Dissecting the transcriptome in cardiovascular disease

The human transcriptome comprises a complex network of coding and non-coding RNAs implicated in a myriad of biological functions. Non-coding RNAs exhibit highly organized spatial and temporal expression patterns and are emerging as critical regulators of differentiation, homeostasis, and pathological states, including in the cardiovascular system. This review defines the current knowledge gaps, unmet methodological needs, and describes the challenges in dissecting and understanding the role and regulation of the non-coding transcriptome in cardiovascular disease. These challenges include poor annotation of the non-coding genome, determination of the cellular distribution of transcripts, assessment of the role of RNA processing and identification of cell-type specific changes in cardiovascular physiology and disease. We highlight similarities and differences in the hurdles associated with the analysis of the non-coding and protein-coding transcriptomes. In addition, we discuss how the lack of consensus and absence of standardized methods affect reproducibility of data. These shortcomings should be defeated in order to make significant scientific progress and foster the development of clinically applicable non-coding RNA-based therapeutic strategies to lessen the burden of cardiovascular disease

Solid-state laser system for laser cooling of Sodium

We demonstrate a frequency-stabilized, all-solid laser source at 589 nm with up to 800 mW output power. The laser relies on sum-frequency generation from two laser sources at 1064 nm and 1319 nm through a PPKTP crystal in a doubly-resonant cavity. We obtain conversion efficiency as high as 2 W/W^2 after optimization of the cavity parameters. The output wavelength is tunable over 60 GHz, which is sufficient to lock on the Sodium D2 line. The robustness, beam quality, spectral narrowness and tunability of our source make it an alternative to dye lasers for atomic physics experiments with Sodium atoms

Pioglitazone Prevents Capillary Rarefaction in Streptozotocin-Diabetic Rats Independently of Glucose Control and Vascular Endothelial Growth Factor Expression

Background/Aims: Reduction of capillary network density occurs early in the development of metabolic syndrome and may be relevant for the precipitation of diabetes. Agonists of the peroxisome proliferator-activated receptor (PPAR)-gamma transcription factor are vasculoprotective, but their capacity for structural preservation of the microcirculation is unclear. Methods: Male Wistar rats were rendered diabetic by streptozotocin and treated with pioglitazone in chow for up to 12 weeks. Capillary density was determined in heart and skeletal muscle after platelet endothelial cell adhesion molecule-1 (PECAM-1) immunostaining. Hallmarks of apoptosis and angiogenesis were determined. Results: Capillary density deteriorated progressively in the presence of hyperglycemia (from 971/mm(2) to 475/mm(2) in quadriceps muscle during 13 weeks). Pioglitazone did not influence plasma glucose, left ventricular weight, or body weight but nearly doubled absolute and relative capillary densities compared to untreated controls (1.2 vs. 0.6 capillaries/myocyte in heart and 1.5 vs. 0.9 capillaries/myocyte in quadriceps muscle) after 13 weeks of diabetes. No antiapoptotic or angiogenic influence of pioglitazone was detected while a reduced expression of hypoxia-inducible factor-3 alpha and PPAR coactivator-1 alpha (PGC-1 alpha) mRNA as well as vascular endothelial growth factor (VEGF) protein possibly occurred as a consequence of improved vascularization. Conclusion: Pioglitazone preserves microvascular structure in diabetes independently of improvements in glycemic control and by a mechanism unrelated to VEGF-mediated angiogenesis. Copyright (C) 2012 S. Karger AG, Base

Role for substance p-based nociceptive signaling in progenitor cell activation and angiogenesis during ischemia in mice and in human subjects

Our data highlight the role of SP in reparative neovascularization. Nociceptive signaling may represent a novel target of regenerative medicine

