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The association between dilated cardiomyopathy (DCM) and low thyroid hormone (TH) levels has been previously described. In these patients abnormal thyroid function is significantly related to impaired left ventricular (LV) function and increased risk of death. Although TH was originally thought to be produced exclusively by the thyroid gland, we recently reported TH biosynthesis in the human ischemic heart.Based on these findings, we evaluated whether the genes required for TH production are also altered in patients with DCM.Twenty-three LV tissue samples were obtained from patients with DCM (n = 13) undergoing heart transplantation and control donors (n = 10), and used for RNA sequencing analysis. The number of LV DCM samples was increased to 23 to determine total T4 and T3 tissue levels by ELISA.We found that all components of TH biosynthesis are expressed in human dilated heart tissue. Expression of genes encoding thyroperoxidase (-2.57-fold, P < 0.05) and dual oxidase 2 (2.64-fold, P < 0.01), the main enzymatic system of TH production, was significantly altered in patients with DCM and significantly associated with LV remodeling parameters. Thyroxine (T4) cardiac tissue levels were significantly increased (P < 0.01), whilst triiodothyronine (T3) levels were significantly diminished (P < 0.05) in the patients.Expression of TH biosynthesis machinery in the heart and total tissue levels of T4 and T3, are altered in patients with DCM. Given the relevance of TH in cardiac pathology, our results provide a basis for new gene-based therapeutic strategies for treating DCM.

Alterations in myocardial structure and reduced cardiomyocyte adhesions have been previously described in dilated cardiomyopathy (DCM). We studied the transcriptome of cell adhesion molecules in these patients and their relationships with left ventricular (LV) function decay. We also visualized the intercalated disc (ID) structure and organization. The transcriptomic profile of 23 explanted LV samples was analyzed using RNA-sequencing (13 DCM, 10 control [CNT]), focusing on cell adhesion genes. Electron microscopy analysis to visualize ID structural differences and immunohistochemistry experiments of ID proteins was also performed. RT-qPCR and western blot experiments were carried out on ID components. We found 29 differentially expressed genes, most of all, constituents of the ID structure. We found that the expression of GJA3, DSP and CTNNA3 was directly associated with LV ejection fraction (r = 0.741, P = 0.004; r = 0.674, P = 0.011 and r = 0.565, P = 0.044, respectively), LV systolic (P = 0.003, P = 0.003, P = 0.028, respectively) and diastolic dimensions (P = 0.006, P = 0.001, P = 0.025, respectively). Electron microscopy micrographs showed a reduced ID convolution index and immunogold labeling of connexin 46 (GJA gene), desmoplakin (DSP gene) and catenin α-3 (CTNNA3 gene) proteins in DCM patients. Moreover, we observed that protein and mRNA levels analyzed by RT-qPCR of these ID components were diminished in DCM group. In conclusion, we report significant gene and protein expression changes and found that the ID components GJA3, DSP and CTNNA3 were highly related to LV function. Microscopic observations indicated that ID is structurally compromised in these patients. These findings give new data for understanding the ventricular depression that characterizes DCM, opening new therapeutic perspectives for these critically diseased patients.

The Macaronesian Scrophularia lowei is hypothesized to have arisen from the widespread S. arguta on the basis of several phylogenetic studies of the genus, but sampling has been limited. Although these two annual species are morphologically distinct, the origin of S. lowei is unclear because genetic studies focused on this Macaronesian species are lacking. We studied 5 S. lowei and 25 S. arguta populations to determine the relationship of both species and to infer the geographical origin of S. lowei. The timing of S. lowei divergence and differentiation was inferred by dating analysis of the ITS region. A phylogenetic analysis of two nuclear (ITS and ETS) and two chloroplast (psbJ-petA and psbA-trnH) DNA regions was performed to study the relationship between the two species, and genetic differentiation was analysed by AMOVA. Haplotype network construction and Bayesian phylogeographic analysis were conducted using chloroplast DNA regions and a spatial clustering analysis was carried out on a combined dataset of all studied regions. Our results indicate that both species constitute a well-supported clade that diverged in the Miocene and differentiated in the Late Miocene-Pleistocene. Although S. lowei constitutes a well-supported clade according to nDNA, cpDNA revealed a close relationship between S. lowei and western Canarian S. arguta, a finding supported by the spatial clustering analysis. Both species have strong population structure, with most genetic variability explained by inter-population differences. Our study therefore supports a recent peripatric speciation of S. lowei-a taxon that differs morphologically and genetically at the nDNA level from its closest relative, S. arguta, but not according to cpDNA, from the closest Macaronesian populations of that species. In addition, a recent dispersal of S. arguta to Madeira from Canary Islands or Selvagens Islands and a rapid morphological differentiation after the colonization to generate S. lowei is the most likely hypothesis to explain the origin of the last taxon.

