We investigated the effect of N-acetyl-l-cysteine (NAC) on the expression of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, antioxidant enzymes, and inflammatory markers in diabetic rat hearts. Metabolic parameters, free 15-F(2t)-isoprostane level, protein expression of NADPH oxidase, superoxide dismutase (SOD), heme oxygenase (HO-1), interleukin-6 (IL-6), and cyclooxygenase-2 (COX-2) were analyzed in control and streptozotocin-induced diabetic rats treated with or without NAC in drinking water for 8 wk. The cardiac protein expression of p67(phox) and p22(phox) was increased in diabetic rats, accompanied by increased NADPH-dependent superoxide production. As a compensatory response to the increased NADPH oxidase, the protein expression of Cu-Zn-SOD and HO-1 and the total SOD activity were also increased in diabetic rat hearts. Consequently, cardiac free 15-F(2t)-isoprostane, an index of oxidative stress, was increased in diabetic rats, indicating that the production of reactive oxygen species becomes excessive in diabetic rat hearts. Cardiac inflammatory markers IL-6 and COX-2 were also increased in diabetic rats. NAC treatment prevented the increased expression of p22(phox) and translocation of p67(phox) to the membrane in diabetic rat hearts. Subsequently, the levels of cardiac free 15-F(2t)-isoprostane, HO-1, Cu-Zn-SOD, total SOD, IL-6, and COX-2 in diabetic rats were decreased by NAC. Consequently, cardiac hypertrophy was attenuated in diabetic rats treated with NAC. The protective effects of NAC on diabetic rat hearts may be attributable to its protection of hearts against oxidative damage induced by the increased NADPH oxidase and to its reduction in cardiac inflammatory mediators IL-6 and COX-2.

The results suggest that PKCbeta(2) overexpression represents a mechanism causing hyperglycemia-mediated myocardial hypertrophy, which can be prevented by the antioxidant N-acetylcysteine.

Novel classes of antimicrobials are needed to address the challenge of multidrug-resistant bacteria such as methicillin-resistant Staphylococcus aureus (MRSA). Using the architecture of the MRSA interactome, we identified pyruvate kinase (PK) as a potential novel drug target based upon it being a highly connected, essential hub in the MRSA interactome. Structural modeling, including X-ray crystallography, revealed discrete features of PK in MRSA, which appeared suitable for the selective targeting of the bacterial enzyme. In silico library screening combined with functional enzymatic assays identified an acyl hydrazone-based compound (IS-130) as a potent MRSA PK inhibitor (50% inhibitory concentration [IC 50 ] of 0.1 M) with >1,000-fold selectivity over human PK isoforms. Medicinal chemistry around the IS-130 scaffold identified analogs that more potently and selectively inhibited MRSA PK enzymatic activity and S. aureus growth in vitro (MIC of 1 to 5 g/ml). These novel anti-PK compounds were found to possess antistaphylococcal activity, including both MRSA and multidrug-resistant S. aureus (MDRSA) strains. These compounds also exhibited exceptional antibacterial activities against other Gram-positive genera, including enterococci and streptococci. PK lead compounds were found to be noncompetitive inhibitors and were bactericidal. In addition, mutants with significant increases in MICs were not isolated after 25 bacterial passages in culture, indicating that resistance may be slow to emerge. These findings validate the principles of network science as a powerful approach to identify novel antibacterial drug targets. They also provide a proof of principle, based upon PK in MRSA, for a research platform aimed at discovering and optimizing selective inhibitors of novel bacterial targets where human orthologs exist, as leads for anti-infective drug development.

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