Xenotransplantation of Human Cardiomyocyte Progenitor Cells Does Not Improve Cardiac Function in a Porcine Model of Chronic Ischemic Heart Failure. Results from a Randomized,Blinded, Placebo Controlled Trial

Background

Recently cardiomyocyte progenitor cells (CMPCs) were successfully isolated from fetal and adult human hearts. Direct intramyocardial injection of human CMPCs (hCMPCs) in experimental mouse models of acute myocardial infarction significantly improved cardiac function compared to controls.

Aim

Here, our aim was to investigate whether xenotransplantation via intracoronary infusion of fetal hCMPCs in a pig model of chronic myocardial infarction is safe and efficacious, in view of translation purposes.

Methods & Results

We performed a randomized, blinded, placebo controlled trial. Four weeks after ischemia/reperfusion injury by 90 minutes of percutaneous left anterior descending artery occlusion, pigs (n = 16, 68.5 ± 5.4 kg) received intracoronary infusion of 10 million fetal hCMPCs or placebo. All animals were immunosuppressed by cyclosporin (CsA). Four weeks after infusion, endpoint analysis by MRI displayed no difference in left ventricular ejection fraction, left ventricular end diastolic and left ventricular end systolic volumes between both groups. Serial pressure volume (PV-)loop and echocardiography showed no differences in functional parameters between groups at any timepoint. Infarct size at follow-up, measured by late gadolinium enhancement MRI showed no difference between groups. Intracoronary pressure and flow measurements showed no signs of coronary obstruction 30 minutes after cell infusion. No premature death occurred in cell treated animals.

Conclusion

Xenotransplantation via intracoronary infusion of hCMPCs is feasible and safe, but not associated with improved left ventricular performance and infarct size compared to placebo in a porcine model of chronic myocardial infarction.     

Intramyocardial Delivery of Mesenchymal Stem Cell-Seeded Hydrogel Preserves Cardiac Function and Attenuates Ventricular Remodeling after Myocardial Infarction

Background

To improve the efficacy of bone marrow-derived mesenchymal stem cell (MSC) therapy targeted to infarcted myocardium, we investigated whether a self-setting silanized hydroxypropyl methylcellulose (Si-HPMC) hydrogel seeded with MSC (MSC+hydrogel) could preserve cardiac function and attenuate left ventricular (LV) remodeling during an 8-week follow-up study in a rat model of myocardial infarction (MI).

Methodology/Principal Finding

Si-HPMC hydrogel alone, MSC alone or MSC+hydrogel were injected into the myocardium immediately after coronary artery ligation in female Lewis rats. Animals in the MSC+hydrogel group showed an increase in cardiac function up to 28 days after MI and a mid-term prevention of cardiac function alteration at day 56. Histological analyses indicated that the injection of MSC+hydrogel induced a decrease in MI size and an increase in scar thickness and ultimately limited the transmural extent of MI. These findings show that intramyocardial injection of MSC+hydrogel induced short-term recovery of ventricular function and mid-term attenuation of remodeling after MI.

Conclusion/Significance

These beneficial effects may be related to the specific scaffolding properties of the Si-HPMC hydrogel that may provide the ability to support MSC injection and engraftment within myocardium.     

Intracoronary Delivery of Human Mesenchymal/Stromal Stem Cells: Insights from Coronary Microcirculation Invasive Assessment in a Swine Model

Background

Mesenchymal stem/stromal cells have unique properties favorable to their use in clinical practice and have been studied for cardiac repair. However, these cells are larger than coronary microvessels and there is controversy about the risk of embolization and microinfarctions, which could jeopardize the safety and efficacy of intracoronary route for their delivery. The index of microcirculatory resistance (IMR) is an invasive method for quantitatively assessing the coronary microcirculation status.

Objectives

To examine heart microcirculation after intracoronary injection of mesenchymal stem/stromal cells with the index of microcirculatory resistance.

Methods

Healthy swine were randomized to receive by intracoronary route either 30x10⁶ MSC or the same solution with no cells (1% human albumin/PBS) (placebo). Blinded operators took coronary pressure and flow measurements, prior to intracoronary infusion and at 5 and 30 minutes post-delivery. Coronary flow reserve (CFR) and the IMR were compared between groups.