Intercalated disks are unique structures in cardiac tissue, in which adherens junctions, desmosomes, and GAP junctions co-localize, thereby facilitating cardiac muscle contraction and function. Protocadherins are involved in these junctions; however, their role in heart physiology is poorly understood. We aimed to analyze the transcriptomic profile of adhesion molecules in patients with ischemic cardiomyopathy (ICM) and relate the changes uncovered with the hemodynamic alterations and functional depression observed in these patients.Twenty-three left ventricular tissue samples from patients diagnosed with ICM (n = 13) undergoing heart transplantation and control donors (CNT, n = 10) were analyzed using RNA sequencing. Forty-two cell adhesion genes involved in cellular junctions were differentially expressed in ICM myocardium. Notably, the levels of protocadherin PCDHGA3 were related with the stroke volume (r = -0.826, P = 0.003), ejection fraction (r = -0.793, P = 0.004) and left ventricular end systolic and diastolic diameters (r = 0.867, P = 0.001; r = 0.781, P = 0.005, respectively).Our results support the importance of intercalated disks molecular alterations, closely involved in the contractile function, highlighting its crucial significance and showing gene expression changes not previously described. Specifically, altered PCDHGA3 gene expression was strongly associated with reduced stroke volume and ventricular dysfunction in ICM, suggesting a relevant role in hemodynamic perturbations and cardiac performance for this unexplored protocadherin.

In dilated cardiomyopathy (DCM), cardiac failure is accompanied by profound alterations of extracellular matrix associated with the progression of cardiac dilation and left ventricular (LV) dysfunction. Recently, we reported alterations of non-fibrillar collagen expression in ischemic cardiomyopathy linked to fibrosis and cardiac remodeling. We suspect that expression changes in genes coding for non-fibrillar collagens may have a potential role in DCM development.This study sought to analyze changes in the expression profile of non-fibrillar collagen genes in patients with DCM and to examine relationships between cardiac remodeling parameters and the expression levels of these genes.Twenty-three human left ventricle tissue samples were obtained from DCM patients (n = 13) undergoing heart transplantation and control donors (n = 10) for RNA sequencing analysis. We found increased mRNA levels of six non-fibrillar collagen genes, such as COL4A5, COL9A1, COL21A1, and COL23A1 (P < 0.05 for all), not previously described in DCM. Protein levels of COL8A1 and COL16A1 (P < 0.05 for both), were correspondingly increased. We also identified TGF-β1 significantly upregulated and related to both COL8A1 and COL16A1. Interestingly, we found a significant relationship between LV mass index and the gene expression level of COL8A1 (r = 0.653, P < 0.05).In our research, we identified new non-fibrillar collagens with altered expression in DCM, being COL8A1 overexpression directly related to LV mass index, suggesting that they may be involved in the progression of cardiac dilation and remodeling.

Disruptions in cardiac ion channels have shown to influence the impaired cardiac contraction in heart failure. We sought to determine the altered gene expression profile of this category in dilated cardiomyopathy (DCM) patients and relate the altered gene expression with the clinical signs present in our patients, such as ventricular dysfunction and sustained monomorphic ventricular tachycardia (SMVT).Left ventricular (LV) tissue samples were used in RNA-sequencing technique to elucidate the transcriptomic changes of 13 DCM patients compared to controls (n = 10). We analyzed the differential gene expression of cardiac ion channels, and we found a total of 34 altered genes. We found that the calcium channel CACNG8 mRNA and protein levels were down-regulated and highly and inversely related with LV ejection fraction (LVEF) (r = -0.78, P

BACKGROUND: The endoplasmic reticulum (ER) is a multifunctional organelle responsible for the synthesis and folding of proteins as well as for signalling and calcium storage, that has been linked to the contraction-relaxation process. Perturbations of its homeostasis activate a stress response in diseases such as heart failure (HF). To elucidate the alterations in ER molecular components, we analyze the levels of ER stress and structure proteins in human dilated (DCM) and ischemic (ICM) cardiomyopathies, and its relationship with patient's functional status. METHODS AND RESULTS: We examined 52 explanted human hearts from DCM (n = 21) and ICM (n = 21) subjects and 10 non-failing hearts as controls. Our results showed specific changes in stress (IRE1, p

