Cardioprotective and Anti-Inflammatory Effects of FAM3D in Myocardial Ischemia-Reperfusion Injury

HomeCirculation ResearchVol. 133, No. 7Cardioprotective and Anti-Inflammatory Effects of FAM3D in Myocardial Ischemia-Reperfusion Injury Free AccessResearch ArticlePDF/EPUBAboutView PDFView EPUBSections ToolsAdd to favoritesDownload citationsTrack citationsPermissions ShareShare onFacebookTwitterLinked InMendeleyReddit Jump toFree AccessResearch ArticlePDF/EPUBCardioprotective and Anti-Inflammatory Effects of FAM3D in Myocardial Ischemia-Reperfusion Injury James Rhee, Rebecca Freeman, Kangsan Roh, Margaret Lyons, Chunyang Xiao, Daniel Zlotoff, Ashish Yeri, Haobo Li, Justin Guerra, J. Sawalla Guseh, Alexandra Kuznetsov, Nicholas Houstis, Jason Roh, Federico Damilano, Xiaojun Liu, Michael Silverman, Raymond Kwong, Saumya Das and Anthony Rosenzweig James RheeJames Rhee Correspondence to: James Rhee, MD, PhD, Massachusetts General Hospital, Boston, MA, Email E-mail Address: [email protected] Department of Medicine, Cardiovascular Research Center and Division of Cardiology (J. Rhee, R.F., K.R., M.L., C.X., D.Z., A.Y., H.L., J.G., J.S.G., A.K., N.H., J. Roh, F.D., X.L., M.S., S.D., A.R.), Massachusetts General Hospital, Boston. Department of Anesthesia, Critical Care, and Pain Medicine (J. Rhee, R.F., K.R., M.L.), Massachusetts General Hospital, Boston. , Rebecca FreemanRebecca Freeman Department of Medicine, Cardiovascular Research Center and Division of Cardiology (J. Rhee, R.F., K.R., M.L., C.X., D.Z., A.Y., H.L., J.G., J.S.G., A.K., N.H., J. Roh, F.D., X.L., M.S., S.D., A.R.), Massachusetts General Hospital, Boston. Department of Anesthesia, Critical Care, and Pain Medicine (J. Rhee, R.F., K.R., M.L.), Massachusetts General Hospital, Boston. , Kangsan RohKangsan Roh https://orcid.org/0000-0001-8067-7772 Department of Medicine, Cardiovascular Research Center and Division of Cardiology (J. Rhee, R.F., K.R., M.L., C.X., D.Z., A.Y., H.L., J.G., J.S.G., A.K., N.H., J. Roh, F.D., X.L., M.S., S.D., A.R.), Massachusetts General Hospital, Boston. Department of Anesthesia, Critical Care, and Pain Medicine (J. Rhee, R.F., K.R., M.L.), Massachusetts General Hospital, Boston. , Margaret LyonsMargaret Lyons Department of Medicine, Cardiovascular Research Center and Division of Cardiology (J. Rhee, R.F., K.R., M.L., C.X., D.Z., A.Y., H.L., J.G., J.S.G., A.K., N.H., J. Roh, F.D., X.L., M.S., S.D., A.R.), Massachusetts General Hospital, Boston. Department of Anesthesia, Critical Care, and Pain Medicine (J. Rhee, R.F., K.R., M.L.), Massachusetts General Hospital, Boston. , Chunyang XiaoChunyang Xiao https://orcid.org/0000-0003-1972-2241 Department of Medicine, Cardiovascular Research Center and Division of Cardiology (J. Rhee, R.F., K.R., M.L., C.X., D.Z., A.Y., H.L., J.G., J.S.G., A.K., N.H., J. Roh, F.D., X.L., M.S., S.D., A.R.), Massachusetts General Hospital, Boston. , Daniel ZlotoffDaniel Zlotoff Department of Medicine, Cardiovascular Research Center and Division of Cardiology (J. Rhee, R.F., K.R., M.L., C.X., D.Z., A.Y., H.L., J.G., J.S.G., A.K., N.H., J. Roh, F.D., X.L., M.S., S.D., A.R.), Massachusetts General Hospital, Boston. , Ashish YeriAshish Yeri Department of Medicine, Cardiovascular Research Center and Division of Cardiology (J. Rhee, R.F., K.R., M.L., C.X., D.Z., A.Y., H.L., J.G., J.S.G., A.K., N.H., J. Roh, F.D., X.L., M.S., S.D., A.R.), Massachusetts General Hospital, Boston. , Haobo LiHaobo Li https://orcid.org/0000-0002-5660-7835 Department of Medicine, Cardiovascular Research Center and Division of Cardiology (J. Rhee, R.F., K.R., M.L., C.X., D.Z., A.Y., H.L., J.G., J.S.G., A.K., N.H., J. Roh, F.D., X.L., M.S., S.D., A.R.), Massachusetts General Hospital, Boston. , Justin GuerraJustin Guerra https://orcid.org/0000-0001-8507-5922 Department of Medicine, Cardiovascular Research Center and Division of Cardiology (J. Rhee, R.F., K.R., M.L., C.X., D.Z., A.Y., H.L., J.G., J.S.G., A.K., N.H., J. Roh, F.D., X.L., M.S., S.D., A.R.), Massachusetts General Hospital, Boston. Institute