FORUM EDITORIAL

New Redox-Related Arrows in the Arsenalof Cardiac Disease Treatment

Jonathan A. Kirk and Nazareno Paolocci

Abstract

While great strides have been made to improve the poor prognosis with cardiac disease, heart failure inparticular, cardiac affections still remain the most prevalent, difficult-to-treat, and costly human pathologies inthe western world. At rest, the heart produces a significant oxidative environment inside diverse cell com-partments, due to its high-energy demand. Cardiac cells have an exquisite control system to deal with thisconstant redox stress. However, persistent hemodynamic alterations can compromise these mechanisms, fuelingfurther myocardial redox imbalance and dysfunction. Still, this would be a one-sided and incomplete view,because the physiological role of reactive oxygen species (ROS) should be considered as well. Indeed, ROS aremultipurpose agents, serving signaling and cell defense tasks too, and, similar to antioxidants, these functionscan be highly compartmentalized within the cell. The present Forum was designed to collect cutting-edgeresearch concerning when and how to effectively counter excessive oxidative burden to preserve cardiacstructure and/or to improve function, under conditions of ordinary or extraordinary stress. Another majorobjective was to unravel old and new intersections between different myocardial processes by which ROS mayact as ‘‘on’’ or ‘‘off’’ switches, and in doing so, dictating function, always with an eye on possible, immediatetherapeutic applications, as suggested by the title of the Forum itself, that is, Cardiac Therapeutics. Antioxid.Redox Signal. 21, 1945–1948.

All substances are poisons; there is none which is not a poison.The right dose differentiates a poison from a remedy.

—Paracelsus (1493–1541)

The previous decade has seen an impressive decline( - 16.7%) in the number of deaths from cardiovascular

diseases (CVD) and heart failure (HF). However, CVD stillaccount for approximately one out of every three deaths in theUnited States, with more than 2150 deaths per day (5).Moreover, total direct and indirect costs for CVD were 315.4billion in 2010, compared with 201.5 billion for all cancers(5). These facts highlight that mortality for CVD and HFremains high and therapeutic treatment is a major socioeco-nomic burden.

The heart depends on highly oxidative metabolism. Thisexposes the organ to the possibility of being ‘‘oxidatively’’challenged and eventually injured, particularly under condi-tions of prolonged hemodynamic stress. However, as recentlystated, chronic oxidative changes in membrane lipids and

proteins found in many chronic diseases might not be theresult of accidental damage. Rather, they represent a mech-anism of adaptation or ‘‘shielding,’’ elaborated by all eu-karyote cells when placed in a chemically or microbiallyhostile environment. Thus, the view that countering oxidativestress should always be beneficial is likely one sided, par-ticularly if one considers that, so far, clinical trials with an-tioxidants have shown no benefit in treating cardiovasculardisorders. This evidence lends further support to the in-creasing awareness that reactive oxygen species (ROS) andreactive nitrogen species (RNS) may serve important sig-naling purposes in the cell, including those present in theheart.

One main impetus of the present Forum on CardiacTherapeutics was the idea of collecting original contributions

Division of Cardiology, Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, Maryland.

ANTIOXIDANTS & REDOX SIGNALINGVolume 21, Number 14, 2014ª Mary Ann Liebert, Inc.DOI: 10.1089/ars.2014.6124

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and updated review articles on new therapeutic avenues, es-pecially on HF treatment, for which changes in cardiac redoxconditions, at either mitochondrial or cytoplasmic level, ROSsignaling, and their scavenging constitute or may constitute amajor distinctive trait (Fig. 1).

