Modulation of Fatty Acid-Related Genes in the Response of H9c2 Cardiac Cells to Palmitate and n-3 Polyunsaturated Fatty Acids

While high levels of saturated fatty acids are associated with impairment of cardiovascular functions, n-3 polyunsaturated fatty acids (PUFAs) have been shown to exert protective effects. However the molecular mechanisms underlying this evidence are not completely understood. In the present study we have used rat H9c2 ventricular cardiomyoblasts as a cellular model of lipotoxicity to highlight the effects of palmitate, a saturated fatty acid, on genetic and epigenetic modulation of fatty acid metabolism and fate, and the ability of PUFAs, eicosapentaenoic acid, and docosahexaenoic acid, to contrast the actions that may contribute to cardiac dysfunction and remodeling. Treatment with a high dose of palmitate provoked mitochondrial depolarization, apoptosis, and hypertrophy of cardiomyoblasts. Palmitate also enhanced the mRNA levels of sterol regulatory element-binding proteins (SREBPs), a family of master transcription factors for lipogenesis, and it favored the expression of genes encoding key enzymes that metabolically activate palmitate and commit it to biosynthetic pathways. Moreover, miR-33a, a highly conserved microRNA embedded in an intronic sequence of the SREBP2 gene, was co-expressed with the SREBP2 messenger, while its target carnitine palmitoyltransferase-1b was down-regulated. Manipulation of the levels of miR-33a and SREBPs allowed us to understand their involvement in cell death and hypertrophy. The simultaneous addition of PUFAs prevented the effects of palmitate and protected H9c2 cells. These results may have implications for the control of cardiac metabolism and dysfunction, particularly in relation to dietary habits and the quality of fatty acid intake

Nitric oxide can function as either a killer molecule or an antiapoptotic effector in cardiomyocytes

AbstractCaspase enzymes are a family of cysteine proteases that play a central role in apoptosis. Recently, it has been demonstrated that caspases can be S-nitrosylated and inhibited by nitric oxide (NO). The present report shows that in chick embryo heart cells (CEHC), NO donor molecules such as S-nitroso-N-acetylpenicillamine (SNAP), S-nitrosoglutathione, spermine-NO or sodium nitroprusside inhibit caspase activity in both basal and staurosporine-treated cells. However, the inhibitory effect of NO donors on caspase activity is accompanied by a parallel cytotoxic effect, that precludes NO to exert its antiapoptotic capability. N-Acetylcysteine (NAC) at a concentration of 10 mM blocks depletion of cellular glutathione and cell death in SNAP-treated CEHC, but it poorly affects the ability of SNAP to inhibit caspase activity. Consequently, in the presence of NAC, SNAP attenuates not only caspase activity but also cell death of staurosporine-treated CEHC. These data show that changes in the redox environment may inhibit NO-mediated toxicity, without affecting the antiapoptotic capability of NO, mediated by inhibition of caspase enzymes. NO may thus be transformed from a killer molecule into an antiapoptotic agent

Evidence that AMP-activated protein kinase can negatively modulate Ornithine decarboxylase activity in cardiac myoblasts

AbstractThe responses of AMP-activated protein kinase (AMPK) and Ornithine decarboxylase (ODC) to isoproterenol have been examined in H9c2 cardiomyoblasts, AMPK represents the link between cell growth and energy availability whereas ODC, the key enzyme in polyamine biosynthesis, is essential for all growth processes and it is thought to have a role in the development of cardiac hypertrophy. Isoproterenol rapidly induced ODC activity in H9c2 cardiomyoblasts by promoting the synthesis of the enzyme protein and this effect was counteracted by inhibitors of the PI3K/Akt pathway. The increase in enzyme activity became significant between 15 and 30min after the treatment. At the same time, isoproterenol stimulated the phosphorylation of AMPKα catalytic subunits (Thr172), that was associated to an increase in acetyl coenzyme A carboxylase (Ser72) phosphorylation. Downregulation of both α1 and α2 isoforms of the AMPK catalytic subunit by siRNA to knockdown AMPK enzymatic activity, led to superinduction of ODC in isoproterenol-treated cardiomyoblasts. Downregulation of AMPKα increased ODC activity even in cells treated with other adrenergic agonists and in control cells. Analogue results were obtained in SH-SY5Y neuroblastoma cells transfected with a shRNA construct against AMPKα. In conclusion, isoproterenol quickly activates in H9c2 cardiomyoblasts two events that seem to contrast one another. The first one, an increase in ODC activity, is linked to cell growth, whereas the second, AMPK activation, is a homeostatic mechanism that negatively modulates the first. The modulation of ODC activity by AMPK represents a mechanism that may contribute to control cell growth processes

Are pre-miR-146a and PTTG1 associated with papillary thyroid cancer?

Papillary thyroid carcinoma (PTC) is the most common endocrine malignancy, with a steadily increasing incidence in the last few decades worldwide. The predisposition to developing this carcinoma by the heterozygous state of rs2910164 within the precursor of the miR-146a has been reported, but recently not confirmed. Interestingly, on the same chromosome, almost 50 kb separate the pre-miR-146a from the pituitary tumor-transforming gene 1 (PTTG1), a proto-oncogene involved in several tumors, including thyroid cancers. In this study, we analyzed, using a case–control design, the genetic association between PTC and the genomic region encompassing pre-miR-146a rs2910164 and PTTG1 rs1862391 and rs2910202. We enrolled 307 affected patients and 206 healthy controls. The possible presence of thyroid nodules in controls was excluded by ultrasonography. All the cases were submitted to single- nucleotide polymorphism (SNP) genotyping of pre-miR-146a and PTTG1, and risk association analyses were carried out. The genotypic and allelic frequencies of pre-miR-146a rs2910164 were not statistically different in the patients and controls, and this SNP was not in linkage disequilibrium with the investigated PTTG1 SNPs. Consistently, meta-analyses, the first including all the affected cases published to date, did not confirm the previously reported association of the heterozygous CG genotype with PTC. The PTTG1 SNPs exhibited the same allelic frequency in the patients and controls and were not associated with the disease. In conclusion, in a well-selected Italian population, neither pre-miR-146a rs2910164 nor PTTG1 rs1862391 and rs2910202 were found to be associated with the risk of developing PTC

Antiapoptotic and Antiautophagic Effects of Eicosapentaenoic Acid in Cardiac Myoblasts Exposed to Palmitic Acid

Apoptosis is a programmed cell death that plays a critical role in cell homeostasis. In particular, apoptosis in cardiomyocytes is involved in several cardiovascular diseases including heart failure. Recently autophagy has emerged as an important modulator of programmed cell death pathway. Recent evidence indicates that saturated fatty acids induce cell death through apoptosis and this effect is specific for palmitate. On the other hand, n-3 polyunsaturated fatty acids (PUFAs) have been implicated in the protection against cardiovascular diseases, cardiac ischemic damage and myocardial dysfunction. In the present study we show that n-3 PUFA eicosapentaenoic acid (EPA) treatment to culture medium of H9c2 rat cardiomyoblasts protects cells against palmitate-induced apoptosis, as well as counteracts palmitate-mediated increase of autophagy. Further investigation is required to establish whether the antiautophagic effect of EPA may be involved in its cytoprotective outcome and to explore the underlying biochemical mechanisms through which palmitate and EPA control the fate of cardiac cells