Mapping genetic modifiers of radiation-induced cardiotoxicity to rat chromosome 3.

Radiation therapy is used in ~50% of cancer patients to reduce the risk of recurrence and in some cases improve survival. Despite these benefits, doses can be limited by toxicity in multiple organs, including the heart. The underlying causes and biomarkers of radiation-induced cardiotoxicity are currently unknown, prompting the need for experimental models with inherent differences in sensitivity and resistance to the development of radiation-induced cardiotoxicity. We have identified the parental SS (Dahl salt-sensitive/Mcwi) rat strain to be a highly-sensitized model of radiation-induced cardiotoxicity. In comparison, substitution of rat chromosome 3 from the resistant BN (Brown Norway) rat strain onto the SS background (SS-3BN consomic) significantly attenuated radiation-induced cardiotoxicity. SS-3BN rats had less radiation-induced cardiotoxicity than SS rats, as measured by survival, pleural and pericardial effusions, echocardiogram parameters, and histological damage. Mast cells, previously shown to have predominantly protective roles in radiation-induced cardiotoxicity, were increased in the more resistant SS-3BN hearts postradiation. RNA sequencing from SS and SS-3BN hearts at 1 wk postradiation revealed 5,098 differentially expressed candidate genes across the transcriptome and 350 differentially expressed genes on rat chromosome 3, which coincided with enrichment of multiple pathways, including mitochondrial dysfunction, sirtuin signaling, and ubiquitination. Upstream regulators of enriched pathways included the oxidative stress modulating transcription factor, Nrf2, which is located on rat chromosome 3. Nrf2 target genes were also differentially expressed in the SS vs. SS-3BN consomic hearts postradiation. Collectively, these data confirm the existence of heritable modifiers in radiation-induced cardiotoxicity and provide multiple biomarkers, pathways, and candidate genes for future analyses. NEW & NOTEWORTHY This novel study reveals that heritable genetic factors have the potential to modify normal tissue sensitivity to radiation. Gene variant(s) on rat chromosome 3 can contribute to enhanced cardiotoxicity displayed in the SS rats vs. the BN and SS-3BN consomic rats. Identifying genes that lead to understanding the mechanisms of radiation-induced cardiotoxicity represents a novel method to personalize radiation treatment, as well as predict the development of radiation-induced cardiotoxicity.

Sleeve Gastrectomy in Obese Zucker Rats Restores Cardiac Function and Geometry Toward a Lean Phenotype Independent of Weight Loss.

OBJECTIVE: Bariatric surgery, including sleeve gastrectomy (SG), significantly improves cardiac geometry and function in patients with heart failure. In this study, we used the obese Zucker rat as an animal model of heart failure with preserved ejection fraction (HFpEF) to test the hypothesis that a SG will improve cardiac function independent of weight loss. METHODS AND RESULTS: Obese, male Zucker rats underwent SG, pair-fed sham, or ad-lib sham surgery. Lean Zucker rats also underwent ad-lib sham surgery. Echocardiograms were performed preoperatively and at 6 weeks postoperatively. Obese SG and obese pair-fed sham rats had similar body weights postoperatively. Obese SG and lean, ad-lib, sham rats had a significant increase in postoperative stroke volume, and left ventricular internal diameter in diastole and systole. SG preserved systolic function and significantly improved isovolumetric relaxation time (13.9 ± 2.4 to 11.1 ± 2.1 ms, P = .02) independent of weight loss. DISCUSSION: SG has a beneficial impact on both systolic and diastolic cardiac function in obese Zucker rats toward a lean phenotype independent of weight loss and caloric restriction. These findings may represent a weight-loss independent mechanism generated from the gastrointestinal tract that has the potential to improve diastolic dysfunction independent of obesity status and translate to patients with HFpEF.

Augmentation of Muscle Blood Flow by Ultrasound Cavitation Is Mediated by ATP and Purinergic Signaling.

