Tipifarnib Reduces Extracellular Vesicles and Protects From Heart Failure.

BACKGROUND: Heart failure (HF) is one of the leading causes of mortality worldwide. Extracellular vesicles, including small extracellular vesicles or exosomes, and their molecular cargo are known to modulate cell-to-cell communication during multiple cardiac diseases. However, the role of systemic extracellular vesicle biogenesis inhibition in HF models is not well documented and remains unclear. METHODS: We investigated the role of circulating exosomes during cardiac dysfunction and remodeling in a mouse transverse aortic constriction (TAC) model of HF. Importantly, we investigate the efficacy of tipifarnib, a recently identified exosome biogenesis inhibitor that targets the critical proteins (Rab27a, nSMase2 [neutral sphingomyelinase 2], and Alix [ALG-2-interacting protein X]) involved in exosome biogenesis for this mouse model of HF. In this study, 10-week-old male mice underwent TAC surgery were randomly assigned to groups with and without tipifarnib treatment (10 mg/kg 3 times/wk) and monitored for 8 weeks, and a comprehensive assessment was conducted through performed echocardiographic, histological, and biochemical studies. RESULTS: TAC significantly elevated circulating plasma exosomes and markedly increased cardiac left ventricular dysfunction, cardiac hypertrophy, and fibrosis. Furthermore, injection of plasma exosomes from TAC mice induced left ventricular dysfunction and cardiomyocyte hypertrophy in uninjured mice without TAC. On the contrary, treatment of tipifarnib in TAC mice reduced circulating exosomes to baseline and remarkably improved left ventricular functions, hypertrophy, and fibrosis. Tipifarnib treatment also drastically altered the miRNA profile of circulating post-TAC exosomes, including miR 331-5p, which was highly downregulated both in TAC circulating exosomes and in TAC cardiac tissue. Mechanistically, miR 331-5p is crucial for inhibiting the fibroblast-to-myofibroblast transition by targeting HOXC8, a critical regulator of fibrosis. Tipifarnib treatment in TAC mice upregulated the expression of miR 331-5p that acts as a potent repressor for one of the fibrotic mechanisms mediated by HOXC8. CONCLUSIONS: Our study underscores the pathological role of exosomes in HF and fibrosis in response to pressure overload. Tipifarnib-mediated inhibition of exosome biogenesis and cargo sorting may serve as a viable strategy to prevent progressive cardiac remodeling in HF.

Epigenetic mechanisms regulate sex differences in cardiac reparative functions of bone marrow progenitor cells.

Historically, a lower incidence of cardiovascular diseases (CVD) and related deaths in women as compared with men of the same age has been attributed to female sex hormones, particularly estrogen and its receptors. Autologous bone marrow stem cell (BMSC) clinical trials for cardiac cell therapy overwhelmingly included male patients. However, meta-analysis data from these trials suggest a better functional outcome in postmenopausal women as compared with aged-matched men. Mechanisms governing sex-specific cardiac reparative activity in BMSCs, with and without the influence of sex hormones, remain unexplored. To discover these mechanisms, Male (M), female (F), and ovariectomized female (OVX) mice-derived EPCs were subjected to a series of molecular and epigenetic analyses followed by in vivo functional assessments of cardiac repair. F-EPCs and OVX EPCs show a lower inflammatory profile and promote enhanced cardiac reparative activity after intra-cardiac injections in a male mouse model of myocardial infarction (MI). Epigenetic sequencing revealed a marked difference in the occupancy of the gene repressive H3K9me3 mark, particularly at transcription start sites of key angiogenic and proinflammatory genes in M-EPCs compared with F-EPCs and OVX-EPCs. Our study unveiled that functional sex differences in EPCs are, in part, mediated by differential epigenetic regulation of the proinflammatory and anti-angiogenic gene CCL3, orchestrated by the control of H3K9me3 by histone methyltransferase, G9a/Ehmt2. Our research highlights the importance of considering the sex of donor cells for progenitor-based tissue repair.

Cell-Specific mRNA Therapeutics for Cardiovascular Diseases and Regeneration.