Among the arrows in the arsenal of HF management is theuse of b-blockers, which help reverse the negative conse-quences of heightened catecholamine stimulation that occursin the disease. However, b-signaling in the heart is complex,and further clinical benefits may be derived from furtheroptimizing this system. Here, Huang et al. have contributedan up-to-date review on G-protein-coupled receptor (GPCR)kinase 2 (GRK2) (7). Animal models reveal that inhibitingGRK2 could offer synergistic benefits with b-blocker ther-apy, but even further benefits could arise from its ‘‘non-canonical’’ roles. These recently elucidated pathways includeGRK2’s ability to translocate to mitochondria in an ERK-dependent manner, where increased GRK2 signaling mayresult in decreased Ca2 + uptake capacity and opening of themitochondria permeability transition pore at a lower Ca2 +

threshold. This new action of GRK2 may account for celldeath, particularly under conditions of ischemia or exagger-ated ROS production and emission.

Ca2 + is arguably the most important second messenger inthe body, and the changes in Ca2 + handling in HF are welldocumented. Ziolo and Houser detail, in their comprehensivereview, the nitroso-redox modifications of the major Ca2 +

handling proteins, and how they are involved in disease (9).As mentioned earlier, clinical trials of antioxidant therapieshave not fared well overall. However, more specific, targeted,and compartmentalized antioxidant approaches should help

in preventing irreversible oxidative modifications due toexcessive ROS or RNS production that can affect, amongothers, cardiac E-C coupling proteins, thus rescuing, at leastin part, Ca2 + handling in chronically stressed hearts. Indeed,Ca2 + levels are directly tied to force production in the heart,and improving the heart’s ability to eject blood efficiently is aprimary concern in a large cohort of HF patients.

Almost all forms of heart disease result in a decrease in thenumber of properly functioning myocytes. With this loss ofmyocytes comes a loss of function, further progressing thedisease and leading further down the road to HF. Onepromising therapeutic approach is to replace these eithermissing or damaged myocytes with new ones. There are twopossible sources of new myocytes: endogenous and exoge-nous. In their timely review, Goichberg et al. discuss theinnate regenerative capacity of the heart (6). There is in-creasing evidence that the mature heart has a population ofcardiac stem cells, but its capacity to regenerate heart func-tion remains under debate. Obviously, as a therapeutic, trig-gering the heart to regenerate itself is enormously appealing,but many challenges remain.

It may also be possible to use an exogenous source of stemcells to repair the myocardium. Accordingly, the review byCho et al. presents a balanced overview of the various ap-proaches and technologies that are being used to develop anddeliver stem cells to the damaged heart (3). Progress is madein developing these cells from many different sources, andfrom these efforts it can be evinced that the likelihood ofearning new and effective therapeutic tools increases, almostday by day. However, both review articles insist on an im-portant point: Regardless of whether stem cells are of

FIG. 1. Schematic show-ing some of the reactiveoxygen species (ROS)-mediated mechanisms ofthe cardiac therapeutics in-troduced in this Forum.Each numeral indicates thereference that details that me-chanism. SR, sarcoplasmic re-ticulum; CA, catecholamines;ULK1, unc-51 like autophagyactivating kinase 1. All otherabbreviations are explained inthe text. To see this illustrationin color, the reader is referredto the web version of this ar-ticle at www.liebertpub.com/ars

1946 KIRK AND PAOLOCCI

endogenous or exogenous origin, it is increasingly evidentthat ROS production accompanies and contributes, at least tosome extent, to cell proliferation, differentiation, and sur-vival. The molecular and redox intricacies that underlie theseeffects remain to be elucidated, providing a fertile terrain forfuture investigation.

Mitochondria constitute *30% by volume of the cardiacmyocyte. Through oxidative phosphorylation, these organ-elles produce the bulk of ATP necessary to maintain myo-cardial function. During this process, they handle amounts ofO2 that are much higher than those used by the brain or theskeletal muscle. The mitochondrial respiratory chain reducesO2 efficiently through cytochrome oxidase, but the electronflux can be diverted to produce the free radical superoxide,O2� - . These mitochondrial-borne ROS are continuously

