ResearchPad - cardiovascular-physiology Default RSS Feed en-us © 2020 Newgen KnowledgeWorks <![CDATA[Plasma Galectin-3 predicts deleterious vascular dysfunction affecting post-myocardial infarction patients: An explanatory study]]> In a previous analysis of a post-myocardial infarction (MI) cohort, abnormally high systemic vascular resistances (SVR) were shown to be frequently revealed by MRI during the healing period, independently of MI severity, giving evidence of vascular dysfunction and limiting further recovery of cardiac function. The present ancillary and exploratory analysis of the same cohort was aimed at characterizing those patients suffering from high SVR remotely from MI with a large a panel of cardiovascular MRI parameters and blood biomarkers.MethodsMRI and blood sampling were performed 2–4 days after a reperfused MI and 6 months thereafter in 121 patients. SVR were monitored with a phase-contrast MRI sequence and patients with abnormally high SVR at 6-months were characterized through MRI parameters and blood biomarkers, including Galectin-3, an indicator of cardiovascular inflammation and fibrosis after MI. SVR were normal at 6-months in 90 patients (SVR-) and abnormally high in 31 among whom 21 already had high SVR at the acute phase (SVR++) while 10 did not (SVR+).ResultsWhen compared with SVR-, both SVR+ and SVR++ exhibited lower recovery in cardiac function from baseline to 6-months, while baseline levels of Galectin-3 were significantly different in both SVR+ (median: 14.4 (interquartile range: 12.3–16.7) ng.mL-1) and SVR++ (13.0 (11.7–19.4) ng.mL-1) compared to SVR- (11.7 (9.8–13.5) ng.mL-1, both p < 0.05). Plasma Galectin-3 was an independent baseline predictor of high SVR at 6-months (p = 0.002), together with the baseline levels of SVR and left ventricular end-diastolic volume, whereas indices of MI severity and left ventricular function were not. In conclusion, plasma Galectin-3 predicts a deleterious vascular dysfunction affecting post-MI patients, an observation that could lead to consider new therapeutic targets if confirmed through dedicated prospective studies. ]]> <![CDATA[Ion currents, action potentials, and noradrenergic responses in rat pulmonary vein and left atrial cardiomyocytes]]> Effects of noradrenaline on action potential (AP) repolarisation in PV cardiomyocytes. A Representative AP recording from a PV cardiomyocyte in control solution and in the presence of 1 μM noradrenaline (NA). B Representative AP recording from another PV cardiomyocyte in control solution and in the presence of 1 μM NA showing lack of AP prolongation. C Before‐and‐after plot of APD30 in PV cardiomyocytes in control solution and after superfusion with 1 μM NA. D Noradrenaline‐induced fold‐changes in APD30 in LA and PV cardiomyocytes.

<![CDATA[The effect of age on cerebral blood flow responses during repeated and sustained stand to sit transitions]]> Aging and age‐related cerebrovascular diseases are associated with impaired cerebrovascular function. The novel finding of this study is that older adults showed higher pressure–flow responses during repeated stand–sit transitions compared to younger adults, and that a high pressure–flow response was associated with low cerebrovascular reactivity to CO2. These results suggest that impaired vascular function and increased arterial stiffness may contribute to the pressure–flow responses observed in the older adults.

<![CDATA[The natural drug DIAVIT is protective in a type II mouse model of diabetic nephropathy]]>

There is evidence to suggest that abnormal angiogenesis, inflammation, and fibrosis drive diabetic nephropathy (DN). However, there is no specific treatment to counteract these processes. We aimed to determine whether DIAVIT, a natural Vaccinium myrtillus (blueberry) and Hippophae Rhamnoides (sea buckthorn) extract, is protective in a model of type II DN. Diabetic db/db mice were administered DIAVIT in their drinking water for 14 weeks. We assessed the functional, structural, and ultra-structural phenotype of three experimental groups (lean+vehicle, db/db+vehicle, db/db+DIAVIT). We also investigated the angiogenic and fibrotic pathways involved in the mechanism of action of DIAVIT. Diabetic db/db mice developed hyperglycaemia, albuminuria, and an increased glomerular water permeability; the latter two were prevented by DIAVIT. db/db mice developed fibrotic glomeruli, endothelial insult, and glomerular ultra-structural changes, which were not present in DIAVIT-treated mice. Vascular endothelial growth factor A (VEGF-A) splicing was altered in the db/db kidney cortex, increasing the pro-angiogenic VEGF-A165 relative to the anti-angiogenic VEGF-A165b. This was partially prevented with DIAVIT treatment. Delphinidin, an anthocyanin abundant in DIAVIT, increased the VEGF-A165b expression relative to total VEGF-A165 in cultured podocytes through phosphorylation of the splice factor SRSF6. DIAVIT, in particular delphinidin, alters VEGF-A splicing in type II DN, rescuing the DN phenotype. This study highlights the therapeutic potential of natural drugs in DN through the manipulation of gene splicing and expression.