Cardiovascular diseases (CVDs) represent a significant global health burden, demanding innovative therapeutic approaches. In recent years, mRNA therapeutics have emerged as a promising strategy to combat CVDs effectively. Unlike conventional small-molecule drugs, mRNA therapeutics enable the direct modulation of cellular functions by delivering specific mRNA molecules to target cells. This approach offers unprecedented advantages, including the ability to harness endogenous cellular machinery for protein synthesis, thus allowing precise control over gene expression without insertion into the genome. This review summarizes the current status of the potential of cell-specific mRNA therapeutics in the context of cardiovascular diseases. First, it outlines the challenges associated with traditional CVD treatments and emphasizes the need for targeted therapies. Subsequently, it elucidates the underlying principles of mRNA therapeutics and the development of advanced delivery systems to ensure cell-specificity and enhanced efficacy. Notably, innovative delivery methods such as lipid nanoparticles and exosomes have shown promise in improving the targeted delivery of mRNA to cardiac cells, activated fibroblasts, and other relevant cell types. Furthermore, the review highlights the diverse applications of cell-specific mRNA therapeutics in addressing various aspects of cardiovascular diseases, including atherosclerosis, myocardial infarction, heart failure, and arrhythmias. By modulating key regulatory genes involved in cardiomyocyte proliferation, inflammation, angiogenesis, tissue repair, and cell survival, mRNA therapeutics hold the potential to intervene at multiple stages of CVD pathogenesis. Despite its immense potential, this abstract acknowledges the challenges in translating cell-specific mRNA therapeutics from preclinical studies to clinical applications like off-target effects and delivery. In conclusion, cell-specific mRNA therapeutics have emerged as a revolutionary gene therapy approach for CVD, offering targeted interventions with the potential to significantly improve patient outcomes.

Lin28a cardiomyocyte-specific modified mRNA translation system induces cardiomyocyte cell division and cardiac repair.

The mammalian heart has a limited regenerative capacity. Previous work suggested the heart can regenerate during development and immediately after birth by inducing cardiomyocyte (CM) proliferation; however, this capacity is lost seven days after birth. modRNA gene delivery, the same technology used successfully in the two mRNA vaccines against SARS-CoV-2, can prompt cardiac regeneration, cardiovascular regeneration and cardiac protection. We recently established a novel CM-specific modRNA translational system (SMRTs) that allows modRNA translation only in CMs. We demonstrated that this system delivers potent intracellular genes (e.g., cell cyclepromoting Pkm2), which are beneficial when expressed in one cell type (i.e., CMs) but not others (non-CMs). Here, we identify Lin28a as an important regulator of the CM cell cycle. We show that Lin28a is expressed in CMs during development and immediately after birth, but not during adulthood. We describe that specific delivery of Lin28a into CM, using CM SMRTs, enables CM cell division and proliferation. Further, we determine that this proliferation leads to cardiac repair and better outcome post MI. Moreover, we identify the molecular pathway of Lin28a in CMs. We also demonstrate that Lin28a suppress Let-7 which is vital for CM proliferation, partially due to its suppressive role on cMYC, HMGA2 and K-RAS.

Modified mRNA Therapeutics for Heart Diseases.

Cardiovascular diseases (CVD) remain a substantial global health problem and the leading cause of death worldwide. Although many conventional small-molecule treatments are available to support the cardiac function of the patient with CVD, they are not effective as a cure. Among potential targets for gene therapy are severe cardiac and peripheral ischemia, heart failure, vein graft failure, and some forms of dyslipidemias. In the last three decades, multiple gene therapy tools have been used for heart diseases caused by proteins, plasmids, adenovirus, and adeno-associated viruses (AAV), but these remain as unmet clinical needs. These gene therapy methods are ineffective due to poor and uncontrolled gene expression, low stability, immunogenicity, and transfection efficiency. The synthetic modified mRNA (modRNA) presents a novel gene therapy approach which provides a transient, stable, safe, non-immunogenic, controlled mRNA delivery to the heart tissue without any risk of genomic integration, and achieves a therapeutic effect in different organs, including the heart. The mRNA translation starts in minutes, and remains stable for 8-10 days (pulse-like kinetics). The pulse-like expression of modRNA in the heart induces cardiac repair, cardiomyocyte proliferation and survival, and inhibits cardiomyocyte apoptosis post-myocardial infarction (MI). Cell-specific (cardiomyocyte) modRNA translation developments established cell-specific modRNA therapeutics for heart diseases. With these laudable characteristics, combined with its expression kinetics in the heart, modRNA has become an attractive therapeutic for the treatment of CVD. This review discusses new developments in modRNA therapy for heart diseases.

Live cell screening identifies glycosides as enhancers of cardiomyocyte cell cycle activity.

