Systemic and heart autonomous effects of sphingosine Δ4 desaturase deficiency in lipotoxic cardiac pathophysiology

ABSTRACTLipotoxic cardiomyopathy (LCM) is characterized by cardiac steatosis, including the accumulation of fatty acids, triglycerides and ceramides. Model systems have shown the inhibition of ceramide biosynthesis to antagonize obesity and improve insulin sensitivity. Sphingosine Δ4 desaturase (encoded by ifc in Drosophila melanogaster) enzymatically converts dihydroceramide into ceramide. Here, we examine ifc mutants to study the effects of desaturase deficiency on cardiac function in Drosophila. Interestingly, ifc mutants exhibited classic hallmarks of LCM: cardiac chamber dilation, contractile defects and loss of fractional shortening. This outcome was phenocopied in global ifc RNAi-mediated knockdown flies. Surprisingly, cardiac-specific ifc knockdown flies exhibited cardiac chamber restriction with no contractile defects, suggesting heart autonomous and systemic roles for ifc activity in cardiac function. Next, we demonstrated that ifc mutants exhibit suppressed Sphingosine kinase 1 (Sk1) expression. Ectopic overexpression of Sk1 was sufficient to prevent cardiac chamber dilation and loss of fractional shortening in ifc mutants. Partial rescue was also observed with cardiac- and fat-body-specific Sk1 overexpression. Finally, we showed that cardiac-specific expression of Drosophila inhibitor of apoptosis (dIAP) also prevented cardiac dysfunction in ifc mutants, suggesting a role for caspase activity in the observed cardiac pathology. Collectively, we show that spatial regulation of sphingosine Δ4 desaturase activity differentially affects cardiac function in heart autonomous and systemic mechanisms through tissue interplay.

Download Full-text

Abstract 20118: Defining the Sufficiency of Atg7 to Suppress Phenylephrine-induced Cardiac Hypertrophy

Circulation ◽

10.1161/circ.130.suppl_2.20118 ◽

2014 ◽

Vol 130 (suppl_2) ◽

Author(s):

Marcus Tjeerdsma ◽

Levi Froke ◽

Jessica Freeling ◽

Scott Pattison

Keyword(s):

Body Weight ◽

Cardiac Hypertrophy ◽

Cardiac Function ◽

Heart Weight ◽

Autophagic Flux ◽

Specific Expression ◽

Hypertrophic Growth ◽

Fractional Shortening

Introduction: Macroautophagy is a process of bulk protein degradation. Our prior work showed that Atg7 expression is sufficient to induce autophagic flux in vitro and in vivo . When Atg7 was co-expressed with CryAB R120G in the heart, cardiac hypertrophy was blunted in heart weight/body weight ratios and fetal gene expression markers. To determine if Atg7 expression is sufficient to limit hypertrophic growth in another model, we tested the effects of Atg7 overexpression with phenylephrine-induced hypertrophy both in vitro and in vivo . Hypothesis: Atg7 will blunt the hypertrophic effects of phenylephrine. Methods: Rat neonatal cardiomyocytes were infected with adenoviruses expressing either LacZ or Atg7 and treated with phenylephrine to induce cardiomyocytes hypertrophy. Osmotic pumps were surgically implanted into control mice and mice with cardiac-specific expression of Atg7 to infuse phenylephrine (PE) or vehicle (saline) for four weeks. Results: PE treatment significantly increased neonatal cardiomyocyte areas in LacZ-expressing cells, while Atg7-expressing cardiomyocytes showed no growth. In mice, all genotypes responded to PE treatment with significantly increased heart weight/body weight ratios and increased fiber size. However, Atg7-expressing hearts differed significantly from control hearts in normalized heart mass following PE delivery. Vehicle treated Atg7-expressing hearts had 17% smaller myofiber cross-sectional areas than those from control genotypes and had a reduced hypertrophic response to PE, relative to controls. Echocardiography showed that Atg7-expressing hearts had significantly elevated cardiac function (% fractional shortening) prior to and throughout the experiment over control hearts (33% vs. 29%). PE significantly increased fractional shortening) from 29% to 36% in control hearts, but failed to significantly elevate cardiac function in Atg7-expressing hearts further (33% vs 35%). Additional assays are underway to understand the Atg7-dependent adaptations to PE. Conclusion: Atg7 expression yields modestly smaller hearts with enhanced cardiac function which may protect them from hypertrophic stresses like phenylephrine.

