Yikui Tian

Development of target-specific liposomes for delivering small molecule drugs after reperfused myocardial infarction.

Although reperfusion is essential in restoring circulation to ischemic myocardium, it also leads to irreversible events including reperfusion injury, decreased cardiac function and ultimately scar formation. Various cell types are involved in the multi-phase repair process including inflammatory cells, vascular cells and cardiac fibroblasts. Therapies targeting these cell types in the infarct border zone can improve cardiac function but are limited by systemic side effects. The aim of this work was to develop liposomes with surface modifications to include peptides with affinity for cell types present in the post-infarct myocardium. To identify peptides specific for the infarct/border zone, we used in vivo phage display methods and an optical imaging approach: fluorescence molecular tomography (FMT). We identified peptides specific for cardiomyocytes, endothelial cells, myofibroblasts, and c-Kit + cells present in the border zone of the remodeling infarct. These peptides were then conjugated to liposomes and in vivo specificity and pharmacokinetics were determined. As a proof of concept, cardiomyocyte specific (I-1) liposomes were used to deliver a PARP-1 (poly [ADP-ribose] polymerase 1) inhibitor: AZ7379. Using a targeted liposomal approach, we were able to increase AZ7379 availability in the infarct/border zone at 24h post-injection as compared with free AZ7379. We observed ~3-fold higher efficiency of PARP-1 inhibition when all cell types were assessed using I-1 liposomes as compared with negative control peptide liposomes (NCP). When analyzed further, I-1 liposomes had 9-fold and 1.5-fold higher efficiencies in cardiomyocytes and macrophages, respectively, as compared with NCP liposomes. In conclusion, we have developed a modular drug delivery system that can be targeted to cell types of therapeutic interest in the infarct border zone.

Adenosine 2B Receptor Activation Reduces Myocardial Reperfusion Injury by Promoting Anti-Inflammatory Macrophages Differentiation via PI3K/Akt Pathway

Background. Activation of the adenosine A2B receptor (A2BR) can reduce myocardial ischemia/reperfusion (IR) injury. However, the mechanism underlying the A2BR-mediated cardioprotection is less clear. The present study was designed to investigate the potential mechanisms of cardioprotection mediated by A2BR. Methods and Results. C57BL/6 mice underwent 40-minute ischemia and 60-minute reperfusion. ATL-801, a potent selective A2BR antagonist, could not block ischemic preconditioning induced protection. BAY 60-6583, a highly selective A2BR agonist, significantly reduced myocardial infarct size, and its protective effect could be blocked by either ATL-801 or wortmannin. BAY 60-6583 increased phosphorylated Akt (p-Akt) levels in the heart at 10 min of reperfusion, and this phosphorylation could also be blocked by ATL-801 or wortmannin. Furthermore, BAY 60-6583 significantly increased M2 macrophages and decreased M1 macrophage and neutrophils infiltration in reperfused hearts, which also could be blocked by wortmannin. Meanwhile, confocal imaging studies showed that the majority of Akt phosphorylation in the heart was colocalized to CD206+ cells in both control and BAY 60-6583 pretreated hearts. Conclusion. Our results indicated that pretreatment with BAY 60-6583 protects the heart against myocardial IR injury by its anti-inflammatory effects, probably by modulating macrophages phenotype switching via a PI3K/Akt pathway.

Acute Hyperglycemia Abolishes Ischemic Preconditioning by Inhibiting Akt Phosphorylation: Normalizing Blood Glucose before Ischemia Restores Ischemic Preconditioning

This study examined the hypothesis that acute hyperglycemia (HG) blocks ischemic preconditioning (IPC) by inhibiting Akt phosphorylation. Brief HG of approximately 400 mg/dL was induced in C57BL/6 mice via intraperitoneal injection of 20% dextrose (2 g/kg). All mice underwent 40 min LAD occlusion and 60 min reperfusion. The IPC protocol was 2 cycles of 5 min ischemia and 5 min reperfusion prior to index ischemia. Results. In control mice, infarct size (IF) was 51.7 ± 2.0 (% risk region). Preconditioning reduced IF by 50% to 25.8 ± 3.2 (P < 0.05 versus control). In HG mice, IF was significantly exacerbated to 58.1 ± 2.3. However, the effect of IPC completely disappeared in HG mice. Normalization of blood glucose with insulin 5 min before IPC recovered the cardioprotective effect. Administration of CCPA before index ischemia mimicked IPC effect. The cardioprotective effect of CCPA, not its chronotropic effect, completely disappeared in HG mice. Phosphorylation of cardiac tissue Akt before index ischemia was enhanced by IPC or CCPA but was significantly inhibited by HG in both groups. Normalization of glucose with insulin reversed the inhibition of Akt phosphorylation by HG. Conclusion. HG abolishes the cardioprotective effect of preconditioning by inhibiting Akt phosphorylation. Normalization of blood glucose with insulin suffices to recover the cardioprotective effect of preconditioning.

