Altering Sphingolipid Metabolism Attenuates Cell Death and Inflammatory Response After Myocardial Infarction

Background: Sphingolipids have recently emerged as a biomarker of recurrence and mortality after myocardial infarction (MI). The increased ceramide levels in mammalian heart tissues during acute MI, as demonstrated by several groups, is associated with higher cell death rates in the left ventricle and deteriorated cardiac function. Ceramidase, the only enzyme known to hydrolyze proapoptotic ceramide, generates sphingosine, which is then phosphorylated by sphingosine kinase to produce the prosurvival molecule sphingosine-1-phosphate. We hypothesized that Acid Ceramidase (AC) overexpression would counteract the negative effects of elevated ceramide and promote cell survival, thereby providing cardioprotection after MI. Methods: We performed transcriptomic, sphingolipid, and protein analyses to evaluate sphingolipid metabolism and signaling post-MI. We investigated the effect of altering ceramide metabolism through a loss (chemical inhibitors) or gain (modified mRNA [modRNA]) of AC function post hypoxia or MI. Results: We found that several genes involved in de novo ceramide synthesis were upregulated and that ceramide (C16, C20, C20:1, and C24) levels had significantly increased 24 hours after MI. AC inhibition after hypoxia or MI resulted in reduced AC activity and increased cell death. By contrast, enhancing AC activity via AC modRNA treatment increased cell survival after hypoxia or MI. AC modRNA-treated mice had significantly better heart function, longer survival, and smaller scar size than control mice 28 days post-MI. We attributed the improvement in heart function post-MI after AC modRNA delivery to decreased ceramide levels, lower cell death rates, and changes in the composition of the immune cell population in the left ventricle manifested by lowered abundance of proinflammatory detrimental neutrophils. Conclusions: Our findings suggest that transiently altering sphingolipid metabolism through AC overexpression is sufficient and necessary to induce cardioprotection post-MI, thereby highlighting the therapeutic potential of AC modRNA in ischemic heart disease.

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Abstract 58: AT2 Receptor Agonism Regulates TIMP1/MMP9 Axis in the Heart Preventing Cardiac Fibrosis and Improving Heart Function After Experimental Myocardial Infarction

Hypertension ◽

10.1161/hyp.62.suppl_1.a58 ◽

2013 ◽

Vol 62 (suppl_1) ◽

Author(s):

Dilyara Lauer ◽

Svetlana Slavic ◽

Manuela Sommerfeld ◽

Christa Thöne-Reineke ◽

Yuliya Sharkovska ◽

...

Keyword(s):

Myocardial Infarction ◽

Left Ventricle ◽

Late Stage ◽

Heart Function ◽

Cardiac Fibroblasts ◽

Collagen Content ◽

Vehicle Group ◽

At2 Receptor ◽

Down Regulation ◽

Tgf Β1

Aims: A selective nonpeptide agonist for the angiotensin AT2 receptor compound 21 (C21) improved cardiac functions 7 days after myocardial infarction (MI). Here, we aimed to investigate what are the cellular mechanisms underlying cardiac protection in the late stage after MI. Methods and Results: MI was induced in Wistar rats by permanent ligation of the left coronary artery. Treatment with C21 (0.03mg/kg i.p. daily) started 6h after MI and continued for 6 weeks. Hemodynamic parameters were measured via transthoracic Doppler echocardiography and intracardiac Samba catheter. The expression of MMP9, TIMP1, TGF-β1 and collagen content were determined in left ventricle. Anti-proteolytic effects were additionally studied in primary cardiac fibroblasts. C21 significantly improved systolic and diastolic function 6 weeks after MI in comparison with the vehicle group as shown by ejection fraction (71.2±4.7 % vs. 53.4±7.0%; p<0.001), fractional shortening (40.8±2.3% vs. 30.9±3.1%; p<0.05), LVIDs (4.4±0.5mm vs. 5.2±0.8mm; p<0.05), LV EDP (16.9±1.2mmHg vs. 22.1±1.4mmHg; p<0.05), E/A ratio, dP/dt max and dP/dt min (p<0.05). Moreover, C21 improved arterial stiffness parameter (AIx) (18±1.2% vs. 25%±1.8, p<0.05) and reduced collagen content (15%; p<0.05) in postinfarcted myocardium. TIMP1 protein expression in the left ventricle was strongly up-regulated (17.7-fold; p<0.05) whereas MMP9 and TGF-β1 were significantly down-regulated (1.5-fold, p<0.05; 3.4-fold p<0.001, respectively) in the treated group. In cardiac fibroblasts, C21 primarily induced TIMP1 expression followed by attenuated MMP9 secretion and TGF-β1 down-regulation. Conclusion: C21 improves heart function in the late stage after MI and prevents cardiac remodeling. Activation of TIMP1 and subsequent inhibition of MMP9-mediated proteolysis as well as down-regulation of TGF-β1 followed by decreased collagen accumulation may attenuate disintegration of the extracellular matrix and reduce fibrosis.