Results

CFR and IMR were done with a variance within the 3 transit time measurements of 6% at rest and 11% at maximal hyperemia. After intracoronary infusion there were no significant differences in CFR. The IMR was significantly higher in MSC-injected animals (at 30 minutes, 14.2U vs. 8.8U, p = 0.02) and intragroup analysis showed a significant increase of 112% from baseline to 30 minutes after cell infusion, although no electrocardiographic changes or clinical deterioration were noted.

Conclusion

Overall, this study provides definitive evidence of microcirculatory disruption upon intracoronary administration of mesenchymal stem/stromal cells, in a large animal model closely resembling human cardiac physiology, function and anatomy.     

Exercise related ventricular arrhythmias are related to cardiac fibrosis in hypertrophic cardiomyopathy mutation carriers

Aims

Hypertrophic cardiomyopathy (HCM) is a frequent cause of sudden cardiac death (SCD) due to exercise-related ventricular arrhythmias (ERVA); however the pathological substrate is uncertain. The aim was to determine the prevalence of ERVA and their relation with fibrosis as determined by cardiac magnetic resonance imaging (CMR) in carriers of an HCM causing mutation.

Methods

We studied the prevalence and origin of ERVA and related these with fibrosis on CMR in a population of 31 HCM mutation carriers.

Results

ERVA occurred in seven patients (23%) who all showed evidence of fibrosis (100% ERVA(+) vs. 58% ERVA(-), p = 0.04). No ventricular tachycardia or ventricular fibrillation occurred. In patients with ERVA, the extent of fibrosis was significantly larger (8 ± 4% vs. 3 ± 4%, p = 0.02). ERVA originated from areas with a high extent of fibrosis or regions directly adjacent to these areas.

Conclusions

ERVA in HCM mutation carriers arose from the area of fibrosis detected by CMR; ERVA seems closely related to cardiac fibrosis. Fibrosis as detected by CMR should be evaluated as an additional risk factor to further delineate risk of SCD in carriers of an HCM causing mutation.     

In Vivo Early Intervention and the Therapeutic Effects of 20(S)-Ginsenoside Rg3 on Hypertrophic Scar Formation

Background

Intra-lesional injections of corticosteroids, interferon, and chemotherapeutic drugs are currently the most popular treatments of hypertrophic scar formation. However, these drugs can only be used after HS is formed, and not during the inflammatory phase of wound healing, which regulates the HS forming process.

Objective

To investigate a new, effective, combining therapeutic and safe drug for early intervention and treatment for hypertrophic scars.

Methods

Cell viability assay and flow cytometric analysis were studied in vitro. Animal studies were done to investigate the combining therapeutic effects of 20(S)-ginsenoside Rg3 (Rg3) on the inflammatory phase of wound healing and HS formation.

Results

In vitro studies showed that Rg3 can inhibit HS fibroblasts proliferation and induce HSF apoptosis in a concentration-dependent manner. In vivo studies demonstrated that Rg3 can limit the exaggerated inflammation, and do not delay the wound healing process, which indicates that Rg3 could be used as an early intervention to reduce HS formation. Topical injection of 4 mg/mL Rg3 can reduce HS formation by 34%. Histological and molecular studies revealed that Rg3 injection inhibits fibroblasts proliferation thus reduced the accumulation of collagen fibers, and down-regulates VEGF expression in the HS tissue.

Conclusion

Rg3 can be employed as an early intervention and a combining therapeutic drug to reduce inflammation and HS formation as well.     

Diclofenac Prolongs Repolarization in Ventricular Muscle with Impaired Repolarization Reserve

Background

The aim of the present work was to characterize the electrophysiological effects of the non-steroidal anti-inflammatory drug diclofenac and to study the possible proarrhythmic potency of the drug in ventricular muscle.

Methods

Ion currents were recorded using voltage clamp technique in canine single ventricular cells and action potentials were obtained from canine ventricular preparations using microelectrodes. The proarrhythmic potency of the drug was investigated in an anaesthetized rabbit proarrhythmia model.

