial mechanical reserve.

Modular synthesis and preliminary biological evaluation of stereochemically diverse 1,3-dioxanes.

Modular synthesis and substrate stereocontrol were combined to furnish 18,000 diverse 1,3-dioxanes whose distribution in chemical space rivals that of a reference set of over 2,000 bioactive small molecules. Library quality was assessed at key synthetic stages, culminating in a detailed postsynthesis analysis of purity, yield, and structural characterizability, and the resynthesis of library subsets that did not meet quality standards. The importance of this analysis-resynthesis process is highlighted by the discovery of new biological probes through organismal and protein binding assays, and by determination of the building block and stereochemical basis for their bioactivity. This evaluation of a portion of the 1,3-dioxane library suggests that many additional probes for chemical genetics will be identified as the entire library becomes biologically annotated.

Structural biasing elements for in-cell histone deacetylase paralog selectivity.

We use the structural dissection of two 1,3-dioxanes with in-cell histone deacetylase (HDAC) paralog selectivity to identify key elements for selective HDAC inhibitors. We demonstrate that o-aminoanilides are inactive toward HDAC6 while apparently inhibiting deacetylases that act upon histone substrates. This finding has important clinical implications for the development of HDAC inhibitor-based treatments that do not interfere with microtubule dynamics associated with HDAC6. We also show that suberoylanilide hydroxamic acid (SAHA) alone is a nonparalog-selective HDAC inhibitor and that the 1,3-dioxane diversity appended to SAHA is essential for HDAC6 paralog selectivity.