Impact of Untreated Obstructive Sleep Apnea on Left and Right Ventricular Myocardial Function and Effects of CPAP Therapy

Background

Obstructive sleep apnea (OSA) has deteriorating effect on LV function, whereas its impact on RV function is controversial. We aimed to determine the effect of OSA and continuous positive airway pressure (CPAP) treatment on left and right ventricular (LV, RV) function using transthoracic echocardiography (TTE) and 2 dimensional speckle tracking (2D ST) analysis of RV deformation capability.

Methods and Results

82 patients with OSA and need for CPAP therapy were prospectively enrolled and underwent TTE at study inclusion and after 6 months of follow up (FU). Multivariate regression analysis revealed an independent association between baseline apical right ventricular longitudinal strain (RV-Sl), BMI and the severity of OSA (apical RV-Sl: P = 0.0002, BMI: P = 0.02). After CPAP therapy, LV functional parameters (LVEF: P<0.0001, LV performance index: P = 0.03, stroke volume: P = 0.042), and apical RV-Sl (P = 0.001) improved significantly. The effect of CPAP therapy was related to severity of OSA (LVEF: AHI 5–14, 66.4±8.8%, 68.5±10.6% [P = ns]; AHI 15–30∶59.8±7.7%, 68.6±9.3% [P = 0.002]; AHI>30∶54.1±12.4%, 68.2±13.6%[P<0.0001]; apical RV-Sl: AHI 5–14: −17.3±8.7%, −16.0±10.8% [P = ns], AHI 15–30: −9.8±6.0%, −15.4±10.9% [P = 0.028], AHI>30: −6.3±5.7%, −17.9±11.2% [P<0.0001]).

Conclusions

OSA seems to have deteriorating effect on LV and RV function. We found a beneficial effect of CPAP on LV and RV functional parameters predominately in patients with severe OSA. 2D speckle tracking might be of value to determine early changes in global and regional right ventricular function.     

Effects of 12 Weeks High-Intensity & Reduced-Volume Training in Elite Athletes

It was investigated if high-intensity interval training (HIT) at the expense of total training volume improves performance, maximal oxygen uptake and swimming economy. 41 elite swimmers were randomly allocated to a control (CON) or HIT group. For 12 weeks both groups trained ∼12 h per week. HIT comprised ∼5 h vs. 1 h and total distance was ∼17 km vs. 35 km per week for HIT and CON, respectively. HIT was performed as 6-10×10-30 s maximal effort interspersed by 2–4 minutes of rest. Performance of 100 m all-out freestyle and 200 m freestyle was similar before and after the intervention in both HIT (60.4±4.0 vs. 60.3±4.0 s; n = 13 and 133.2±6.4 vs. 132.6±7.7 s; n = 14) and CON (60.2±3.7 vs. 60.6±3.8 s; n = 15 and 133.5±7.0 vs. 133.3±7.6 s; n = 15). Maximal oxygen uptake during swimming was similar before and after the intervention in both the HIT (4.0±0.9 vs. 3.8±1.0 l O₂×min⁻¹; n = 14) and CON (3.8±0.7 vs. 3.8±0.7 l O₂×min⁻¹; n = 11) group. Oxygen uptake determined at fixed submaximal speed was not significantly affected in either group by the intervention. Body fat % tended to increase (P = 0.09) in the HIT group (15.4±1.6% vs. 16.3±1.6%; P = 0.09; n = 16) and increased (P<0.05) in the CON group (13.9±1.5% vs. 14.9±1.5%; n = 17). A distance reduction of 50% and a more than doubled HIT amount for 12 weeks did neither improve nor compromise performance or physiological capacity in elite swimmers.     

Right Ventricular Function Quantification in Takotsubo Cardiomyopathy Using Two-Dimensional Strain Echocardiography

Aims

This study sought to characterize global and regional right ventricular (RV) myocardial function in patients with Takotsubo cardiomyopathy (TC) using 2D strain imaging.

Methods

We compared various parameters of RV and left ventricular (LV) systolic function between 2 groups of consecutive patients with TC at initial presentation and upon follow-up. Group 1 had RV involvement and group 2 did not have RV involvement.

Results

At initial presentation, RV peak systolic longitudinal strain (RVPSS) and RV fractional area change (RVFAC) were significantly lower in group 1 (−13.2±8.6% vs. −21.8±5.4%, p = 0.001; 30.7±9.3% vs. 43.5±6.3%, p = 0.001) and improved significantly upon follow-up. Tricuspid annular plane systolic excursion (TAPSE) did not differ significantly at initial presentation between both groups (14.8±4.1 mm vs. 17.9±3.5 mm, p = 0.050). Differences in regional systolic RV strain were only observed in the mid and apical segments. LV ejection fraction (LVEF) and LV global strain were significantly lower in group 1 (36±8% vs. 46±10%, p = 0.006 and −5.5±4.8% vs. −10.2±6.2%, p = 0.040) at initial presentation. None of the parameters were significantly different between the 2 groups upon follow-up. A RVPSS cut-off value of >−19.1% had a sensitivity of 85% and a specificity of 71% to discriminate between the 2 groups.