Promoting cardiomyocyte proliferation is a promising strategy to regenerate the heart. Yet, so far, it is poorly understood how cardiomyocyte proliferation is regulated, and no factor identified to promote mammalian cardiomyocyte proliferation has been translated into medical practice. Therefore, finding a novel factor will be vital. Here, we established a live cell screening based on mouse embryonic stem cell-derived cardiomyocytes expressing a non-functional human geminin deletion mutant fused to Azami Green (CM7/1-hgem-derived cardiomyocytes). We screened for a subset of compounds of the small molecule library Spectrum Collection and identified 19 potential inducers of stem cell-derived cardiomyocyte proliferation. Furthermore, the pro-proliferative potential of identified candidate compounds was validated in neonatal and adult rat cardiomyocytes as well as human induced pluripotent stem cell-derived cardiomyocytes. 18 of these compounds promoted mitosis and cytokinesis in neonatal rat cardiomyocytes. Among the top four candidates were two cardiac glycosides, peruvoside and convallatoxin, the flavonoid osajin, and the selective α-adrenoceptor antagonist and imidazoline I1 receptor ligand efaroxan hydrochloride. Inhibition of PTEN and GSK-3ß enhanced cell cycle re-entry and progression upon stimulation with cardiac glycosides and osajin, while inhibition of IP3 receptors inhibited the cell cycle-promoting effect of cardiac glycosides. Collectively, we established a screening system and identified potential compounds to promote cardiomyocyte proliferation. Our data suggest that modulation of calcium handling and metabolism promotes cardiomyocyte proliferation, and cardiac glycosides might, besides increasing myocardial contraction force, contribute to cardiac repair by inducing cardiomyocyte proliferation.

Diabetes impairs cardioprotective function of endothelial progenitor cell-derived extracellular vesicles via H3K9Ac inhibition.

Background and Purpose: Myocardial infarction (MI) in diabetic patients results in higher mortality and morbidity. We and others have previously shown that bone marrow-endothelial progenitor cells (EPCs) promote cardiac neovascularization and attenuate ischemic injury. Lately, small extracellular vesicles (EVs) have emerged as major paracrine effectors mediating the benefits of stem cell therapy. Modest clinical outcomes of autologous cell-based therapies suggest diabetes-induced EPC dysfunction and may also reflect their EV derivatives. Moreover, studies suggest that post-translational histone modifications promote diabetes-induced vascular dysfunctions. Therefore, we tested the hypothesis that diabetic EPC-EVs may lose their post-injury cardiac reparative function by modulating histone modification in endothelial cells (ECs). Methods: We collected EVs from the culture medium of EPCs isolated from non-diabetic (db/+) and diabetic (db/db) mice and examined their effects on recipient ECs and cardiomyocytes in vitro, and their reparative function in permanent ligation of left anterior descending (LAD) coronary artery and ischemia/reperfusion (I/R) myocardial ischemic injuries in vivo. Results: Compared to db/+ EPC-EVs, db/db EPC-EVs promoted EC and cardiomyocyte apoptosis and repressed tube-forming capacity of ECs. In vivo, db/db EPC-EVs depressed cardiac function, reduced capillary density, and increased fibrosis compared to db/+ EPC-EV treatments after MI. Moreover, in the I/R MI model, db/+ EPC-EV-mediated acute cardio-protection was lost with db/db EPC-EVs, and db/db EPC-EVs increased immune cell infiltration, infarct area, and plasma cardiac troponin-I. Mechanistically, histone 3 lysine 9 acetylation (H3K9Ac) was significantly decreased in cardiac ECs treated with db/db EPC-EVs compared to db/+ EPC-EVs. The H3K9Ac chromatin immunoprecipitation sequencing (ChIP-Seq) results further revealed that db/db EPC-EVs reduced H3K9Ac level on angiogenic, cell survival, and proliferative genes in cardiac ECs. We found that the histone deacetylase (HDAC) inhibitor, valproic acid (VPA), partly restored diabetic EPC-EV-impaired H3K9Ac levels, tube formation and viability of ECs, and enhanced cell survival and proliferative genes, Pdgfd and Sox12, expression. Moreover, we observed that VPA treatment improved db/db EPC-mediated post-MI cardiac repair and functions. Conclusions: Our findings unravel that diabetes impairs EPC-EV reparative function in the ischemic heart, at least partially, through HDACs-mediated H3K9Ac downregulation leading to transcriptional suppression of angiogenic, proliferative and cell survival genes in recipient cardiac ECs. Thus, HDAC inhibitors may potentially be used to restore the function of diabetic EPC and other stem cells for autologous cell therapy applications.

