Coronary flow capacity and survival prediction after revascularization: physiological basis and clinical implications.

BACKGROUND AND AIMS: Coronary flow capacity (CFC) is associated with an observed 10-year survival probability for individual patients before and after actual revascularization for comparison to virtual hypothetical ideal complete revascularization. METHODS: Stress myocardial perfusion (mL/min/g) and coronary flow reserve (CFR) per pixel were quantified in 6979 coronary artery disease (CAD) subjects using Rb-82 positron emission tomography (PET) for CFC maps of artery-specific size-severity abnormalities expressed as percent left ventricle with prospective follow-up to define survival probability per-decade as fraction of 1.0. RESULTS: Severely reduced CFC in 6979 subjects predicted low survival probability that improved by 42% after revascularization compared with no revascularization for comparable severity (P = .0015). For 283 pre-and-post-procedure PET pairs, severely reduced regional CFC-associated survival probability improved heterogeneously after revascularization (P < .001), more so after bypass surgery than percutaneous coronary interventions (P < .001) but normalized in only 5.7%; non-severe baseline CFC or survival probability did not improve compared with severe CFC (P = .00001). Observed CFC-associated survival probability after actual revascularization was lower than virtual ideal hypothetical complete post-revascularization survival probability due to residual CAD or failed revascularization (P < .001) unrelated to gender or microvascular dysfunction. Severely reduced CFC in 2552 post-revascularization subjects associated with low survival probability also improved after repeat revascularization compared with no repeat procedures (P = .025). CONCLUSIONS: Severely reduced CFC and associated observed survival probability improved after first and repeat revascularization compared with no revascularization for comparable CFC severity. Non-severe CFC showed no benefit. Discordance between observed actual and virtual hypothetical post-revascularization survival probability revealed residual CAD or failed revascularization.

Co-Delivery of Therapeutics and Bioactive Gas Using a Novel Liposomal Platform for Enhanced Treatment of Acute Arterial Injury.

Atherosclerosis is a complex, multi-stage disease characterized by pathological changes across the vascular wall. Endothelial dysfunction, inflammation, hypoxia, and vascular smooth muscle cell proliferation contribute to its progression. An effective strategy capable of delivering pleiotropic treatment to the vascular wall is essential to limit neointimal formation. Echogenic liposomes (ELIP), which can encapsulate bioactive gases and therapeutic agents, have the potential to deliver enhanced penetration and treatment efficacy for atherosclerosis. In this study, liposomes loaded with nitric oxide (NO) and rosiglitazone, a peroxisome proliferator-activated receptor agonist, were prepared using hydration, sonication, freeze-thawing, and pressurization. The efficacy of this delivery system was evaluated in a rabbit model of acute arterial injury induced by balloon injury to the common carotid artery. Intra-arterial administration of rosiglitazone/NO co-encapsulated liposomes (R/NO-ELIP) immediately following injury resulted in reduced intimal thickening after 14 days. The anti-inflammatory and anti-proliferative effects of the co-delivery system were investigated. These liposomes were echogenic, enabling ultrasound imaging to assess their distribution and delivery. R/NO-ELIP delivery exhibited a greater attenuation (88 ± 15%) of intimal proliferation when compared to NO-ELIP (75 ± 13%) or R-ELIP (51 ± 6%) delivery alone. The study demonstrates the potential of echogenic liposomes as a promising platform for ultrasound imaging and therapeutic delivery.

Biomechanical Evaluation of Mitral Valve Repair: Virtual Chordal Transposition to Restore Anterior Leaflet Prolapse.