Conclusion

In TC, RVFAC, RVPSS, LVEF and LV global strain differed significantly between patients with and without RV dysfunction, whereas TAPSE did not. 2 D strain imaging was feasible for the assessment of RV dysfunction in TC and could discriminate between patients with and without RV involvement in a clinically meaningful way.     

Acoustic Radiation Force Impulse Imaging for Noninvasive Evaluation of Renal Parenchyma Elasticity: Preliminary Findings

Objective

To evaluate the diagnostic value of acoustic radiation force impulse (ARFI) to test the elasticity of renal parenchyma by measuring the shear wave velocity (SWV) which might be used to detect chronic kidney disease (CKD).

Methods

327 healthy volunteers and 64 CKD patients were enrolled in the study. The potential influencing factors and measurement reproducibility were evaluated in the healthy volunteers. Correlations between SWV and laboratory tests were analyzed in CKD patients.?Receiver-operating characteristic curve (ROC) analyses were performed to assess the diagnostic performance of ARFI.

Results

The SWV of healthy volunteers correlated significantly to age (r = −0.22, P<0.001, n = 327) and differed significantly between men and women (2.06±0.48 m/s vs. 2.2±0.52 m/s, P = 0.018, n = 327). However, it did not correlate significantly to height, weight, body mass index, waistline, kidney dimension and the depth for SWV measurement (n = 30). Inter- and intraobserver agreement expressed as intraclass coefficient correlation were 0.64 (95% CI: 0.13 to 0.82, P = 0.011) and 0.6 (95% CI: 0.31 to 0.81, P = 0.001) (n = 40). The mean SWV in healthy volunteers was 2.15±0.51 m/s, while was 1.81±0.43 m/s, 1.79±0.29 m/s, 1.81±0.44 m/s, 1.64±0.55 m/s, and 1.36±0.17 m/s for stage 1, 2, 3, 4 and 5 in CKD patients respectively. The SWV was significantly higher for healthy volunteers compared with each stage in CKD patients. ARFI could not predict the different stages of CKD except stage 5. In CKD patients, SWV correlated to e-GFR (r = 0.3, P = 0.018), to urea nitrogen (r =  −0.3, P = 0.016), and to creatinine (r =  −0.41, P = 0.001). ROC analyses indicated that the area under the ROC curve was 0.752 (95% CI: 0.704 to 0.797) (P<0.001). The cut-off value for predicting CKD was 1.88 m/s (sensitivity 71.87% and specificity 69.69%).

Conclusion

ARFI may be a potentially useful tool in detecting CKD.     

De Novo Lipogenesis and Cholesterol Synthesis in Humans with Long-Standing Type 1 Diabetes Are Comparable to Non-Diabetic Individuals

Background

Synthesis of lipid species, including fatty acids (FA) and cholesterol, can contribute to pathological disease. The purpose of this study was to investigate FA and cholesterol synthesis in individuals with type 1 diabetes, a group at elevated risk for vascular disease, using stable isotope analysis.

Methods

Individuals with type 1 diabetes (n = 9) and age-, sex-, and BMI-matched non-diabetic subjects (n = 9) were recruited. On testing day, meals were provided to standardize food intake and elicit typical feeding responses. Blood samples were analyzed at fasting (0 and 24 h) and postprandial (2, 4, 6, and 8 hours after breakfast) time points. FA was isolated from VLDL to estimate hepatic FA synthesis, whereas free cholesterol (FC) and cholesteryl ester (CE) was isolated from plasma and VLDL to estimate whole-body and hepatic cholesterol synthesis, respectively. Lipid synthesis was measured using deuterium incorporation and isotope ratio mass spectrometry.

Results

Fasting total hepatic lipogenesis (3.91±0.90% vs. 5.30±1.22%; P = 0.41) was not significantly different between diabetic and control groups, respectively, nor was synthesis of myristic (28.60±4.90% vs. 26.66±4.57%; P = 0.76), palmitic (12.52±2.75% vs. 13.71±2.64%; P = 0.65), palmitoleic (3.86±0.91% vs. 4.80±1.22%; P = 0.65), stearic (5.55±1.04% vs. 6.96±0.97%; P = 0.29), and oleic acid (1.45±0.28% vs. 2.10±0.51%; P = 0.21). Postprandial lipogenesis was also not different between groups (P = 0.38). Similarly, fasting synthesis of whole-body FC (8.2±1.3% vs. 7.3±0.8%/day; P = 0.88) and CE (1.9±0.4% vs. 2.0±0.3%/day; P = 0.96) and hepatic FC (8.2±2.0% vs. 8.1±0.8%/day; P = 0.72) was not significantly different between diabetic and control subjects.