Download Full-text

Cardiac overexpression of Mammalian enabled (Mena) exacerbates heart failure in mice

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.00342.2013 ◽

2013 ◽

Vol 305 (6) ◽

pp. H875-H884 ◽

Cited By ~ 6

Author(s):

Stephen L. Belmonte ◽

Rashmi Ram ◽

Deanne M. Mickelsen ◽

Frank B. Gertler ◽

Burns C. Blaxall

Keyword(s):

Heart Failure ◽

Cardiac Myocyte ◽

Cardiac Injury ◽

Actin Dynamics ◽

Cardiac Pathology ◽

Cardiac Pathophysiology ◽

Precise Role ◽

Functional Deterioration ◽

Fractional Shortening

Mammalian enabled (Mena) is a key regulator of cytoskeletal actin dynamics, which has been implicated in heart failure (HF). We have previously demonstrated that cardiac Mena deletion produced cardiac dysfunction with conduction abnormalities and hypertrophy. Moreover, elevated Mena expression correlates with HF in human and animal models, yet the precise role of Mena in cardiac pathophysiology is unclear. In these studies, we evaluated mice with cardiac myocyte-specific Mena overexpression (TTA/TgTetMena) comparable to that observed in cardiac pathology. We found that the hearts of TTA/TgTetMena mice were functionally and morphologically comparable to wild-type littermates, except for mildly increased heart mass in the transgenic mice. Interestingly, TTA/TgTetMena mice were particularly susceptible to cardiac injury, as these animals experienced pronounced decreases in ejection fraction and fractional shortening as well as heart dilatation and hypertrophy after transverse aortic constriction (TAC). By “turning off” Mena overexpression in TTA/TgTetMena mice either immediately prior to or immediately after TAC surgery, we discovered that normalizing Mena levels eliminated cardiac hypertrophy in TTA/TgTetMena animals but did not preclude post-TAC cardiac functional deterioration. These findings indicate that hearts with increased levels of Mena fare worse when subjected to cardiac injury and suggest that Mena contributes to HF pathophysiology.

Download Full-text

Fat-body brummer lipase determines survival and cardiac function during starvation in Drosophila melanogaster

iScience ◽

10.1016/j.isci.2021.102288 ◽

2021 ◽

Vol 24 (4) ◽

pp. 102288

Author(s):

Annelie Blumrich ◽

Georg Vogler ◽

Sandra Dresen ◽

Soda Balla Diop ◽

Carsten Jaeger ◽

...

Keyword(s):

Drosophila Melanogaster ◽

Cardiac Function ◽

Fat Body

Download Full-text

Abstract 269: Miofibroblast Tgf-beta Signaling in a Proteotoxic Mouse Model

Circulation Research ◽

10.1161/res.119.suppl_1.269 ◽

2016 ◽

Vol 119 (suppl_1) ◽

Author(s):

Bidur Bhandary ◽

Qinghang Meng ◽

Hanna Osinska ◽

Kritton Shay-Winkler ◽

James Gulick ◽

...

Keyword(s):

Heart Disease ◽

Mouse Model ◽

Cardiac Function ◽

Transforming Growth Factor Beta ◽

Cardiac Fibrosis ◽

Transforming Growth Factor ◽

Prolonged Survival ◽

Tgfβ Signaling ◽

Specific Expression ◽

Western Blots

Introduction: Transforming Growth Factor Beta (TGFβ) is an important cytokine in mediating the fibrogenic response and, in particular, cardiac fibrosis. Extensive fibrosis accompanies the cardiac remodeling that occurs during development of the protein conformation-based disease caused by cardiomyocyte-specific expression of a mutant, small, heat shock-like protein and chaperone, aB crystallin (CryABR120G). During the onset of fibrosis, fibroblasts are activated to the so-called “myofibroblast” state and TGFβ binding is thought to mediate an essential signaling pathway underlying this process. Our central hypothesis is that TGFβ signaling processes that result in significant cardiac fibrosis in a mouse model of proteotoxic heart disease are mediated by cardiac fibroblasts, rather than cardiomyocytes. Here, we have partially ablated TGFβ signaling only in cardiac myofibroblasts to observe if cardiac fibrosis is reduced. Aims and Methods: The objective of this study was to understand the contributions of fibroblast-derived TGFβ signaling to the development of cardiac fibrosis in a proteotoxic mouse model that results in significant cardiac fibrosis. To test the hypothesis we partially deleted the myofibroblast specific canonical and non-canonical signaling by crossing CryAB R120G mice with Tgfbr1 or Tgfbr2 floxed mice. The double transgene containing mice were further crossed with activated myofibroblast specific Cre mice in which Cre expression was driven off the periostin promoter. Echocardiography, Masson’s Trichome staining, PCR arrays, IHC and western blots were performed to characterize the fibrotic progression in CryAB R120G transgenic mice. Results: We observed that myofibroblast-targeted partial knockdown of Tgf βr1 signaling prolonged survival, modestly reducing fibrosis and improving cardiac function . Similarly, Tgf βr2 partial knockdown prolonged survival, modestly reducing fibrosis without improving cardiac function during fibrosis development in CryAB R120G mice. Conclusion: These findings suggest that, in a model of proteotoxic heart disease, myofibroblast based TGFβ signaling in the heart may contribute to cardiac hypertrophy/dysfunction but cannot account entirely for the fibrotic response.