Results

Action potentials were slightly lengthened in ventricular muscle but were shortened in Purkinje fibers by diclofenac (20 µM). The maximum upstroke velocity was decreased in both preparations. Larger repolarization prolongation was observed when repolarization reserve was impaired by previous BaCl₂ application. Diclofenac (3 mg/kg) did not prolong while dofetilide (25 µg/kg) significantly lengthened the QT_c interval in anaesthetized rabbits. The addition of diclofenac following reduction of repolarization reserve by dofetilide further prolonged QT_c. Diclofenac alone did not induce Torsades de Pointes ventricular tachycardia (TdP) while TdP incidence following dofetilide was 20%. However, the combination of diclofenac and dofetilide significantly increased TdP incidence (62%). In single ventricular cells diclofenac (30 µM) decreased the amplitude of rapid (I_Kr) and slow (I_Ks) delayed rectifier currents thereby attenuating repolarization reserve. L-type calcium current (I_Ca) was slightly diminished, but the transient outward (I_to) and inward rectifier (I_K1) potassium currents were not influenced.

Conclusions

Diclofenac at therapeutic concentrations and even at high dose does not prolong repolarization markedly and does not increase the risk of arrhythmia in normal heart. However, high dose diclofenac treatment may lengthen repolarization and enhance proarrhythmic risk in hearts with reduced repolarization reserve.     

Aedes Mosquito Saliva Modulates Rift Valley Fever Virus Pathogenicity

Background

Rift Valley fever (RVF) is a severe mosquito-borne disease affecting humans and domestic ruminants. Mosquito saliva contains compounds that counteract the hemostatic, inflammatory, and immune responses of the host. Modulation of these defensive responses may facilitate virus infection. Indeed, Aedes mosquito saliva played a crucial role in the vector''s capacity to effectively transfer arboviruses such as the Cache Valley and West Nile viruses. The role of mosquito saliva in the transmission of Rift Valley fever virus (RVFV) has not been investigated.

Objective

Using a murine model, we explored the potential for mosquitoes to impact the course of RVF disease by determining whether differences in pathogenesis occurred in the presence or absence of mosquito saliva and salivary gland extract.

Methods

C57BL/6NRJ male mice were infected with the ZH548 strain of RVFV via intraperitoneal or intradermal route, or via bites from RVFV-exposed mosquitoes. The virus titers in mosquitoes and mouse organs were determined by plaque assays.

Findings

After intraperitoneal injection, RVFV infection primarily resulted in liver damage. In contrast, RVFV infection via intradermal injection caused both liver and neurological symptoms and this route best mimicked the natural infection by mosquitoes. Co-injections of RVFV with salivary gland extract or saliva via intradermal route increased the mortality rates of mice, as well as the virus titers measured in several organs and in the blood. Furthermore, the blood cell counts of infected mice were altered compared to those of uninfected mice.

Interpretation

Different routes of infection determine the pattern in which the virus spreads and the organs it targets. Aedes saliva significantly increases the pathogenicity of RVFV.     

Cell Therapy Attenuates Cardiac Dysfunction Post Myocardial Infarction: Effect of Timing,Routes of Injection and a Fibrin Scaffold

Background

Cell therapy approaches for biologic cardiac repair hold great promises, although basic fundamental issues remain poorly understood. In the present study we examined the effects of timing and routes of administration of bone marrow cells (BMC) post-myocardial infarction (MI) and the efficacy of an injectable biopolymer scaffold to improve cardiac cell retention and function.

Methodology/Principal Findings

^99mTc-labeled BMC (6×10⁶ cells) were injected by 4 different routes in adult rats: intravenous (IV), left ventricular cavity (LV), left ventricular cavity with temporal aorta occlusion (LV⁺) to mimic coronary injection, and intramyocardial (IM). The injections were performed 1, 2, 3, or 7 days post-MI and cell retention was estimated by γ-emission counting of the organs excised 24 hs after cell injection. IM injection improved cell retention and attenuated cardiac dysfunction, whereas IV, LV or LV* routes were somewhat inefficient (<1%). Cardiac BMC retention was not influenced by timing except for the IM injection that showed greater cell retention at 7 (16%) vs. 1, 2 or 3 (average of 7%) days post-MI. Cardiac cell retention was further improved by an injectable fibrin scaffold at day 3 post-MI (17 vs. 7%), even though morphometric and function parameters evaluated 4 weeks later displayed similar improvements.