Repeatability and variability of myocardial perfusion imaging techniques in mice: Comparison of arterial spin labeling and first-pass contrast-enhanced MRI

Preclinical imaging of myocardial blood flow (MBF) can elucidate molecular mechanisms underlying cardiovascular disease. We compared the repeatability and variability of two methods, first‐pass MRI and arterial spin labeling (ASL), for imaging MBF in mice.

Atorvastatin at Reperfusion Reduces Myocardial Infarct Size in Mice by Activating eNOS in Bone Marrow-Derived Cells

Background The current study was designed to test our hypothesis that atorvastatin could reduce infarct size in intact mice by activating eNOS, specifically the eNOS in bone marrow-derived cells. C57BL/6J mice (B6) and congenic eNOS knockout (KO) mice underwent 45 min LAD occlusion and 60 min reperfusion. Chimeric mice, created by bone marrow transplantation between B6 and eNOS KO mice, underwent 40 min LAD occlusion and 60 min reperfusion. Mice were treated either with vehicle or atorvastatin in 5% ethanol at a dose of 10 mg/kg IV 5 min before initiating reperfusion. Infarct size was evaluated by TTC and Phthalo blue staining. Results Atorvastatin treatment reduced infarct size in B6 mice by 19% (p<0.05). In eNOS KO vehicle-control mice, infarct size was comparable to that of B6 vehicle-control mice (p = NS). Atorvastatin treatment had no effect on infarct size in eNOS KO mice (p = NS). In chimeras, atorvastatin significantly reduced infarct size in B6/B6 (donor/recipient) mice and B6/KO mice (p<0.05), but not in KO/KO mice or KO/B6 mice (p = NS). Conclusions The results demonstrate that acute administration of atorvastatin significantly reduces myocardial ischemia/reperfusion injury in an eNOS-dependent manner, probably through the post-transcriptional activation of eNOS in bone marrow-derived cells.

Pharmacologic immunomodulation via adenosine 2a receptor stimulation improves LV remodeling and systolic strain in regions adjacent to the infarct as assessed by cardiac MRI

Background Agonists of the Adenosine 2a Receptor (A2aR) are used clinically for perfusion imaging, but they are also potent immunomodulators that reduce infarct size in animal models when administered at reperfusion. The long term effects of sustained A2aR agonist administration initiated after myocardial infarction (MI) are largely unexplored. We hypothesized that sustained administration of an A2aR agonist would inhibit post-infarct left ventricular (LV) remodeling and improve contractile strain in the LV.

Improved Ultrasound-Mediated Molecular Imaging of Previously Ischemic Mouse Myocardium Using Dual-Targeted Microbubbles and Constant Infusion

Molecularly targeted ultrasound contrast agents have been shown to identify previously-ischemic regions of the myocardium by targeting cell adhesion molecules that are displayed on activated endothelial surfaces in response to ischemia/reperfusion injury. An improved method for targeted microbubble (MB) delivery using constant infusion and dual targeted microbubbles is presented to enable better characterization of microbubble interaction with activated endothelium. Ten male C57BL/6 mice underwent 15-minute Left Anterior Descending (LAD) coronary occlusion followed by 2-hour reperfusion. Immediately after the 2-hour reperfusion, mouse hearts were imaged with a Sequoia scanner and 15L8 transducer. All mice received three of five MB preparations in random order: Sialyl Lewis X MB (MB_X), anti-P-Selectin (MB_P), anti-VCAM MB (MB_V), Sialyl Lewis X + anti-VCAM MB (MB_D), and isotype control MB (MB_I). Subsequently, mouse hearts were excised and stained with TTC & Phthalo blue dyes to assess infarct size and area at risk, respectively. Video data indicated that the binding of targeted MB to ischemic regions was significantly higher for MB_D, MB_X, MB_P, and MB_V, compared to MB_I (p<;0.05). Furthermore, normalized intensity for MB_D was higher than both MB_X and MB_V (p<;0.05). Using Phthalo blue dye as the gold standard, receiver operating curve analysis demonstrated accurate detection of both ischemic and non-ischemic regions using MB_D (AUC = 0.90), MB_X (AUC = 0.89), MB_P (AUC = 0.87), and MB_V (AUC = 0.97).