Background: Surgical management of an anterior leaflet prolapse remains comparatively challenging and has led to the lack of any firmly established standard repair techniques, as seen in cases of posterior leaflet prolapse. Chordal transposition repair is widely acknowledged as a remarkably durable technique that utilizes the patient's native chordae. This study aims to evaluate and predict the biomechanical and functional characteristics of a normal mitral valve (MV) model and a pathological MV model featuring anterior ruptured mitral chordae tendineae (RMCT), and to assess the effectiveness of the chordal transposition repair in the pathological MV model. Methods: There are four stages in the proposed virtual MV repair evaluation protocol: (1) modeling the virtual pathological MV model with an anterior (A2) RMCT; (2) performing chordal transposition as the virtual MV repair procedure; (3) dynamic finite element simulation of the normal (control) MV model, the pre-repair (pathological) MV model, and the post-repair (chorda transposition) MV model; (4) assessment and comparison of the physiological and biomechanical features among the normal, pre-repair, and post-repair cases. Results: The pathological MV model with anterior RMCT clearly demonstrated a substantial flail, a marked increase in chordal stresses on the two intact chordae adjacent to the ruptured A2 chordae, and severe anterior leaflet prolapse due to the A2 chordal rupture. The virtual chordal transposition demonstrated remarkable efficacy in mitigating the stress concentrations in the leaflet and chordae, restoring leaflet coaptation, and resolving anterior leaflet prolapse. Conclusions: This virtual MV surgery strategy offers a valuable means to predict, evaluate, and quantify functional and biomechanical improvements before and after MV repair, thereby empowering informed decision-making in the planning of chordal transposition interventions.

Intravenous infusion of xenon-containing liposomes generates rapid antidepressant-like effects.

AIM: Similar to ketamine, xenon gas acts as a glutamatergic N-methyl-d-aspartate receptor antagonist, but devoid of propensity to cause untoward effects. Herein, we loaded xenon gas into a liposomal carrier called xenon-containing liposomes (Xe-liposome) for systemic delivery, and investigated its effect as an antidepressant and also analyzed synaptic biomarkers including brain-derived neurotrophic factor (BDNF), protein kinase B (AKT), mammalian target of rapamycin (mTOR), protein kinase C (PKC) and extracellular signal-regulated kinase-1/2 (ERK1/2) in blood and brain. METHODS: Xe-liposomes (15 µl/mg) were prepared by a pressurized freeze-thaw method, and injected via the lateral tail vein (0.6 mL/rat) in male Wistar rats. The uncaging of xenon gas from circulating Xe-liposome was facilitated by continuous ultrasound application externally on the neck over the internal common carotid artery. One-hour after Xe-liposome infusion, animals were assessed for depression-like behaviors using a forced swimming test (FST), and spontaneous locomotor activity. Blood, as well as frontal cortex and hippocampal samples were obtained for immunoblotting and/or enzyme-linked immune sorbent assays. RESULTS: Acute intravenous infusion of Xe-liposome, at 6 mg/kg, showed an increase in swimming time in the FST (p < 0.006), indicating antidepressant-like phenotypes. Higher doses of Xe-liposomes (9 mg/kg) failed to improve swimming duration. This behavioral discrepancy was not associated with locomotion aberrations, as gross activity of rats remained similar for both doses. In biochemical analyses of frontal cortex, protein levels of BDNF increased by 64%, and enhanced phosphorylation of AKT (43%) and mTOR (93%) was observed at the 6 mg/kg dose level of Xe-liposomes, while these biomarkers and phosphorylated PKC and ERK1/2 levels remained unchanged at the higher dose. Moreover, Xe-liposomal treatment did not change the plasma and protein levels of BDNF, and phosphorylated AKT, mTOR, PKC and ERK1/2 hippocampal expressions. CONCLUSION: Xe-liposomes mediate a rapid antidepressant-like effect through activation of AKT/mTOR/BDNF signaling pathway.

Visualization of plaque distribution in a curved artery: three-dimensional intravascular ultrasound imaging.

Intravascular ultrasound (IVUS) imaging provides an excellent tool for evaluation of the type, morphology, extent, and severity of an atheromatous plaque. 3 D IVUS imaging offers additive information pertaining to morphology of the arterial structures and volumetric plaque distributions. A new 3 D IVUS visualization technique was developed to provide 3 D structural information of a curved artery. A virtual 3 D curved arterial phantom consisting of varying cross-sectional shapes, wall thicknesses, and acoustic intensity information was utilized to validate the nonlinear interpolation technique to create intermediary 2 D IVUS images. IVUS imaging was performed for the iliofemoral arterial segment of an atherosclerotic Yucatan miniswine model. These in-vivo IVUS data were utilized for intermediary IVUS image generation and volumetric 3 D IVUS visualization. Smooth transitional changes of cross-sectional shape, wall thickness and grayscale intensity were found between the intermediary images and the original arterial phantom slices. The 3 D IVUS imaging of the unfolded curved iliofemoral artery provided realistic 3 D luminal surface images of the arteries with physiologic grayscale intensity information. This unique 3 D IVUS imaging technique may help with assessment of 3 D plaque distribution across the curved arterial structure, and improve 3 D visualization of atheromatous components.