Conclusions/Significance

These results show that cells injected post-MI display comparable tissue distribution profile regardless of the route of injection and that there is no time effect for cardiac cell accumulation for injections performed 1 to 3 days post-MI. As expected the IM injection is the most efficient for cardiac cell retention, it can be further improved by co-injection with a fibrin scaffold and it significantly attenuates cardiac dysfunction evaluated 4 weeks post myocardial infarction. These pharmacokinetic data obtained under similar experimental conditions are essential for further development of these novel approaches.     

Remote magnetic navigation vs. manual navigation for ablation of ventricular tachycardia: a meta-analysis

Background

The purpose of this study was to prospectively evaluate the efficacy and safety of remote magnetic navigation (RMN) in comparison with manual catheter navigation (MCN) in performing ventricular tachycardia ablation.

Methods

An electronic search was performed using PubMed (1948–2013) and EMBASE (1974–2013) studies comparing RMN with MCN which were published prior to 31 December 2013. Outcomes of interest were as follows: acute success, recurrence rate, complications, total procedure and fluoroscopic times. Standard mean difference (SMD) and its 95 % confidence interval (CI) were used for continuous outcomes; odds ratios (OR) were reported for dichotomous variables.

Results

Four non-randomised studies, including a total of 328 patients, were identified. RMN was deployed in 191 patients. Acute success and long-term freedom from arrhythmias were not significantly different between the RMN and control groups (OR 1.845, 95 % CI 0.731–4.659, p = 0.195 and OR 0.676, 95 % CI 0.383–1.194, p = 0.177, respectively). RMN was associated with less peri-procedural complications (OR 0.279, 95 % CI 0.092–0.843, p = 0.024). Shorter procedural and fluoroscopy times were achieved (95 % CI -0.487 to -0.035, p = 0.024 and 95 % CI -1.467 to -0.984, p<0.001, respectively).

Conclusion

The acute and long-term success rates for VT ablation are equal between RMN and MCN, whereas the RMN-guided procedure can be performed with a lower complication rate and less procedural and fluoroscopic times. More prospective randomised trials will be needed to better evaluate the superior role of RMN for catheter ablation of ventricular tachycardia.     

HSPB1, HSPB6, HSPB7 and HSPB8 protect against RhoA GTPase-induced remodeling in tachypaced atrial myocytes

Background

We previously demonstrated the small heat shock protein, HSPB1, to prevent tachycardia remodeling in in vitro and in vivo models for Atrial Fibrillation (AF). To gain insight into its mechanism of action, we examined the protective effect of all 10 members of the HSPB family on tachycardia remodeling. Furthermore, modulating effects of HSPB on RhoA GTPase activity and F-actin stress fiber formation were examined, as this pathway was found of prime importance in tachycardia remodeling events and the initiation of AF.

Methods and Results

Tachypacing (4 Hz) of HL-1 atrial myocytes significantly and progressively reduced the amplitude of Ca²⁺ transients (CaT). In addition to HSPB1, also overexpression of HSPB6, HSPB7 and HSPB8 protected against tachypacing-induced CaT reduction. The protective effect was independent of HSPB1. Moreover, tachypacing induced RhoA GTPase activity and caused F-actin stress fiber formation. The ROCK inhibitor Y27632 significantly prevented tachypacing-induced F-actin formation and CaT reductions, showing that RhoA activation is required for remodeling. Although all protective HSPB members prevented the formation of F-actin stress fibers, their mode of action differs. Whilst HSPB1, HSPB6 and HSPB7 acted via direct prevention of F-actin formation, HSPB8-protection was mediated via inhibition of RhoA GTPase activity.

Conclusion

Overexpression of HSPB1, as well as HSPB6, HSPB7 and HSPB8 independently protect against tachycardia remodeling by attenuation of the RhoA GTPase pathway at different levels. The cardioprotective role for multiple HSPB members indicate a possible therapeutic benefit of compounds able to boost the expression of single or multiple members of the HSPB family.     

Crucial Residue Involved in L-Lactate Recognition by Human Monocarboxylate Transporter 4 (hMCT4)

Background

Monocarboxylate transporters (MCTs) transport monocarboxylates such as lactate, pyruvate and ketone bodies. These transporters are very attractive therapeutic targets in cancer. Elucidations of the functions and structures of MCTs is necessary for the development of effective medicine which targeting these proteins. However, in comparison with MCT1, there is little information on location of the function moiety of MCT4 and which constituent amino acids govern the transport function of MCT4. The aim of the present work was to determine the molecular mechanism of L-lactate transport via hMCT4.