Download Full-text

Abstract 14391: Transient Cell Cycle Induction in Cardiomyocytes is a Promising Approach to Treat Ischemia Induced Heart Failure

Circulation ◽

10.1161/circ.142.suppl_3.14391 ◽

2020 ◽

Vol 142 (Suppl_3) ◽

Author(s):

Riham Abouleisa ◽

Qinghui Ou ◽

Xian-liang Tang ◽

Mitesh Solanki ◽

Yiru Guo ◽

...

Keyword(s):

Cell Cycle ◽

Cardiac Function ◽

Single Cell ◽

Ischemia Reperfusion Injury ◽

Ischemia Reperfusion ◽

Cardiomyocyte Proliferation ◽

Specific Expression ◽

Control Virus

Rationale: The regenerative capacity of the heart to repair itself after myocardial infarction (MI)is limited. Our previous study showed that ectopic introduction of Cdk1/CyclinB1 andCdk4/CyclinD1 complexes (4F) promotes cardiomyocyte proliferation in vitro and in vivo andimproves cardiac function after MI. However, its clinical application is limited due to the concernsfor tumorigenic potential in other organs. Objectives: To first, identify on a single cell transcriptomic basis the necessary reprogrammingsteps that cardiomyocytes need to undertake to progress through the proliferation processfollowing 4F overexpression, and then, to determine the pre-clinical efficacy of transient andcardiomyocyte specific expression of 4F in improving cardiac function after MI in small and largeanimals. Methods and Results: Temporal bulk and single cell RNAseq of mature hiPS-CMs treated with4F or LacZ control for 24, 48, or 72 h revealed full cell cycle reprogramming in 15% of thecardiomyocyte population which was associated with sarcomere disassembly and metabolicreprogramming. Transient overexpression of 4F specifically in cardiomyocytes was achievedusing non-integrating lentivirus (NIL) driven by TNNT2 (TNNT2-4F-NIL). One week after inductionof ischemia-reperfusion injury in rats or pigs, TNNT2-4F-NIL or control virus was injectedintramyocardially. Compared with controls, rats or pigs treated with TNNT2-4F-NIL showed a 20-30% significant improvement in ejection fraction and scar size four weeks after treatment, asassessed by echocardiography and histological analysis. Quantification of cardiomyocyteproliferation in pigs using a novel cytokinesis reporter showed that ~10% of the cardiomyocyteswithin the injection site were labelled as daughter cells following injection with TNNT2-4F-NILcompared with ~0.5% background labelling in control groups. Conclusions: We provide the first understanding of the process of forced cardiomyocyteproliferation and advanced the clinical applicability of this approach through minimization ofoncogenic potential of the cell cycle factors using a novel transient and cardiomyocyte-specificviral construct.

Download Full-text

Abstract 791: A Critical Role for Bmper (BMP-binding Endothelial cell precursor-derived Regulator) in Cardiac Muscle Mass Regulation during Development and Pressure Overload Induced Hypertrophy

Circulation ◽

10.1161/circ.116.suppl_16.ii_152-c ◽

2007 ◽

Vol 116 (suppl_16) ◽

Author(s):

Monte Willis ◽

Rongqin Ren ◽

Cam Patterson

Keyword(s):