Gas chromatography/mass spectrometry measurement of xenon in gas-loaded liposomes for neuroprotective applications.

RATIONALE: We have produced a liposomal formulation of xenon (Xe-ELIP) as a neuroprotectant for inhibition of brain damage in stroke patients. This mandates development of a reliable assay to measure the amount of dissolved xenon released from Xe-ELIP in water and blood samples. METHODS: Gas chromatography/mass spectrometry (GC/MS) was used to quantify xenon gas released into the headspace of vials containing Xe-ELIP samples in water or blood. In order to determine blood concentration of xenon in vivo after Xe-ELIP administration, 6 mg of Xe-ELIP lipid was infused intravenously into rats. Blood samples were drawn directly from a catheterized right carotid artery. After introduction of the samples, each vial was allowed to equilibrate to 37°C in a water bath, followed by 20 minutes of sonication prior to headspace sampling. Xenon concentrations were calculated from a gas dose-response curve and normalized using the published xenon water-gas solubility coefficient. RESULTS: The mean corrected percent of xenon from Xe-ELIP released into water was 3.87 ± 0.56% (SD, n = 8), corresponding to 19.3 ± 2.8 µL/mg lipid, which is consistent with previous independent Xe-ELIP measurements. The corresponding xenon content of Xe-ELIP in rat blood was 23.38 ± 7.36 µL/mg lipid (n = 8). Mean rat blood xenon concentration after intravenous administration of Xe-ELIP was 14 ± 10 µM, which is approximately 15% of the estimated neuroprotective level. CONCLUSIONS: Using this approach, we have established a reproducible method for measuring dissolved xenon in fluids. These measurements have established that neuroprotective effects can be elicited by less than 20% of the calculated neuroprotective xenon blood concentration. More work will have to be done to establish the protective xenon pharmacokinetic range. Copyright © 2016 John Wiley & Sons, Ltd.

Computational virtual evaluation of the effect of annuloplasty ring shape.

Mitral regurgitation (MR) is a result of mitral valve (MV) pathology. Its etiology can be categorized as degenerative or functional MR. Ring annuloplasty aims to reconfigure a dilated mitral annulus to its normal size and shape. We investigated the effect of annuloplasty ring shape on MR outcome using our established 3-dimensional (3-D) echocardiography-based computational MV evaluation protocols. Virtual patient MV models were created from 3-D transesophageal echocardiographic data in patients with MR because of mitral annular dilation. Two distinct annuloplasty rings (Physio II and GeoForm) were designed and virtually implanted to the patient MVs. Dynamic finite element simulations of MV function were performed for each MV after virtual ring annuloplasty of either ring, and physiologic and biomechanical characteristics of MV function were compared. Excessive stress values appeared primarily in the midanterior and midposterior regions, and lack of leaflet coaptation was found in pre-annuloplasty patient MVs. Both rings demonstrated marked reduction of stresses and efficient leaflet coaptation. The Physio II ring demonstrated more evenly distributed stress reduction across the leaflets and annulus compared with the GeoForm ring. Conversely, the highly nonplanar curvature of the GeoForm ring more effectively increased leaflet coaptation compared with the Physio II ring. This indicates that the shape of annuloplasty ring affects post-annuloplasty physiologic and biomechanical conditions, which can lead to tissue alteration over a longer period after ring annuloplasty. This virtual ring annuloplasty simulation strategy provides detailed physiologic and biomechanical information and may help better plan the optimal ring selection and improved patient-specific MV repairs.

In vitro thrombolytic efficacy of echogenic liposomes loaded with tissue plasminogen activator and octafluoropropane gas.