Direct reprogramming induces vascular regeneration post muscle ischemic injury.

Reprogramming non-cardiomyocytes (non-CMs) into cardiomyocyte (CM)-like cells is a promising strategy for cardiac regeneration in conditions such as ischemic heart disease. Here, we used a modified mRNA (modRNA) gene delivery platform to deliver a cocktail, termed 7G-modRNA, of four cardiac-reprogramming genes-Gata4 (G), Mef2c (M), Tbx5 (T), and Hand2 (H)-together with three reprogramming-helper genes-dominant-negative (DN)-TGFß, DN-Wnt8a, and acid ceramidase (AC)-to induce CM-like cells. We showed that 7G-modRNA reprogrammed 57% of CM-like cells in vitro. Through a lineage-tracing model, we determined that delivering the 7G-modRNA cocktail at the time of myocardial infarction reprogrammed ∼25% of CM-like cells in the scar area and significantly improved cardiac function, scar size, long-term survival, and capillary density. Mechanistically, we determined that while 7G-modRNA cannot create de novo beating CMs in vitro or in vivo, it can significantly upregulate pro-angiogenic mesenchymal stromal cells markers and transcription factors. We also demonstrated that our 7G-modRNA cocktail leads to neovascularization in ischemic-limb injury, indicating CM-like cells importance in other organs besides the heart. modRNA is currently being used around the globe for vaccination against COVID-19, and this study proves this is a safe, highly efficient gene delivery approach with therapeutic potential to treat ischemic diseases.

Therapeutic Delivery of Pip4k2c-Modified mRNA Attenuates Cardiac Hypertrophy and Fibrosis in the Failing Heart.

Heart failure (HF) remains a major cause of morbidity and mortality worldwide. One of the risk factors for HF is cardiac hypertrophy (CH), which is frequently accompanied by cardiac fibrosis (CF). CH and CF are controlled by master regulators mTORC1 and TGF-ß, respectively. Type-2-phosphatidylinositol-5-phosphate-4-kinase-gamma (Pip4k2c) is a known mTORC1 regulator. It is shown that Pip4k2c is significantly downregulated in the hearts of CH and HF patients as compared to non-injured hearts. The role of Pip4k2c in the heart during development and disease is unknown. It is shown that deleting Pip4k2c does not affect normal embryonic cardiac development; however, three weeks after TAC, adult Pip4k2c-/- mice has higher rates of CH, CF, and sudden death than wild-type mice. In a gain-of-function study using a TAC mouse model, Pip4k2c is transiently upregulated using a modified mRNA (modRNA) gene delivery platform, which significantly improve heart function, reverse CH and CF, and lead to increased survival. Mechanistically, it is shown that Pip4k2c inhibits TGFß1 via its N-terminal motif, Pip5k1α, phospho-AKT 1/2/3, and phospho-Smad3. In sum, loss-and-gain-of-function studies in a TAC mouse model are used to identify Pip4k2c as a potential therapeutic target for CF, CH, and HF, for which modRNA is a highly translatable gene therapy approach.

Cardiovascular Manifestations of COVID-19 Infection.

SARS-CoV-2 induced the novel coronavirus disease (COVID-19) outbreak, the most significant medical challenge in the last century. COVID-19 is associated with notable increases in morbidity and death worldwide. Preexisting conditions, like cardiovascular disease (CVD), diabetes, hypertension, and obesity, are correlated with higher severity and a significant increase in the fatality rate of COVID-19. COVID-19 induces multiple cardiovascular complexities, such as cardiac arrest, myocarditis, acute myocardial injury, stress-induced cardiomyopathy, cardiogenic shock, arrhythmias and, subsequently, heart failure (HF). The precise mechanisms of how SARS-CoV-2 may cause myocardial complications are not clearly understood. The proposed mechanisms of myocardial injury based on current knowledge are the direct viral entry of the virus and damage to the myocardium, systemic inflammation, hypoxia, cytokine storm, interferon-mediated immune response, and plaque destabilization. The virus enters the cell through the angiotensin-converting enzyme-2 (ACE2) receptor and plays a central function in the virus's pathogenesis. A systematic understanding of cardiovascular effects of SARS-CoV2 is needed to develop novel therapeutic tools to target the virus-induced cardiac damage as a potential strategy to minimize permanent damage to the cardiovascular system and reduce the morbidity. In this review, we discuss our current understanding of COVID-19 mediated damage to the cardiovascular system.