Conclusions

Despite long-standing disease, lipogenesis and cholesterol synthesis was not different in individuals with type 1 diabetes compared to healthy non-diabetic humans.     

Alterations of Neuromuscular Function after the World's Most Challenging Mountain Ultra-Marathon

We investigated the physiological consequences of the most challenging mountain ultra-marathon (MUM) in the world: a 330-km trail run with 24000 m of positive and negative elevation change. Neuromuscular fatigue (NMF) was assessed before (Pre-), during (Mid-) and after (Post-) the MUM in experienced ultra-marathon runners (n = 15; finish time  = 122.43 hours ±17.21 hours) and in Pre- and Post- in a control group with a similar level of sleep deprivation (n = 8). Blood markers of muscle inflammation and damage were analyzed at Pre- and Post-. Mean ± SD maximal voluntary contraction force declined significantly at Mid- (−13±17% and −10±16%, P<0.05 for knee extensor, KE, and plantar flexor muscles, PF, respectively), and further decreased at Post- (−24±13% and −26±19%, P<0.01) with alteration of the central activation ratio (−24±24% and −28±34% between Pre- and Post-, P<0.05) in runners whereas these parameters did not change in the control group. Peripheral NMF markers such as 100 Hz doublet (KE: −18±18% and PF: −20±15%, P<0.01) and peak twitch (KE: −33±12%, P<0.001 and PF: −19±14%, P<0.01) were also altered in runners but not in controls. Post-MUM blood concentrations of creatine kinase (3719±3045 Ul·¹), lactate dehydrogenase (1145±511 UI·L⁻¹), C-Reactive Protein (13.1±7.5 mg·L⁻¹) and myoglobin (449.3±338.2 µg·L⁻¹) were higher (P<0.001) than at Pre- in runners but not in controls. Our findings revealed less neuromuscular fatigue, muscle damage and inflammation than in shorter MUMs. In conclusion, paradoxically, such extreme exercise seems to induce a relative muscle preservation process due likely to a protective anticipatory pacing strategy during the first half of MUM and sleep deprivation in the second half.     

Calpain Inhibition Reduces Amplitude and Accelerates Decay of the Late Sodium Current in Ventricular Myocytes from Dogs with Chronic Heart Failure

Calpain is an intracellular Ca²⁺ -activated protease that is involved in numerous Ca²⁺ dependent regulation of protein function in many cell types. This paper tests a hypothesis that calpains are involved in Ca²⁺ -dependent increase of the late sodium current (I_NaL) in failing heart. Chronic heart failure (HF) was induced in 2 dogs by multiple coronary artery embolization. Using a conventional patch-clamp technique, the whole-cell I_NaL was recorded in enzymatically isolated ventricular cardiomyocytes (VCMs) in which I_NaL was activated by the presence of a higher (1μM) intracellular [Ca²⁺] in the patch pipette. Cell suspensions were exposed to a cell- permeant calpain inhibitor MDL-28170 for 1–2 h before I_NaL recordings. The numerical excitation-contraction coupling (ECC) model was used to evaluate electrophysiological effects of calpain inhibition in silico. MDL caused acceleration of I_NaL decay evaluated by the two-exponential fit (τ₁ = 42±3.0 ms τ₂ = 435±27 ms, n = 6, in MDL vs. τ₁ = 52±2.1 ms τ₂ = 605±26 control no vehicle, n = 11, and vs. τ₁ = 52±2.8 ms τ₂ = 583±37 ms n = 7, control with vehicle, P<0.05 ANOVA). MDL significantly reduced I_NaL density recorded at –30 mV (0.488±0.03, n = 12, in control no vehicle, 0.4502±0.0210, n = 9 in vehicle vs. 0.166±0.05pA/pF, n = 5, in MDL). Our measurements of current-voltage relationships demonstrated that the I_NaL density was decreased by MDL in a wide range of potentials, including that for the action potential plateau. At the same time the membrane potential dependency of the steady-state activation and inactivation remained unchanged in the MDL-treated VCMs. Our ECC model predicted that calpain inhibition greatly improves myocyte function by reducing the action potential duration and intracellular diastolic Ca²⁺ accumulation in the pulse train.

Conclusions

Calpain inhibition reverses I_NaL changes in failing dog ventricular cardiomyocytes in the presence of high intracellular Ca²⁺. Specifically it decreases I_NaL density and accelerates I_NaL kinetics resulting in improvement of myocyte electrical response and Ca²⁺ handling as predicted by our in silico simulations.     

