Tafazzin knockdown in mice leads to a developmental cardiomyopathy with early diastolic dysfunction preceding myocardial noncompaction.
ABSTRACT: Barth syndrome is a rare, multisystem disorder caused by mutations in tafazzin that lead to cardiolipin deficiency and mitochondrial abnormalities. Patients most commonly develop an early-onset cardiomyopathy in infancy or fetal life.Knockdown of tafazzin (TAZKD) in a mouse model was induced from the start of gestation via a doxycycline-inducible shRNA transgenic approach. All liveborn TAZKD mice died within the neonatal period, and in vivo echocardiography revealed prenatal loss of TAZKD embryos at E12.5-14.5. TAZKD E13.5 embryos and newborn mice demonstrated significant tafazzin knockdown, and mass spectrometry analysis of hearts revealed abnormal cardiolipin profiles typical of Barth syndrome. Electron microscopy of TAZKD hearts demonstrated ultrastructural abnormalities in mitochondria at both E13.5 and newborn stages. Newborn TAZKD mice exhibited a significant reduction in total mitochondrial area, smaller size of individual mitochondria, reduced cristae density, and disruption of the normal parallel orientation between mitochondria and sarcomeres. Echocardiography of E13.5 and newborn TAZKD mice showed good systolic function, but early diastolic dysfunction was evident from an abnormal flow pattern in the dorsal aorta. Strikingly, histology of E13.5 and newborn TAZKD hearts showed myocardial thinning, hypertrabeculation and noncompaction, and defective ventricular septation. Altered cellular proliferation occurring within a narrow developmental window accompanied the myocardial hypertrabeculation-noncompaction.In this murine model, tafazzin deficiency leads to a unique developmental cardiomyopathy characterized by ventricular myocardial hypertrabeculation-noncompaction and early lethality. A central role of cardiolipin and mitochondrial functioning is strongly implicated in cardiomyocyte differentiation and myocardial patterning required for heart development. (J Am Heart Assoc. 2012;1:jah3-e000455 doi: 10.1161/JAHA.111.000455.).
Project description:Barth syndrome is an X-linked genetic disorder caused by mutations in the tafazzin (taz) gene and characterized by dilated cardiomyopathy, exercise intolerance, chronic fatigue, delayed growth, and neutropenia. Tafazzin is a mitochondrial transacylase required for cardiolipin remodeling. Although tafazzin function has been studied in non-mammalian model organisms, mammalian genetic loss of function approaches have not been used. We examined the consequences of tafazzin knockdown on sarcomeric mitochondria and cardiac function in mice. Tafazzin knockdown resulted in a dramatic decrease of tetralinoleoyl cardiolipin in cardiac and skeletal muscles and accumulation of monolysocardiolipins and cardiolipin molecular species with aberrant acyl groups. Electron microscopy revealed pathological changes in mitochondria, myofibrils, and mitochondrion-associated membranes in skeletal and cardiac muscles. Echocardiography and magnetic resonance imaging revealed severe cardiac abnormalities, including left ventricular dilation, left ventricular mass reduction, and depression of fractional shortening and ejection fraction in tafazzin-deficient mice. Tafazzin knockdown mice provide the first mammalian model system for Barth syndrome in which the pathophysiological relationships between altered content of mitochondrial phospholipids, ultrastructural abnormalities, myocardial and mitochondrial dysfunction, and clinical outcome can be completely investigated.
Project description:Barth syndrome is a complex metabolic disorder caused by mutations in the mitochondrial transacylase tafazzin. Recently, an inducible tafazzin shRNA knockdown mouse model was generated to deconvolute the complex bioenergetic phenotype of this disease. To investigate the underlying cause of hemodynamic dysfunction in Barth syndrome, we interrogated the cardiac structural and signaling lipidome of this mouse model as well as its myocardial bioenergetic phenotype. A decrease in the distribution of cardiolipin molecular species and robust increases in monolysocardiolipin and dilysocardiolipin were demonstrated. Additionally, the contents of choline and ethanolamine glycerophospholipid molecular species containing precursors for lipid signaling at the sn-2 position were altered. Lipidomic analyses revealed specific dysregulation of HETEs and prostanoids, as well as oxidized linoleic and docosahexaenoic metabolites. Bioenergetic interrogation uncovered differential substrate utilization as well as decreases in Complex III and V activities. Transgenic expression of cardiolipin synthase or iPLA2? ablation in tafazzin-deficient mice did not rescue the observed phenotype. These results underscore the complex nature of alterations in cardiolipin metabolism mediated by tafazzin loss of function. Collectively, we identified specific lipidomic, bioenergetic, and signaling alterations in a murine model that parallel those of Barth syndrome thereby providing novel insights into the pathophysiology of this debilitating disease.
