Yoriko Uozumi

Taurine depletion caused by knocking out the taurine transporter gene leads to cardiomyopathy with cardiac atrophy

The sulfur-containing beta-amino acid, taurine, is the most abundant free amino acid in cardiac and skeletal muscle. Although its physiological function has not been established, it is thought to play an important role in ion movement, calcium handling, osmoregulation and cytoprotection. To begin examining the physiological function of taurine, we generated taurine transporter- (TauT-) knockout mice (TauTKO), which exhibited a deficiency in myocardial and skeletal muscle taurine content compared with their wild-type littermates. The TauTKO heart underwent ventricular remodeling, characterized by reductions in ventricular wall thickness and cardiac atrophy accompanied with the smaller cardiomyocytes. Associated with the structural changes in the heart was a reduction in cardiac output and increased expression of heart cardiac failure (fetal) marker genes, such as ANP, BNP and beta-MHC. Moreover, ultrastructural damage to the myofilaments and mitochondria was observed. Further, the skeletal muscle of the TauTKO mice also exhibited decreased cell volume, structural defects and a reduction of exercise endurance capacity. Importantly, the expression of Hsp70, ATA2 and S100A4, which are upregulated by osmotic stress, was elevated in both heart and skeletal muscle of the TauTKO mice. Taurine depletion causes cardiomyocyte atrophy, mitochondrial and myofiber damage and cardiac dysfunction, effects likely related to the actions of taurine. Our data suggest that multiple actions of taurine, including osmoregulation, regulation of mitochondrial protein expression and inhibition of apoptosis, collectively ensure proper maintenance of cardiac and skeletal muscular structure and function.

Atrogin-1 ubiquitin ligase is upregulated by doxorubicin via p38-MAP kinase in cardiac myocytes

AIMS Doxorubicin (DOX) is one of the most effective anti-neoplastic agents; however, its clinical use is limited by drug-induced cardiomyopathy. The molecular mechanisms responsible for this toxicity remain to be fully addressed. In the present study, we investigated the involvement of atrogin-1, one of the muscle-specific ubiquitin ligases, in DOX-induced cardiotoxicity. METHODS AND RESULTS This method involved intraperitoneal administration of DOX-induced atrogin-1 in the hearts and skeletal muscles of C57BL/6 mice. Consistently, atrogin-1 mRNA was upregulated with DOX treatment in cultured rat neonatal cardiomyocytes. Adenoviral transfer of atrogin-1 induced a reduction in cell size that was ameliorated by the ubiquitin proteasome inhibitor, MG-132. The transduction of constitutively active Akt (caAkt), a serine/threonine protein kinase, inhibited the DOX-mediated induction of atrogin-1. The phosphorylation status of Akt and its downstream target, FOXO, was not affected by DOX. DOX treatment did not activate the atrogin-1 promoter that contains FOXO-binding sites, suggesting that DOX induced atrogin-1 without modulating the Akt/FOXO pathway; importantly, DOX activated p38-mitogen-activated protein kinase (MAPK) and c-Jun N-terminal kinase (JNK). Furthermore, pharmacological inhibition of p38-MAPK, but not JNK, abrogated DOX-mediated induction of atrogin-1. Finally, adenoviral transfer of caAkt inhibited the DOX-induced p38-MAPK activation. CONCLUSIONS DOX induces atrogin-1 through a p38-MAPK-dependent pathway in cardiac myocytes. Constitutive activation of Akt negatively regulates DOX-mediated atrogin-1 induction by inhibiting p38-MAPK activity as a novel mechanism.

Cardiac and skeletal muscle abnormality in taurine transporter-knockout mice

Taurine, a sulfur-containing β-amino acid, is highly contained in heart and skeletal muscle. Taurine has a variety of biological actions, such as ion movement, calcium handling and cytoprotection in the cardiac and skeletal muscles. Meanwhile, taurine deficiency leads various pathologies, including dilated cardiomyopathy, in cat and fox. However, the essential role of taurine depletion on pathogenesis has not been fully clarified. To address the physiological role of taurine in mammalian tissues, taurine transporter-(TauT-) knockout models were recently generated. TauTKO mice exhibited loss of body weight, abnormal cardiac function and the reduced exercise capacity with tissue taurine depletion. In this chapter, we summarize pathological profile and histological feature of heart and skeletal muscle in TauTKO mice.