Regular Cocaine Use Is Associated with Increased Systolic Blood Pressure,Aortic Stiffness and Left Ventricular Mass in Young Otherwise Healthy Individuals

Background

The cardiovascular impact of cocaine use in otherwise healthy individuals who consider themselves ‘social’ users is not well established.

Methods/Results

Twenty regular cocaine users and 20 control subjects were recruited by word-of-mouth. Cardiovascular magnetic resonance was performed to assess cardiac and vascular structure and function. Cocaine users had higher systolic blood pressure compared to non-users (134±11 vs 126±11 mmHg, p = 0.036), a finding independent of age, body surface area, smoking and alcohol consumption. Cocaine use was associated with increased arterial stiffness - reflected by reduced aortic compliance (1.3±0.2 vs 1.7±0.5 cm²×10⁻².mmHg⁻¹, p = 0.004), decreased distensibility (3.8±0.9 vs 5.1±1.4 mmHg⁻¹.10⁻³, p = 0.001), increased stiffness index (2.6±0.6 vs 2.1±0.6, p = 0.005), and higher pulse wave velocity (5.1±0.6 vs 4.4±0.6 m.s⁻¹, p = 0.001). This change in aortic stiffness was independent of vessel wall thickness. Left ventricular mass was 18% higher in cocaine users (124±25 vs 105±16 g, p = 0.01), a finding that was independent of body surface area, and left atrial diameter was larger in the user group than controls (3.8±0.6 vs 3.5±0.3 cm, p = 0.04). The increased left ventricular mass, systolic blood pressure and vascular stiffness measures were all associated with duration and/or frequency of cocaine use. No late gadolinium enhancement or segmental wall motion abnormalities were seen in any of the subjects.

Conclusions

Compared with the non-user control cohort, cocaine users had increased aortic stiffness and systolic blood pressure, associated with greater left ventricular mass. These measures are all well known risk factors for premature cardiovascular events, highlighting the dangers of cocaine use, even in a ‘social’ setting, and have important public health implications.     

Comparison of “Live High-Train Low” in Normobaric versus Hypobaric Hypoxia

We investigated the changes in both performance and selected physiological parameters following a Live High-Train Low (LHTL) altitude camp in either normobaric hypoxia (NH) or hypobaric hypoxia (HH) replicating current “real” practices of endurance athletes. Well-trained triathletes were split into two groups (NH, n = 14 and HH, n = 13) and completed an 18-d LHTL camp during which they trained at 1100–1200 m and resided at an altitude of 2250 m (P_iO₂  = 121.7±1.2 vs. 121.4±0.9 mmHg) under either NH (hypoxic chamber; F_iO₂ 15.8±0.8%) or HH (real altitude; barometric pressure 580±23 mmHg) conditions. Oxygen saturations (S_pO₂) were recorded continuously daily overnight. P_iO₂ and training loads were matched daily. Before (Pre-) and 1 day after (Post-) LHTL, blood samples, VO_2max, and total haemoglobin mass (Hb_mass) were measured. A 3-km running test was performed near sea level twice before, and 1, 7, and 21 days following LHTL. During LHTL, hypoxic exposure was lower for the NH group than for the HH group (220 vs. 300 h; P<0.001). Night S_pO₂ was higher (92.1±0.3 vs. 90.9±0.3%, P<0.001), and breathing frequency was lower in the NH group compared with the HH group (13.9±2.1 vs. 15.5±1.5 breath.min⁻¹, P<0.05). Immediately following LHTL, similar increases in VO_2max (6.1±6.8 vs. 5.2±4.8%) and Hb_mass (2.6±1.9 vs. 3.4±2.1%) were observed in NH and HH groups, respectively, while 3-km performance was not improved. However, 21 days following the LHTL intervention, 3-km run time was significantly faster in the HH (3.3±3.6%; P<0.05) versus the NH (1.2±2.9%; ns) group. In conclusion, the greater degree of race performance enhancement by day 21 after an 18-d LHTL camp in the HH group was likely induced by a larger hypoxic dose. However, one cannot rule out other factors including differences in sleeping desaturations and breathing patterns, thus suggesting higher hypoxic stimuli in the HH group.     

Long-Term Prognostic Value of Restitution Slope in Patients with Ischemic and Dilated Cardiomyopathies

Background

An action potential duration (APD) restitution curve with a steep slope ≥1 has been associated with increased susceptibility for malignant ventricular arrhythmias. We aimed to evaluate the “restitution hypothesis” and tested ventricular APD restitution slope as well as effective refractory period (ERP)/APD ratio for long-term prognostic value in patients with ischemic (ICM) or dilated cardiomyopathy (DCM).