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Abstract 17357: Direct Cardiac Reprogramming Using Combinatorial Modified mRNA

Circulation ◽

10.1161/circ.142.suppl_3.17357 ◽

2020 ◽

Vol 142 (Suppl_3) ◽

Author(s):

Keerat Kaur ◽

Asharee Mahmoud ◽

Hanna Girard ◽

Ann Anu Kurian ◽

Magdalena Zak ◽

...

Keyword(s):

Myocardial Infarction ◽

Gene Delivery ◽

High Efficiency ◽

De Novo ◽

Heart Function ◽

Lineage Tracing ◽

Dominant Negative ◽

Post Myocardial Infarction ◽

Transduction Efficiency

Introduction: Despite various clinical modalities, ischemic heart disease remains among the leading causes of mortality and morbidity worldwide. The elemental problem is the immense loss of cardiomyocytes (CMs) post-myocardial infarction (MI). Reprogramming non- cardiomyocytes (non-CMs) into cardiomyocyte (CM)-like cells in vivo is a promising strategy for cardiac regeneration: however, the traditional viral delivery method hampered its application into clinical settings due to low and erratic transduction efficiency. Methods: We used a modified mRNA (modRNA) gene delivery platform to deliver different stoichiometry of cardiac-reprogramming genes (Gata4, Mef2c, Tbx5 and Hand2) together with reprogramming helper genes (Dominant Negative (DN)-TGFβ, DN- Wnt8a and Acid ceramidase (AC)), named 7G, to induce direct cardiac reprogramming post myocardial infarction (MI). Results: Here, we identified 7G modRNA cocktail as an important regulator ofthe cardiac reprogramming. Cardiac transfection with 7G modRNA doubled cardiac reprogramming efficiency (57%) in comparison to Gata4, Mef2C and Tbx5 (GMT) alone (28%) in vitro . By inducing MI in our lineage tracing model, we showed that one-time delivery of the 7G-modRNA cocktail reprogrammed ~25% of the non-CMs in the scar area to CM- like cells. Furthermore, 7G modRNA treated mice showed significantly improved cardiac function, longer survival, reduced scar size and greater capillary density than control mice 28 days post-MI. We attributed the improvement in heart function post modRNA delivery of 7G or 7G with increased Hand2 ratio (7G-GMT Hx2) to significant upregulation of 15 key angiogenic factors without any signs of angioma or edema. Conclusions: 7G or 7G GMT HX2 modRNA cocktails boosts de novo CM-like cells and promotes cardiovascular regeneration post-MI. Thus, we highlight that this gene delivery approach not only has high efficiency but also high margin of safety for translation to clinics.

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P2572Human iPSC-derived cardiomyocytes preserve murine heart function after myocardial infarction unlike adipose tissue-derived stromal cells

European Heart Journal ◽

10.1093/eurheartj/ehz748.0899 ◽

2019 ◽

Vol 40 (Supplement_1) ◽

Author(s):

J Dulak ◽

J Stepniewski ◽

M Tomczyk ◽

K Andrysiak ◽

I Kraszewska ◽

...