Echogenic liposomes loaded with the thrombolytic recombinant tissue-type plasminogen activator (rt-PA) are under development for the treatment of ischemic stroke. These agents are designed to co-encapsulate cavitation nuclei to promote bubble activity in response to ultrasound exposure, and to enable localized delivery of thrombolytic. Stable cavitation improves the efficacy of the thrombolytic through enhanced fluid mixing. Echogenic liposomes that encapsulate air-filled microbubbles nucleate scant stable cavitation activity in response to 120 kHz intermittent ultrasound exposure, and have demonstrated thrombolytic efficacy equivalent to rt-PA alone. It was hypothesized that encapsulating octafluoropropane (OFP) gas within rt-PA-loaded liposomes instead of air will enhance ultrasound-mediated stable cavitation activity and increase thrombolytic efficacy compared to previous studies. The thrombolytic efficacy and cavitation activity nucleated from liposomes that encapsulate OFP microbubbles and rt-PA (OFP t-ELIP) was evaluated in vitro. Human whole blood clots were exposed to human fresh-frozen plasma alone, rt-PA (0, 0.32, 1.58, and 3.15 µg ml-1), or OFP t-ELIP at equivalent enzymatic activity, with and without exposure to intermittent ultrasound. Further, numerical simulations were performed to gain insight into the mechanisms of cavitation nucleation. Sustained ultraharmonic activity was nucleated from OFP t-ELIP when exposed to ultrasound. Furthermore, the thrombolytic efficacy was enhanced compared to rt-PA alone at concentrations of 1.58 µg ml-1 and 3.15 µg ml-1 (p < 0.05). These results indicate that OFP t-ELIP can nucleate sustained stable cavitation activity and enhance the efficacy of thrombolysis.

Loss of gas from echogenic liposomes exposed to pulsed ultrasound.

The destruction of echogenic liposomes (ELIP) in response to pulsed ultrasound excitations has been studied acoustically previously. However, the mechanism underlying the loss of echogenicity due to cavitation nucleated by ELIP has not been fully clarified. In this study, an ultra-high speed imaging approach was employed to observe the destruction phenomena of single ELIP exposed to ultrasound bursts at a center frequency of 6 MHz. We observed a rapid size reduction during the ultrasound excitation in 139 out of 397 (35%) ultra- high-speed recordings. The shell dilation rate, which is defined as the microbubble wall velocity divided by the instantaneous radius, [Formula: see text] /R, was extracted from the radius versus time response of each ELIP, and was found to be correlated with the deflation. Fragmentation and surface mode vibrations were also observed and are shown to depend on the applied acoustic pressure and initial radius. Results from this study can be utilized to optimize the theranostic application of ELIP, e.g. by tuning the size distribution or the excitation frequency.

Ultrasound-enhanced bevacizumab release from echogenic liposomes for inhibition of atheroma progression.

CONTEXT: Bevacizumab (BEV) is a monoclonal antibody to vascular endothelial growth factor (VEGF) that ameliorates atheroma progression by inhibiting neovascularization. OBJECTIVE: We aimed to determine whether BEV release from echogenic liposomes (BEV-ELIP) could be enhanced by color Doppler ultrasound (US) and whether the released BEV inhibits VEGF expression by endothelial cells in vitro. MATERIALS AND METHODS: BEV-ELIP samples were subjected to 6 MHz color Doppler ultrasound (MI = 0.4) for 5 min. We assessed release of BEV with a direct ELISA and with fluoresceinated BEV (FITC-BEV) loaded into ELIP by the same method. Human umbilical vein endothelial cell (HUVEC) cultures were stimulated to express VEGF by 10 nM phorbol-12-myristate 13-acetate (PMA). Cell-associated VEGF levels were determined using a cell-based ELISA. RESULTS: Overall, US caused an additional 100 µg of BEV to be released or exposed per BEV-ELIP aliquot within 60 min BEV-ELIP treated with US inhibited VEGF expression by 90% relative to non-treated controls and by 70% relative to BEV-ELIP without US. Also, US-treated BEV-ELIP inhibited HUVEC proliferation by 64% relative to untreated controls and by 45% relative to BEV-ELIP without US. DISCUSSION AND CONCLUSION: We have demonstrated that BEV-ELIP retains its VEGF-binding activity in a liposomal formulation and that clinical Doppler US can significantly increase that activity, both by releasing free BEV and by enhancing the surface exposure of the immunoreactive antibody.

Personalized Computational Modeling of Mitral Valve Prolapse: Virtual Leaflet Resection.