Methodology/Principal Findings

Monophasic action potentials were recorded in patients with ICM (n = 32) and DCM (n = 42) undergoing routine programmed ventricular stimulation (PVS). Left ventricular ejection fraction was 32±7% and 28±9%, respectively. APD and ERP were measured at baseline stimulation (S₁) and upon introduction of one to three extrastimuli (S₂–S₄). ERP/APD ratios and the APD restitution curve were calculated and the maximum restitution slope was determined. After a mean follow-up of 6.1±3.0 years, the combined end-point of mortality and and/or implantable cardioverter-defibrillator shock was not predicted by restitution slope or ERP/APD ratios. Comparing S₂ vs. S₃ vs. S₄ extrastimuli for restitution slope (1.5±0.6 vs. 1.4±0.4 vs. 1.3±0.5; p = NS), additional extrastimuli did not lead to a steepening restitution slope. ERP/APD ratio decreased with additional extrastimuli (0.98±0.09 [S₁] vs. 0.97±0.10 [S₂] vs. 0.93±0.11 [S₃]; p = 0.03 S₁ vs. S₃). Positive PVS was strongly predictive of outcome (p = 0.006).

Conclusions/Significance

Neither ventricular APD restitution slope nor ERP/APD ratios predict outcome in patients with ICM or DCM.     

SORL1 Is Genetically Associated with Late-Onset Alzheimer’s Disease in Japanese,Koreans and Caucasians

To discover susceptibility genes of late-onset Alzheimer’s disease (LOAD), we conducted a 3-stage genome-wide association study (GWAS) using three populations: Japanese from the Japanese Genetic Consortium for Alzheimer Disease (JGSCAD), Koreans, and Caucasians from the Alzheimer Disease Genetic Consortium (ADGC). In Stage 1, we evaluated data for 5,877,918 genotyped and imputed SNPs in Japanese cases (n = 1,008) and controls (n = 1,016). Genome-wide significance was observed with 12 SNPs in the APOE region. Seven SNPs from other distinct regions with p-values <2×10⁻⁵ were genotyped in a second Japanese sample (885 cases, 985 controls), and evidence of association was confirmed for one SORL1 SNP (rs3781834, P = 7.33×10⁻⁷ in the combined sample). Subsequent analysis combining results for several SORL1 SNPs in the Japanese, Korean (339 cases, 1,129 controls) and Caucasians (11,840 AD cases, 10,931 controls) revealed genome wide significance with rs11218343 (P = 1.77×10⁻⁹) and rs3781834 (P = 1.04×10⁻⁸). SNPs in previously established AD loci in Caucasians showed strong evidence of association in Japanese including rs3851179 near PICALM (P = 1.71×10⁻⁵) and rs744373 near BIN1 (P = 1.39×10⁻⁴). The associated allele for each of these SNPs was the same as in Caucasians. These data demonstrate for the first time genome-wide significance of LOAD with SORL1 and confirm the role of other known loci for LOAD in Japanese. Our study highlights the importance of examining associations in multiple ethnic populations.     

Association between Myocardial Triglyceride Content and Cardiac Function in Healthy Subjects and Endurance Athletes

Ectopic fat accumulation plays important roles in various metabolic disorders and cardiovascular diseases. Recent studies reported that myocardial triglyceride (TG) content measured by proton magnetic resonance spectroscopy (¹H-MRS) is associated with aging, diabetes mellitus, and cardiac dysfunction. However, myocardial TG content in athletes has not yet been investigated. We performed ¹H-MRS and cardiac magnetic resonance imaging in 10 male endurance athletes and 15 healthy male controls. Serum markers and other clinical parameters including arterial stiffness were measured. Cardiopulmonary exercise testing was also performed. There were no significant differences in clinical characteristics including age, anthropometric parameters, blood test results, or arterial stiffness between the two groups. Peak oxygen uptakes, end–diastolic volume (EDV), end–systolic volume (ESV), left ventricular (LV) mass, peak ejection rates and peak filling rates were significantly higher in the athlete group than in the control group (all P<0.02). Myocardial TG content was significantly lower in the athlete group than in the control group (0.60±0.20 vs. 0.89±0.41%, P<0.05). Myocardial TG content was negatively correlated with EDV (r = −0.47), ESV (r = −0.64), LV mass (r = −0.44), and epicardial fat volume (r = 0.47) (all P<0.05). In conclusion, lower levels of myocardial TG content were observed in endurance athletes and were associated with morphological changes related to physiological LV alteration in athletes, suggesting that metabolic imaging for measurement of myocardial TG content by ¹H-MRS may be a useful technique for noninvasively assessing the “athlete’s heart”.     