for Heart and Brain Health, University of Michigan Medical Center, Ann Arbor (J.G., A.R.). , J. Sawalla GusehJ. Sawalla Guseh https://orcid.org/0000-0003-0992-5635 Department of Medicine, Cardiovascular Research Center and Division of Cardiology (J. Rhee, R.F., K.R., M.L., C.X., D.Z., A.Y., H.L., J.G., J.S.G., A.K., N.H., J. Roh, F.D., X.L., M.S., S.D., A.R.), Massachusetts General Hospital, Boston. , Alexandra KuznetsovAlexandra Kuznetsov https://orcid.org/0000-0002-6074-5819 Department of Medicine, Cardiovascular Research Center and Division of Cardiology (J. Rhee, R.F., K.R., M.L., C.X., D.Z., A.Y., H.L., J.G., J.S.G., A.K., N.H., J. Roh, F.D., X.L., M.S., S.D., A.R.), Massachusetts General Hospital, Boston. , Nicholas HoustisNicholas Houstis Department of Medicine, Cardiovascular Research Center and Division of Cardiology (J. Rhee, R.F., K.R., M.L., C.X., D.Z., A.Y., H.L., J.G., J.S.G., A.K., N.H., J. Roh, F.D., X.L., M.S., S.D., A.R.), Massachusetts General Hospital, Boston. , Jason RohJason Roh https://orcid.org/0000-0002-6999-6868 Department of Medicine, Cardiovascular Research Center and Division of Cardiology (J. Rhee, R.F., K.R., M.L., C.X., D.Z., A.Y., H.L., J.G., J.S.G., A.K., N.H., J. Roh, F.D., X.L., M.S., S.D., A.R.), Massachusetts General Hospital, Boston. , Federico DamilanoFederico Damilano Department of Medicine, Cardiovascular Research Center and Division of Cardiology (J. Rhee, R.F., K.R., M.L., C.X., D.Z., A.Y., H.L., J.G., J.S.G., A.K., N.H., J. Roh, F.D., X.L., M.S., S.D., A.R.), Massachusetts General Hospital, Boston. , Xiaojun LiuXiaojun Liu Department of Medicine, Cardiovascular Research Center and Division of Cardiology (J. Rhee, R.F., K.R., M.L., C.X., D.Z., A.Y., H.L., J.G., J.S.G., A.K., N.H., J. Roh, F.D., X.L., M.S., S.D., A.R.), Massachusetts General Hospital, Boston. , Michael SilvermanMichael Silverman https://orcid.org/0000-0002-5885-1866 Department of Medicine, Cardiovascular Research Center and Division of Cardiology (J. Rhee, R.F., K.R., M.L., C.X., D.Z., A.Y., H.L., J.G., J.S.G., A.K., N.H., J. Roh, F.D., X.L., M.S., S.D., A.R.), Massachusetts General Hospital, Boston. , Raymond KwongRaymond Kwong https://orcid.org/0000-0001-8212-0759 Cardiovascular Medicine, Brigham and Women's Hospital, Boston, MA (R.K.). , Saumya DasSaumya Das https://orcid.org/0000-0002-4521-4606 Department of Medicine, Cardiovascular Research Center and Division of Cardiology (J. Rhee, R.F., K.R., M.L., C.X., D.Z., A.Y., H.L., J.G., J.S.G., A.K., N.H., J. Roh, F.D., X.L., M.S., S.D., A.R.), Massachusetts General Hospital, Boston. and Anthony RosenzweigAnthony Rosenzweig Anthony Rosenzweig, MD, University of Michigan Medical Center, Ann Arbor, MI, Email E-mail Address: [email protected] https://orcid.org/0000-0001-6387-0386 Department of Medicine, Cardiovascular Research Center and Division of Cardiology (J. Rhee, R.F., K.R., M.L., C.X., D.Z., A.Y., H.L., J.G., J.S.G., A.K., N.H., J. Roh, F.D., X.L., M.S., S.D., A.R.), Massachusetts General Hospital, Boston. Institute for Heart and Brain Health, University of Michigan Medical Center, Ann Arbor (J.G., A.R.). Originally published28 Aug 2023https://doi.org/10.1161/CIRCRESAHA.123.322640Circulation Research. 2023;133:651–653Other version(s) of this articleYou are viewing the most recent version of this article. Previous versions: August 28, 2023: Ahead of Print Pathophysiological mechanisms underlying adverse cardiac remodeling after myocardial ischemia-reperfusion injury are incompletely understood. Although large infarct size and impaired function at the time of myocardial infarction (MI) are strong predictors of eventual heart failure, even patients with initially preserved function can experience late adverse remodeling that leads to heart failure.1 Here, we performed plasma proteomics in a subgroup of patients enrolled in the OMEGA-REMODEL trial2 presenting with acute ST-elevation MI and successfully reperfused with primary percutaneous coronary intervention. Adverse or favorable cardiac remodeling after