BACKGROUND: Augmentation of tissue blood flow by therapeutic ultrasound is thought to rely on convective shear. Microbubble contrast agents that undergo ultrasound-mediated cavitation markedly amplify these effects. We hypothesized that purinergic signaling is responsible for shear-dependent increases in muscle perfusion during therapeutic cavitation. METHODS: Unilateral exposure of the proximal hindlimb of mice (with or without ischemia produced by iliac ligation) to therapeutic ultrasound (1.3 MHz, mechanical index 1.3) was performed for 10 minutes after intravenous injection of 2×108 lipid microbubbles. Microvascular perfusion was evaluated by low-power contrast ultrasound perfusion imaging. In vivo muscle ATP release and in vitro ATP release from endothelial cells or erythrocytes were assessed by a luciferin-luciferase assay. Purinergic signaling pathways were assessed by studying interventions that (1) accelerated ATP degradation; (2) inhibited P2Y receptors, adenosine receptors, or KATP channels; or (3) inhibited downstream signaling pathways involving endothelial nitric oxide synthase or prostanoid production (indomethacin). Augmentation in muscle perfusion by ultrasound cavitation was assessed in a proof-of-concept clinical trial in 12 subjects with stable sickle cell disease. RESULTS: Therapeutic ultrasound cavitation increased muscle perfusion by 7-fold in normal mice, reversed tissue ischemia for up to 24 hours in the murine model of peripheral artery disease, and doubled muscle perfusion in patients with sickle cell disease. Augmentation in flow extended well beyond the region of ultrasound exposure. Ultrasound cavitation produced an ≈40-fold focal and sustained increase in ATP, the source of which included both endothelial cells and erythrocytes. Inhibitory studies indicated that ATP was a critical mediator of flow augmentation that acts primarily through either P2Y receptors or adenosine produced by ectonucleotidase activity. Combined indomethacin and inhibition of endothelial nitric oxide synthase abolished the effects of therapeutic ultrasound, indicating downstream signaling through both nitric oxide and prostaglandins. CONCLUSIONS: Therapeutic ultrasound using microbubble cavitation to increase muscle perfusion relies on shear-dependent increases in ATP, which can act through a diverse portfolio of purinergic signaling pathways. These events can reverse hindlimb ischemia in mice for >24 hours and increase muscle blood flow in patients with sickle cell disease. CLINICAL TRIAL REGISTRATION: URL: http://clinicaltrials.gov. Unique identifier: NCT01566890.

Treating a 20 mm Hg gradient alleviates myocardial hypertrophy in experimental aortic coarctation.

BACKGROUND: Children with coarctation of the aorta (CoA) can have a hyperdynamic and remodeled left ventricle (LV) from increased afterload. Literature from an experimental model suggests the putative 20 mm Hg blood pressure gradient (BPG) treatment guideline frequently implemented in CoA studies may permit irreversible vascular changes. LV remodeling from pressure overload has been studied, but data are limited following correction and using a clinically representative BPG. MATERIALS AND METHODS: Rabbits underwent CoA at 10 weeks to induce a 20 mm Hg BPG using permanent or dissolvable suture thereby replicating untreated and corrected CoA, respectively. Cardiac function was evaluated at 32 weeks by magnetic resonance imaging using a spoiled cine GRE sequence (TR/TE/FA 8/2.9/20), 14 × 14-cm FOV, and 3-mm slice thickness. Images (20 frames/cycle) were acquired in 6-8 short axis views from the apex to the mitral valve annulus. LV volume, ejection fraction (EF), and mass were quantified. RESULTS: LV mass was elevated for CoA (5.2 ± 0.55 g) versus control (3.6 ± 0.16 g) and corrected (4.0 ± 0.44 g) rabbits, resulting in increased LV mass/volume ratio for CoA rabbits. A trend toward increased EF and stroke volume was observed but did not reach significance. Elevated EF by volumetric analysis in CoA rabbits was supported by concomitant increases in total aortic flow by phase-contrast magnetic resonance imaging. CONCLUSIONS: The indices quantified trended toward a persistent hyperdynamic LV despite correction, but differences were not statistically significant versus control rabbits. These findings suggest the current putative 20 mm Hg BPG for treatment may be reasonable from the LV's perspective.

Clinical and imaging predictors of 1-year and long-term mortality in light chain (AL) amyloidosis: a 5-year follow-up study.

Light chain amyloidosis (AL) involves multiorgan failure induced by amyloidogenic light chain proteins, and is associated with high mortality. We aimed to identify clinical, laboratory, and imaging parameters that would predict 1-year and long-term AL mortality. Forty-four biopsy-proven AL patients (61.5 ± 12 years, 20 females) underwent clinical evaluation including laboratory assays, echocardiography, and contrast cardiac magnetic resonance imaging (CMR, n = 31) prior to chemotherapy. Patients were prospectively followed for median duration of 62.7 months (interquartile range 35.5 months). Clinical and laboratory parameters were compared between 1-year survivors and nonsurvivors. Univariate Kaplan-Meier survival plots were calculated followed by stepwise logistic regression analysis to assess independent predictors of long-term survival. Eighteen (40.9 %) patients died within 1 year and an additional 10 subjects died during long-term follow-up. Patients who expired within 1 year presented with more advanced class of heart failure, higher alkaline phosphatase and uric acid, lower limb lead voltage on electrocardiography, shorter left ventricular ejection time (ET) on echocardiography, and a higher proportion of late gadolinium enhancement on CMR. On multivariable analysis, only ET ≤240 ms on echocardiography (hazard ratio (HR) 5.07, 95 % confidence interval (CI) 1.83-14.1, P = 0.002) and New York Heart Association functional class II-IV presentation (HR 1.0058, 95 % CI 1.0014-1.0103, P = 0.01) were independent predictors of AL mortality. In conclusion, AL amyloidosis is associated with high 1-year and long-term mortality. Among clinical, laboratory, and imaging parameters tested, an echocardiographic finding of ET ≤240 ms has independent and additive prognostic value to clinical heart failure evaluation in determining long-term survival of AL patients. This result may be important in the early identification of patients at risk.