Experimental approach

Transport of L-lactate via hMCT4 was determined by using hMCT4 cRNA-injected Xenopus laevis oocytes. hMCT4 mediated L-lactate uptake in oocytes was measured in the absence and presence of chemical modification agents and 4,4′-diisothiocyanostilbene-2,2′-disulphonate (DIDS). In addition, L-lactate uptake was measured by hMCT4 arginine mutants. Immunohistochemistry studies revealed the localization of hMCT4.

Results

In hMCT4-expressing oocytes, treatment with phenylglyoxal (PGO), a compound specific for arginine residues, completely abolished the transport activity of hMCT4, although this abolishment was prevented by the presence of L-lactate. On the other hand, chemical modifications except for PGO treatment had no effect on the transport activity of hMCT4. The transporter has six conserved arginine residues, two in the transmembrane-spanning domains (TMDs) and four in the intracellular loops. In hMCT4-R278 mutants, the uptake of L-lactate is void of any transport activity without the alteration of hMCT4 localization.

Conclusions

Our results suggest that Arg-278 in TMD8 is a critical residue involved in substrate, L-lactate recognition by hMCT4.     

Efficacy of Mycobacterium indicus pranii Immunotherapy as an Adjunct to Chemotherapy for Tuberculosis and Underlying Immune Responses in the Lung

Background

The 9-month-long chemotherapy of tuberculosis often results in poor compliance and emergence of drug-resistant strains. So, improved therapeutic strategy is urgently needed. Immunotherapy could be beneficial for the effective management of the disease. Previously we showed the protective efficacy of Mycobacterium indicus pranii (MIP) when given as prophylactic vaccine in animal models of tuberculosis.

Methods

We sought to investigate whether MIP can be used as an adjunct to the chemotherapy in guinea pig models of tuberculosis. Efficacy of MIP was evaluated when given subcutaneously or by aerosol.

Results

MIP-therapy as an adjunct to the chemotherapy was found to be effective in accelerating bacterial killing and improving organ pathology. MIP-immunotherapy resulted in higher numbers of activated antigen-presenting cells and lymphocytes in the infected lungs and also modulated the granulomatous response. Early increase in protective Th1 immune response was observed in the immunotherapy group. Following subsequent doses of MIP, decrease in the inflammatory response and increase in the immunosuppressive response was observed, which resulted in the improvement of lung pathology.

Conclusion

MIP immunotherapy is a valuable adjunct to chemotherapy for tuberculosis. Aerosol route of immunotherapy can play a crucial role for inducing immediate local immune response in the lung.     

Observational Cohort Study of Ventricular Arrhythmia in Adults with Marfan Syndrome Caused by FBN1 Mutations

Background

Marfan syndrome is associated with ventricular arrhythmia but risk factors including FBN1 mutation characteristics require elucidation.

Methods and Results

We performed an observational cohort study of 80 consecutive adults (30 men, 50 women aged 42±15 years) with Marfan syndrome caused by FBN1 mutations. We assessed ventricular arrhythmia on baseline ambulatory electrocardiography as >10 premature ventricular complexes per hour (>10 PVC/h), as ventricular couplets (Couplet), or as non-sustained ventricular tachycardia (nsVT), and during 31±18 months of follow-up as ventricular tachycardia (VT) events (VTE) such as sudden cardiac death (SCD), and sustained ventricular tachycardia (sVT). We identified >10 PVC/h in 28 (35%), Couplet/nsVT in 32 (40%), and VTE in 6 patients (8%), including 3 with SCD (4%). PVC>10/h, Couplet/nsVT, and VTE exhibited increased N-terminal pro–brain natriuretic peptide serum levels(P<.001). All arrhythmias related to increased NT-proBNP (P<.001), where PVC>10/h and Couplet/nsVT also related to increased indexed end-systolic LV diameters (P = .024 and P = .020), to moderate mitral valve regurgitation (P = .018 and P = .003), and to prolonged QTc intervals (P = .001 and P = .006), respectively. Moreover, VTE related to mutations in exons 24–32 (P = .021). Kaplan–Meier analysis corroborated an association of VTE with increased NT-proBNP (P<.001) and with mutations in exons 24–32 (P<.001).

