Guifu Wu

Enhanced External Counterpulsation Inhibits Intimal Hyperplasia by Modifying Shear Stress–Responsive Gene Expression in Hypercholesterolemic Pigs

Background— Enhanced external counterpulsation (EECP) is a circulation assist device that may improve endothelial dysfunction by increasing shear stress. Chronic exposure of vascular endothelial cells and vascular smooth muscle cells to relatively high physiological shear stress has antiproliferative and vasoprotective effects. The present study hypothesizes that EECP inhibits intimal hyperplasia and atherogenesis by modifying shear stress–responsive gene expression. Methods and Results— Thirty-five male pigs were randomly assigned to 3 groups: high-cholesterol diet (n=11), high-cholesterol diet plus EECP (n=17), and usual diet (control; n=7). The coronary arteries and aortas were collected for histopathological study and immunohistochemical and Western blot analysis. The peak diastolic arterial wall shear stress during EECP increased significantly compared with before EECP (49.62±10.71 versus 23.92±7.28 dyne/cm2; P<0.001). Intimal hyperplasia was observed in the coronary arteries of the high-cholesterol diet group, whereas in animals receiving EECP, the intima-to-media area ratio was significantly decreased by 41.59% (21.27±10.00% versus 36.41±16.69%; P=0.008). Hypercholesterolemia attenuated the protein expression of endothelial NO synthase and enhanced the phosphorylation of extracellular signal-regulated kinases 1/2. EECP treatment alleviated these adverse changes. Conclusions— EECP reduces hypercholesterolemia-induced endothelial damage, arrests vascular smooth muscle cell proliferation and migration, decreases proliferating cell nuclear antigen proliferative index, suppresses extracellular matrix formation, and eventually inhibits intimal hyperplasia and the development of atherosclerosis by increasing the arterial wall shear stress, which in turn activates the endothelial NO synthase/NO pathway and probably suppresses extracellular signal-regulated kinases 1/2 overactivation.

PR39 Inhibits Apoptosis in Hypoxic Endothelial Cells Role of Inhibitor Apoptosis Protein-2

Background—PR39 is a proline- and arginine-rich peptide implicated in wound healing and myocardial ischemia protection. To determine the potential mechanisms of PR39 in ischemia, we examined the role of PR39 in hypoxia-induced apoptosis in vascular endothelial cells. Methods and Results—Hypoxia results in an increase of apoptosis in bovine aortic endothelial cells (BAECs), as determined by terminal deoxynucleotidyl transferase–mediated dUTP biotin nick-end labeling (TUNEL) analysis and caspase-3 activity. Hypoxia induced 66.2±2.7% TUNEL-positive cells, whereas in the presence of synthesized PR39 peptide, TUNEL-positive cells were reduced to 29.6±1.9% (P <0.05). After 24 hours of hypoxia, the addition of PR39 reduced caspase-3 activity to 3.17±0.47 pMol/min from 10.52±0.55 pMol/min in hypoxic BAECs. Moreover, PR39 increased inhibitor of apoptosis protein-2 (IAP-2) gene and protein expression by 3-fold in a time- and dose-dependent manner. The induction of IAP-2 by PR39 conferred an increase in IAP-2 gene transcription and IAP-2 mRNA stability. Furthermore, inhibiting IAP-2 with second mitochondria-derived activator of caspase (Smac) and with small interfering RNA targeting IAP-2 abrogated the ability of PR39 to reduce caspase-3 activity. Conclusions—We provide the first direct evidence for PR39 as an antiapoptotic factor in endothelial cells during hypoxia. These data suggest that PR39 inhibits hypoxia-induced apoptosis and decreases caspase-3 activity in endothelial cells through an increase of IAP-2 expression.

Exercise-induced expression of VEGF and salvation of myocardium in the early stage of myocardial infarction

The mechanism of exercise-induced benefit and angiogenesis in ischemic heart disease remains poorly defined. This study was designed to investigate the effects of exercise training on the expression of angiogenic factors and angiogenesis in the infarcted myocardium [myocarial infaction (MI)]. Sixty-three male FVB mice were used for study and were divided into subgroups to test the response to exercise: the time-dependent expression of angiogenic factors to exercise training in normal (group 1; n = 12) and infarcted myocardium (group 2; n = 15) and the exercise-induced angiogenic response in normal and infarcted myocardium (group 3; n = 20) as well as the impact of exercise preconditioning on infarcted myocardium (group 4; n = 26). Exercise training consisted of daily treadmill exercise for 1 h for 3 days. Expression of VEGF and its receptors Flt-1 and Flk-1 was upregulated by exercise training in mice with MI. Exercise-induced VEGF expression in the MI group was higher than that in the sham (control) group. Cell proliferation assessment showed a significantly higher (P < 0.05) number of bromodeoxyuridine-positive cells in post-MI mice in the exercise group as opposed to post-MI mice in the sedentary group. 2,3,5-Triphenyltetrazolium chloride staining revealed a profound difference in the size of MI (18.25 +/- 2.93%) in the exercise group versus the sedentary group (29.26 +/- 7.64%, P = 0.02). Moreover, exercise preconditioning before MI promoted VEGF expression at both mRNA and protein levels. In conclusion, activation of VEGF and its receptors occurs in the infarcted mice heart in response to exercise, which results in decreased infarct size and improved angiogenesis.