Project description:Mitochondria fulfill vital metabolic functions and act as crucial cellular signaling hubs, integrating their metabolic status into the cellular context. Here, we show that defective cardiolipin remodeling, upon loss of the cardiolipin acyl transferase tafazzin, decreases HIF-1? signaling in hypoxia. Tafazzin deficiency does not affect posttranslational HIF-1? regulation but rather HIF-1? gene expression, a dysfunction recapitulated in iPSC-derived cardiomyocytes from Barth syndrome patients with tafazzin deficiency. RNA-seq analyses confirmed drastically altered signaling in tafazzin mutant cells. In hypoxia, tafazzin-deficient cells display reduced production of reactive oxygen species (ROS) perturbing NF-?B activation and concomitantly HIF-1? gene expression. Tafazzin-deficient mice hearts display reduced HIF-1? levels and undergo maladaptive hypertrophy with heart failure in response to pressure overload challenge. We conclude that defective mitochondrial cardiolipin remodeling dampens HIF-1? signaling due to a lack of NF-?B activation through reduced mitochondrial ROS production, decreasing HIF-1? transcription.
Project description:Quantitative and qualitative alterations of mitochondrial cardiolipin have been implicated in the pathogenesis of Barth syndrome, an X-linked cardioskeletal myopathy caused by a deficiency in tafazzin, an enzyme in the cardiolipin remodeling pathway. We have generated and previously reported a tafazzin-deficient Drosophila model of Barth syndrome that is characterized by low cardiolipin concentration, abnormal cardiolipin fatty acyl composition, abnormal mitochondria, and poor motor function. Here, we first show that tafazzin deficiency in Drosophila disrupts the final stage of spermatogenesis, spermatid individualization, and causes male sterility. This phenotype can be genetically suppressed by inactivation of the gene encoding a calcium-independent phospholipase A(2), iPLA2-VIA, which also prevents cardiolipin depletion/monolysocardiolipin accumulation, although in wild-type flies inactivation of the iPLA2-VIA does not affect the molecular composition of cardiolipin. Furthermore, we show that treatment of Barth syndrome patients' lymphoblasts in tissue culture with the iPLA(2) inhibitor, bromoenol lactone, partially restores their cardiolipin homeostasis. Taken together, these findings establish a causal role of cardiolipin deficiency in the pathogenesis of Barth syndrome and identify iPLA2-VIA as an important enzyme in cardiolipin deacylation, and as a potential target for therapeutic intervention.
Project description:Aim: Tafazzin knockdown (TazKD) in mice is widely used to create an experimental model of Barth syndrome (BTHS) that exhibits dilated cardiomyopathy and impaired exercise capacity. Peroxisome proliferator-activated receptors (PPARs) are a group of nuclear receptor proteins that play essential roles as transcription factors in the regulation of carbohydrate, lipid, and protein metabolism. We hypothesized that the activation of PPAR signaling with PPAR agonist bezafibrate (BF) may ameliorate impaired cardiac and skeletal muscle function in TazKD mice. This study examined the effects of BF on cardiac function, exercise capacity, and metabolic status in the heart of TazKD mice. Additionally, we elucidated the impact of PPAR activation on molecular pathways in TazKD hearts. Methods: BF (0.05% w/w) was given to TazKD mice with rodent chow. Cardiac function in wild type-, TazKD-, and BF-treated TazKD mice was evaluated by echocardiography. Exercise capacity was evaluated by exercising mice on the treadmill until exhaustion. The impact of BF on metabolic pathways was evaluated by analyzing the total transcriptome of the heart by RNA sequencing. Results: The uptake of BF during a 4-month period at a clinically relevant dose effectively protected the cardiac left ventricular systolic function in TazKD mice. BF alone did not improve the exercise capacity however, in combination with everyday voluntary running on the running wheel BF significantly ameliorated the impaired exercise capacity in TazKD mice. Analysis of cardiac transcriptome revealed that BF upregulated PPAR downstream target genes involved in a wide spectrum of metabolic (energy and protein) pathways as well as chromatin modification and RNA processing. In addition, the Ostn gene, which encodes the metabolic hormone musclin, is highly induced in TazKD myocardium and human failing hearts, likely as a compensatory response to diminished bioenergetic homeostasis in cardiomyocytes. Conclusion: The PPAR agonist BF at a clinically relevant dose has the therapeutic potential to attenuate cardiac dysfunction, and possibly exercise intolerance in BTHS. The role of musclin in the failing heart should be further investigated.