Estrogen-dependent dynamic profile of eNOS-DNA associations in prostate cancer

In previous work we have documented the nuclear translocation of endothelial NOS (eNOS) and its participation in combinatorial complexes with Estrogen Receptor Beta (ERβ) and Hypoxia Inducible Factors (HIFs) that determine localized chromatin remodeling in response to estrogen (E2) and hypoxia stimuli, resulting in transcriptional regulation of genes associated with adverse prognosis in prostate cancer (PCa). To explore the role of nuclear eNOS in the acquisition of aggressive phenotype in PCa, we performed ChIP-Sequencing on chromatin-associated eNOS from cells from a primary tumor with poor outcome and from metastatic LNCaP cells. We found that: 1. the eNOS-bound regions (peaks) are widely distributed across the genome encompassing multiple transcription factors binding sites, including Estrogen Response Elements. 2. E2 increased the number of peaks, indicating hormone-dependent eNOS re-localization. 3. Peak distribution was similar with/without E2 with ≈ 55% of them in extragenic DNA regions and an intriguing involvement of the 5′ domain of several miRs deregulated in PCa. Numerous potentially novel eNOS-targeted genes have been identified suggesting that eNOS participates in the regulation of large gene sets. The parallel finding of downregulation of a cluster of miRs, including miR-34a, in PCa cells associated with poor outcome led us to unveil a molecular link between eNOS and SIRT1, an epigenetic regulator of aging and tumorigenicity, negatively regulated by miR-34a and in turn activating eNOS. E2 potentiates miR-34a downregulation thus enhancing SIRT1 expression, depicting a novel eNOS/SIRT1 interplay fine-tuned by E2-activated ER signaling, and suggesting that eNOS may play an important role in aggressive PCa

Gene Deletion of the Kinin Receptor B1 Attenuates Cardiac Inflammation and Fibrosis During the Development of Experimental Diabetic Cardiomyopathy

Objective: Diabetic cardiomyopathy is associated with increased mortality in patients with diabetes mellitus. The underlying pathology of this disease is still under discussion. We studied the role of the kinin B1 receptor on the development of experimental diabetic cardiomyopathy. Research Design and Methods: We utilized B1 receptor knockout mice and investiged cardiac inflammation, fibrosis and oxidative stress after induction of streptozotocin (STZ)-induced diabetes mellitus. Furthermore, the left ventricular function was measured by pressure-volume loops after 8 weeks of diabetes mellitus. Results: B1 receptor knockout mice showed an attenuation of diabetic cardiomyopathy with improved systolic and diastolic function in comparison with diabetic control mice. This was associated with a decreased activation state of the MAP kinase p38, less oxidative stress as well as normalized cardiac inflammation, shown by fewer invading cells and, no increase in matrix metalloproteinase-9 as well as the chemokine CXCL-5. Furthermore, the pro-fibrotic connective tissue growth factor was normalized, leading to a reduction in cardiac fibrosis despite severe hyperglycemia in mice lacking the B1 receptor. Conclusion: These findings suggest that the B1 receptor is detrimental in diabetic cardiomyopathy in that it mediates inflammatory and fibrotic processes. These insights might have useful implications on future studies utilizing B1 receptor antagonists for treatment of human diabetic cardiomyopathy

Intramyocardial Transplantation of Undifferentiated Rat Induced Pluripotent Stem Cells Causes Tumorigenesis in the Heart

BACKGROUND: Induced pluripotent stem cells (iPSCs) are a novel candidate for use in cardiac stem cell therapy. However, their intrinsic tumorigenicity requires further investigation prior to use in a clinical setting. In this study we investigated whether undifferentiated iPSCs are tumorigenic after intramyocardial transplantation into immunocompetent allogeneic recipients. METHODOLOGY/PRINCIPAL FINDINGS: We transplanted 2 × 10(4), 2 × 10(5), or 2 × 10(6) cells from the established rat iPSC line M13 intramyocardially into intact or infarcted hearts of immunocompetent allogeneic rats. Transplant duration was 2, 4, or 6 weeks. Histological examination with hematoxylin-eosin staining confirmed that undifferentiated rat iPSCs could generate heterogeneous tumors in both intracardiac and extracardiac sites. Furthermore, tumor incidence was independent of cell dose, transplant duration, and the presence or absence of myocardial infarction. CONCLUSIONS/SIGNIFICANCE: Our study demonstrates that allogeneic iPSC transplantation in the heart will likely result in in situ tumorigenesis, and that cells leaked from the beating heart are a potential source of tumor spread, underscoring the importance of evaluating the safety of future iPSC therapy for cardiac disease