Posterior leaflet prolapse following chordal elongation or rupture is one of the primary valvular diseases in patients with degenerative mitral valves (MVs). Quadrangular resection followed by ring annuloplasty is a reliable and reproducible surgical repair technique for treatment of posterior leaflet prolapse. Virtual MV repair simulation of leaflet resection in association with patient-specific 3D echocardiographic data can provide quantitative biomechanical and physiologic characteristics of pre- and post-resection MV function. We have developed a solid personalized computational simulation protocol to perform virtual MV repair using standard clinical guidelines of posterior leaflet resection with annuloplasty ring implantation. A virtual MV model was created using 3D echocardiographic data of a patient with posterior chordal rupture and severe mitral regurgitation. A quadrangle-shaped leaflet portion in the prolapsed posterior leaflet was removed, and virtual plication and suturing were performed. An annuloplasty ring of proper size was reconstructed and virtual ring annuloplasty was performed by superimposing the ring and the mitral annulus. Following the quadrangular resection and ring annuloplasty simulations, patient-specific annular motion and physiologic transvalvular pressure gradient were implemented and dynamic finite element simulation of MV function was performed. The pre-resection MV demonstrated a substantial lack of leaflet coaptation which directly correlated with the severe mitral regurgitation. Excessive stress concentration was found along the free marginal edge of the posterior leaflet involving the chordal rupture. Following the virtual resection and ring annuloplasty, the severity of the posterior leaflet prolapse markedly decreased. Excessive stress concentration disappeared over both anterior and posterior leaflets, and complete leaflet coaptation was effectively restored. This novel personalized virtual MV repair strategy has great potential to help with preoperative selection of the patient-specific optimal MV repair techniques, allow innovative surgical planning to expect improved efficacy of MV repair with more predictable outcomes, and ultimately provide more effective medical care for the patient.

Three-dimensional echocardiography-based prediction of posterior leaflet resection.

Clinical long-term outcomes have shown that partial leaflet resection followed by ring annuloplasty is a reliable and reproducible surgical repair technique for treatment of mitral valve (MV) leaflet prolapse. We report a 61-year-old male for three-dimensional transesophageal echocardiography (3DTEE)-based virtual posterior leaflet resection and ring annuloplasty. Severe mitral regurgitation was found and computational evaluation demonstrated substantial leaflet malcoaptation and high stress concentration. Following virtual resection and ring annuloplasty, posterior leaflet prolapse markedly decreased, sufficient leaflet coaptation was restored, and high stress concentration disappeared. Virtual MV repair strategies using 3DTEE have the potential to help optimize MV repair.

A novel liposomal nanomedicine for nitric oxide delivery and breast cancer treatment.

Breast cancer is the most common type of cancer occurring among women in the United States. Nitric oxide (NO) is endogenous signaling molecules that regulate biological processes. NO has the potential to induce either cancer progression or cancer cell apoptosis depending on intra-tumoral NO concentration. High levels of NO have a cytotoxic effect on cancer cells. A novel cytotoxic gas delivery system has been developed using NO-loaded echogenic liposomes (ELIP) for breast cancer treatment. Empty ELIP and NO-ELIP were prepared using the previously developed freezing-under-pressure method with modified lipid composition. Echogenicity of NO-ELIP was measured to determine the stability of NO-ELIP. Two types of breast cancer cell (BCC) lines, MDA-MB-231 and MDA-MB-468, were utilized. MTT assay was performed after NO-ELIP treatment to determine BCC viability. Echogenicity data demonstrated improved stability of NO-ELIP with the use of BSA for resuspension of NO-ELIP. Cell death induced by NO-ELIP was not from lipid cytotoxicity but from NO. The cytotoxic effect of NO-ELIP on BCC was highly dependent on NO-ELIP concentration. NO-ELIP in concentration of 1.0-2.0 mg/ml induced dramatically decreased BCC viability. This novel cytotoxic gas delivery nanomedicine using liposomal carriers, NO-ELIP, has the potential to provide improved therapeutic effect for breast cancer treatment.

Lipid complex of apolipoprotein A-I mimetic peptide 4F is a novel platform for paraoxonase-1 binding and enhancing its activity and stability.