<![CDATA[Evaluation of upconverting nanoparticles towards heart theranostics]]>

Restricted and controlled drug delivery to the heart remains a challenge giving frequent off-target effects as well as limited retention of drugs in the heart. There is a need to develop and optimize tools to allow for improved design of drug candidates for treatment of heart diseases. Over the last decade, novel drug platforms and nanomaterials were designed to confine bioactive materials to the heart. Yet, the research remains in its infancy, not only in the development of tools but also in the understanding of effects of these materials on cardiac function and tissue integrity. Upconverting nanoparticles are nanomaterials that recently accelerated interest in theranostic nanomedicine technologies. Their unique photophysical properties allow for sensitive in vivo imaging that can be combined with spatio-temporal control for targeted release of encapsulated drugs.

Here we synthesized upconverting NaYF4:Yb,Tm nanoparticles and show for the first time their innocuity in the heart, when injected in the myocardium or in the pericardial space in mice. Nanoparticle retention and upconversion in the cardiac region did not alter heart rate variability, nor cardiac function as determined over a 15-day time course ensuing the sole injection. Altogether, our nanoparticles show innocuity primarily in the pericardial region and can be safely used for controlled spatiotemporal drug delivery. Our results support the use of upconverting nanoparticles as potential theranostics tools overcoming some of the key limitations associated with conventional experimental cardiology.

<![CDATA[Association between sublingual microcirculation, tissue perfusion and organ failure in major trauma: A subgroup analysis of a prospective observational study]]>


Previous studies described impaired microvascular perfusion and tissue oxygenation as reliable predictors of Multiple Organ Failure in major trauma. However, this relationship has been incompletely investigated. The objective of this analysis is to further evaluate the association between organ dysfunction and microcirculation after trauma.

Materials and methods

This is a retrospective subgroup analysis on 28 trauma patients enrolled for the Microcirculation DAIly MONitoring in critically ill patients study (NCT 02649088). Patients were divided in two groups according with their Sequential Organ Failure Assessment (SOFA) score at day 4. At admission and every 24 hours, the sublingual microcirculation was evaluated with Sidestream Darkfield Imaging (SDF) and peripheral tissue perfusion was assessed with Near Infrared Spectroscopy (NIRS) and Vascular Occlusion Test (VOT). Simultaneously, hemodynamic, clinical/laboratory parameters and main organ supports were collected.


Median SOFA score at Day 4 was 6.5. Accordingly, patients were divided in two groups: D4-SOFA ≤6.5 and D4-SOFA >6.5. The Length of Stay in Intensive Care was significantly higher in patients with D4-SOFA>6.5 compared to D4-SOFA≤6.5 (p = 0.013). Total Vessel Density of small vessels was significantly lower in patients with high D4-SOFA score at Day 1 (p = 0.002) and Day 2 (p = 0.006) after admission; the Perfused Vessel Density was lower in patients with high D4-SOFA score at Day 1 (p = 0.007) and Day 2 (p = 0.033). At Day 1, NIRS monitoring with VOT showed significantly faster tissue oxygen saturation downslope (p = 0.018) and slower upslope (p = 0.04) in patients with high D4-SOFA.


In our cohort of major traumas, sublingual microcirculation and peripheral microvascular reactivity were significantly more impaired early after trauma in those patients who developed more severe organ dysfunctions. Our data would support the hypothesis that restoration of macrocirculation can be dissociated from restoration of peripheral and tissue perfusion, and that microvascular alterations can be associated with organ failure.