Mitochondrial DNA variant for complex I reveals a role in diabetic cardiac remodeling.

Myocardial remodeling and dysfunction are serious complications of type 2 diabetes mellitus (T2DM). Factors controlling their development are not well established. To specifically address the role of the mitochondrial genome, we developed novel conplastic rat strains, i.e. strains with the same nuclear genome but a different mitochondrial genome. The new animals were named T2DN(mtFHH) and T2DN(mtWistar), where the acronym T2DN denotes their common nuclear genome (type 2 diabetic nephropathy (T2DN) rats) and mtFHH or mtWistar the origin of their mitochondria, Fawn Hooded Hypertensive (FHH) or Wistar rats, respectively. The T2DN(mtFHH) and T2DN(mtWistar) showed a similar progression of diabetes as determined by HbA1c, cholesterol, and triglycerides with normal blood pressure, thus enabling investigation of the specific role of the mitochondrial genome in cardiac function without the confounding effects of obesity or hypertension found in other models of diabetes. Echocardiographic analysis of 12-week-old animals showed no abnormalities, but at 12 months of age the T2DN(mtFHH) showed left ventricular remodeling that was verified by histology. Decreased complex I and complex IV but not complex II activity within the electron transport chain was found only in T2DN(mtFHH), which was not explained by differences in protein content. Decreased cardiac ATP levels in T2DN(mtFHH) were in agreement with a lower ATP synthetic capacity by isolated mitochondria. Together, our data provide experimental evidence that mtDNA sequence variations have an additional role in energetic heart deficiency. The mitochondrial DNA background may explain the increased susceptibility of certain T2DM patients to develop myocardial dysfunction.

Increased Susceptibility for Adverse Reactions to Ultrasound Enhancing Agents in Sickle Cell Disease.

BACKGROUND: Pain-related adverse events (AEs) to ultrasound enhancing agents (UEAs) have been reported in patients with sickle cell disease (SCD). The aims of this study were to characterize the scope of these AEs in the SCD population and to investigate potential mechanisms on the basis of pathways involved in SCD vaso-occlusive crisis (VOC) and pain. METHODS: The prevalence and classification of AEs were analyzed from two clinical trials in which high-dose Definity infusions were used in patients with SCD (n = 55) or matched control subjects (n = 43) to study muscle or myocardial microvascular perfusion. Because complement (C') activation can trigger VOC in SCD, C' activation and surface adhesion of C' proteins on lipid UEAs were studied in vitro. C'-mediated UEA attachment to bone marrow immune cells was assessed using flow cytometry in a murine SCD model (Townes mice). Blood from patients receiving Definity was obtained to measure specific lysophospholipid metabolites of lipids in Definity thought to mediate SCD pain. RESULTS: Moderate or greater AEs, all of which were nociceptive (back or bone pain), occurred in one control subject and nine SCD subjects (2% vs 16%, P = .02). Patients with SCD who had AEs tended to have more severe manifestations of SCD. Three of the subjects with SCD had previously received Definity without complications. In patients with SCD, four AEs were classified as severe in intensity and as serious AEs on the basis of need for medical intervention. AEs were described to be similar to SCD-related pain, but there was no evidence for VOC, hemolysis, hypotension, or hypoxemia. At baseline, markers of C' activation were greater in patients with SCD than control subjects. However, after administration of lipid UEAs, SCD and control subjects were similar with regard to C' activation response, anaphylatoxin production, bone marrow microbubble retention, and production of lysophospholipids. There was a trend toward increased deposition of C3b and C3bi on lipid UEAs exposed to serum from patients with SCD. CONCLUSIONS: Patients with SCD are particularly susceptible to nociceptive AEs when given Definity at high doses. The mechanism for these AEs remains unclear but most are not related to the triggering of classic VOC.