scavenged, keeping ROS emission at low levels. However, incardiac diseases, mitochondrial function is altered, leading toconcomitant lower energetics and increased ROS emission.The removal of senescent mitochondria is essential, becausethey are cytotoxic, and fission/fusion and mitophagy mech-anisms have been developed to efficiently remove them.Here, Chen et al. present data showing that the geneticknockdown of mitofusin 2, a key protein for mitochondrialfusion, results in a cardiomyopathy phenotype, in both dro-sophila and mice (2). They demonstrate that Mrf2, bytransducing PINK1-Parkin mitophagy signaling, contributesto the integrity of mitochondrial renewal, preventing theaccumulation of mtDNA with double-stranded breaks. Thelatter event is per se sufficient to cause cardiomyopathy, andthe authors suggested that this linearization of mtDNA mayintervene as a causative factor in neurological degenerativedisorders such as Parkinson’s and Alzheimer’s diseases,diseases in which defects in mitochondria protein qualitycontrol are known to occur. However, Chen’s study hints atthe possibility that pathogeric aging could also be due to aprogressive decline in the mechanisms deputed to the effec-tive removal of fragmented mitochondria.

The therapeutic targeting of statins is inhibition of HMG-CoA reductase, which is the rate-limiting enzyme for cho-lesterol biosynthesis. However, in both experimental andhuman myocardial infarction (MI) settings, statins affordprotection in a manner that is independent from changes incholesterol levels. Andres et al. aimed at unraveling themechanism underpinning statin-afforded acute cardiopro-tection. The authors showed that it involves the activation ofthe protein Parkin, known to stimulate mitophagy (1); in fact,simvastatin reduced infarct size in wild-type mice but not inparkin KO mice. They went on to determine the mechanismsaccounting for simvastatin’s salutary action, and reportedthat this statin prevents the accumulation of mevalonate. Thisdepletion attenuated Akt/mTOR signaling and disinhibitedULK1, ultimately resulting in increased macroautophagy.The status quo ante was rescued by mevalonate supple-mentation, thus validating that simvastatin beneficial effectsare related to the inhibition of HMG-CoA reductase activity.Intriguingly, inhibiting the latter may also result in loss incoenzyme Q10 (CoQ), the levels of which also depend onmevalonate’s bioavailability. However, in another twist, theauthors observed that while CoQ supplementation preventedmitophagy in cell cultures, it had minimal impact whensuppressing mTOR signaling. Hence, depletion of CoQ andconsequent enhanced mitophagy is a main mechanism by

which simvastatin induces cardiac protection. These findings,however, remain to be reconciled with existing evidenceshowing that CoQ administration could be beneficial in pa-tients with diabetes and certain neurodegenerative disorders,due its antioxidant or mitochondria-protecting effects orboth.

MI is still the main cause of death in the Western world.de Castro Bras et al. here reported surprising results showingthat the matrix metalloproteinase 9 (MMP9), a gelatinase thatcontrols the turnover of the extracellular matrix, is a novelregulator of mitochondrial activity in post-MI conditions (4).Usually, citrate synthase (CS), the rate-limiting mitochon-drial enzyme that mediates the first step of the tricarboxylicacid cycle, is decreased in the heart after infarction. Deletingthe MMP9 gene prevents this effect, and the authors dem-onstrated that MMP9 directly cleaves CS, both in vitro andin vivo. Although how exactly the expression/activity of acollagen turnover protein and enzymes involved in mito-chondrial oxidative metabolism intersect requires extensiveadditional investigation, these investigators provide intriguinginitial evidence that in MMP9 null mice manganese super-oxide dismutase levels are increased as compared with theircontrol littermates, suggesting that mitochondrial functioncould be somehow improved in the null mice. This studyunravels a new crossroad between enzymes involved in en-ergy production, extracellular matrix remodeling, and mito-chondrial ROS scavenging, likely extending the possibleapplications of MMP inhibitors, beyond their consolidatedrole in preventing adverse left ventricle remodeling after MI.