Delivery of Modified mRNA in a Myocardial Infarction Mouse Model.

Myocardial infarction (MI) is a leading cause of morbidity and mortality in the Western world. In the past decade, gene therapy has become a promising treatment option for heart disease, owing to its efficiency and exceptional therapeutic effects. In an effort to repair the damaged tissue post-MI, various studies have employed DNA-based or viral gene therapy but have faced considerable hurdles due to the poor and uncontrolled expression of the delivered genes, edema, arrhythmia, and cardiac hypertrophy. Synthetic modified mRNA (modRNA) presents a novel gene therapy approach that offers high, transient, safe, nonimmunogenic, and controlled mRNA delivery to the heart tissue without any risk of genomic integration. Due to these remarkable characteristics combined with its bell-shaped pharmacokinetics in the heart, modRNA has become an attractive approach for the treatment of heart disease. However, to increase its effectiveness in vivo, a consistent and reliable delivery method needs to be followed. Hence, to maximize modRNA delivery efficiency and yield consistency in modRNA use for in vivo applications, an optimized method of preparation and delivery of modRNA intracardiac injection in a mouse MI model is presented. This protocol will make modRNA delivery more accessible for basic and translational research.

Optimization of 5' Untranslated Region of Modified mRNA for Use in Cardiac or Hepatic Ischemic Injury.

Modified mRNA (modRNA) is a gene-delivery platform for transiently introducing a single gene or several genes of interest to different cell types and tissues. modRNA is considered to be a safe vector for gene transfer, as it negligibly activates the innate immune system and does not compromise the genome integrity. The use of modRNA in basic and translational science is rising, due to the clinical potential of modRNA. We are currently using modRNA to induce cardiac regeneration post-ischemic injury. Major obstacles in using modRNA for cardiac ischemic disease include the need for the direct and single administration of modRNA to the heart and the inefficient translation of modRNA due to its short half-life. Modulation of the 5' untranslated region (5' UTR) to enhance translation efficiency in ischemic cardiac disease has great value, as it can reduce the amount of modRNA needed per delivery and will achieve higher and longer protein production post-single delivery. Here, we identified that 5' UTR, from the fatty acid metabolism gene carboxylesterase 1D (Ces1d), enhanced the translation of firefly luciferase (Luc) modRNA by 2-fold in the heart post-myocardial infarction (MI). Moreover, we identified, in the Ces1d, a specific RNA element (element D) that is responsible for the improvement of modRNA translation and leads to a 2.5-fold translation increment over Luc modRNA carrying artificial 5' UTR, post-MI. Importantly, we were able to show that 5' UTR Ces1d also enhances modRNA translation in the liver, but not in the kidney, post-ischemic injury, indicating that Ces1d 5' UTR and element D may play a wider role in translation of protein under an ischemic condition.

Pkm2 Regulates Cardiomyocyte Cell Cycle and Promotes Cardiac Regeneration.

BACKGROUND: The adult mammalian heart has limited regenerative capacity, mostly attributable to postnatal cardiomyocyte cell cycle arrest. In the last 2 decades, numerous studies have explored cardiomyocyte cell cycle regulatory mechanisms to enhance myocardial regeneration after myocardial infarction. Pkm2 (Pyruvate kinase muscle isoenzyme 2) is an isoenzyme of the glycolytic enzyme pyruvate kinase. The role of Pkm2 in cardiomyocyte proliferation, heart development, and cardiac regeneration is unknown. METHODS: We investigated the effect of Pkm2 in cardiomyocytes through models of loss (cardiomyocyte-specific Pkm2 deletion during cardiac development) or gain using cardiomyocyte-specific Pkm2 modified mRNA to evaluate Pkm2 function and regenerative affects after acute or chronic myocardial infarction in mice. RESULTS: Here, we identify Pkm2 as an important regulator of the cardiomyocyte cell cycle. We show that Pkm2 is expressed in cardiomyocytes during development and immediately after birth but not during adulthood. Loss of function studies show that cardiomyocyte-specific Pkm2 deletion during cardiac development resulted in significantly reduced cardiomyocyte cell cycle, cardiomyocyte numbers, and myocardial size. In addition, using cardiomyocyte-specific Pkm2 modified RNA, our novel cardiomyocyte-targeted strategy, after acute or chronic myocardial infarction, resulted in increased cardiomyocyte cell division, enhanced cardiac function, and improved long-term survival. We mechanistically show that Pkm2 regulates the cardiomyocyte cell cycle and reduces oxidative stress damage through anabolic pathways and ß-catenin. CONCLUSIONS: We demonstrate that Pkm2 is an important intrinsic regulator of the cardiomyocyte cell cycle and oxidative stress, and highlight its therapeutic potential using cardiomyocyte-specific Pkm2 modified RNA as a gene delivery platform.

