Shizu Oyamada

Chymase Inhibition Reduces Infarction and Matrix Metalloproteinase-9 Activation and Attenuates Inflammation and Fibrosis after Acute Myocardial Ischemia/Reperfusion

Chymase is activated after acute myocardial ischemia/reperfusion (AMI-R) and is associated with an early activation of matrix metalloproteinase-9 (MMP-9), which increases infarct size after experimental AMI, and late fibrosis. We assessed the effect of chymase inhibition on myocardial protection and early signs of fibrosis after AMI-R. Fourteen pigs underwent AMI-R and received intravenously either vehicle (V; n = 7) or chymase inhibitor (CM; n = 7). Separately, rat myocardial fibroblast was incubated with vehicle (n = 4), low-dose chymase (n = 4), high-dose chymase (n = 4), or high-dose chymase plus chymase inhibitor (n = 4). Infarct size (V, 41 ± 5; CM, 24 ± 5; P < 0.01) and serum troponin T (P = 0.03) at the end of reperfusion were significantly reduced in CM. Chymase activity in both the area at risk (AAR) (P = 0.01) and nonischemic area (P = 0.02) was significantly lower in CM. Myocardial levels of pro, cleaved, and cleaved/pro-MMP-9 in the AAR were significantly lower in CM than V (P < 0.01, < 0.01, and = 0.02, respectively), whereas phospho-endothelial nitric-oxide synthase (eNOS) (P < 0.01) and total eNOS (P = 0.03) were significantly higher in CM. Apoptotic cells (P = 0.05), neutrophils (P < 0.05), and MMP-9-colocalizing mast cells (P < 0.05) in the AAR were significantly reduced in CM. Interleukin-18 (P < 0.05) and intercellular adhesion molecule-1 (P < 0.05) mRNA levels were significantly lower in CM. In cultured cardiac fibrosis, Ki-67-positive cells were significantly higher in the high-dose chymase groups (P < 0.03). This study demonstrates that chymase inhibition plays crucial roles in myocardial protection related to MMP-9, inflammatory markers, and the eNOS pathway. It may also attenuate fibrosis induced by activated chymase after AMI-R.

Paclitaxel/sirolimus combination coated drug-eluting stent: in vitro and in vivo drug release studies.

Paclitaxel and sirolimus are the two major drugs for the treatment of coronary arterial disease in current drug-eluting stents. The two drugs can effectively inhibit the in-stent restenosis through their independent pathways and show synergistic effect in preventing tumor tissue growth. We hypothesize that the combination of the two drugs in a drug-eluting stent (DES) can also effectively suppress the neointima growth in the stented artery. The present work was focused on the investigation of paclitaxel/sirolimus combination release profiles from a novel biodegradable polymer (poly (D, L-lactide-co-glycolide)/amorphous calcium phosphate, PLGA/ACP) coated stent both in vitro and in vivo. For the in vitro, the drug releasing profiles were characterized by measuring the drug concentration in a drug release medium (Dulbeccos phosphate buffered saline, DPBS, pH 7.4) at predetermined time points. For the in vivo, a rat aorta stenting model was employed. The results showed that both paclitaxel and sirolimus had a two-phase release profile both in vitro and in vivo, which is similar to the drug release profile of their individual coated DESs, and there is no evident of interference between two drugs. The data suggest that paclitaxel and sirolimus can be combined pharmacokinetically in a DES for the treatment of coronary arterial diseases.

Is hyperglycemia bad for the heart during acute ischemia

OBJECTIVE This study investigates the impact of diabetes on myocardium in the setting of acute ischemia-reperfusion in a porcine model. METHODS In normoglycemic (ND group) and alloxan-induced diabetic (DM group) male Yucatan pigs, the left anterior descending coronary artery territory was made ischemic and then reperfused. Hemodynamic values and myocardial function were measured. Monastryl blue and triphenyl tetrazolium chloride staining were used to assess size of the areas at risk and infarction. Glycogen content was assessed using periodic acid-Schiff staining. Cell death and survival signaling pathways were assessed by immunoblotting. RESULTS Mean arterial pressure and developed left ventricular pressure were lower in the DM group (P < .05). Whereas global left ventricular function was worse in the DM group (P < .05), regional function in the area at risk was improved on the horizontal axis (P < .05). Mean infarct size was smaller in the DM versus the ND group (19% vs 43%; P < .05), whereas the area at risk was similar in both groups (34% vs 36%; P = .7). Ischemic myocardium in the DM group displayed more prominent staining for glycogen compared with the ND group. In the area at risk, expression of cell survival proteins including phosphorylated endothelial nitric oxide synthase (0.17 ± 0.04 vs 0.04 ± 0.01; P < .05), heat shock protein 27 (0.7 ± 0.2 vs 0.3 ± 0.1; P < .05), nuclear factor-κB (0.14 ± 0.02 vs 0.03 ± 0.01; P < .05), and mammalian target of rapamycin (0.35 ± 0.05 vs 0.15 ± 0.02; P < .05) were higher in DM animals, whereas in nonischemic tissue, expression of these proteins was similar or lower in the DM group. CONCLUSIONS Although type I diabetes worsens global left ventricular function, it is protective in the ischemic area, leading to increased expression of cell survival proteins and decreased infarct size.