<![CDATA[SIRT5 deficiency suppresses mitochondrial ATP production and promotes AMPK activation in response to energy stress]]>

Sirtuin 5 (SIRT5) is a member of the NAD+-dependent sirtuin family of protein deacylase that catalyzes removal of post-translational modifications, such as succinylation, malonylation, and glutarylation on lysine residues. In light of the SIRT5's roles in regulating mitochondrion function, we show here that SIRT5 deficiency leads to suppression of mitochondrial NADH oxidation and inhibition of ATP synthase activity. As a result, SIRT5 deficiency decreases mitochondrial ATP production, increases AMP/ATP ratio, and subsequently activates AMP-activated protein kinase (AMPK) in cultured cells and mouse hearts under energy stress conditions. Moreover, Sirt5 knockout attenuates transverse aortic constriction (TAC)-induced cardiac hypertrophy and cardiac dysfunction in mice, which is associated with decreased ATP level, increased AMP/ATP ratio and enhanced AMPK activation. Our study thus uncovers an important role of SIRT5 in regulating cellular energy metabolism and AMPK activation in response to energy stress.

<![CDATA[Prediction of all-cause mortality after liver transplantation using left ventricular systolic and diastolic function assessment]]>

Although pretransplant cardiac dysfunction is considered a major predictor of poor outcomes after liver transplantation (LT), the ability of left ventricular (LV) systolic/diastolic function (LVSF/LVDF), together or individually, to predict mortality after LT is poorly characterized. We retrospectively evaluated pretransplant clinical and Doppler echocardiographic data of 839 consecutive LT recipients from 2009 to 2012 aged 18–60 years. The primary endpoint was all-cause mortality at 4 years. The overall survival rate was 91.2%. In multivariate Cox analysis, reduced LV ejection fraction (LVEF, P = 0.014) and decreased transmitral E/A ratio(P = 0.022) remained significant prognosticators. In LVSF analysis, patients with LVEF≤60% (quartile [Q]1) had higher mortality than those with LVEF>60% (hazard ratio = 1.90, 95% confidence interval = 1.15–3.15, P = 0.012). In LVDF analysis, patients with an E/A ratio<0.9(Q1) had a 2.19-fold higher risk of death (95% confidence interval = 1.11–4.32, P = 0.024) than those with an E/A ratio>1.4(Q4). In combined LVDF and LVSF analysis, patients with an E/A ratio<0.9 and LVEF≤60% had poorer survival outcomes than patients with an E/A ratio≥0.9 and LVEF>60% (79.5% versus 93.3%, P = 0.001). Patients with an early mitral inflow velocity/annular velocity (E/e’ ratio)>11.5(Q4) and LV stroke volume index (LVSVI)<33mL/m2(Q1) showed worse survival than those with an E/e’ ratio≤11.5 and LVSVI ≥33mL/m2(78.4% versus 92.2%, P = 0.003). A combination of LVSF and LVDF is a better predictor of survival than LVSF or LVDF alone.

<![CDATA[Regeneration-associated cell transplantation contributes to tissue recovery in mice with acute ischemic stroke]]>

Various cell-based therapeutic strategies have been investigated for vascular and tissue regeneration after ischemic stroke. We have developed a novel cell population, called regeneration-associated cells (RACs), by quality- and quantity-controlled culture of unfractionated mononuclear cells. RACs were trans-arterially injected into 10-week-old syngeneic male mice at 1, 3, 5 or 7 days after permanent middle cerebral artery occlusion (MCAO) to determine the optimal timing for administration in terms of outcome at day 21. Next, we examined the effects of RACs injection at day 1 after MCAO on neurological deficits, infarct volume, and mediators of vascular regeneration and anti-inflammation at days 7 and 21. Infarct volume at day 21 was significantly reduced by transplantation of RACs at day 1 or 3. RACs injected at day 1 reduced the infarct volume at day 7 and 21. Angiogenesis and anti-inflammatory mediators, VEGF and IL-10, were increased at day 7, and VEGF was still upregulated at day 21. We also observed significantly enhanced ink perfusion in vivo, tube formation in vitro, and definitive endothelial progenitor cell colonies in colony assay. These results suggest that RAC transplantation in MCAO models promoted significant recovery of neural tissues through intensified anti-inflammatory and angiogenic effects.