Carotid plaque regression following 6-month statin therapy assessed by 3T cardiovascular magnetic resonance: comparison with ultrasound intima media thickness.

BACKGROUND: Cardiovascular magnetic resonance (CMR) allows volumetric carotid plaque measurement that has advantage over 2-dimensional ultrasound (US) intima-media thickness (IMT) in evaluating treatment response. We tested the hypothesis that 6-month statin treatment in patients with carotid plaque will lead to plaque regression when measured by 3 Tesla CMR but not by IMT. METHODS: Twenty-six subjects (67 ± 2 years, 7 females) with known carotid plaque (> 1.1 mm) and coronary or cerebrovascular atherosclerotic disease underwent 3T CMR (T1, T2, proton density and time of flight sequences) and US at baseline and following 6 months of statin therapy (6 had initiation, 7 had increase and 13 had maintenance of statin dosing). CMR plaque volume (PV) was measured in the region 12 mm below and up to 12 mm above carotid flow divider using software. Mean posterior IMT in the same region was measured. Baseline and 6-month CMR PV and US IMT were compared. Change in lipid rich/necrotic core (LR/NC) and calcification plaque components from CMR were related to change in PV. RESULTS: Low-density lipoprotein cholesterol decreased (86 ± 6 to 74 ± 4 mg/dL, p = 0.046). CMR PV decreased 5.8 ± 2% (1036 ± 59 to 976 ± 65 mm3, p = 0.018). Mean IMT was unchanged (1.12 ± 0.06 vs. 1.14 ± 0.06 mm, p = NS). Patients with initiation or increase of statins had -8.8 ± 2.8% PV change (p = 0.001) while patients with maintenance of statin dosing had -2.7 ± 3% change in PV (p = NS). There was circumferential heterogeneity in CMR plaque thickness with greatest thickness in the posterior carotid artery, in the region opposite the flow divider. Similarly there was circumferential regional difference in change of plaque thickness with significant plaque regression in the anterior carotid region in region of the flow divider. Change in LR/NC (R = 0.62, p = 0.006) and calcification (R = 0.45, p = 0.03) correlated with PV change. CONCLUSIONS: Six month statin therapy in patients with carotid plaque led to reduced plaque volume by 3T CMR, but ultrasound posterior IMT did not show any change. The heterogeneous spatial distribution of plaque and regional differences in magnitude of plaque regression may explain the difference in findings and support volumetric measurement of plaque. 3T CMR has potential advantage over ultrasound IMT to assess treatment response in individuals and may allow reduced sample size, duration and cost of clinical trials of plaque regression.

Ultrasonographic monitoring of fetal development in unrestrained bonobos (Pan paniscus) at the Milwaukee County Zoo.

The bonobo, Pan paniscus, is one of the most endangered primate species. In the context of the Bonobo Species Survival Plan(®), the Milwaukee County Zoo established a successful breeding group. Although the bonobo serves as a model species for human evolution, no prenatal growth curves are available. To develop growth graphs, the animals at the Milwaukee County Zoo were trained by positive reinforcement to allow for ultrasound exams without restraint. With this method, the well being of mother and fetus were maintained and ultrasound exams could be performed frequently. The ovulation date of the four animals in the study was determined exactly so that gestational age was known for each examination. Measurements of biparietal diameter (BPD), head circumference (HC), abdominal circumference (AC), and femur length (FL) were used to create growth curves. Prenatal growth of P. paniscus was compared with the data of humans and the common chimpanzee, P. troglodytes. With respect to cranial structures, such as BPD and HC, humans have significant acceleration of growth compared with P. paniscus and P. troglodytes. In P. paniscus, growth of AC was similar to HC throughout pregnancy, whereas in humans AC only reaches the level of HC close to term. Growth rate of FL was similar in humans and the two Pan species until near day 180 post-ovulation. After that, the Pan species FL growth slowed compared with human FL. The newly developed fetal growth curves of P. paniscus will assist in monitoring prenatal development and predicting birth dates of this highly endangered species.

Time-efficient patient-specific quantification of regional carotid artery fluid dynamics and spatial correlation with plaque burden.