Enhanced External Counterpulsation Attenuates Atherosclerosis Progression Through Modulation of Proinflammatory Signal Pathway

Objective—Shear stress may be the most crucial local factor affecting atherogenesis. The present study investigated the effect of exposure to increased shear stress promoted by enhanced external counterpulsation (EECP) on the progression of atherosclerosis and the underlying inflammation-related molecular mechanisms in a porcine model of hypercholesterolemia. Methods and Results—Hypercholesterolemic pigs were subjected to a 7-week EECP intervention while being fed a high-cholesterol diet. EECP resulted in a 34.38% increase of mean wall shear stress and a significantly lower pulsatility index in the brachial artery. The animals receiving EECP showed a marked reduction in atherosclerotic lesion size in the coronary artery and abdominal aorta compared with the hypercholesterolemic control group, associated with a decrease in macrophage accumulation. The expression of a set of genes involved in inflammation (including C-reactive protein [CRP], complement 3a, vascular cell adhesion molecule-1 [VCAM-1], and inducible nitric oxide synthase), mitogen-activated protein kinase (MAPK)-p38 phosphorylation, and nuclear factor-&kgr;B (NF-&kgr;B) activation, was attenuated. Conclusion—These findings suggested that long-term EECP exerts a retarding effect on atherosclerosis by downregulating proinflammatory gene expression. The underlying mechanisms are related to chronic exposure to increased pulsatile shear stress promoted by EECP; this exposure suppresses the overactivation of the MAPK-P38/NF-&kgr;B/VCAM-1 signaling pathway induced by hypercholesterolemia.

Autologous Cardiomyotissue Implantation Promotes Myocardial Regeneration, Decreases Infarct Size, and Improves Left Ventricular Function

Background— Cell therapy for myocardial infarction (MI) may be limited by poor cell survival and lack of transdifferentiation. We report a novel technique of implanting whole autologous myocardial tissue from preserved myocardial regions into infarcted regions. Methods and Results— Fourteen rats were used to optimize cardiomyotissue size with peritoneal wall implantation (300 &mgr;m identified as optimal size). Thirty-nine pigs were used to investigate cardiomyotissue implantation in MI induced by left anterior descending balloon occlusion (10 animals died; male-to-female transplantation for tracking with in situ hybridization for Y chromosome, n=4 [2 donors and 2 MI animals]; acute MI implantation cohort at 1 hour, n=13; and healed MI implantation at 2 weeks, n=12). Assessment included echocardiography, magnetic resonance imaging, hemodynamics, triphenyltetrazolium chloride staining, and histological and molecular analyses. Tracking studies demonstrated viable implants with donor cells interspersed in the adjacent myocardium with gap junctions and desmosomes. In the acute MI cohort, treated animals compared with controls had improved perfusion by magnetic resonance imaging (1.2±0.01 versus 0.86±0.05; P<0.01), decreased MI size (magnetic resonance imaging: left ventricle, 2.2±0.5% versus 5.4±1.5%, P=0.04; triphenyltetrazolium chloride: anterior wall, 10.3±4.6% versus 28.9±5.8%, P<0.03), and improved contractility (dP/dt, 1235±215 versus 817±817; P<0.05). In the healed MI cohort, treated animals had less decline in ejection fraction between 2 and 4 week assessment (−3±4% versus −13±−4%; P<0.05), less decline in ±dP/dt, and smaller MI (triphenyltetrazolium chloride, 21±11% versus 3±8%; P=0.006) than control animals. Infarcts in the treated animals contained more mdr-1+ cells and fewer c-kit+ cells with a trend for decreased expression of matrix metalloproteinase-2 and increased expression of tissue inhibitor of metalloproteinase-2. Conclusion— Autologous cardiomyotissue implanted in an MI area remains viable, exhibits electromechanical coupling, decreases infarct size, and improves left ventricular function.