Cohen RA, Colucci WS. Cytosolic H2O2 mediates hypertrophy, apoptosis, and decreased SERCA activity in mice with chronic hemodynamic overload. Am J Physiol Heart Circ Physiol 306: H1453-H1463, 2014. First published March 14, 2014 doi:10.1152/ajpheart.00084.2014.-Oxidative stress in the myocardium plays an important role in the pathophysiology of hemodynamic overload. The mechanism by which reactive oxygen species (ROS) in the cardiac myocyte mediate myocardial failure in hemodynamic overload is not known. Accordingly, our goals were to test whether myocyte-specific overexpression of peroxisomal catalase (pCAT) that localizes in the sarcoplasm protects mice from hemodynamic overload-induced failure and prevents oxidation and inhibition of sarco(endo)plasmic reticulum Ca 2ϩ -ATPase (SERCA), an important sarcoplasmic protein. Chronic hemodynamic overload was caused by ascending aortic constriction (AAC) for 12 wk in mice with myocyte-specific transgenic expression of pCAT. AAC caused left ventricular hypertrophy and failure associated with a generalized increase in myocardial oxidative stress and specific oxidative modifications of SERCA at cysteine 674 and tyrosine 294/5. pCAT overexpression ameliorated myocardial hypertrophy and apoptosis, decreased pathological remodeling, and prevented the progression to heart failure. Likewise, pCAT prevented oxidative modifications of SERCA and increased SERCA activity without changing SERCA expression. Thus cardiac myocyte-restricted expression of pCAT effectively ameliorated the structural and functional consequences of chronic hemodynamic overload and increased SERCA activity via a post-translational mechanism, most likely by decreasing inhibitory oxidative modifications. In pressure overload-induced heart failure cardiac myocyte cytosolic ROS play a pivotal role in mediating key pathophysiologic events including hypertrophy, apoptosis, and decreased SERCA activity. apoptosis; catalase; H2O2; hypertrophy; SERCA THE ROLE OF REACTIVE OXYGEN species (ROS) in mediating myocardial failure in response to chronic hemodynamic overload is well appreciated (29,32,34). Systemic administration of small molecule antioxidants ameliorated pathological remodeling and failure in mice with hemodynamic overload due to chronic aortic constriction (8,35). Transgenic total body overexpression of mitochondrial catalase (mCAT) likewise decreased left ventricular (LV) hypertrophy and failure in mice with aortic constriction (6), further suggesting that mitochondria are a source of ROS in hemodynamic overload. However, several nonmitochondrial sources of ROS in the myocardium have been implicated in hemodynamic overload including uncoupled endothelial nitric oxide synthase (23, 33), NADPH oxidases (4), and xanthine oxidase (21). Because these nonmitochondrial sources of ROS are located in the sarcoplasm, we reasoned that their effects should be susceptible to catalase that is targeted to the sarcoplasm of the cardiac myocyte. Accordingly, our first goal was to test whether LV hypertrophy and failur...

We analyzed 49 patients with multiple myeloma (MM) using propidium iodide (PI) staining and flow cytometry to assess DNA content of MM cells in bone marrow. In 12 patients the presence or absence of DNA content aneuploidy could not be determined by analysis of single-parameter DNA histograms alone. Using forward light scatter measurement, resolution of ploidy was readily obtained in all but three cases. Assuming the DNA content of the low light scatter population to be diploid, we were able to classify 7 cases as hypodiploid and 39 as hyperdiploid or diploid. Hyperdiploid or diploid patients survived a median of 24 months and hypodiploid patients only 2.5 months. We conclude that multiparameter analysis of DNA content and light scatter provides easier analysis for the presence or absence of aneuploidy. Patients with DNA content hypodiploidy have a short survival.

Background: Sarco(endo)plasmic reticulum calcium ATPase (SERCA) is regulated by oxidative post-translational modifications at cysteine 674 (C674). Since sarcoplasmic reticulum (SR) calcium has been shown to play a critical role in mediating mitochondrial dysfunction in response to reactive oxygen species (ROS), we hypothesized that SERCA oxidation at C674 would modulate the effects of ROS on mitochondrial calcium and mitochondria-dependent apoptosis in cardiac myocytes. Methods: Adult rat ventricular myocytes (ARVM) expressing wild-type (WT) SERCA2b or a redox-insensitive mutant in which C674 is replaced by serine (C674S) were exposed to H 2 O 2 (100 μ M). Free mitochondrial calcium concentration was measured in ARVM using a genetically-targeted fluorescent probe and SR calcium content was assessed by measuring caffeine-stimulated release. Mice with heterozygous knock-in of the SERCA C674S mutation (SKI) were subjected to chronic ascending aortic constriction (AAC). Results: In ARVM expressing WT SERCA, H 2 O 2 caused a 25% increase in mitochondrial calcium concentration that was associated with a 50% decrease in SR calcium content, both of which were prevented by the ryanodine receptor inhibitor tetracaine. In cells expressing the C674S mutant, basal SR calcium content was decreased by 31% and the H 2 O 2 -stimulated rise in mitochondrial calcium concentration was attenuated by 40%. In WT cells, H 2 O 2 caused cytochrome c release and apoptosis, both of which were prevented in C674S-expressing cells. In myocytes from SKI mice, basal SERCA activity and SR calcium content were decreased. To test the effect of C674 oxidation on apoptosis in vivo , SKI mice were subjected to chronic AAC. In WT mice, AAC caused myocyte apoptosis, LV dilation and systolic failure - all of which were inhibited in SKI mice. Conclusions: Redox activation of SERCA C674 regulates basal SR calcium content thereby mediating the pathologic ROS-stimulated rise in mitochondrial calcium required for myocyte apoptosis and myocardial failure.

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