Cardiac Function ◽

Cardiac Muscle ◽

Muscle Mass ◽

Cardiac Development ◽

Critical Role ◽

Pressure Overload ◽

Posterior Wall ◽

Fractional Shortening

Bone morphogenetic proteins (BMPs) of the TGF-beta superfamily, have been implicated in multiple processes during cardiac development. Our laboratory recently described an unprecedented role for Bmper in antagonizing BMP-2, BMP-4, and BMP-6. To determine the role of Bmper on cardiac development in vivo, we created Bmper null (Bmper −/−) mice by replacing exons 1 and 2 with GFP. Since Bmper −/− mice are perinatally lethal, we determined pre-natal cardiac function of Bmper −/− mice in utero just before birth. By echocardiography, E18.5 Bmper −/− embryos had decreased cardiac function (24.2 +/− 8.1% fractional shortening) compared to Bmper +/− and Bmper +/+ siblings (52.2 +/− 1.6% fractional shortening) (N=4/group). To further characterize the role of Bmper on cardiac function in adult mice, we performed echocardiography on 8-week old male and female Bmper +/− and littermate control Bmper +/+. Bmper +/− mice had an approximately 15% decrease in anterior and posterior wall thickness compared to sibling Bmper +/+ mice at baseline (n=10/group). Cross-sectional areas of Bmper +/− cardiomyocytes were approximately 20% less than wild type controls, indicating cardiomyocyte hypoplasia in adult Bmper +/− mice at baseline. Histologically, no significant differences were identified in representative H&E and trichrome stained adult Bmper +/− and Bmper +/+ cardiac sections at baseline. To determine the effects of Bmper expression on the development of cardiac hypertrophy, both Bmper +/− and Bmper +/+ sibling controls underwent transaortic constriction (TAC), followed by weekly echocardiography. While a deficit was identified in Bmper +/− mice at baseline, both anterior and posterior wall thicknesses increased after TAC, such that identical wall thicknesses were identified in Bmper +/− and Bmper +/+ mice 1–4 weeks after TAC. Notably, cardiac function (fractional shortening %) and histological evaluation revealed no differences between Bmper +/− and Bmper +/+ any time after TAC. These studies identify for the first time that Bmper expression plays a critical role in regulating cardiac muscle mass during development, and that Bmper regulates the development of hypertrophy in response to pressure overload in vivo.

Download Full-text

Rescue of Prothrombin-deficiency by Transgene Expression in Mice

Thrombosis and Haemostasis ◽

10.1055/s-0037-1613344 ◽

2002 ◽

Vol 88 (12) ◽

pp. 984-991 ◽

Cited By ~ 20

Author(s):

William Sun ◽

Mallory Coleman ◽

David Witte ◽

Sandra Degen

Keyword(s):

Transgenic Mice ◽

Transgene Expression ◽

Embryonic Lethality ◽

The Other ◽

Model Systems ◽

Bleeding Events ◽

Specific Expression ◽

Spontaneous Bleeding ◽

Knock Out ◽

Transgenic Expression

SummaryProthrombin has diverse biological functions in addition to its well established role in blood coagulation. In order to study these functions in more detail mouse model systems are needed. Since deficiency of prothrombin in mice results in partial embryonic lethality and neonatal death, alternative approaches are required to study the biology of prothrombin in the adult mouse. The liver is the major site of synthesis of prothrombin and therefore liver-specific promoters were used to express prothrombin in transgenic mice. Mice generated from crosses with these transgenic mice and mice hemizygous for the knock-out allele were used to test whether liver-specific expression is sufficient to correct the phenotype of null mice and whether liver-specific expression is sufficient for the development and survival of mice to adulthood. The mouse albumin promoter/enhancer was used initially for transgene expression without success in obtaining transgene positive, endogenous prothrombin null mice. Two lines of transgene positive, endogenous prothrombin deficient mice were obtained using the mouse transthyretin (TTR) promoter/enhancer driving expression of a human prothrombin cDNA. One line was able to rescue both the embryonic and the neonatal lethality while the other line was only able to correct the embryonic lethality. Expression of prothrombin was restricted to the liver and stomach in one line and to the liver, pancreas, stomach and kidney in the other line of mice. Thrombin activity for one line was determined to be at 5-10% of wildtype levels. These mice developed normally and did not have spontaneous bleeding events unless traumatized. Therefore, transgenic expression of human prothrombin is sufficient for the rescue of the lethality found for prothrombin deficiency in mice.

Download Full-text

Deep Characterization of Circular RNAs from Human Cardiovascular Cell Models and Cardiac Tissue

Cells ◽

10.3390/cells9071616 ◽

2020 ◽

Vol 9 (7) ◽

pp. 1616

Author(s):

Tobias Jakobi ◽

Dominik Siede ◽

Jessica Eschenbach ◽

Andreas W. Heumüller ◽

Martin Busch ◽

...