Conclusions

Marfan syndrome with causative FBN1 mutations is associated with an increased risk for arrhythmia, and affected persons may require life-long monitoring. Ventricular arrhythmia on electrocardiography, signs of myocardial dysfunction and mutations in exons 24–32 may be risk factors of VTE.     

Anti-Angiogenic Effects of a Mutant Endostatin: A New Prospect for Treating Retinal and Choroidal Neovascularization

Purpose

Pathological fundus angiogenesis is a major cause of vision loss in retina diseases. Endostatin, a C-terminal fragment of collagen XVIII, is an endogenous anti-angiogenic protein. The present study aimed to investigate the in vitro and in vivo anti-angiogenic properties of two proteins: an N-terminal H1D/H3D mutant endostatin (M-ES) and a polyethylene glycol propionaldehyde (PEG) covalent M-ES (PEG-M-ES).

Methods

M-ES and PEG-M-ES properties were characterized in vitro using a zinc ion binding assay and a stability test. Activity assays, including migration, proliferation, and tube formation assays, were performed with human retinal microvascular endothelial cells (HRMECs) and human umbilical vein endothelial cells (HUVECs). Mouse oxygen-induced retinopathy (OIR) and choroidal neovascularization (CNV) models were used to evaluate in vivo anti-angiogenic effects. In addition, a rabbit model was used to study the retinal pharmacokinetic profile following an intravitreal injection.

Results

The results indicated that the H1D/H3D mutations of endostatin reduced the zinc binding capacity of M-ES and facilitated PEG covalent binding. PEG-M-ES was more stable and persisted longer in the retina compared with M-ES. The in vitro studies demonstrated that M-ES and PEG-M-ES inhibited HRMEC and HUVEC proliferation, migration, and tube formation more efficiently than ES. In vivo, a single intravitreal injection of M-ES and PEG-M-ES significantly decreased neovascularization in both the OIR and CNV animal models.

Conclusion

The present study demonstrated for the first time that PEG-M-ES exhibits a long-term inhibitory effect on neovascularization in vitro and in vivo. These data suggest that PEG-M-ES may represent an innovative therapeutic strategy to prevent fundus neovascularization.     

Hemochromatosis Enhances Tumor Progression via Upregulation of Intracellular Iron in Head and Neck Cancer

Introduction

Despite improvements in treatment strategies for head and neck squamous cell carcinoma (HNSCC), outcomes have not significantly improved; highlighting the importance of identifying novel therapeutic approaches to target this disease. To address this challenge, we proceeded to evaluate the role of iron in HNSCC.

Experimental Design

Expression levels of iron-related genes were evaluated in HNSCC cell lines using quantitative RT-PCR. Cellular phenotypic effects were assessed using viability (MTS), clonogenic survival, BrdU, and tumor formation assays. The prognostic significance of iron-related proteins was determined using immunohistochemistry.

Results

In a panel of HNSCC cell lines, hemochromatosis (HFE) was one of the most overexpressed genes involved in iron regulation. In vitro knockdown of HFE in HNSCC cell lines significantly decreased hepcidin (HAMP) expression and intracellular iron level. This in turn, resulted in a significant decrease in HNSCC cell viability, clonogenicity, DNA synthesis, and Wnt signalling. These cellular changes were reversed by re-introducing iron back into HNSCC cells after HFE knockdown, indicating that iron was mediating this phenotype. Concordantly, treating HNSCC cells with an iron chelator, ciclopirox olamine (CPX), significantly reduced viability and clonogenic survival. Finally, patients with high HFE expression experienced a reduced survival compared to patients with low HFE expression.

Conclusions

Our data identify HFE as potentially novel prognostic marker in HNSCC that promotes tumour progression via HAMP and elevated intracellular iron levels, leading to increased cellular proliferation and tumour formation. Hence, these findings suggest that iron chelators might have a therapeutic role in HNSCC management.     