Keyword(s):

Myocardial Infarction ◽

Adipose Tissue ◽

Stromal Cells ◽

Luciferase Activity ◽

Therapeutic Potential ◽

Heart Function ◽

Rational Approach ◽

Acute Mi ◽

In Vivo Administration

Abstract Introduction Despite progress in pharmacological treatment of myocardial infarction (MI), there is still an immense need for novel therapies for this life-threatening condition. Accordingly, cell-based therapies have been extensively investigated with most studies focusing on mesenchymal stromal cells. However due to their inability to differentiate into cardiomyocytes as well as limited survival upon in vivo administration, no effective treatment of MI has been developed. In contrast, application of hiPSC-derived cardiomyocytes (hiPSC-CM) represent biologically rational approach with pre-clinical studies confirming their therapeutic potential in various models of MI. However further optimization is required due to limited survival of hiPSC-CM upon in vivo administration. Therefore, we evaluated the therapeutic potential of genetically modified hiPSC-CM in murine model of acute MI and compared it to the effect of adipose tissue-derived stromal cells (ADSC). Methods In the first step hiPSC overexpressing GFP, luciferase (Luc) and pro-angiogenic and cardioprotective factors: heme oxygenase-1 (HO-1, heme degrading enzyme) or stromal cell-derived factor-1 (SDF-1, pro-angiogenic chemokine) were subjected to cardiac differentiation which yielded in each group 70–90% cardiac troponin T-positive contracting cells. hiPSC-CM (5x105 in 10 μl) were administered into NOD-SCID mice which underwent permanent ligation of left anterior descending (LAD) coronary artery. The cells were injected into the peri-infarct zone. Mice subjected to sham operation as well as injected with saline after MI were used as controls. The ultrasonography of hearts was performed on day 7, 14, 28 and 42 whereas the presence of hiPSC-CM was monitored using IVIS Spectrum system upon administration of luciferin and analysed in sections of collected hearts. The same experimental scheme was used to assess therapeutic potential of ADSC (CD105+CD73+CD90+CD44+CD146-CD34-) overexpressing luciferase and GFP. Results Ultrasonography demonstrated that upon delivery of hiPSC-CM the left ventricle ejection fraction (LVEF) was very significantly higher in comparison to control group injected with saline after induction of MI. In contrast, no improvement of LVEF was observed after administration of ADSC. Interestingly, measurements of luciferase activity revealed the strongest bioluminescent signal in the hearts of mice transplanted with iPSC-CM-HO1 42 days after MI. Importantly, the survival of hiPSC-CM in murine myocardium six weeks upon administration was further confirmed with immunofluorescent analysis of heart sections using human specific anti-Ku80 antibody. Again, luciferase activity was not observed upon delivery of ADSC. Conclusion These results strongly indicate that administration of hiPSC-CM, unlike ADSC, preserve murine heart function in acute MI model. Additionally, overexpression of HO-1 may positively influence their survival upon in vivo delivery into infarcted tissue. Acknowledgement/Funding The National Centre for Research and Development (STRATEGMED 2/269415/11/NCBR/2015), National Science Centre of Poland (HARMONIA 2014/14/M/NZ1/00010)

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Inhibition of Myocardial Ischemia/Reperfusion Injury by Exosomes Secreted from Mesenchymal Stem Cells

Stem Cells International ◽

10.1155/2016/4328362 ◽

2016 ◽

Vol 2016 ◽

pp. 1-8 ◽

Cited By ~ 18

Author(s):

Heng Zhang ◽

Meng Xiang ◽

Dan Meng ◽

Ning Sun ◽

Sifeng Chen

Keyword(s):

Stem Cells ◽

Mesenchymal Stem Cells ◽

Left Ventricle ◽

Reperfusion Injury ◽

Ischemia Reperfusion Injury ◽

Therapeutic Potential ◽

Heart Function ◽

Ischemia Reperfusion ◽

Left Ventricular ◽

Area At Risk

Exosomes secreted by mesenchymal stem cells have shown great therapeutic potential in regenerative medicine. In this study, we performed meta-analysis to assess the clinical effectiveness of using exosomes in ischemia/reperfusion injury based on the reports published between January 2000 and September 2015 and indexed in the PUBMED and Web of Science databases. The effect of exosomes on heart function was evaluated according to the following parameters: the area at risk as a percentage of the left ventricle, infarct size as a percentage of the area at risk, infarct size as a percentage of the left ventricle, left ventricular ejection fraction, left ventricular fraction shortening, end-diastolic volume, and end-systolic volume. Our analysis indicated that the currently available evidence confirmed the therapeutic potential of mesenchymal stem cell-secreted exosomes in the improvement of heart function. However, further mechanistic studies, therapeutic safety, and clinical trials are required for optimization and validation of this approach to cardiac regeneration after ischemia/reperfusion injury.