Hypertension is the second most common cause of HF,accounting for *25% of HF cases. This syndrome affectsone out of every three U.S. adults older than the age of 20 (5),and in the elderly, almost 70% of HF cases are due to hy-pertension. The hemodynamic stress imparted by increasedafterload in hypertensive patients initially leads to a status ofconcentric/compensated hypertrophy, but with the persis-tence of elevated peripheral vascular resistance in time, itwould inevitably result in HF. The addition of small ubi-quitin-like modifier type 1 (SUMO-1) to lysine residues 480and 585 stabilizes SERCA2a, enhancing its activity. Thispost-translational modification (SUMOylation) increases cellsurvival in the presence of oxygen/glucose deprivation,suggesting it as a part of the ischemic tolerance package. Intheir paper, Lee et al. extended the protective coverage ex-erted by SUMO-1 to mouse hearts subject to pressure over-load (8), showing that SUMO-1 gene transfer blocks thetransition from compensated hypertrophy to HF. The authorsfound that the overexpression of SUMO leads to a down-regulation of Nox4, an effect achieved via the SUMOylationof c-Jun, which is a key transcriptional regulator of Nox4expression. Hence, this study brings to the fore that amongthe protective actions exerted by this post-translationalmodification, there is a reduction in the extent of oxidativestress that, in addition to preserved SERCA2 function,maintains function in pressure-overloaded hearts.

These reviews and articles can be viewed integratively toprovide several ‘‘take-home messages.’’ First, processesgoverning myocardial biology and function that at firstglance look functionally distant from each other are, in re-ality, tightly interconnected. For instance, this is the case ofextracellular matrix turnover and mitochondrial oxidativemetabolism. ROS are ubiquitous and, in some cases, readily

NOVEL CARDIAC THERAPEUTICS 1947

diffusible; thus, it is a legitimate speculation that thesemolecules are on/off nodes in these novel networks.

Second, we should definitely abandon the view of ROSscavengers as mono-dimensional or mono-directional agents;rather, their actions should always be contextualized. Indeed,forcing their expression may be beneficial in some com-partments, but suppress important salutary effects in others.

Third, the mitochondrial environment is a main ‘‘maker’’of the overall cellular redox conditions, dictating the mag-nitude and frequency of emitted ROS pulses that are alsoimplicated in signaling. When partially damaged, these ma-chines need parts replaced, but when overtly broken and nolonger reparable, the organelles’ pieces should be taken apartto avoid exacerbation of oxidative stress and the occurrenceof cardiomyopathy. Studies included here reinstate mito-chondria as dictators of both cell life and death, emphasizingtheir pathogenetic role even when their own life cycle isalready over. Thus, interventions aimed at maintainingproper myocardial redox balance (and with that the functionof many functionally relevant redox switches) cannot pre-scind from taking care of healthy or diseased mitochondria inthe first place.

Finally, ROS are a ‘‘pharmakon,’’ in the Greek meaningof the term, that is, remedy and poison simultaneously,exclusively based on their dosage. Differently from drugs,however, they are endogenous and constantly produced,either as byproducts of oxidative metabolism or as catabo-lites of endogenous messengers. Since they serve essentialdefensive purposes too, they should not be indiscriminatelysuppressed. Thus, any therapeutic attempts for cardiac dis-eases that also harbor ‘‘altered redox balance’’ should beprimarily directed to the root causes of a given disease.Countering excessive ROS emission in this context shouldbe a very specifically targeted task, taking place during thedue course of the affection, in a highly compartmentalizedmanner, and certainly not when oxidative damage is alreadya postfactual bystander. In essence, ROS are not only in-evitable by-products of any aerobic life, including ours, butalso sentinels of it and ultimate signaling molecules in ourbodies. Hence, we should learn more about their physio-logical aspects, including their defensive capacity andability to control cell differentiation and contacts, to turn apossible poison into a possible remedy, even under cardiacpathological conditions.

Acknowledgments

The authors acknowledge the American Heart Association(Scientist Development Grant 20380148, to J.A.K., andGrant-in-Aid 17070027, to N.P.).