<![CDATA[Prognostic value of uPAR expression and angiogenesis in primary and metastatic melanoma]]>

Angiogenesis is important for the progression of cutaneous melanoma. Here, we analyzed the prognostic impact of the angiogenic factor urokinase plasminogen activator resecptor (uPAR), vascular proliferation index (VPI) and tumor necrosis as a measure of hypoxia in a patient series of nodular melanomas (n = 255) and matched loco-regional metastases (n = 78). Expression of uPAR was determined by immunohistochemistry and VPI was assessed by dual immunohistochemistry using Factor-VIII/Ki67 staining. Necrosis was recorded based on HE-slides. As novel findings, high uPAR expression and high VPI were associated with each other, and with increased tumor thickness, presence of tumor necrosis, tumor ulceration, increased mitotic count and reduced cancer specific survival in primary melanoma. In matched cases, VPI was decreased in metastases, whereas the frequency of necrosis was increased. Our findings demonstrate for the first time the impact on melanoma specific survival of uPAR expression and VPI in primary tumors, and of increased necrosis as an indicator of tumor hypoxia in loco-regional metastases. These findings support the importance of tumor angiogenesis in melanoma aggressiveness, and suggest uPAR as an indicator of vascular proliferation and a potential biomarker in melanoma.

<![CDATA[Phorbol ester-induced angiogenesis of endothelial progenitor cells: The role of NADPH oxidase-mediated, redox-related matrix metalloproteinase pathways]]>

Endothelial progenitor cells (EPCs) may contribute to ischemia-induced angiogenesis in atherosclerotic diseases. The protein kinase C (PKC) family is involved in the regulation of angiogenesis, however the role of PKCα in EPCs during angiogenesis is unclear. The aim of this study was to evaluate the role of PKCα in EPCs during angiogenesis. Phorbol-12-myristate-13-acetate (PMA), a PKCα activator, significantly increased the activity and expression of matrix metalloproteinases (MMP) -2 and -9 in human (late outgrowth) EPCs in vitro. The MMPs promoted the migratory function and vascular formation of EPCs, which then contributed to neovascularization in a mouse hindlimb-ischemia model. Reactive oxygen species derived from nicotinamide adenine dinucleotide phosphate (NADPH) oxidase enhanced the expression of MMPs to increase the bioactivity of EPCs during angiogenesis. The mitogen-activated protein kinase (MAPK) signal pathway was associated with the activation of NADPH oxidase. PMA extensively activated the extracellular signal–regulated kinase (Erk) signal pathway to increase the expression of MMP-9. PMA also activated the p38, Erk, and c-Jun N-terminal kinase signal pathways to increase the expression of MMP-2. PMA-stimulated EPCs enhanced neovascularization in a mouse model of hindlimb ischemia via nuclear factor-κB translocation to up-regulation of the expression of MMP-2 and MMP-9. PMA could activate PKCα and promote the angiogenesis capacity of human EPCs via NADPH oxidase-mediated, redox-related, MMP-2 and MMP-9 pathways. The PKCα-activated, NADPH oxidase-mediated, redox-related MMP pathways could contribute to the function of human EPCs for ischemia-induced neovascularization, which may provide novel insights into the potential modification of EPCs for therapeutic angiogenesis.

<![CDATA[Engineering vasculature: Architectural effects on microcapillary-like structure self-assembly]]>

One of the greatest obstacles to clinical translation of bone tissue engineering is the inability to effectively and efficiently vascularize scaffolds. The goal of this work was to explore systematically whether architecture, at a scale of hundreds of microns, can be used to direct the growth of microcapillary-like structures into the core of scaffolds. Biphasic bioceramic patterned architectures were produced using silicone molds of 3D printed parts. Grooves and ridges were designed to have widths of 330 μm and 660 μm, with periodicities respectively of 1240 μm and 630 μm. Groove depth was varied between 150 μm and 585 μm. Co-cultures of human dermal microvascular endothelial cells (HDMECs) and human osteoblasts (hOBs) were used to grow microcapillary-like structures on substrates. Bioceramic architecture was found to significantly affect microcapillary-like structure location and orientation. Microcapillary-like structures were found to form predominantly in grooves or between convexities. For all patterned samples, the CD31 (endothelial cell marker) signal was at least 2.5 times higher along grooves versus perpendicular to grooves. In addition, the average signal was at least two times higher within grooves than outside grooves for all samples. Grooves with a width of 330 μm and a depth of 300 μm resulted in the formation of individual, highly aligned microcapillary-like structures with lengths around 5 mm. Extensive literature has focused on the role of nano- and micro-topography (on the scale below tens of microns) on cellular response. However, the idea that architecture at a scale much larger than a cell could be used to modulate angiogenesis has not been systematically investigated. This work shows the crucial influence of architecture on microcapillary-like structure self-assembly at the scale of hundreds of microns. Elucidating the precise correspondence between architecture and microcapillary-like structure organization will ultimately allow the engineering of microvasculature by tuning local scaffold design to achieve desirable microvessel properties.