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Ceramide synthases at the centre of sphingolipid metabolism and biology

Biochemical Journal ◽

10.1042/bj20111626 ◽

2012 ◽

Vol 441 (3) ◽

pp. 789-802 ◽

Cited By ~ 250

Author(s):

Thomas D. Mullen ◽

Yusuf A. Hannun ◽

Lina M. Obeid

Keyword(s):

Cell Death ◽

De Novo ◽

Acyl Chain ◽

Sphingolipid Metabolism ◽

Sphingoid Base ◽

Salvage Pathway ◽

Organismal Biology ◽

Acyl Chain Length ◽

Specific Manner ◽

Ceramide Synthases

Sphingolipid metabolism in metazoan cells consists of a complex interconnected web of numerous enzymes, metabolites and modes of regulation. At the centre of sphingolipid metabolism reside CerSs (ceramide synthases), a group of enzymes that catalyse the formation of ceramides from sphingoid base and acyl-CoA substrates. From a metabolic perspective, these enzymes occupy a unique niche in that they simultaneously regulate de novo sphingolipid synthesis and the recycling of free sphingosine produced from the degradation of pre-formed sphingolipids (salvage pathway). Six mammalian CerSs (CerS1–CerS6) have been identified. Unique characteristics have been described for each of these enzymes, but perhaps the most notable is the ability of individual CerS isoforms to produce ceramides with characteristic acyl-chain distributions. Through this control of acyl-chain length and perhaps in a compartment-specific manner, CerSs appear to regulate multiple aspects of sphingolipid-mediated cell and organismal biology. In the present review, we discuss the function of CerSs as critical regulators of sphingolipid metabolism, highlight their unique characteristics and explore the emerging roles of CerSs in regulating programmed cell death, cancer and many other aspects of biology.

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Resveratrol Targets Sphingolipid Metabolism to Induce Growth Inhibition in FLT3 ITD Acute Myeloid Leukemia

Proceedings ◽

10.3390/proceedings2019040004 ◽

2019 ◽

Vol 40 (1) ◽

pp. 4

Author(s):

Ersöz ◽

Adan

Keyword(s):

Growth Inhibition ◽

Cell Fate ◽

De Novo ◽

Therapeutic Potential ◽

Annexin V ◽

Sphingolipid Metabolism ◽

Ceramide Synthase ◽

Cytotoxic Effects ◽

Sphingosine 1 Phosphate ◽

First Time

Sphingolipids are important signaling lipids which play crucial roles to determine the cell fate. Ceramide, apoptotic central molecule of sphingolipid metabolism, which is produced through de novo pathway by serine palmitoyl transferase (SPT) and can be converted to antiapoptotic sphingosine-1-phosphate (S1P) and glucosyl ceramide (GC) by sphingosine kinase (SK) and glucosyl ceramide synthase (GCS), respectively. It is aimed to investigate therapeutic potential of resveratrol on FLT3-ITD (Internal Tandem Duplication) AML cells and to identify potential mechanism behind resveratrol-mediated growth inhibition by targeting of ceramide metabolism. The cytotoxic effects of resveratrol, SPT inhibitor (myricoin), SK-1 inhibitor (SKI II), GCS inhibitor (PDMP), resveratrol: SPT inhibitor, resveratrol: SK-1 inhibitor and resveratrol: GCS inhibitor combinations on MOLM-13 and MV4-11 FLT3 ITD AML cells were investigated by cell proliferation assay. Apoptosis was evaluated by annexin V/PI double staining. There were synergistic cytotoxic effects of resveratrol with co-administration of SPT inhibitor, SK-1 inhibitor and GCS inhibitor and apoptosis was synergistically induced for resveratrol and its combinations. This preliminary data showed for the first time that resveratrol might inhibit the growth of FLT3 ITD AML cells through targeting ceramide metabolism.