PURPOSE: Low wall shear stress (WSS) and high oscillatory shear index (OSI) influence plaque formation, yet little is known about their role in progression/regression of established plaques because of lack of practical means to calculate them in individual patients. Our aim was to use computational fluid dynamics (CFD) models of patients with carotid plaque undergoing statin treatment to calculate WSS and OSI in a time-efficient manner, and determine their relationship to plaque thickness (PT), plaque composition (PC), and regression. METHODS: Eight patients (68 +/- 9 yr, one female) underwent multicontrast 3 T MRI at baseline and six-month post statin treatment. PT and PC were measured in carotid segments (common-CC, bifurcation-B, internal-IC) and circumferentially in nonoverlapping 600 angles and correlated with CFD models created from MRI, ultrasound, and blood pressure. RESULTS: PT was highest in B (2.42 +/- 0.98 versus CC: 1.60 +/- 0.47, IC: 1.62 +/- 0.52 mm, p < 0.01). Circumferentially, plaque was greatest opposite the flow divider (p < 0.01), where the lowest WSS and highest OSI were observed. In B and IC, PT was inversely related to WSS (R = -0.28 and -0.37, p < 0.01) and directly related to OSI (R = 0.22 and 0.52, p < 0.05). The total plaque volume changed from 1140 +/- 437 to 974 +/- 587 mm3 at six months (p = 0.1). Baseline WSS, but not OSI, correlated with changes in PT, necrotic tissue, and hemorrhage in B and IC, but not CC. CFD modeling took 49 +/- 18 h per patient. CONCLUSIONS: PT and PC correspond to adverse WSS and OSI in B and IC, and WSS is modestly but significantly related to changes in PT after short-term statin treatment. Regional hemodynamics from CFD can feasibly augment routine clinical imaging for comprehensive plaque evaluation.

Acquired Immunity Is Not Essential for Radiation-Induced Heart Dysfunction but Exerts a Complex Impact on Injury.

While radiation therapy (RT) can improve cancer outcomes, it can lead to radiation-induced heart dysfunction (RIHD) in patients with thoracic tumors. This study examines the role of adaptive immune cells in RIHD. In Salt-Sensitive (SS) rats, image-guided whole-heart RT increased cardiac T-cell infiltration. We analyzed the functional requirement for these cells in RIHD using a genetic model of T- and B-cell deficiency (interleukin-2 receptor gamma chain knockout (IL2RG-/-)) and observed a complex role for these cells. Surprisingly, while IL2RG deficiency conferred protection from cardiac hypertrophy, it worsened heart function via echocardiogram three months after a large single RT dose, including increased end-systolic volume (ESV) and reduced ejection fraction (EF) and fractional shortening (FS) (p < 0.05). Fractionated RT, however, did not yield similarly increased injury. Our results indicate that T cells are not uniformly required for RIHD in this model, nor do they account for our previously reported differences in cardiac RT sensitivity between SS and SS.BN3 rats. The increasing use of immunotherapies in conjunction with traditional cancer treatments demands better models to study the interactions between immunity and RT for effective therapy. We present a model that reveals complex roles for adaptive immune cells in cardiac injury that vary depending on clinically relevant factors, including RT dose/fractionation, sex, and genetic background.

Left atrial volume measurement with automated border detection by 3-dimensional echocardiography: comparison with Magnetic Resonance Imaging.

OBJECTIVE: Left atrial size is an important marker for adverse cardiovascular events. There is general consensus that left atrial volume index (LAVI) is the best measurement of size. The current LAVI measurement techniques are laborious. Semi-automated measurement with a 3-dimensional echocardiography (3DE) system may be a practical clinical alternative to measure LAVI, but it has not been adequately evaluated against Magnetic Resonance Imaging (MRI) gold standard. The aim of this study was to compare the accuracy of a commercially available 3D algorithm for measurement of LAVI against LAVI obtained from MRI and Area Length Method (ALM). DESIGN: In 27 consecutive subjects referred for cardiac MRI (age 54 +/- 13 years, 63% male), LAVI was measured using 3 imaging modalities: 3DE, ALM, MRI and the results were correlated. ALM was measured using standard American Society of Echocardiography guidelines. The time required to measure LAVI by 3DE and ALM were compared. RESULTS: There was a significant correlation in systolic and diastolic LA volumes and left atrial ejection fraction between 3DE and MRI (r = 0.86 for systole, r = 0.76 for diastole, r = 0.88 for ejection fraction, P < 0.0001 for all). There was also significant correlation of diastolic volumes between 3DE and ALM (r = 0.77, P < 0.0001). The time to obtain LAVI was shorter using 3DE versus ALM (56 +/- 8 vs 135 +/- 55 seconds, P < 0.0001). CONCLUSION: Three-dimensional echocardiography with semiautomatic border detection is a practical alternative for obtaining the left atrial volume in a time-efficient manner compared to the current standard.

Contrast-enhanced ultrasound detects changes in microvascular blood flow in adults with sickle cell disease.