In vitro and in vivo degradation of poly(D, L-lactide-co-glycolide)/ amorphous calcium phosphate copolymer coated on metal stents

The purpose of this study was to optimize a novel biodegradable polymer for drug eluting stent (DES) applications. Degradation profiles of different poly(D,L-lactide-co-glycolide)/amorphous calcium phosphate (PLGA/ACP) composites coated on stents were studied both in vitro and in vivo for three months. For the in vitro study, stents were immersed into the phosphate buffered saline (37 °C, pH 7.4) with constant shaking. The polymer weight loss was measured weekly and morphological changes were analyzed. The results demonstrated that approximately 60% of polymer was degraded within the three-month period and there was no significant difference between the different PLGA/ACP composites. However, the composite of 50% PLGA (65/35) with 50% ACP showed a slightly faster degradation rate than other composites. Morphologically, all stent surfaces changed from a micro-porous before degradation to a corrugated solid micro-net-like structure at two months post degradation. Based on in vitro results, 65% PLGA (65/35) with 35% ACP) coated stents were selected and implanted into rat aortas (n = 12) for the in vivo study. Microscopic observation showed that no composite was found on any of the implanted stents at 12 weeks post implantation, which indicated the selected PLGA/ACP composite is desired for DES applications.

Temporal and Spatial Changes in Collateral Formation and Function During Chronic Myocardial Ischemia

BACKGROUND We investigated time dependence and spatial progression of cardiac function and angiogenesis signaling in a porcine model of chronic myocardial ischemia. STUDY DESIGN Yorkshire mini-swine (n = 7/group) were subjected to chronic myocardial ischemia by placing an ameroid constrictor on the left circumflex coronary artery under general anesthesia. Swine were sacrificed after either 4 or 7 weeks of ischemia. Myocardial function, angiographic evidence of angiogenesis, microvessel function, molecular signaling, and levels of apoptosis and oxidative stress were assessed. RESULTS Flow reserve was significantly increased at 7 versus 4 weeks. Myocardial function (+dP/dt) improved 1.5-fold by 7 weeks. In the ischemic territory, microvessels at 4 weeks displayed abnormal contraction responses to serotonin, which diminished at 7 weeks. Delta-like ligand 4 protein expression decreased at 7 weeks; expression of vascular endothelial growth factor (VEGF) and phospho-endothelial nitric acid synthase (eNOS) increased. The number of apoptotic cells was decreased at 7 weeks, and antiapoptotic markers heat shock protein (HSP) 27 and HSP 90 were upregulated at 7 weeks. There was an increase in proliferating endothelial cells at 7 weeks as compared with 4 weeks. In the adjacent normal ventricle, microvessels demonstrated smaller contraction responses to endothelin-1 and serotonin at 7 weeks. There was an increase in protein peroxidation in the ischemic territory at 7 weeks. CONCLUSIONS Over time, myocardial perfusion, function, and angiogenic signaling improved in the ischemic myocardium and adjacent normal territory compared with what is observed shortly after coronary occlusion.

Myocardial therapeutic angiogenesis: a review of the state of development and future obstacles.

A significant percentage of patients have coronary artery disease that is too advanced or diffuse for percutaneous or surgical intervention. Therapeutic angiogenesis is a treatment modality to induce vessel formation that is being developed for patients with advanced coronary disease not amenable to currently available interventions. A number of approaches to induce coronary collateralization are being developed. These include gene, protein, cellular and miRNA modalities, each of which have advantages and disadvantages. At this time, no modality has emerged as the single clear choice, and combination therapies may provide synergistic benefits. However, there have been a number of recent studies advancing our knowledge as to how we can refine procollateralizing treatments. In this article, we will examine some recent successes and future obstacles in the effort to bring therapeutic angiogenesis to patients.

Effect of Dimerized Thrombin Fragment TP508 on Acute Myocardial Ischemia Reperfusion Injury in Hypercholesterolemic Swine

The thrombin-related peptide TP508 is a 23-amino acid monomer that represents a portion of the receptor binding domain in the thrombin molecule. TP508 is also known to readily convert to a dimer in an aqueous environment. In this study the dimeric form of TP508 was investigated in a porcine model of acute myocardial ischemia reperfusion injury (and compared with its monomer). Twenty-four hypercholesterolemic pigs underwent 60 min of mid-left anterior descending coronary artery occlusion followed by 120 min of reperfusion and received either vehicle (n = 6), TP508 monomer (n = 6), or two different doses of dimer (n = 6). Infarct size was significantly reduced in the monomer and two dimer groups compared with vehicle. Improvement in both endothelium-dependent and -independent coronary microvascular relaxations was also observed in treated groups. In addition, the expression of 27-kDa heat shock protein, αB-crystalline, and phosphorylated B-cell lymphoma 2 (Ser70) in the ischemic area at risk were higher in treated groups than in vehicle, whereas the expression of cleaved poly-ADP ribose polymerase was lower in treated groups. Finally, there were fewer apoptotic cells in treated groups than in vehicle. This study suggests that TP508 dimer provides a myocardial-protective effect on acute ischemia reperfusion injury in hypercholesterolemic swine, similar to TP508 monomer, by up-regulating cell survival pathways or down-regulating apoptotic pathways.

TRANS-ILIAC RAT AORTA STENTING MODEL: A NOVEL HIGH THROUGHPUT PRECLINICAL STENT MODEL FOR RESTENOSIS AND THROMBOSIS

Methods: 74 metal stents (316L stainless steel 13mm, VasoTech, Inc.) coated with various polymers were pre-mounted on 1.5mm x 15mm balloon catheters and were implanted into aspirin treated Sprague-Dawley rats (475±35g) initially using direct access of abdominal aorta (Group A, n=7) then using trans-iliac approach (cut-down, Group B, n=67). The surviving rats were sacrificed at either one month or three months post implantation and the stented arteries were harvested, pressure fixed and analyzed.