Improved Clinical Outcomes of High Risk β Thalassemia Major Patients Undergoing a HLA Matched Related Allogeneic Stem Cell Transplant with a Treosulfan Based Conditioning Regimen and Peripheral Blood Stem Cell Grafts

Improving clinical outcomes among high risk Class III β thalassemia major patients (Class IIIHR) receiving an allogeneic SCT remains a challenge. From October, 2009 a treosulfan based regimen (TreoFluT) was used for all consecutive Class III patients (n = 50). The clinical outcomes were compared with the historical conventional busulfan (BuCy) based regimen (n = 139). Use of TreoFluT was associated with a significantly reduced incidence of sinusoidal obstruction syndrome (SOS) among Class IIIHR cases (78% to 30%; P = 0.000) and early TRM (46% to 13%; p = 0.005). There was also a trend towards better engraftment in the Class IIIHR subset (P = 0.055). However, the use of bone marrow (BM) as source of stem cells along with the TreoFluT regimen was associated with 50% early mixed chimerism which reduced to 8.5% with the use of a peripheral blood stem cell graft (PBSC). Use of a PBSC graft was not associated with a significant increase in the incidence of acute or chronic graft versus host disease (GVHD). The overall and event free survival was significantly better among the Class IIIHR subset with the use of TreoFluT Vs. BuCy (86.6±7.3 Vs. 39.4±6.8%; P = 0.002 and 77.8±8.8 Vs. 32.4±6.5%; P = 0.003 respectively). A TreoFluT conditioning regimen with a PBSC graft can significantly improve clinical outcomes of Class IIIHR patients.     

Pacing-Induced Regional Differences in Adenosine Receptors mRNA Expression in a Swine Model of Dilated Cardiomyopathy

The adenosinergic system is essential in the mediation of intrinsic protection and myocardial resistance to insult; it may be considered a cardioprotective molecule and adenosine receptors (ARs) represent potential therapeutic targets in the setting of heart failure (HF). The aim of the study was to test whether differences exist between mRNA expression of ARs in the anterior left ventricle (LV) wall (pacing site: PS) compared to the infero septal wall (opposite region: OS) in an experimental model of dilated cardiomyopathy. Cardiac tissue was collected from LV PS and OS of adult male minipigs with pacing-induced HF (n = 10) and from a control group (C, n = 4). ARs and TNF–α mRNA expression was measured by Real Time-PCR and the results were normalized with the three most stably expressed genes (GAPDH, HPRT1, TBP). Immunohistochemistry analysis was also performed. After 3 weeks of pacing higher levels of expression for each analyzed AR were observed in PS except for A₁R (A₁R: C = 0.6±0.2, PS = 0.1±0.04, OS = 0.04±0.01, p<0.0001 C vs. PS and OS respectively; A_2AR: C = 1.04±0.59, PS = 2.62±0.79, OS = 2.99±0.79; A_2BR: C = 1.2±0.1, PS = 5.59±2.3, OS = 1.59±0.46; A₃R: C = 0.76±0.18, PS = 8.40±3.38, OS = 4.40±0.83). Significant contractile impairment and myocardial hypoperfusion were observed at PS after three weeks of pacing as compared to OS. TNF-α mRNA expression resulted similar in PS (6.3±2.4) and in OS (5.9±2.7) although higher than in control group (3.4±1.5). ARs expression was mainly detected in cardiomyocytes. This study provided new information on ARs local changes in the setting of LV dysfunction and on the role of these receptors in relation to pacing-induced abnormalities of myocardial perfusion and contraction. These results suggest a possible therapeutic role of adenosine in patients with HF and dyssynchronous LV contraction.     

Low-Intensity Pulsed Ultrasound Induces Angiogenesis and Ameliorates Left Ventricular Dysfunction in a Porcine Model of Chronic Myocardial Ischemia

Background

Although a significant progress has been made in the management of ischemic heart disease (IHD), the number of severe IHD patients is increasing. Thus, it is crucial to develop new, non-invasive therapeutic strategies. In the present study, we aimed to develop low-intensity pulsed ultrasound (LIPUS) therapy for the treatment of IHD.

Methods and Results

We first confirmed that in cultured human endothelial cells, LIPUS significantly up-regulated mRNA expression of vascular endothelial growth factor (VEGF) with a peak at 32-cycle (P<0.05). Then, we examined the in vivo effects of LIPUS in a porcine model of chronic myocardial ischemia with reduced left ventricular ejection fraction (LVEF) (n = 28). The heart was treated with either sham (n = 14) or LIPUS (32-cycle with 193 mW/cm² for 20 min, n = 14) at 3 different short axis levels. Four weeks after the treatment, LVEF was significantly improved in the LIPUS group (46±4 to 57±5%, P<0.05) without any adverse effects, whereas it remained unchanged in the sham group (46±5 to 47±6%, P = 0.33). Capillary density in the ischemic region was significantly increased in the LIPUS group compared with the control group (1084±175 vs. 858±151/mm², P<0.05). Regional myocardial blood flow was also significantly improved in the LIPUS group (0.78±0.2 to 1.39±0.4 ml/min/g, P<0.05), but not in the control group (0.84±0.3 to 0.97±0.4 ml/min/g). Western blot analysis showed that VEGF, eNOS and bFGF were all significantly up-regulated only in the LIPUS group.