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.

mRNA-Based Protein Replacement Therapy for the Heart.

Myocardial infarction (MI) and heart failure (HF) are the leading causes of death in the United States and in most other industrialized nations. MI leads to a massive loss of cardiomyocytes (CMs), which are replaced with non-CM cells, leading to scarring and, in most cases, HF. The adult mammalian heart has a low intrinsic regenerative capacity, mainly because of cell-cycle arrest in CMs. No effective treatment promoting heart regeneration is currently available. Recent efforts to use DNA-based or viral gene therapy approaches to induce cardiac regeneration post-MI or in HF conditions have encountered major challenges, mostly because of the poor and uncontrolled delivery of the introduced genes. Modified mRNA (modRNA) is a safe, non-immunogenic, efficient, transient, local, and controlled nucleic acid delivery system that can overcome the obstacles to DNA-based or viral approaches for cardiac gene delivery. We here review the use of modRNA in cardiac therapy, to induce cardioprotection and vascular or cardiac regeneration after MI. We discuss the current challenges in modRNA-based cardiac treatment, which will need to be overcome for the application of such treatment to ischemic heart disease.

Ablation of a Single N-Glycosylation Site in Human FSTL 1 Induces Cardiomyocyte Proliferation and Cardiac Regeneration.

Adult mammalian hearts have a very limited regeneration capacity, due largely to a lack of cardiomyocyte (CM) proliferation. It was recently reported that epicardial, but not myocardial, follistatin-like 1 (Fstl1) activates CM proliferation and cardiac regeneration after myocardial infarction (MI). Furthermore, bacterially synthesized human FSTL 1 (hFSTL1) was found to induce CM proliferation, whereas hFSTL1 synthesized in mammals did not, suggesting that post-translational modifications (e.g., glycosylation) of the hFSTL1 protein affect its regenerative activity. We used modified mRNA (modRNA) technology to investigate the possible role of specific hFSTL1 N-glycosylation sites in the induction, by hFSTL1, of CM proliferation and cardiac regeneration. We found that the mutation of a single site (N180Q) was sufficient and necessary to increase the proliferation of rat neonatal and mouse adult CMs in vitro and after MI in vivo, respectively. A single administration of the modRNA construct encoding the N180Q mutant significantly increased cardiac function, decreased scar size, and increased capillary density 28 days post-MI. Overall, our data suggest that the delivery of N180Q hFSTL1 modRNA to the myocardium can mimic the beneficial effect of epicardial hFSTL1, triggering marked CM proliferation and cardiac regeneration in a mouse MI model.

PPARß/δ: Linking Metabolism to Regeneration.

In contrast to the general belief that regeneration is a rare event, mainly occurring in simple organisms, the ability of regeneration is widely distributed in the animal kingdom. Yet, the efficiency and extent of regeneration varies greatly. Humans can recover from blood loss as well as damage to tissues like bone and liver. Yet damage to the heart and brain cannot be reversed, resulting in scaring. Thus, there is a great interest in understanding the molecular mechanisms of naturally occurring regeneration and to apply this knowledge to repair human organs. During regeneration, injury-activated immune cells induce wound healing, extracellular matrix remodeling, migration, dedifferentiation and/or proliferation with subsequent differentiation of somatic or stem cells. An anti-inflammatory response stops the regenerative process, which ends with tissue remodeling to achieve the original functional state. Notably, many of these processes are associated with enhanced glycolysis. Therefore, peroxisome proliferator-activated receptor (PPAR) ß/δ­which is known to be involved for example in lipid catabolism, glucose homeostasis, inflammation, survival, proliferation, differentiation, as well as mammalian regeneration of the skin, bone and liver­appears to be a promising target to promote mammalian regeneration. This review summarizes our current knowledge of PPARß/δ in processes associated with wound healing and regeneration.