<![CDATA[Regeneration-associated cells improve recovery from myocardial infarction through enhanced vasculogenesis, anti-inflammation, and cardiomyogenesis]]>


Considering the impaired function of regenerative cells in myocardial infarction (MI) patients with comorbidities and associated risk factors, cell therapy to enhance the regenerative microenvironment was designed using regeneration-associated cells (RACs), including endothelial progenitor cells (EPCs) and anti-inflammatory cells.


RACs were prepared by quality and quantity control culture of blood mononuclear cells (QQMNCs). Peripheral blood mononuclear cells (PBMNCs) were isolated from Lewis rats and conditioned for 5 days using a medium containing stem cell factors, thrombopoietin, Flt-3 ligand, vascular endothelial growth factor, and interleukin-6 to generate QQMNCs.


A 5.3-fold increase in the definitive colony-forming EPCs and vasculogenic EPCs was observed, in comparison to naïve PBMNCs. QQMNCs were enriched with EPCs (28.9-fold, P<0.0019) and M2 macrophages (160.3-fold, P<0.0002). Genes involved in angiogenesis (angpt1, angpt2, and vegfb), stem/progenitors (c-kit and sca-1), and anti-inflammation (arg-1, erg-2, tgfb, and foxp3) were upregulated in QQMNCs. For in vivo experiments, cells were administered into syngeneic rat models of MI. QQMNC-transplanted group (QQ-Tx) preserved cardiac function and fraction shortening 28 days post-MI in comparison with PBMNCs-transplanted (PB-Tx) (P<0.0001) and Control (P<0.0008) groups. QQ-Tx showed enhanced angiogenesis and reduced interstitial left ventricular fibrosis, along with a decrease in neutrophils and an increase in M2 macrophages in the acute phase of MI. Cell tracing studies revealed that intravenously administered QQMNCs preferentially homed to ischemic tissues via blood circulation. QQ-Tx showed markedly upregulated early cardiac transcriptional cofactors (Nkx2-5, 29.8-fold, and Gata-4, 5.2-fold) as well as c-kit (4.5-fold) while these markers were downregulated in PB-Tx. In QQ-Tx animals, de novo blood vessels formed a “Biological Bypass”, observed macroscopically and microscopically, while PB-Tx and Control-Tx groups showed severe fibrotic adhesion to the surrounding tissues, but no epicardial blood vessels.


QQMNCs conferred potent angiogenic and anti-inflammatory properties to the regenerative microenvironment, enhancing myocardiogenesis and functional recovery of rat MI hearts.

<![CDATA[Distinct genes and pathways associated with transcriptome differences in early cardiac development between fast- and slow-growing broilers]]>

Modern fast-growing broilers are susceptible to cardiac dysfunctions because their relatively small hearts cannot adequately meet the increased need of pumping blood through a large body mass. To improve cardiac health in broilers through breeding, we need to identify the genes and pathways that contribute to imbalanced cardiac development and occurrence of heart dysfunction. Two broiler lines–Ross 708 and Illinois–were included in this study as models of modern fast-growing and heritage slow-growing broilers, respectively. The left ventricular transcriptome were compared between the two broiler lines at day 6 and 21 post hatch through RNA-seq analysis to identify genes and pathways regulating compromised cardiac development in modern broilers. Number of differentially expressed genes (DEGs, p<0.05) between the two broiler lines increased from 321 at day 6 to 819 at day 21. As the birds grew, Ross broilers showed more DEGs (n = 1879) than Illinois broilers (n = 1117). Both broilers showed significant change of muscle related genes and immune genes, but Ross broilers showed remarkable change of expression of several lipid transporter genes including APOA4, APOB, APOH, FABP4 and RBP7. Ingenuity pathway analysis (IPA) suggested that increased cell apoptosis and inhibited cell cycle due to increased lipid accumulation, oxidative stress and endoplasmic reticulum stress may be related to the increased cardiac dysfunctions in fast-growing broilers. Cell cycle regulatory pathways like “Mitotic Roles of Polo-like Kinases” are ranked as the top changed pathways related to the cell apoptosis. These findings provide further insight into the cardiac dysfunction in modern broilers and also potential targets for improvement of their cardiac health through breeding.