Conclusions

These results suggest that the LIPUS therapy is promising as a new, non-invasive therapy for IHD.     

Periaortic Brown Adipose Tissue as a Major Determinant of [18F]-Fluorodeoxyglucose Vascular Uptake in Atherosclerosis-Prone,ApoE?/? Mice

Background

[¹⁸F]-fluorodeoxyglucose (FDG) has been suggested for the clinical and experimental imaging of inflammatory atherosclerotic lesions. Significant FDG uptake in brown adipose tissue (BAT) has been observed both in humans and mice. The objective of the present study was to investigate the influence of periaortic BAT on apolipoprotein E-deficient (apoE^−/−) mouse atherosclerotic lesion imaging with FDG.

Methods

ApoE^−/− mice (36±2 weeks-old) were injected with FDG (12±2 MBq). Control animals (Group A, n = 7) were injected conscious and kept awake at room temperature (24°C) throughout the accumulation period. In order to minimize tracer activity in periaortic BAT, Group B (n = 7) and C (n = 6) animals were injected under anaesthesia at 37°C and Group C animals were additionally pre-treated with propranolol. PET/CT acquisitions were performed prior to animal euthanasia and ex vivo analysis of FDG biodistribution.

Results

Autoradiographic imaging indicated higher FDG uptake in atherosclerotic lesions than in the normal aortic wall (all groups, P<0.05) and the blood (all groups, P<0.01) which correlated with macrophage infiltration (R = 0.47; P<0.001). However, periaortic BAT uptake was either significantly higher (Group A, P<0.05) or similar (Group B and C, P = NS) to that observed in atherosclerotic lesions and was shown to correlate with in vivo quantified aortic FDG activity.

Conclusion

Periaortic BAT FDG uptake was identified as a confounding factor while using FDG for the non-invasive imaging of mouse atherosclerotic lesions.     

Gene Expression Profiling in the Type 1 Diabetes Rat Diaphragm

Background

Respiratory muscle contractile performance is impaired by diabetes, mechanisms of which included altered carbohydrate and lipid metabolism, oxidative stress and changes in membrane electrophysiology. The present study examined to what extent these cellular perturbations involve changes in gene expression.

Methodology/Principal Findings

Diaphragm muscle from streptozotocin-diabetic rats was analyzed with Affymetrix gene expression arrays. Diaphragm from diabetic rats had 105 genes with at least ±2-fold significantly changed expression (55 increased, 50 decreased), and these were assigned to gene ontology groups based on over-representation analysis using DAVID software. There was increased expression of genes involved in palmitoyl-CoA hydrolase activity (a component of lipid metabolism) (P = 0.037, n = 2 genes, fold change 4.2 to 27.5) and reduced expression of genes related to carbohydrate metabolism (P = 0.000061, n = 8 genes, fold change −2.0 to −8.5). Other gene ontology groups among upregulated genes were protein ubiquitination (P = 0.0053, n = 4, fold change 2.2 to 3.4), oxidoreductase activity (P = 0.024, n = 8, fold change 2.1 to 6.0), and morphogenesis (P = 0.012, n = 10, fold change 2.1 to 4.3). Other downregulated gene groups were extracellular region (including extracellular matrix and collagen) (P = 0.00032, n = 13, fold change −2.2 to −3.7) and organogenesis (P = 0.032, n = 7, fold change −2.1 to −3.7). Real-time PCR confirmed the directionality of changes in gene expression for 30 of 31 genes tested.

Conclusions/Significance

These data indicate that in diaphragm muscle type 1 diabetes increases expression of genes involved in lipid energetics, oxidative stress and protein ubiquitination, decreases expression of genes involved in carbohydrate metabolism, and has little effect on expression of ion channel genes. Reciprocal changes in expression of genes involved in carbohydrate and lipid metabolism may change the availability of energetic substrates and thereby directly modulate fatigue resistance, an important issue for a muscle like the diaphragm which needs to contract without rest for the entire lifetime of the organism.     

A Genome-Wide Association Study Reveals Variants in ARL15 that Influence Adiponectin Levels