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Thrombin inhibits Bim (Bcl-2-interacting mediator of cell death) expression and prevents serum-withdrawal-induced apoptosis via protease-activated receptor 1

Biochemical Journal ◽

10.1042/bj20030346 ◽

2003 ◽

Vol 375 (1) ◽

pp. 99-109 ◽

Cited By ~ 33

Author(s):

Claire J. CHALMERS ◽

Kathryn BALMANNO ◽

Kathryn HADFIELD ◽

Rebecca LEY ◽

Simon J. COOK

Keyword(s):

Cell Death ◽

Cell Survival ◽

De Novo ◽

Fibroblast Cell ◽

Mitogen Activated Protein Kinase ◽

Serum Withdrawal ◽

Mitogen Activated Protein ◽

Protease Activated Receptor 1 ◽

Induced Apoptosis ◽

Ccl39 Cells

To investigate the role of thrombin in regulating apoptosis, we have used CCl39 cells, a fibroblast cell line in which thrombin-induced cell proliferation has been extensively studied. Withdrawal of serum from CCl39 cells resulted in a rapid apoptotic response that was completely prevented by the inclusion of thrombin. The protective effect of thrombin was reversed by pertussis toxin, suggesting that cell-survival signalling pathways are activated via a Gi or Go heterotrimeric GTPase. Serum-withdrawal-induced death required de novo gene expression and was preceded by the rapid de novo expression of the pro-apoptotic ‘BH3-only’ protein Bim (Bcl-2-interacting mediator of cell death). Thrombin strongly inhibited the up-regulation of both Bim protein and Bim mRNA. The ability of thrombin to repress Bim expression, and to protect cells from apoptosis, was reversed by U0126, a MEK1/2 [MAPK (mitogen-activated protein kinase) or ERK (extracellular-signal-regulated kinase) 1/2] inhibitor, or LY294002, a phosphoinositide 3′-kinase (PI3K) inhibitor, suggesting that both the Raf→MEK→ERK1/2 and PI3K pathways co-operate to repress Bim and promote cell survival. A PAR1p (protease-activated receptor 1 agonist peptide) was also able to protect cells from serum-withdrawal-induced apoptosis, suggesting that thrombin acts via PAR1 to prevent apoptosis.

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Considering Cause and Effect of Immune Cell Aging on Cardiac Repair after Myocardial Infarction

Cells ◽

10.3390/cells9081894 ◽

2020 ◽

Vol 9 (8) ◽

pp. 1894 ◽

Cited By ~ 1

Author(s):

Stephanie W. Tobin ◽

Faisal J. Alibhai ◽

Richard D. Weisel ◽

Ren-Ke Li

Keyword(s):

Myocardial Infarction ◽

Immune System ◽

Immune Cell ◽

Heart Function ◽

Cardiac Repair ◽

Cell Aging ◽

Cell Behavior ◽

Complex Nature ◽

Older Individuals ◽

Hematopoietic Stem

The importance of the immune system for cardiac repair following myocardial infarction is undeniable; however, the complex nature of immune cell behavior has limited the ability to develop effective therapeutics. This limitation highlights the need for a better understanding of the function of each immune cell population during the inflammatory and resolution phases of cardiac repair. The development of reliable therapies is further complicated by aging, which is associated with a decline in cell and organ function and the onset of cardiovascular and immunological diseases. Aging of the immune system has important consequences on heart function as both chronic cardiac inflammation and an impaired immune response to cardiac injury are observed in older individuals. Several studies have suggested that rejuvenating the aged immune system may be a valid therapeutic candidate to prevent or treat heart disease. Here, we review the basic patterns of immune cell behavior after myocardial infarction and discuss the autonomous and nonautonomous manners of hematopoietic stem cell and immune cell aging. Lastly, we identify prospective therapies that may rejuvenate the aged immune system to improve heart function such as anti-inflammatory and senolytic therapies, bone marrow transplant, niche remodeling and regulation of immune cell differentiation.