<![CDATA[Response to Mechanical Stress Is Mediated by the TRPA Channel Painless in the Drosophila Heart]]>

Mechanotransduction modulates cellular functions as diverse as migration, proliferation, differentiation, and apoptosis. It is crucial for organ development and homeostasis and leads to pathologies when defective. However, despite considerable efforts made in the past, the molecular basis of mechanotransduction remains poorly understood. Here, we have investigated the genetic basis of mechanotransduction in Drosophila. We show that the fly heart senses and responds to mechanical forces by regulating cardiac activity. In particular, pauses in heart activity are observed under acute mechanical constraints in vivo. We further confirm by a variety of in situ tests that these cardiac arrests constitute the biological force-induced response. In order to identify molecular components of the mechanotransduction pathway, we carried out a genetic screen based on the dependence of cardiac activity upon mechanical constraints and identified Painless, a TRPA channel. We observe a clear absence of in vivo cardiac arrest following inactivation of painless and further demonstrate that painless is autonomously required in the heart to mediate the response to mechanical stress. Furthermore, direct activation of Painless is sufficient to produce pauses in heartbeat, mimicking the pressure-induced response. Painless thus constitutes part of a mechanosensitive pathway that adjusts cardiac muscle activity to mechanical constraints. This constitutes the first in vivo demonstration that a TRPA channel can mediate cardiac mechanotransduction. Furthermore, by establishing a high-throughput system to identify the molecular players involved in mechanotransduction in the cardiovascular system, our study paves the way for understanding the mechanisms underlying a mechanotransduction pathway.

<![CDATA[Vasculoprotective Effects of 3-Hydroxybenzaldehyde against VSMCs Proliferation and ECs Inflammation]]>

3-hydroxybenzaldehyde (3-HBA) is a precursor compound for phenolic compounds like Protocatechuic aldehyde (PCA). From recent reports, PCA has shown vasculoprotective potency, but the effects of 3-HBA remain unclear. The aim of this study is to investigate the vasculoprotective effects of 3-HBA in endothelial cells, vascular smooth muscle cells and various animal models. We tested effects of 3-HBA in both vitro and vivo. 3-HBA showed that it prevents PDGF-induced vascular smooth muscle cells (VSMCs) migration and proliferation from MTS, BrdU assays and inhibition of AKT phosphorylation. It arrested S and G0/G1 phase of VSMC cell cycle in PI staining and it also showed inhibited expression levels of Rb1 and CD1. In human umbilical vein endothelial cells (HUVECs), 3-HBA inhibited inflammatory markers and signaling molecules (VCAM-1, ICAM-1, p-NF-κB and p-p38). For ex vivo, 3-HBA has shown dramatic effects in suppressing the sprouting from aortic ring of Spargue Dawley (SD) rats. In vivo data supported the vasculoprotective effects of 3-HBA as it inhibited angiogenesis from Matrigel Plug assay in C57BL6 mouse, prevented ADP-induced thrombus generation, increased blood circulation after formation of thrombus, and attenuated neointima formation induced by common carotid artery balloon injury of SD rats. 3-HBA, a novel therapeutic agent, has shown vasculoprotective potency in both in vitro and in vivo.

<![CDATA[Combinatorial Effects of Double Cardiomyopathy Mutant Alleles in Rodent Myocytes: A Predictive Cellular Model of Myofilament Dysregulation in Disease]]>

Inherited cardiomyopathy (CM) represents a diverse group of cardiac muscle diseases that present with a broad spectrum of symptoms ranging from benign to highly malignant. Contributing to this genetic complexity and clinical heterogeneity is the emergence of a cohort of patients that are double or compound heterozygotes who have inherited two different CM mutant alleles in the same or different sarcomeric gene. These patients typically have early disease onset with worse clinical outcomes. Little experimental attention has been directed towards elucidating the physiologic basis of double CM mutations at the cellular-molecular level. Here, dual gene transfer to isolated adult rat cardiac myocytes was used to determine the primary effects of co-expressing two different CM-linked mutant proteins on intact cardiac myocyte contractile physiology. Dual expression of two CM mutants, that alone moderately increase myofilament activation, tropomyosin mutant A63V and cardiac troponin mutant R146G, were shown to additively slow myocyte relaxation beyond either mutant studied in isolation. These results were qualitatively similar to a combination of moderate and strong activating CM mutant alleles αTmA63V and cTnI R193H, which approached a functional threshold. Interestingly, a combination of a CM myofilament deactivating mutant, troponin C G159D, together with an activating mutant, cTnIR193H, produced a hybrid phenotype that blunted the strong activating phenotype of cTnIR193H alone. This is evidence of neutralizing effects of activating/deactivating mutant alleles in combination. Taken together, this combinatorial mutant allele functional analysis lends molecular insight into disease severity and forms the foundation for a predictive model to deconstruct the myriad of possible CM double mutations in presenting patients.