The adipocyte-derived protein adiponectin is highly heritable and inversely associated with risk of type 2 diabetes mellitus (T2D) and coronary heart disease (CHD). We meta-analyzed 3 genome-wide association studies for circulating adiponectin levels (n = 8,531) and sought validation of the lead single nucleotide polymorphisms (SNPs) in 5 additional cohorts (n = 6,202). Five SNPs were genome-wide significant in their relationship with adiponectin (P≤5×10⁻⁸). We then tested whether these 5 SNPs were associated with risk of T2D and CHD using a Bonferroni-corrected threshold of P≤0.011 to declare statistical significance for these disease associations. SNPs at the adiponectin-encoding ADIPOQ locus demonstrated the strongest associations with adiponectin levels (P-combined = 9.2×10⁻¹⁹ for lead SNP, rs266717, n = 14,733). A novel variant in the ARL15 (ADP-ribosylation factor-like 15) gene was associated with lower circulating levels of adiponectin (rs4311394-G, P-combined = 2.9×10⁻⁸, n = 14,733). This same risk allele at ARL15 was also associated with a higher risk of CHD (odds ratio [OR] = 1.12, P = 8.5×10⁻⁶, n = 22,421) more nominally, an increased risk of T2D (OR = 1.11, P = 3.2×10⁻³, n = 10,128), and several metabolic traits. Expression studies in humans indicated that ARL15 is well-expressed in skeletal muscle. These findings identify a novel protein, ARL15, which influences circulating adiponectin levels and may impact upon CHD risk.     

KCa3.1 Channel-Blockade Attenuates Airway Pathophysiology in a Sheep Model of Chronic Asthma

Background

The Ca²⁺-activated K⁺ channel K_Ca3.1 is expressed in several structural and inflammatory airway cell types and is proposed to play an important role in the pathophysiology of asthma. The aim of the current study was to determine whether inhibition of K_Ca3.1 modifies experimental asthma in sheep.

Methodology and Principal Findings

Atopic sheep were administered either 30 mg/kg Senicapoc (ICA-17073), a selective inhibitor of the K_Ca3.1-channel, or vehicle alone (0.5% methylcellulose) twice daily (orally). Both groups received fortnightly aerosol challenges with house dust mite allergen for fourteen weeks. A separate sheep group received no allergen challenges or drug treatment. In the vehicle-control group, twelve weeks of allergen challenges resulted in a 60±19% increase in resting airway resistance, and this was completely attenuated by treatment with Senicapoc (0.25±12%; n = 10, P = 0.0147). The vehicle-control group had a peak-early phase increase in lung resistance of 82±21%, and this was reduced by 58% with Senicapoc treatment (24±14%; n = 10, P = 0.0288). Senicapoc-treated sheep also demonstrated reduced airway hyperresponsiveness, requiring a significantly higher dose of carbachol to increase resistance by 100% compared to allergen-challenged vehicle-control sheep (20±5 vs. 52±18 breath-units of carbachol; n = 10, P = 0.0340). Senicapoc also significantly reduced eosinophil numbers in bronchoalveolar lavage taken 48 hours post-allergen challenge, and reduced vascular remodelling.

Conclusions

These findings suggest that K_Ca3.1-activity contributes to allergen-induced airway responses, inflammation and vascular remodelling in a sheep model of asthma, and that inhibition of K_Ca3.1 may be an effective strategy for blocking allergen-induced airway inflammation and hyperresponsiveness in humans.     

Self-Assembling Nanoparticles Containing Dexamethasone as a Novel Therapy in Allergic Airways Inflammation

Nanocarriers can deliver a wide variety of drugs, target them to sites of interest, and protect them from degradation and inactivation by the body. They have the capacity to improve drug action and decrease undesirable systemic effects. We have previously developed a well-defined non-toxic PEG-dendritic block telodendrimer for successful delivery of chemotherapeutics agents and, in these studies, we apply this technology for therapeutic development in asthma. In these proof-of-concept experiments, we hypothesized that dexamethasone contained in self-assembling nanoparticles (Dex-NP) and delivered systemically would target the lung and decrease allergic lung inflammation and airways hyper-responsiveness to a greater degree than equivalent doses of dexamethasone (Dex) alone. We found that ovalbumin (Ova)-exposed mice treated with Dex-NP had significantly fewer total cells (2.78±0.44×10⁵ (n = 18) vs. 5.98±1.3×10⁵ (n = 13), P<0.05) and eosinophils (1.09±0.28×10⁵ (n = 18) vs. 2.94±0.6×10⁵ (n = 12), p<0.05) in the lung lavage than Ova-exposed mice alone. Also, lower levels of the inflammatory cytokines IL-4 (3.43±1.2 (n = 11) vs. 8.56±2.1 (n = 8) pg/ml, p<0.05) and MCP-1 (13.1±3.6 (n = 8) vs. 28.8±8.7 (n = 10) pg/ml, p<0.05) were found in lungs of the Dex-NP compared to control, and they were not lower in the Dex alone group. In addition, respiratory system resistance was lower in the Dex-NP compared to the other Ova-exposed groups suggesting a better therapeutic effect on airways hyperresponsiveness. Taken together, these findings from early-stage drug development studies suggest that the encapsulation and protection of anti-inflammatory agents such as corticosteroids in nanoparticle formulations can improve efficacy. Further development of novel drugs in nanoparticles is warranted to explore potential treatments for chronic inflammatory diseases such as asthma.