Keyword(s):

Cardiac Tissue ◽

Umbilical Vein ◽

Expression Profiles ◽

Developed Countries ◽

Model Systems ◽

Start Codon ◽

Circular Rnas ◽

Specific Expression ◽

Cardiac Model ◽

Depth Analysis

For decades, cardiovascular disease (CVD) has been the leading cause of death throughout most developed countries. Several studies relate RNA splicing, and more recently also circular RNAs (circRNAs), to CVD. CircRNAs originate from linear transcripts and have been shown to exhibit tissue-specific expression profiles. Here, we present an in-depth analysis of sequence, structure, modification, and cardiac circRNA interactions. We used human induced pluripotent stem cell-derived cardiac myocytes (hiPSC-CMs), human healthy and diseased (ischemic cardiomyopathy, dilated cardiomyopathy) cardiac tissue, and human umbilical vein endothelial cells (HUVECs) to profile circRNAs. We identified shared circRNAs across all samples, as well as model-specific circRNA signatures. Based on these circRNAs, we identified 63 positionally conserved and expressed circRNAs in human, pig, and mouse hearts. Furthermore, we found that the sequence of circRNAs can deviate from the sequence derived from the genome sequence, an important factor in assessing potential functions. Integration of additional data yielded evidence for m6A-methylation of circRNAs, potentially linked to translation, as well as, circRNAs overlapping with potential Argonaute 2 binding sites, indicating potential association with the RISC complex. Moreover, we describe, for the first time in cardiac model systems, a sub class of circRNAs containing the start codon of their primary transcript (AUG circRNAs) and observe an enrichment for m6A-methylation for AUG circRNAs.

Download Full-text

Imprinting of insulin-like growth factor 2 is modulated during hematopoiesis

Blood ◽

10.1182/blood.v96.9.3023.h8003023_3023_3028 ◽

2000 ◽

Vol 96 (9) ◽

pp. 3023-3028 ◽

Cited By ~ 1

Author(s):

Ian M. Morison ◽

Michael R. Eccles ◽

Anthony E. Reeve

Keyword(s):

Growth Factor ◽

Mononuclear Cells ◽

Model Systems ◽

Paternal Allele ◽

Parental Origin ◽

Insulin Like Growth Factor ◽

Specific Expression ◽

Peripheral Blood Mononuclear ◽

Multistep Process

The transcription of insulin-like growth factor 2 (IGF-2) is affected by genomic imprinting, a multistep process through which the parental origin of a gene influences its transcription. The maternal copy of IGF-2 is silenced in most human tissues, but in the choroid plexus and the adult liver both alleles of IGF-2 are expressed. This study shows that though in peripheral blood mononuclear cells IGF-2shows paternal allele-specific expression, in total bone marrow both alleles are transcribed. This modulation of imprinting is not attributable to use of the P1 promoter, because transcription from the P3 promoter occurred from both alleles. These results suggest that transcriptional recognition of the IGF-2 imprint can be modulated during hematopoiesis and may facilitate the development of in vitro model systems to study the transcriptional recognition of a genomic imprint.

Download Full-text

Bone marrow stem and progenitor cell contribution to neovasculogenesis is dependent on model system with SDF-1 as a permissive trigger

Blood ◽

10.1182/blood-2009-03-211342 ◽

2009 ◽

Vol 114 (19) ◽

pp. 4310-4319 ◽

Cited By ~ 43

Author(s):

Gerard J. Madlambayan ◽

Jason M. Butler ◽

Koji Hosaka ◽

Marda Jorgensen ◽

Dongtao Fu ◽

...

Keyword(s):

Bone Marrow ◽

Neutralizing Antibodies ◽

Model Systems ◽

Experimental Methodology ◽

Specific Expression ◽

Hematopoietic Stem ◽

Lung Carcinoma Cells ◽

Myeloid Progenitor ◽

Myeloid Progenitor Cells

Abstract Adult bone marrow (BM) contributes to neovascularization in some but not all settings, and reasons for these discordant results have remained unexplored. We conducted novel comparative studies in which multiple neovascularization models were established in single mice to reduce variations in experimental methodology. In different combinations, BM contribution was detected in ischemic retinas and, to a lesser extent, Lewis lung carcinoma cells, whereas B16 melanomas showed little to no BM contribution. Using this spectrum of BM contribution, we demonstrate the necessity for site-specific expression of stromal-derived factor-1α (SDF-1α) and its mobilizing effects on BM. Blocking SDF-1α activity with neutralizing antibodies abrogated BM-derived neovascularization in lung cancer and retinopathy. Furthermore, secondary transplantation of single hematopoietic stem cells (HSCs) showed that HSCs are a long-term source of neovasculogenesis and that CD133+CXCR4+ myeloid progenitor cells directly participate in new blood vessel formation in response to SDF-1α. The varied BM contribution seen in different model systems is suggestive of redundant mechanisms governing postnatal neovasculogenesis and provides an explanation for contradictory results observed in the field.

Download Full-text