In patients with sickle cell disease (SCD), poor outcome measures compromise the potential success of clinical trials. Contrast-enhanced ultrasound (CEUS) is a technique that can non-invasively quantify deep tissue microvascular blood flow. We tested the hypothesis that CEUS of forearm skeletal muscle could be used to: 1) assess microvascular abnormalities that occur during vaso-occlusive crisis; and 2) test new therapies for SCD that are targeted to improving the status of the microcirculation. We performed a prospective study, CEUS perfusion imaging of resting forearm muscle was performed in adults with SCD: 1) during and after a pain episode, and 2) before, during, and after a 24-hour infusion of the investigative agent, regadenoson, an adenosine A2A agonist. CEUS destruction-replenishment time-intensity data were analyzed to measure microvascular blood flow, as well as its components, microvascular blood volume and flux rate. Serial CEUS measurements were obtained in 32 adults with SCD. For the studies during crisis, there was a 30% reduction in microvascular blood flow compared to steady-state (p = 0.031), a reduction that was largely due to microvascular flux rate. For the regadenoson group, a non-significant 25% increase in flux rate and 9% increase in microvascular blood flow compared to baseline were detected during infusion. In a study of adults with SCD, CEUS detected changes in microvascular blood flow associated with vaso-occlusive crises. No changes were found during an infusion of the adenosine A2A agonist, regadenoson. This study provides preliminary evidence that CEUS could detect blood flow changes consistent with SCD physiology.

Telomerase Deficiency Predisposes to Heart Failure and Ischemia-Reperfusion Injury.

Introduction: Elevated levels of mitochondrial reactive oxygen species (ROS) contribute to the development of numerous cardiovascular diseases. TERT, the catalytic subunit of telomerase, has been shown to translocate to mitochondria to suppress ROS while promoting ATP production. Acute overexpression of TERT increases survival and decreases infarct size in a mouse model of myocardial infarct, while decreased telomerase activity predisposes to mitochondrial defects and heart failure. In the present study, we examined the role of TERT on cardiac structure and function under basal conditions and conditions of acute or prolonged stress in a novel rat model of TERT deficiency. Methods: Cardiac structure and function were evaluated via transthoracic echocardiogram. Langendorff preparations were used to test the effects of acute global ischemia reperfusion injury on cardiac function and infarction. Coronary flow and left ventricular pressure were measured during and after ischemia/reperfusion (I/R). Mitochondrial DNA integrity was measured by PCR and mitochondrial respiration was assessed in isolated mitochondria using an Oxygraph. Angiotensin II infusion was used as an established model of systemic stress. Results: No structural changes (echocardiogram) or coronary flow/left ventricle pressure (isolated hearts) were observed in TERT-/- rats at baseline; however, after I/R, coronary flow was significantly reduced in TERT-/- compared to wild type (WT) rats, while diastolic Left Ventricle Pressure was significantly elevated (n = 6 in each group; p < 0.05) in the TERT-/-. Interestingly, infarct size was less in TERT-/- rats compared to WT rats, while mitochondrial respiratory control index decreased and mitochondrial DNA lesions increased in TERT-/- compared to WT. Angiotensin II treatment did not alter cardiac structure or function; however, it augmented the infarct size significantly more in TERT-/- compared to the WT. Conclusion: Absence of TERT activity increases susceptibility to stress like cardiac injury. These results suggest a critical role of telomerase in chronic heart disease.

Cardiac Remodeling and Reversible Pulmonary Hypertension During Pneumonitis in Rats after 13-Gy Partial-Body Irradiation with Minimal Bone Marrow Sparing: Effect of Lisinopril.

Total-body irradiation causes acute and delayed toxicity to hematopoietic, pulmonary, cardiac, gastrointestinal, renal, and other organ systems. Angiotensin-converting enzyme inhibitors mitigate many of the delayed injuries to these systems. The purpose of this study was to define echocardiographic features in rats at two times after irradiation, the first before lethal radiation pneumonitis (50 d) and the second after recovery from pneumonitis but before lethal radiation nephropathy (100 d), and to determine the actions of the angiotensin-converting enzyme inhibitor lisinopril. Four groups of female WAG/RijCmcr rats at 11-12 wk of age were studied: nonirradiated, nonirradiated plus lisinopril, 13-Gy partial-body irradiation sparing one hind leg (leg-out partial-body irradiation), and 13-Gy leg-out partial-body irradiation plus lisinopril. Lisinopril was started 7 d after radiation. Echocardiograms were obtained at 50 and 100 d, and cardiac histology was assessed after 100 d. Irradiation without lisinopril demonstrated echocardiographic transient pulmonary hypertension by 50 d which was largely resolved by 100 d in survivors. Irradiated rats given lisinopril showed no increase in pulmonary artery pressures at 50 d but exhibited left ventricular remodeling. By 100 d these rats showed some signs of pulmonary hypertension. Lisinopril alone had no impact on echocardiographic end points at either time point in nonirradiated rats. Mild increases in mast cells and fibrosis in the heart were observed after 100 d following 13-Gy leg-out partial-body irradiation. These data demonstrate irradiation-induced pulmonary hypertension which was reversed in survivors of pneumonitis. Lisinopril modified cardiovascular remodeling to enhance survival in this model from 41% to 86% (p = 0.0013).