Leukemia inhibitory factor induces endothelial differentiation in cardiac stem Cells

The importance of interleukin 6 (IL-6)-related cytokines in cardiac homeostasis has been studied extensively; however, little is known about their biological significance in cardiac stem cells. Here we describe that leukemia inhibitory factor (LIF), a member of IL-6-related cytokines, activated STAT3 and ERK1/2 in cardiac Sca-1+ stem cells. LIF stimulation resulted in the induction of endothelial cell-specific genes, including VE-cadherin, Flk-1, and CD31, whereas neither smooth muscle nor cardiac muscle marker genes such as GATA4, GATA6, Nkx-2.5, and calponin were up-regulated. Immunocytochemical examination showed that about 25% of total cells were positively stained with anti-CD31 antibody 14 days after LIF stimulation. Immunofluorescent microscopic analyses identified the Sca-1+ cells that were also positively stained with anti-von Willebrand factor antibody, indicating the differentiating process of Sca-1+ cells into the endothelial cells. IL-6, which did not activate STAT3 and ERK1/2, failed to induce the differentiation of cardiac stem cells into the endothelial cells. In cardiac stem cells, the transduction with dominant negative STAT3 abrogated the LIF-induced endothelial differentiation. And the inhibition of ERK1/2 with the MEK1/2 inhibitor U0126 also prevented the differentiation of Sca-1+ cells into endothelial cells. Thus, both STAT3 and ERK1/2 are required for LIF-mediated endothelial differentiation in cardiac stem cells. Collectively, it is proposed that LIF regulates the commitment of cardiac stem cells into the endothelial cell lineage, contributing to neovascularization in the process of tissue remodeling and/or regeneration.

Myogenic differentiation induces taurine transporter in association with taurine-mediated cytoprotection in skeletal muscles

Skeletal muscle homoeostasis is maintained by a variety of cytoprotective mechanisms. Since ablation of the TauT (taurine transporter) gene results in susceptibility to exercise-induced muscle weakness in vivo, it has been suggested that TauT is essential for skeletal muscle function. However, the regulatory mechanisms of TauT expression remain to be elucidated. In the present study, we demonstrated that TauT was up-regulated during myogenesis in C2C12 cells. Treatment with bFGF (basic fibroblast growth factor), which inhibited muscle differentiation, abrogated myogenic induction of TauT. The promoter activities of TauT were up-regulated during muscle differentiation in C2C12 cells. Database analyses identified an MEF2 (myocyte enhancer binding factor 2) consensus sequence at -844 in the rat TauT gene. Truncation of the promoter region containing the MEF2 site significantly reduced the promoter activity, demonstrating the functional importance of the MEF2 site. Electrophoretic mobility-shift assays confirmed that MEF2 bound to the MEF2 consensus sequence and that DNA-protein complex levels were increased during differentiation. Promoter analyses using mutated promoter-reporter plasmids demonstrated that this site was functional. Importantly, transfection with a MyoD expression vector markedly enhanced TauT promoter activity in the (non-myogenic) 10T1/2 cells. Moreover, co-transfection with an MEF2 expression vector augmented MyoD-induced TauT promoter activity, suggesting that MEF2 is required for full activation of TauT expression. Finally, we examined the effects of taurine on myotube atrophy to clarify the biological significance of the up-regulation of TauT, and demonstrated that taurine attenuated muscle atrophy induced by dexamethasone. TauT expression is regulated under the control of the myogenic programme, and we propose that this is the mechanism for taurine-mediated resistance to muscle atrophy.

Myogenic Induction of Taurine Transporter Prevents Dexamethasone-Induced Muscle Atrophy

We demonstrate for the first time that TauT expression is upregulated during differentiation in C2C12 cells. In addition, taurine have a protective effect on muscle atrophy. Thus, taurine may be a therapeutic agent for cachexic state of diverse pathology.

Molecular mechanisms of cardioprotection by taurine on ischemia-induced apoptosis in cultured cardiomyocytes.

An integral part of the pathogenesis of heart failure is myocyte loss. The traditional explanation for myocyte loss was cell necrosis but there has been a surge of evidence affirming the role of apoptosis in the genesis of heart failure (Garg et al., 2005). Evidence for apoptotic cell death was shown in clinical cases of myocardial infarction, as well as in rabbit, rat, and mouse models of continuous ischemia or ischemia/reperfusion (Garg et al., 2005). It has been shown that the mitochondrial pathways participate in apoptosis induced by ischemia (Garg et al., 2005). Taurine (2-aminoethanesulfonic acid), the -amino acid, is one of the factors that regulates the degree of apoptosis during ischemia (Roysommuti et al., 2003; Schaffer et al., 2003). However, little is known about the cytoprotective signalling pathways mediate this response. We have previously reported that isolated neonatal cardiomyocytes become resistant to ischemia-induced apoptosis when exposed to medium containing 20 mM taurine (Takahashi et al., 2003). In this study, the interaction between taurine and mitochondria-mediated apoptosis is investigated in a newly developed simulated ischemia model utilizing isolated cardiomyocytes that are incubated with medium containing and lacking taurine and then sealed within cultured flasks (Takahashi et al., 2003, Takatani et al., 2004a,b).

TauT gene expression is regulated by TonEBP and plays a role in cell survival.

In summary, this study shows that TonEBP regulates TauT expression in response to hypertonicity, and that the TonEBP/TauT pathway plays an important role in cytoprotection from hypertonic stress. The activation of TonEBP/TauT pathway could contribute to the protective signal pathway against hypertonicity-related tissue damage.