High density lipoprotein (HDL) associated paraoxonase-1 (PON1) is crucial for the anti-oxidant, anti-inflammatory, and anti-atherogenic properties of HDL. Discoidal apolipoprotein (apo)A-I:1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) complex has been shown to be the most effective in binding PON1, stabilizing it, and enhancing its lactonase and inhibitory activity of low density lipoprotein oxidation. Based on our earlier study demonstrating that apoA-I mimetic peptide 4F forms discoidal complex with 1,2-dimyristoyl-sn-glycero-3-phosphocholine, we hypothesized that lipid complexes of 4F would be able to bind PON1 and enhance its activity and stability. To test our hypothesis, we have expressed and purified a recombinant PON1 (rPON1) and studied its interaction with 4F:POPC complex. Our studies show significant increase, compared to the control, in the paraoxonase activity and stability of rPON1 in the presence of 4F:POPC complex. We propose that 4F:POPC complex is a novel platform for PON1 binding, increasing its stability, and enhancing its enzyme activity. We propose a structural model for the 4F:POPC:PON1 ternary complex that is consistent with our results and published observations.

Thrombolytic efficacy of tissue plasminogen activator-loaded echogenic liposomes in a rabbit thrombus model.

INTRODUCTION: Ultrasound (US)-enhanced thrombolytic treatment protocols currently in clinical trials for stroke applications involve systemic administration of tissue plasminogen activator (tPA; Alteplase), which carries a risk of adverse bleeding events. The present study aimed to compare the thrombolytic efficacy of a tPA-loaded echogenic liposome (ELIP) formulation with insonification protocols causing rapid fragmentation or acoustically-driven diffusion. MATERIALS AND METHODS: Thrombi were induced in the abdominal aortas of male New Zealand white rabbits (2-3kg) using thrombin and a sclerosing agent (sodium ricinoleate) after aortic denudation with a balloon catheter. Thrombolytic and cavitation nucleation agents (200µg of tPA alone, tPA mixed with 50µg of a microbubble contrast agent, or tPA-loaded ELIP) were bolus- injected proximal to the clot through a catheter introduced into the abdominal aorta from the carotid artery. Clots were exposed to transabdominal color Doppler US (6MHz) for 30 minutes at a low mechanical index (MI=0.2) to induce sustained bubble activity (acoustically-driven diffusion), or for 2 minutes at an MI of 0.4 to cause ELIP fragmentation. Degree of recanalization was determined by Doppler flow measurements distal to the clots. RESULTS: All treatments showed thrombolysis, but tPA-loaded ELIP was the most efficacious regimen. Both US treatment strategies enhanced thrombolytic activity over control conditions. CONCLUSIONS: The thrombolytic efficacy of tPA-loaded ELIP is comparable to other clinically described effective treatment protocols, while offering the advantages of US monitoring and enhanced thrombolysis from a site-specific delivery agent.

Structural insight into the role of the human melanocortin 3 receptor cysteine residues on receptor function.

Melanocortin-3 receptor (MC3R), expressed in the hypothalamus and limbic systems of the brain, as well as by peripheral sites, plays an important role in the regulation of energy homeostasis and other physiological functions. Past work shows that MC3R-deficiency resulted in fat mass increase, feeding efficiency increase, hyperleptinemia and mild hyperinsulinemia in mice and human. MC3R belongs to G-protein coupled receptor (GPCR) family and many studies indicate that some cysteine residues in GPCR play key roles in maintaining receptor tertiary structure and function. In this study, we examined the role of cysteine residues in MC3R on receptor function. Human MC3R (hMC3R) has eighteen cysteine residues where they are located in the extracellular loops (ELs), the transmembrane domains (TMs) and the intracellular loops (ILs). We replaced these cysteines with serine and expressed these receptors in HEK-293 cells which lack endogenous MC3R. Our results indicate that five cysteines in eighteen of the hMC3R are important for hMC3R function. Mutations, C305S, C311S, and C313S in EL3, resulted in significant decrease in receptor expression and receptor function while two other mutations C115S and C162S in TM3 significantly decreased NDP-MSH binding affinity and potency. These results suggest that extracellular cysteine residue 305, 311 and 313 are crucial for receptor expression and the transmembrane cysteine residue, C115 and 162 are important for ligand binding and signaling. These findings provide important insights into the importance of cysteine residues of hMC3R on receptor tertiary structure and function.

In vivo therapeutic gas delivery for neuroprotection with echogenic liposomes.