Cardiomyocyte-Specific Snrk Prevents Inflammation in the Heart.

Background The SNRK (sucrose-nonfermenting-related kinase) enzyme is critical for cardiac function. However, the underlying cause for heart failure observed in Snrk cardiac conditional knockout mouse is unknown. Methods and Results Previously, 6-month adult mice knocked out for Snrk in cardiomyocytes (CMs) displayed left ventricular dysfunction. Here, 4-month adult mice, on angiotensin II (Ang II) infusion, show rapid decline in cardiac systolic function, which leads to heart failure and death in 2 weeks. These mice showed increased expression of nuclear factor κ light chain enhancer of activated B cells (NF-κB), inflammatory signaling proteins, proinflammatory proteins in the heart, and fibrosis. Interestingly, under Ang II infusion, mice knocked out for Snrk in endothelial cells did not show significant systolic or diastolic dysfunction. Although an NF-κB inflammation signaling pathway was increased in Snrk knockout endothelial cells, this did not lead to fibrosis or mortality. In hearts of adult mice knocked out for Snrk in CMs, we also observed NF-κB pathway activation in CMs, and an increased presence of Mac2+ macrophages was observed in basal and Ang II-infused states. In vitro analysis of Snrk knockdown HL-1 CMs revealed similar upregulation of the NF-κB signaling proteins and proinflammatory proteins that was exacerbated on Ang II treatment. The Ang II-induced NF-κB pathway-mediated proinflammatory effects were mediated in part through protein kinase B or AKT, wherein AKT inhibition restored the proinflammatory signaling protein levels to baseline in Snrk knockdown HL-1 CMs. Conclusions During heart failure, SNRK acts as a cardiomyocyte-specific repressor of cardiac inflammation and fibrosis.

Intraventricular dyssynchrony in light chain amyloidosis: a new mechanism of systolic dysfunction assessed by 3-dimensional echocardiography.

BACKGROUND: Light chain amyloidosis (AL) is a rare but often fatal disease due to intractable heart failure. Amyloid deposition leads to diastolic dysfunction and often preserved ejection fraction. We hypothesize that AL is associated with regional systolic dyssynchrony. The aim is to compare left ventricular (LV) regional synchrony in AL subjects versus healthy controls using 16-segment dyssynchrony index measured from 3-dimension-al (3D) echocardiography. METHODS: Cardiac 3D echocardiography full volumes were acquired in 10 biopsy-proven AL subjects (60 +/- 3 years, 5 females) and 10 healthy controls (52 +/- 1 years, 5 females). The LV was subdivided into 16 segments and the time from end-diastole to the minimal systolic volume for each of the 16 segments was expressed as a percent of the cycle length. The standard deviations of these times provided a 16-segment dyssynchrony index (16-SD%). 16-SD% was compared between healthy and AL subjects. RESULTS: Left ventricular ejection fraction was comparable (control vs. AL: 62.4 +/- 0.6 vs. 58.6 +/- 2.8%, p = NS). 16-SD% was significantly higher in AL versus healthy subjects (5.93 +/- 4.4 vs. 1.67 +/- 0.87%, p = 0.003). 16-SD% correlated with left ventricular mass index (R 0.45, p = 0.04) but not to left ventricular ejection fraction. CONCLUSION: Light chain amyloidosis is associated with left ventricular regional systolic dyssynchrony. Regional dyssynchrony may be an unrecognized mechanism of heart failure in AL subjects.

Sucrose Nonfermenting-Related Kinase Enzyme-Mediated Rho-Associated Kinase Signaling is Responsible for Cardiac Function.