The Non-Peptidic Part Determines the Internalization Mechanism and Intracellular Trafficking of Peptide Amphiphiles

Background

Peptide amphiphiles (PAs) are a class of amphiphilic molecules able to self-assemble into nanomaterials that have shown efficient in vivo targeted delivery. Understanding the interactions of PAs with cells and the mechanisms of their internalization and intracellular trafficking is critical in their further development for therapeutic delivery applications.

Methodology/Principal Findings

PAs of a novel, cell- and tissue-penetrating peptide were synthesized possessing two different lipophilic tail architectures and their interactions with prostate cancer cells were studied in vitro. Cell uptake of peptides was greatly enhanced post-modification. Internalization occurred via lipid-raft mediated endocytosis and was common for the two analogs studied. On the contrary, we identified the non-peptidic part as the determining factor of differences between intracellular trafficking and retention of PAs. PAs composed of di-stearyl lipid tails linked through poly(ethylene glycol) to the peptide exhibited higher exocytosis rates and employed different recycling pathways compared to ones consisting of di-palmitic-coupled peptides. As a result, cell association of the former PAs decreased with time.

Conclusions/Significance

Control over peptide intracellular localization and retention is possible by appropriate modification with synthetic hydrophobic tails. We propose this as a strategy to design improved peptide-based delivery systems.     

Comparison of NOGA Endocardial Mapping and Cardiac Magnetic Resonance Imaging for Determining Infarct Size and Infarct Transmurality for Intramyocardial Injection Therapy Using Experimental Data

Objectives

We compared the accuracy of NOGA endocardial mapping for delineating transmural and non-transmural infarction to the results of cardiac magnetic resonance imaging (cMRI) with late gadolinium enhancement (LE) for guiding intramyocardial reparative substance delivery using data from experimental myocardial infarction studies.

Methods

Sixty domestic pigs underwent diagnostic NOGA endocardial mapping and cMRI-LE 60 days after induction of closed-chest reperfused myocardial infarction. The infarct size was determined by LE of cMRI and by delineation of the infarct core on the unipolar voltage polar map. The sizes of the transmural and non-transmural infarctions were calculated from the cMRI transmurality map using signal intensity (SI) cut-offs of>75% and>25% and from NOGA bipolar maps using bipolar voltage cut-off values of <0.8 mV and <1.9 mV. Linear regression analysis and Bland-Altman plots were used to determine correlations and systematic differences between the two images. The overlapping ratios of the transmural and non-transmural infarcted areas were calculated.

Results

Infarct size as determined by 2D NOGA unipolar voltage polar mapping correlated with the 3D cMRI-LE findings (r = 0.504, p<0.001) with a mean difference of 2.82% in the left ventricular (LV) surface between the two images. Polar maps of transmural cMRI and bipolar maps of NOGA showed significant association for determining of the extent of transmural infarction (r = 0.727, p<0.001, overlap ratio of 81.6±11.1%) and non-transmural infarction (r = 0.555, p<0.001, overlap ratio of 70.6±18.5%). NOGA overestimated the transmural scar size (6.81% of the LV surface) but slightly underestimated the size of the non-transmural infarction (−3.04% of the LV surface).

Conclusions

By combining unipolar and bipolar voltage maps, NOGA endocardial mapping is useful for accurate delineation of the targeted zone for intramyocardial therapy and is comparable to cMRI-LE. This may be useful in patients with contraindications for cMRI who require targeted intramyocardial regenerative therapy.     

Xanthohumol Prevents Atherosclerosis by Reducing Arterial Cholesterol Content via CETP and Apolipoprotein E in CETP-Transgenic Mice

Background

Xanthohumol is expected to be a potent anti-atherosclerotic agent due to its inhibition of cholesteryl ester transfer protein (CETP). In this study, we hypothesized that xanthohumol prevents atherosclerosis in vivo and used CETP-transgenic mice (CETP-Tg mice) to evaluate xanthohumol as a functional agent.