<![CDATA[New approach for visualization of relationships between RR and JT intervals]]>

This paper presents the concept of perfect matrices of Lagrange differences which are used to analyze relationships between RR and JT intervals during the bicycle ergometry exercise. The concept of the perfect matrix of Lagrange differences, its parameters, the construction of the load function and the corresponding optimization problem, the introduction of internal and external smoothing, embedding of the scalar parameter time series into the phase plane—all these computational techniques allow visualization of complex dynamical processes taking place in the cardiovascular system during the load and the recovery processes. Detailed analysis is performed with one person’s RR and JT records only—but the presented techniques open new possibilities for novel interpretation of the dynamics of the cardiovascular system.

<![CDATA[Fine-Tuning of Pten Localization and Phosphatase Activity Is Essential for Zebrafish Angiogenesis]]>

The lipid- and protein phosphatase PTEN is an essential tumor suppressor that is highly conserved among all higher eukaryotes. As an antagonist of the PI3K/Akt cell survival and proliferation pathway, it exerts its most prominent function at the cell membrane, but (PIP3-independent) functions of nuclear PTEN have been discovered as well. PTEN subcellular localization is tightly controlled by its protein conformation. In the closed conformation, PTEN localizes predominantly to the cytoplasm. Opening up of the conformation of PTEN exposes N-terminal and C-terminal regions of the protein that are required for both interaction with the cell membrane and translocation to the nucleus. Lack of Pten leads to hyperbranching of the intersegmental vessels during zebrafish embryogenesis, which is rescued by expression of exogenous Pten. Here, we observed that expression of mutant PTEN with an open conformation rescued the hyperbranching phenotype in pten double homozygous embryos and suppressed the increased p-Akt levels that are characteristic for embryos lacking Pten. In addition, in pten mutant and wild type embryos alike, open conformation PTEN induced stalled intersegmental vessels, which fail to connect with the dorsal longitudinal anastomotic vessel. Functional hyperactivity of open conformation PTEN in comparison to wild type PTEN seems to result predominantly from its enhanced recruitment to the cell membrane. Enhanced recruitment of phosphatase inactive mutants to the membrane did not induce the stalled vessel phenotype nor did it rescue the hyperbranching phenotype in pten double homozygous embryos, indicating that PTEN phosphatase activity is indispensable for its regulatory function during angiogenesis. Taken together, our data suggest that PTEN phosphatase activity needs to be carefully fine-tuned for normal embryogenesis and that the control of its subcellular localization is a key mechanism in this process.

<![CDATA[Nitric Oxide Transport in Normal Human Thoracic Aorta: Effects of Hemodynamics and Nitric Oxide Scavengers]]>

Despite the crucial role of nitric oxide (NO) in the homeostasis of the vasculature, little quantitative information exists concerning NO transport and distribution in medium and large-sized arteries where atherosclerosis and aneurysm occur and hemodynamics is complex. We hypothesized that local hemodynamics in arteries may govern NO transport and affect the distribution of NO in the arteries, hence playing an important role in the localization of vascular diseases. To substantiate this hypothesis, we presented a lumen/wall model of the human aorta based on its MRI images to simulate the production, transport and consumption of NO in the arterial lumen and within the aortic wall. The results demonstrated that the distribution of NO in the aorta was quite uneven with remarkably reduced NO bioavailability in regions of disturbed flow, and local hemodynamics could affect NO distribution mainly via flow dependent NO production rate of endothelium. In addition, erythrocytes in the blood could moderately modulate NO concentration in the aorta, especially at the endothelial surface. However, the reaction of NO within the wall could only slightly affect NO concentration on the luminal surface, but strongly reduce NO concentration within the aortic wall. A strong positive correlation was revealed between wall shear stress and NO concentration, which was affected by local hemodynamics and NO reaction rate. In conclusion, the distribution of NO in the aorta may be determined by local hemodynamics and modulated differently by NO scavengers in the lumen and within the wall.