BACKGROUND: Ischemia-related neurological injury is a primary cause of stroke disability. Studies have demonstrated that xenon (Xe) may have potential as an effective and nontoxic neuroprotectant. Xe delivery is, however, hampered by lack of suitable administration methods. We have developed a pressurization-freeze method to encapsulate Xe into echogenic liposomes (Xe-ELIP) and have modulated local gas release with transvascular ultrasound exposure. METHODS AND RESULTS: Fifteen microliters of Xe were encapsulated into each 1 mg of liposomes (70% Xe and 30% argon). Xe delivery from Xe-ELIP into cells and consequent neuroprotective effects were evaluated with oxygen/glucose-deprived and control neuronal cells in vitro. Xe-ELIP were administered into Sprague-Dawley rats intravenously or intra-arterially after right middle cerebral artery occlusion. One-megahertz low-amplitude (0.18 MPa) continuous wave ultrasound directed onto the internal carotid artery triggered Xe release from circulating Xe-ELIP. Effects of Xe delivery on ischemia-induced neurological injury and disability were evaluated. Xe-ELIP delivery to oxygen/glucose-deprived neuronal cells improved cell viability in vitro and resulted in a 48% infarct volume decrease in vivo. Intravenous Xe-ELIP administration in combination with the ultrasound directed onto the carotid artery enhanced local Xe release from circulating Xe-ELIP and demonstrated 75% infarct volume reduction. This was comparable to the effect after intra-arterial administration. Behavioral tests on limb placement and grid and beam walking correlated with infarct reduction. CONCLUSIONS: This novel methodology may provide a noninvasive strategy for ultrasound-enhanced local therapeutic gas delivery for cerebral ischemia-related injury while minimizing systemic side effects.

Anomalous right coronary artery origin in a high school athlete.

Coronary anomalies are the cause of 12% of sudden deaths among athletes. Similarly anomalous coronary origin from the opposite sinus is often found at autopsy. The use of echocardiography to screen for these types of defects may provide a potentially life-saving diagnosis. The authors present a case that highlights the utility of echocardiography as part of a comprehensive screening program for athletes.

Delivery of stem cells to porcine arterial wall with echogenic liposomes conjugated to antibodies against CD34 and intercellular adhesion molecule-1.

In atherosclerosis, the loss of vascular stem cells via apoptosis impairs the capacity of the vascular wall to repair or regenerate the tissue damaged by atherogenic factors. Recruitment of exogenous stem cells to the plaque tissue may repopulate vascular cells and help repair the arterial tissue. Ultrasound-enhanced liposomal targeting may provide a feasible method for stem cell delivery into atheroma. Bifunctional echogenic immunoliposomes (BF-ELIP) were generated by covalently coupling two antibodies to liposomes; the first one specific for CD34 antigens on the surface of stem cells and the second directed against the intercellular adhesion molecule-1 (ICAM-1) antigens on the inflammatory endothelium covering atheroma. CD34+ stem cells from adult bone marrow were incubated on the ICAM-1-expressing endothelium of the aorta of swine fed high cholesterol diets, which was preloaded with BF-ELIP. Significantly increased stem cell adherence and penetration were detected in particular in the aortic segments treated with 1 MHz low-amplitude continuous wave ultrasound. Fluorescence and scanning electron microscopy confirmed the presence of BF-ELIP-bound CD34+ cells in the intimal compartment of the atheromatous arterial wall. Ultrasound treatment increased the number of endothelial cell progenitors migrating into the intima. Thus, under ultrasound enhancement, BF-ELIP bound CD34+ stem cells selectively bind to the ICAM-1 expressing endothelium of atherosclerotic lesions.

Cardiovascular therapeutic uses of targeted ultrasound contrast agents.

The therapeutic use of ultrasound contrast agents (UCAs) is an emerging methodology with high potential for enhanced directed therapeutic gene, bioactive gas, drug, and stem cell delivery. Ultrasound-targeted microbubble destruction has already demonstrated feasibility for plasmid DNA delivery. Similarly, therapeutic ultrasound for thrombolysis treatment has been taken into the clinical setting, and the addition of UCAs for therapeutic delivery or enhanced effect through cavitation is a natural progression to this investigation. However, as with any new technique, safety needs to be first demonstrated before translation into clinical practice. This review article will focus on the development of UCAs for cardiac and vascular therapeutics as well as the limitations/concerns for the use of therapeutic ultrasound in clinical medicine in order to lay a foundation for investigators planning to enter this exciting field or for those who want to broaden their understanding.