MI was defined as a 20% increase or decrease, respectively, in left ventricular end-systolic volume index from 2 to 4 weeks to 6 months after MI as assessed by cardiac magnetic resonance imaging (MRI). We selected 11 adverse and 10 favorable remodelers who were matched for initial left ventricular mass, function, and infarct size, and shared similar demographic and clinical characteristics. Plasma samples taken at the time of the initial MRI were analyzed with the 1.3K SomaScan platform. The volcano plot (Figure [A]) shows the 14 candidate molecules (green dots) whose difference in abundance between favorable and adverse remodelers satisfied threshold values for significance (unadjusted P<0.05) and fold change (>1.4). The cytokine FAM3D (Family with sequence similarity 3D) was elevated in favorable remodelers and exhibited the highest overall fold change. This secreted factor binds to formyl-peptide receptors expressed predominantly on neutrophils and monocytes, and regulates their trafficking.3 ELISA for FAM3D closely correlated with SomaScan measurements and confirmed the difference between the 2 groups (Figure [B]).Download figureDownload PowerPointFigure. FAM3D (Family with sequence similarity 3D) is elevated in favorable remodelers and its overexpression confers cardioprotective effects. A, Stratification of post myocardial infarction (MI) remodelers by change in left ventricular end-systolic volume index (LVESVI) as assessed by serial cardiac magnetic resonance imaging (MRI). Volcano plot showing differentially expressed proteins between adverse and favorable remodelers. B, Correlations between aptamer-based SomaScan and ELISA (Abnova) measurements of FAM3D and between plasma FAM3D and troponin-I levels 8 hours after cardiac reperfusion. C, Western blots of murine heart and splenic FAM3D 24 hours after cardiac ischemia-reperfusion injury (IRI), with quantification of splenic FAM3D protein (normalized to vinculin, Image Lab software, Biorad) and RNA; n=4 mice per group. P-values were calculated with Mann-Whitney U test. D, Mice were injected with either adenoviral GFP (green fluorescent protein; n=6) or FAM3D (n=12; 1e12 ifu/mouse) 1 week before IRI, or vehicle (n=4) or recombinant FAM3D (0.4 μg/gBW, n=4) 30 minutes before IRI. Heart sections collected after 24 hours of reperfusion were quantified for areas of underperfusion lacking fluorescent microspheres (area at risk [AAR], grayscale images) and infarct areas (red 2,3,5-triphenyltetrazolium chloride-stained images). Hearts collected after 12 hours of reperfusion were stained for neutrophils in red (anti-mouse Gr-1 [Ly-6G/Ly-6C]); blue=DAPI; turquoise=mouse alpha-actinin (Leica LAS X, ImageJ software). n=5 per group; scale bars=1 mm. E, Human peripheral neutrophils were treated with 100 nM serum amyloid A and 100 nM recombinant FAM3D, either alone or in combination, for 3 hours. **P<0.01, ***P<0.001 using Kruskal–Wallis test. F, Fractional shortening and M-mode echocardiographs (GE VividE90) from mice injected with recombinant FAM3D (1 μg/gBW IP, daily for 3 days) and adenoviral FAM3D 24 hours after reperfusion; n=8 per group.Based on our human findings, we looked at FAM3D (or Oit1) in adult mice after 30 minutes of left anterior descending coronary artery ligation followed by various periods of reperfusion. Plasma FAM3D levels after 8 hours of reperfusion correlated with troponin-I levels (Figure [B]). Comprehensive organ harvest 24 hours after reperfusion revealed the spleen as a likely source of increased circulating FAM3D (Figure [C]), consistent with prior studies showing cross-talk between ischemic myocardium and hematopoietic tissues,4 although the difference in protein did not reach statistical significance.We tested the effects of FAM3D overexpression both before and after ischemia-reperfusion injury. Mice were injected with adenovirus encoding CMV-driven FAM3D (1e12 ifu/mouse), which is taken up by multiple organs and achieves FAM3D plasma levels (≈1 nM) about 3-fold higher than our human favorable