References

1. Andres AM, Hernandez G, Lee P, Huang C, Ratliff EP, SinJ, Thornton CA, Damasco MV, and Gottlieb RA. Mitophagyis required for acute cardioprotection by simvastatin. Anti-oxid Redox Signal 21:1960–1973, 2014.

2. Chen Y, Sparks M, Bhandari P, Matkovich SJ, and DornGW II. Mitochondrial genome linearization is a causativefactor for cardiomyopathy in mice and Drosophila. AntioxidRedox Signal 21:1949–1959, 2014.

3. Cho G-S, Fernandez L, and Kwon C. Regenerative medicinefor the heart: perspectives on stem-cell therapy. AntioxidRedox Signal 21:2018–2031, 2014.

4. de Castro Bras LE, Cates CA, Deleon-Pennell KY, Ma Y,Iyer RP, Halade GV, Yabluchanskiy A, Fields GB, Wein-traub ST, and Lindsey ML. Citrate synthase is a novel in vivomatrix metalloproteinase-9 substrate that regulates mito-chondrial function in the postmyocardial infarction leftventricle. Antioxid Redox Signal 21:1974–1985, 2014.

5. Go AS, Mozaffarian D, Roger VL, Benjamin EJ, Berry JD,Blaha MJ, Dai S, Ford ES, Fox CS, Franco S, Fullerton HJ,Gillespie C, Hailpern SM, Heit JA, Howard VJ, HuffmanMD, Judd SE, Kissela BM, Kittner SJ, Lackland DT,Lichtman JH, Lisabeth LD, Mackey RH, Magid DJ, MarcusGM, Marelli A, Matchar DB, McGuire DK, Mohler ER, 3rd,Moy CS, Mussolino ME, Neumar RW, Nichol G, PandeyDK, Paynter NP, Reeves MJ, Sorlie PD, Stein J, Towfighi A,Turan TN, Virani SS, Wong ND, Woo D, Turner MB,American Heart Association Statistics Committee, andStroke Statistics Subcommittee. Heart disease and strokestatistics—2014 update: a report from the American HeartAssociation. Circulation 129: e28–e292, 2014.

6. Goichberg P, Chang J, Liao R, and Leri A. Cardiac stem cells:biology and clinical applications. Antioxid Redox Signal21:2002–2017, 2014.

7. Huang ZM, Gao E, Chuprun JK, and Koch WJ. GRK2 in theheart: a GPCR kinase and beyond. Antioxid Redox Signal21:2032–2043, 2014.

8. Lee A, Jeong D, Mitsuyama S, Oh JG, Liang L, Ikeda Y,Sadoshima J, Hajjar RJ, and Kho C. The role of SUMO-1 incardiac oxidative stress and hypertrophy. Antioxid RedoxSignal 21:1986–2001, 2014.

9. Ziolo MT and Houser SR. Abnormal CA2 + cycling in failingventricular myocytes: role of NOS1-mediated nitroso-redoxbalance. Antioxid Redox Signal 21:2044–2059, 2014.

Address correspondence to:Dr. Nazareno PaolocciDivision of Cardiology

Department of MedicineJohns Hopkins University School of Medicine

Baltimore, MD 21205

E-mail: npaoloc1@jhmi.edu

Date of first submission to ARS Central, September 10, 2014;date of acceptance, September 11, 2014.

Abbreviations Used

CA¼ catecholaminesCoQ¼ coenzyme Q10

CS¼ citrate synthaseCVD¼ cardiovascular disease

GRK2¼G-protein-coupled receptor (GPCR) kinase 2HF¼ heart failureMI¼myocardial infarction

MMP9¼matrix metalloproteinase 9RNS¼ reactive nitrogen speciesROS¼ reactive oxygen species

SUMO-1¼ small ubiquitin-like modifier type 1ULK1¼ unc-51 like autophagy activating kinase 1

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