BACKGROUND: Cardiac metabolism is critical for the functioning of the heart, and disturbance in this homeostasis is likely to influence cardiac disorders or cardiomyopathy. Our laboratory has previously shown that SNRK (sucrose nonfermenting related kinase) enzyme, which belongs to the AMPK (adenosine monophosphate-activated kinase) family, was essential for cardiac metabolism in mammals. Snrk global homozygous knockout (KO) mice die at postnatal day 0, and conditional deletion of Snrk in cardiomyocytes (Snrk cmcKO) leads to cardiac failure and death by 8 to 10 months. METHODS AND RESULTS: We performed additional cardiac functional studies using echocardiography and identified further cardiac functional deficits in Snrk cmcKO mice. Nuclear magnetic resonance-based metabolomics analysis identified key metabolic pathway deficits in SNRK knockdown cardiomyocytes in vitro. Specifically, metabolites involved in lipid metabolism and oxidative phosphorylation are altered, and perturbations in these pathways can result in cardiac function deficits and heart failure. A phosphopeptide-based proteomic screen identified ROCK (Rho-associated kinase) as a putative substrate for SNRK, and mass spec-based fragment analysis confirmed key amino acid residues on ROCK that are phosphorylated by SNRK. Western blot analysis on heart lysates from Snrk cmcKO adult mice and SNRK knockdown cardiomyocytes showed increased ROCK activity. In addition, in vivo inhibition of ROCK partially rescued the in vivo Snrk cmcKO cardiac function deficits. CONCLUSIONS: Collectively, our data suggest that SNRK in cardiomyocytes is responsible for maintaining cardiac metabolic homeostasis, which is mediated in part by ROCK, and alteration of this homeostasis influences cardiac function in the adult heart.

Whole-thorax irradiation induces hypoxic respiratory failure, pleural effusions and cardiac remodeling.

To study the mechanisms of death following a single lethal dose of thoracic radiation, WAG/RijCmcr (Wistar) rats were treated with 15 Gy to the whole thorax and followed until they were morbid or sacrificed for invasive assays at 6 weeks. Lung function was assessed by breathing rate and arterial oxygen saturation. Lung structure was evaluated histologically. Cardiac structure and function were examined by echocardiography. The frequency and characteristics of pleural effusions were determined. Morbidity from 15 Gy radiation occurred in all rats 5 to 8 weeks after exposure, coincident with histological pneumonitis. Increases in breathing frequencies peaked at 6 weeks, when profound arterial hypoxia was also recorded. Echocardiography analysis at 6 weeks showed pulmonary hypertension and severe right ventricular enlargement with impaired left ventricular function and cardiac output. Histologic sections of the heart revealed only rare foci of lymphocytic infiltration. Total lung weight more than doubled. Pleural effusions were present in the majority of the irradiated rats and contained elevated protein, but low lactate dehydrogenase, when compared with serum from the same animal. Pleural effusions had a higher percentage of macrophages and large monocytes than neutrophils and contained mast cells that are rarely present in other pathological states. Lethal irradiation to rat lungs leads to hypoxia with infiltration of immune cells, edema and pleural effusion. These changes may contribute to pulmonary vascular and parenchymal injury that result in secondary changes in heart structure and function. We report that conditions resembling congestive heart failure contribute to death during radiation pneumonitis, which indicates new targets for therapy.

Simvastatin mitigates increases in risk factors for and the occurrence of cardiac disease following 10 Gy total body irradiation.

The ability of simvastatin to mitigate the increases in risk factors for and the occurrence of cardiac disease after 10 Gy total body irradiation (TBI) was determined. This radiation dose is relevant to conditioning for stem cell transplantation and threats from radiological terrorism. Male rats received single dose TBI of 10 Gy. Age-matched, sham-irradiated rats served as controls. Lipid profile, heart and liver morphology and cardiac mechanical function were determined for up to 120 days after irradiation. TBI resulted in a sustained increase in total- and LDL-cholesterol (low-density lipoprotein-cholesterol), and triglycerides. Simvastatin (10 mg/kg body weight/day) administered continuously from 9 days after irradiation mitigated TBI-induced increases in total- and LDL-cholesterol and triglycerides, as well as liver injury. TBI resulted in cellular peri-arterial fibrosis, whereas control hearts had less collagen and fibrosis. Simvastatin mitigated these morphological injuries. TBI resulted in cardiac mechanical dysfunction. Simvastatin mitigated cardiac mechanical dysfunction 20-120 days following TBI. To determine whether simvastatin affects the ability of the heart to withstand stress after TBI, injury from myocardial ischemia/reperfusion was determined in vitro. TBI increased the severity of an induced myocardial infarction at 20 and 80 days after irradiation. Simvastatin mitigated the severity of this myocardial infarction at 20 and 80 days following TBI. It is concluded simvastatin mitigated the increases in risk factors for cardiac disease and the extent of cardiac disease following TBI. This statin may be developed as a medical countermeasure for the mitigation of radiation-induced cardiac disease.