remodelers, 1 week before ischemia-reperfusion injury. They showed dramatically reduced infarcts 24 hours after reperfusion despite identical areas of underperfusion as control animals (area at risk) (Figure [D], top). Infarct reduction was also observed in mice receiving recombinant FAM3D 30 minutes before ischemia-reperfusion injury (Figure [D], middle). Neutrophils constitute the initial inflammatory response to reperfusion injury and FAM3D overexpression significantly decreased their myocardial infiltration 12 hours after reperfusion (Figure [D], bottom). Neutrophils treated in vitro with SAA (serum amyloid A), an acute phase reactant elevated in MI that binds to formyl-peptide receptor,5 displayed a marked induction of proinflammatory cytokines, which trended downwards with coadministered FAM3D (Figure [E]). Finally, to assess the efficacy of a more clinically relevant intervention, mice were injected with FAM3D 24 hours after reperfusion. While initial fractional shortening was similarly impaired in both groups, FAM3D enhanced cardiac functional recovery by 4 weeks (Figure [F]). Taken together, these data show that FAM3D plays an important role in limiting cardiac injury, curbing inflammation, and promoting recovery of heart function after ischemic injury.ARTICLE INFORMATIONData AvailabilityAll methods and reagents are available on request. Proteomic datasets, analysis, and human study details can be found at Zenodo.org (https://zenodo.org/record/8132248). All animal studies (14- to 16-week-old male C57BL/6J mice were purchased from Jackson Laboratory) complied with the National Institutes of Health Guide for the Care and Use of Laboratory Animals and were approved by the MGH Animal Care and Use Committee. All surgeries and analyses were performed by investigators blinded to treatment. For SOMAscan, median normalized relative abundances of the 1305 analytes were imported into R (version 3.3.2) using the SomaDataIO package, and were analyzed using empirical Bayesian Analysis (LIMMA, Bioconductor package) that accounted for distribution of global protein expression, outliers, and likely false positives. Data are presented as mean±SD and analyzed using GraphPad Prism 8. P<0.05 considered significant using Student t test unless otherwise indicated.Sources of FundingThis work was supported by grants from the National Institutes of Health ([NIH]; A. Rosenzweig [R01AG061034, R35HL155318], J. Rhee [K08HL140200]), the American Heart Association (A. Rosenzweig [AHA MERIT Award]), and the Foundation for Anesthesia Education and Research (J. Rhee [FAER Mentored Research Training Grant]).Nonstandard Abbreviations and AcronymsFAM3DFamily with sequence similarity 3DMImyocardial infarctionSAAserum amyloid ADisclosures None.FootnotesFor Sources of Funding and Disclosures, see page 653.Correspondence to: James Rhee, MD, PhD, Massachusetts General Hospital, Boston, MA, Email jrhee@partners.orgAnthony Rosenzweig, MD, University of Michigan Medical Center, Ann Arbor, MI, Email anthros@med.umich.eduREFERENCES1. Schachinger V, Assmus B, Erbs S, Elsasser A, Haberbosch W, Hambrecht R, Yu J, Corti R, Mathey DG, Hamm CW, et al; REPAIR-AMI investigators. 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Authors of the article cited in the comment will be invited to reply, as appropriate.Comments and feedback on AHA/ASA Scientific Statements and Guidelines should be directed to the AHA/ASA Manuscript Oversight Committee via its Correspondence page.Sign In to Submit a Response to This Article Previous Back to top Next FiguresReferencesRelatedDetails September 15, 2023Vol 133, Issue 7 Advertisement Article InformationMetrics © 2023 American Heart Association, Inc.https://doi.org/10.1161/CIRCRESAHA.123.322640PMID: 37638415 Originally publishedAugust 28, 2023 Keywordsformyl peptide receptorheart attackheart failureinflammationischemia-reperfusion injurymyocardial infarctionventricular remodelingPDF download Advertisement SubjectsBiomarkersHeart FailureInflammationMyocardial InfarctionRemodeling