Methodology/Principal Findings

Two strains of mice, CETP-Tg and C57BL/6N (wild-type), were fed a high cholesterol diet with or without 0.05% (w/w) xanthohumol ad libitum for 18 weeks. In CETP-Tg mice, xanthohumol significantly decreased accumulated cholesterol in the aortic arch and increased HDL cholesterol (HDL-C) when compared to the control group (without xanthohumol). Xanthohumol had no significant effect in wild-type mice. CETP activity was significantly decreased after xanthohumol addition in CETP-Tg mice compared with the control group and it inversely correlated with HDL-C (%) (P<0.05). Furthermore, apolipoprotein E (apoE) was enriched in serum and the HDL-fraction in CETP-Tg mice after xanthohumol addition, suggesting that xanthohumol ameliorates reverse cholesterol transport via apoE-rich HDL resulting from CETP inhibition.

Conclusions

Our results suggest xanthohumol prevents cholesterol accumulation in atherogenic regions by HDL-C metabolism via CETP inhibition leading to apoE enhancement.     

Sodium Current Reduction Unmasks a Structure-Dependent Substrate for Arrhythmogenesis in the Normal Ventricles

Background

Organ-scale arrhythmogenic consequences of source-sink mismatch caused by impaired excitability remain unknown, hindering the understanding of pathophysiology in disease states like Brugada syndrome and ischemia.

Objective

We sought to determine whether sodium current (I_Na) reduction in the structurally normal heart unmasks a regionally heterogeneous substrate for the induction of sustained arrhythmia by premature ventricular contractions (PVCs).

Methods

We conducted simulations in rabbit ventricular computer models with 930 unique combinations of PVC location (10 sites) and coupling interval (250–400 ms), I_Na reduction (30 or 40% of normal levels), and post-PVC sinus rhythm (arrested or persistent). Geometric characteristics and source-sink mismatch were quantitatively analyzed by calculating ventricular wall thickness and a newly formulated 3D safety factor (SF), respectively.

Results

Reducing I_Na to 30% of its normal level created a substrate for sustained arrhythmia induction by establishing large regions of critical source-sink mismatch (SF<1) for ectopic wavefronts propagating from thin to thick tissue. In the same simulations but with 40% of normal I_Na, PVCs did not induce reentry because the volume of tissue with SF<1 was >95% smaller. Likewise, when post-PVC sinus activations were persistent instead of arrested, no ectopic excitations initiated sustained reentry because sinus activation breakthroughs engulfed the excitable gap.

Conclusion

Our new SF formulation can quantify ectopic wavefront propagation robustness in geometrically complex 3D tissue with impaired excitability. This novel methodology was applied to show that I_Na reduction precipitates source-sink mismatch, creating a potent substrate for sustained arrhythmia induction by PVCs originating near regions of ventricular wall expansion, such as the RV outflow tract.     

Anti-Thymocyte Globulin Induces Neoangiogenesis and Preserves Cardiac Function after Experimental Myocardial Infarction

Rationale

Acute myocardial infarction (AMI) followed by ventricular remodeling is the major cause of congestive heart failure and death in western world countries.

Objective

Of relevance are reports showing that infusion of apoptotic leucocytes or anti-lymphocyte serum after AMI reduces myocardial necrosis and preserves cardiac function. In order to corroborate this therapeutic mechanism, the utilization of an immunosuppressive agent with a comparable mechanism, such as anti-thymocyte globulin (ATG) was evaluated in this study.

Methods and Results

AMI was induced in rats by ligation of the left anterior descending artery. Initially after the onset of ischemia, rabbit ATG (10 mg/rat) was injected intravenously. In vitro and in vivo experiments showed that ATG induced a pronounced release of pro-angiogenic and chemotactic factors. Moreover, paracrine factors released from ATG co-incubated cell cultures conferred a down-regulation of p53 in cardiac myocytes. Rats that were injected with ATG evidenced higher numbers of CD68+ macrophages in the ischemic myocardium. Animals injected with ATG evidenced less myocardial necrosis, showed a significant reduction of infarct dimension and an improvement of post-AMI remodeling after six weeks (infarct dimension 24.9% vs. 11.4%, p<0.01). Moreover, a higher vessel density in the peri-infarct region indicated a better collateralization in rats that were injected with ATG.

Conclusions

These data indicate that ATG, a therapeutic agent successfully applied in clinical transplant immunology, triggered cardioprotective effects after AMI that salvaged ischemic myocardium by down-regulation of p53. This might have raised the resistance against apoptotic cell death during ischemia. The combination of these mechanisms seems to be causative for improved cardiac function and less ventricular remodeling after experimental AMI.