Cardioprotective Effect of Nicorandil, a Mitochondrial ATP-Sensitive Potassium Channel Opener, Prolongs Survival in HSPB5 R120G Transgenic Mice

BACKGROUND: Transgenic (TG) mice with overexpression of an arg120gly (R120G) missense mutation in HSPB5 display desmin-related cardiomyopathy, which is characterized by formation of aggresomes. It is also known that progressive mitochondrial abnormalities and apoptotic cell death occur in the hearts of R120G TG mice. The role of mitochondrial dysfunction and apoptosis in disease progression, however, remains uncertain. METHODS AND RESULTS: Mitochondrial abnormalities and apoptotic cell death induced by overexpression of HSPB5 R120G were analyzed in neonatal rat cardiomyocytes. Overexpression of mutant HSPB5 led to development of aggresomes with a concomitant reduction in cell viability in the myocytes. Overexpression of mutant HSPB5 induced a reduction in the cytochrome c level in the mitochondrial fraction and a corresponding increase in the cytoplasmic fraction in the myocytes. Down-regulation of BCL2 and up-regulation of BAX were detected in the myocytes expressing the mutant HSPB5. Concomitant with mitochondrial abnormality, the activation of caspase-3 and increased apoptotic cell death was observed. Cell viability was dose-dependently recovered in myocytes overexpressing HSPB5 R120G by treatment with nicorandil a mitochondrial ATP-sensitive potassium channel opener. Nicorandil treatment also inhibited the increase in BAX, the decrease in BCL2, activation of caspase-3 and apoptotic cell death by mutant HSPB5. To confirm the results of the in-vitro study, we analyzed the effect of nicorandil in HSPB5 R120G TG mice. Nicorandil treatment appeared to reduce mitochondrial impairment and apoptotic cell death and prolonged survival in HSPB5 R120G TG mice. CONCLUSIONS: Nicorandil may prolong survival in HSPB5 R120G TG mice by protecting against mitochondrial impairments

Significance of Thymosin β4 and Implication of PINCH-1-ILK-α-Parvin (PIP) Complex in Human Dilated Cardiomyopathy

Myocardial remodeling is a major contributor in the development of heart failure (HF) after myocardial infarction (MI). Integrin-linked kinase (ILK), LIM-only adaptor PINCH-1, and α-parvin are essential components of focal adhesions (FAs), which are highly expressed in the heart. ILK binds tightly to PINCH-1 and α-parvin, which regulates FA assembly and promotes cell survival via the activation of the kinase Akt. Mice lacking ILK, PINCH or α-parvin have been shown to develop severe defects in the heart, suggesting that these proteins play a critical role in heart function. Utilizing failing human heart tissues (dilated cardiomyopathy, DCM), we found a 2.27-fold (p<0.001) enhanced expression of PINCH, 4 fold for α-parvin, and 10.5 fold (p<0.001) for ILK as compared to non-failing (NF) counterparts. No significant enhancements were found for the PINCH isoform PINCH-2 and parvin isoform β-parvin. Using a co-immunoprecipitation method, we also found that the PINCH-1-ILK-α-parvin (PIP) complex and Akt activation were significantly up-regulated. These observations were further corroborated with the mouse myocardial infarction (MI) and transaortic constriction (TAC) model. Thymosin beta4 (Tβ4), an effective cell penetrating peptide for treating MI, was found to further enhance the level of PIP components and Akt activation, while substantially suppressing NF-κB activation and collagen expression—the hallmarks of cardiac fibrosis. In the presence of an Akt inhibitor, wortmannin, we show that Tβ4 had a decreased effect in protecting the heart from MI. These data suggest that the PIP complex and activation of Akt play critical roles in HF development. Tβ4 treatment likely improves cardiac function by enhancing PIP mediated Akt activation and suppressing NF-κB activation and collagen-mediated fibrosis. These data provide significant insight into the role of the PIP-Akt pathway and its regulation by Tβ4 treatment in post-MI