Dilated cardiomyopathy (DCM) is a public health problem with no available curative treatment, and mitochondrial dysfunction plays a critical role in its development. The present study is the first to analyze the mitochondrial proteome in cardiac tissue of patients with DCM to identify potential molecular targets for its therapeutic intervention.16 left ventricular (LV) samples obtained from explanted human hearts with DCM (n = 8) and control donors (n = 8) were extracted to perform a proteomic approach to investigate the variations in mitochondrial protein expression. The proteome of the samples was analyzed by quantitative differential electrophoresis and Mass Spectrometry. These changes were validated by classical techniques and by novel and precise selected reaction monitoring analysis and RNA sequencing approach increasing the total heart samples up to 25. We found significant alterations in energy metabolism, especially in molecules involved in substrate utilization (ODPA, ETFD, DLDH), energy production (ATPA), other metabolic pathways (AL4A1) and protein synthesis (EFTU), obtaining considerable and specific relationships between the alterations detected in these processes. Importantly, we observed that the antioxidant PRDX3 overexpression is associated with impaired ventricular function. PRDX3 is significantly related to LV end systolic and diastolic diameter (r = 0.73, p value

BACKGROUND: The atrium is the major site of ANP synthesis, which has been said to increase in heart failure as a result of increased production in the left ventricular (LV) myocardium. This is a key issue related to its diagnostic and prognostic capabilities. We aimed to evaluate protein levels of proANP and ANP and the enzymes that cleave the natriuretic peptides, corin and furin, in the LV tissue of heart transplant patients with dilated (DCM) and ischemic (ICM) cardiomyopathy compared with control donors (CNT). We also evaluate mRNA levels of ANP gene (NPPA) by RNA sequencing in the same tissue. METHODS AND RESULTS: Seventy-three human LV tissue samples from ICM (n=30) and DCM (n=33) patients and CNT (n=10) were analyzed by Western blot and RNA sequencing. Comparing protein levels according to etiology, neither DCM nor ICM showed levels of proANP or ANP different from those of CNT. However, NPPA was increased in both groups compared to CNT (DCM 32 fold, p

BACKGROUND: Dilated cardiomyopathy (DCM) is characterized by idiopathic dilation and systolic contractile dysfunction of the cardiac chambers. The present work aimed to study the alterations in gene expression of ion channels involved in cardiomyocyte function. METHODS AND RESULTS: Microarray profiling using the Affymetrix Human Gene® 1.0 ST array was performed using 17 RNA samples, 12 from DCM patients undergoing cardiac transplantation and 5 control donors (CNT). The analysis focused on 7 cardiac ion channel genes, since this category has not been previously studied in human DCM. SCN2B was upregulated, while KCNJ5, KCNJ8, CLIC2, CLCN3, CACNB2, and CACNA1C were downregulated. The RT-qPCR (21 DCM and 8 CNT samples) validated the gene expression of SCN2B (p < 0.0001), KCNJ5 (p < 0.05), KCNJ8 (p < 0.05), CLIC2 (p < 0.05), and CACNB2 (p < 0.05). Furthermore, we performed an IPA analysis and we found a functional relationship between the different ion channels studied in this work. CONCLUSION: This study shows a differential expression of ion channel genes involved in cardiac contraction in DCM that might partly underlie the changes in left ventricular function observed in these patients. These results could be the basis for new genetic therapeutic approaches.

Recent studies in human fibroblasts have provided a new general paradigm of tumor suppression according to which oncogenic signaling produces DNA damage and this, in turn, results in ATM/p53-dependent cellular senescence. Here, we have tested this model in a variety of murine experimental systems. Overexpression of oncogenic Ras in murine fibroblasts efficiently induced senescence but this occurred in the absence of detectable DNA damage signaling, thus suggesting a fundamental difference between human and murine cells. Moreover, lung adenomas initiated by endogenous levels of oncogenic K-Ras presented abundant senescent cells, but undetectable DNA damage signaling. Accordingly, K-Ras-driven adenomas were also senescent in Atm-null mice, and the tumorigenic progression of these lesions was only modestly accelerated by Atm-deficiency. Finally, we have examined chemically-induced fibrosarcomas, which possess a persistently activated DNA damage response and are highly sensitive to the activity of p53. We found that the absence of Atm favored genomic instability in the resulting tumors, but did not affect the persistent DNA damage response and did not impair p53-dependent tumor suppression. All together, we conclude that oncogene-induced senescence in mice may occur in the absence of a detectable DNA damage response. Regarding murine Atm, our data suggest that it plays a minor role in oncogene-induced senescence or in p53-dependent tumor suppression, being its tumor suppressive activity probably limited to the maintenance of genomic stability.