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Biomechanical assessment of remote and postinfarction scar remodeling following myocardial infarction

Scientific Reports ◽

10.1038/s41598-019-53351-7 ◽

2019 ◽

Vol 9 (1) ◽

Cited By ~ 1

Author(s):

Mihaela Rusu ◽

Katrin Hilse ◽

Alexander Schuh ◽

Lukas Martin ◽

Ioana Slabu ◽

...

Keyword(s):

Myocardial Infarction ◽

Heart Failure ◽

Type I Collagen ◽

De Novo ◽

Heart Function ◽

Type Iii Collagen ◽

Type I ◽

Compensatory Mechanisms ◽

Biomechanical Assessment ◽

Type V

AbstractThe importance of collagen remodeling following myocardial infarction (MI) is extensively investigated, but little is known on the biomechanical impact of fibrillar collagen on left ventricle post-MI. We aim to identify the significant effects of the biomechanics of types I, III, and V collagen on physio-pathological changes of murine hearts leading to heart failure. Immediately post-MI, heart reduces its function (EF = 40.94 ± 2.12%) while sarcomeres’ dimensions are unchanged. Strikingly, as determined by immunohistochemistry staining, type V collagen fraction significantly grows in remote and scar for sustaining de novo-types I and III collagen fibers’ assembly while hindering their enzymatic degradation. Thereafter, the compensatory heart function (EF = 63.04 ± 3.16%) associates with steady development of types I and III collagen in a stiff remote (12.79 ± 1.09 MPa) and scar (22.40 ± 1.08 MPa). In remote, the soft de novo-type III collagen uncoils preventing further expansion of elongated sarcomeres (2.7 ± 0.3 mm). Once the compensatory mechanisms are surpassed, the increased turnover of stiff type I collagen (>50%) lead to a pseudo-stable biomechanical regime of the heart (≅9 MPa) with reduced EF (50.55 ± 3.25%). These end-characteristics represent the common scenario evidenced in patients suffering from heart failure after MI. Our pre-clinical data advances the understanding of the cause of heart failure induced in patients with extended MI.

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Role of Prokineticin Receptor-1 in Epicardial Progenitor Cells

Journal of Developmental Biology ◽

10.3390/jdb1010020 ◽

2013 ◽

Vol 1 (1) ◽

pp. 20-31 ◽

Cited By ~ 4

Author(s):

Thu Nguyen ◽

Adelin Gasser ◽

Canan Nebigil

Keyword(s):

Myocardial Infarction ◽

Cell Fate ◽

De Novo ◽

Heart Function ◽

Mouse Heart ◽

Stem Cell Maintenance ◽

Prokineticin 2 ◽

Tm Domain ◽

Definition Of

G protein-coupled receptors (GPCRs) form a large class of seven transmembrane (TM) domain receptors. The use of endogenous GPCR ligands to activate the stem cell maintenance or to direct cell differentiation would overcome many of the problems currently encountered in the use of stem cells, such as rapid in vitro differentiation and expansion or rejection in clinical applications. This review focuses on the definition of a new GPCR signaling pathway activated by peptide hormones, called “prokineticins”, in epicardium-derived cells (EPDCs). Signaling via prokineticin-2 and its receptor, PKR1, is required for cardiomyocyte survival during hypoxic stress. The binding of prokineticin-2 to PKR1 induces proliferation, migration and angiogenesis in endothelial cells. The expression of prokineticin and PKR1 increases during cardiac remodeling after myocardial infarction. Gain of function of PKR1 in the adult mouse heart revealed that cardiomyocyte-PKR1 signaling activates EPDCs in a paracrine fashion, thereby promoting de novo vasculogenesis. Transient PKR1 gene therapy after myocardial infarction in mice decreases mortality and improves heart function by promoting neovascularization, protecting cardiomyocytes and mobilizing WT1+ cells. Furthermore, PKR1 signaling promotes adult EPDC proliferation and differentiation to adopt endothelial and smooth muscle cell fate, for the induction of de novo vasculogenesis. PKR1 is expressed in the proepicardium and epicardial cells derived from mice kidneys. Loss of PKR1 causes deficits in EPDCs in the neonatal mice hearts and kidneys and impairs vascularization and heart and kidney function. Taken together, these data indicate a novel role for PKR1 in heart-kidney complex via EPDCs.

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