Project description:Mitochondria fulfill vital metabolic functions and act as crucial cellular signaling hubs integrating their metabolic status into the cellular context. Here, we show that defective cardiolipin-remodeling, upon loss of the cardiolipin acyl transferase Tafazzin, mutes HIF-1a signaling in hypoxia. Tafazzin-deficiency does not affect posttranslational HIF-1a regulation but rather HIF-1a gene-expression, a dysfunction recapitulated in iPSCs-derived cardiomyocytes from Barth Syndrome patients with Tafazzin-deficiency. RNAseq analyses confirmed drastically altered signaling in Tafazzin mutant cells. In hypoxia, Tafazzin-deficient cells display reduced production of reactive oxygen species (ROS) perturbing NF-kB activation and concomitantly HIF-1a gene-expression. In agreement, Tafazzin-deficient mice hearts display reduced HIF-1a levels and undergo maladaptive hypertrophy with heart failure in response to pressure overload challenge. We conclude that defective mitochondrial cardiolipin-remodeling dampens HIF-1a signaling through inactivation of a non-canonical signaling pathway: Lack of NF-kB activation through reduced mitochondrial ROS production diminishes HIF-1a transcription. Overall design: Cells lacking tafazzin were subjected to hypoxia and compared to the ones at normoxia or isogenic WT cells either at hypoxia or nomoxia
Project description:The phospholipid, cardiolipin, is essential for maintaining mitochondrial structure and optimal function. Cardiolipin-deficiency in humans, Barth syndrome, is characterized by exercise intolerance, dilated cardiomyopathy, neutropenia, and 3-methyl-glutaconic aciduria. The causative gene is the mitochondrial acyl-transferase, tafazzin, that is essential for remodeling acyl chains of cardiolipin. We sought to determine metabolic rates in tafazzin-deficient mice during resting and exercise, and investigate the impact of cardiolipin-deficiency on mitochondrial respiratory chain activities. Tafazzin-knockdown in mice markedly impaired oxygen consumption rates during an exercise, without any significant effect on resting metabolic rates. CL-deficiency resulted in significant reduction of mitochondrial respiratory reserve capacity in neonatal cardiomyocytes that is likely to be caused by diminished activity of complex-III, which requires CL for its assembly and optimal activity. Our results may provide mechanistic insights of Barth syndrome pathogenesis.
Project description:Cardiolipin is a mitochondria-specific phospholipid known to be intimately involved with apoptosis. However, the lack of appropriate cellular models to date restricted analysis of its role in cell death. The maturation of cardiolipin requires the transacylase tafazzin, which is mutated in the human disorder Barth syndrome. Using Barth syndrome patient-derived cells and HeLa cells in which tafazzin was knocked down, we show that cardiolipin is required for apoptosis in the type II mitochondria-dependent response to Fas stimulation. Cardiolipin provides an anchor and activating platform for caspase-8 translocation to, and embedding in, the mitochondrial membrane, where it oligomerizes and is further activated, steps that are necessary for an efficient type II apoptotic response.
Project description:Mitochondria constantly adapt to the metabolic needs of a cell. This mitochondrial plasticity is critical to T cells, which modulate metabolism depending on antigen-driven signals and environment. We show here that de novo synthesis of the mitochondrial membrane-specific lipid cardiolipin maintains CD8<sup>+</sup> T cell function. T cells deficient for the cardiolipin-synthesizing enzyme PTPMT1 had reduced cardiolipin and responded poorly to antigen because basal cardiolipin levels were required for activation. However, neither de novo cardiolipin synthesis, nor its Tafazzin-dependent remodeling, was needed for T cell activation. In contrast, PTPMT1-dependent cardiolipin synthesis was vital when mitochondrial fitness was required, most notably during memory T cell differentiation or nutrient stress. We also found CD8<sup>+</sup> T cell defects in a small cohort of patients with Barth syndrome, where TAFAZZIN is mutated, and in a Tafazzin-deficient mouse model. Thus, the dynamic regulation of a single mitochondrial lipid is crucial for CD8<sup>+</sup> T cell immunity.
Project description:Barth syndrome (BTHS) is a cardiomyopathy caused by the loss of tafazzin, a mitochondrial acyltransferase involved in the maturation of the glycerophospholipid cardiolipin. It has remained enigmatic as to why a systemic loss of cardiolipin leads to cardiomyopathy. Using a genetic ablation of tafazzin function in the BTHS mouse model, we identified severe structural changes in respiratory chain supercomplexes at a pre-onset stage of the disease. This reorganization of supercomplexes was specific to cardiac tissue and could be recapitulated in cardiomyocytes derived from BTHS patients. Moreover, our analyses demonstrate a cardiac-specific loss of succinate dehydrogenase (SDH), an enzyme linking the respiratory chain with the tricarboxylic acid cycle. As a similar defect of SDH is apparent in patient cell-derived cardiomyocytes, we conclude that these defects represent a molecular basis for the cardiac pathology in Barth syndrome.