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Silica nanoparticles (SiNPs) are among the non-toxic nanoparticles (NPs) that have magnetic capabilities. It is hypothesized that SiNPs may be able to reduce toxic effects exerted by a mixture of lead (Pb) and mercury (Hg) in African catfish Clarias gariepinus. The in vitro magnetic potential of SiNPs to absorb Pb and Hg was tested. Fish (N = 240) were divided into four groups in triplicates for 30 days. The first group served as control and the second group (SiNPs) was exposed to 1/10 of 96 h LC50 of SiNPs (14.45 mg/L). The third group (HMM) was exposed to 1/10 of 96-h LC50 of a mixture of mercury chloride (HgCl

Lumpy skin disease (LSD) is a devastating, emerging viral disease of cattle. It causes significant economic losses due to trade restrictions that are placed on infected animals and the biological effects of the disease: infertility, dramatic loss in milk production, induction of abortion and mortality. It is caused by lumpy skin disease virus (LSDV), which belongs to the Poxviridae family. Vaccination has been determined to be the most effective way to control LSD infection among livestock. However, some adverse effects have been reported in animals vaccinated with live vaccines. To the best of our knowledge, this is the first study to report the systemic lesions that are associated with LSD vaccination in xenogeneic animals. The aim of our study was to compare the immunogenicity and pathogenicity of a live attenuated vaccine of Romanian strain of sheeppox virus (SPPV) through study of two different routes of administration in xenogeneic animals (mice). Swiss male mice were inoculated with two doses of SPPV vaccine by two different routes intranasal (IN, through nebulisation), and intraperitoneal (IP) injection) and the levels of immunoglobulins and histopathological findings were reported. Our results showed marked increases in levels of immunoglobulins (Ig) dependent on the administration route: IgG in IP-inoculated mice and IgA in IN-vaccinated mice. IgM levels became markedly high after vaccination via both routes. Histologically, nebulisation of mice with SPPV vaccine caused more pulmonary lesions than did IP injection and promoted the proliferation of megakaryocytes in splenic tissues. In contrast, IP injection had less effect on pulmonary tissues and induced activation of extramedullary haematopoiesis (EH) in the hepatic tissues. LSD vaccination in xenogeneic animals caused serious systemic complications and the severity of the lesions caused to tissue depended on the route of administration.

To date, the lowest protective SGLT2 inhibitor dose is unknown. We initially performed a dose-response pilot study in normal rats. Based on the results of this pilot study we compared the cardio-renal effects of the SGLT-2 inhibitor empagliflozin, with placebo or telmisartan in rats with 5/6 nephrectomy (5/6 Nx) on a high salt diet (HSD). The experimental set up was as follows: Sham operation (Sham) with normal diet and placebo; 5/6 Nx with 2% HSD and placebo; 5/6 Nx with HSD and empagliflozin (0.6 mg/kg/day, bid); 5/6 Nx with HSD and telmisartan (5 mg/kg/day, qd). Empagliflozin treatment increased urinary glucose excretion, in parallel to empagliflozin plasma levels, in a dose-dependent manner starting at doses of 1 mg/kg in the pilot study. 5/6Nx rats on HSD treated with this low empagliflozin dose showed significantly reduced cardiac (−34.85%; P

Parkinson’s disease (PD) is a neurodegenerative ailment that worsens over time. PD is characterized by severe motor and non-motor symptoms. Because there is no cure for Parkinson's disease, current treatments heavily rely on relieving symptoms. The current research explores the histopathological changes in the different brain regions including the cerebral cortex, hippocampus, and striatum after subcutaneous rotenone administration every other day for 21 days in adult male Wistar rats. Substantial morphological changes were noticed in cell area. Sections examined showed severe neuronal loss and excessive appearance of abnormal damaged, shrunken red neurons, intercellular edema and vacuolation of intercellular neuropile and loss of myelinated fibers. After the last rotenone injection, chrysin and L-dopa were given for 28 days. The findings showed that chrysin treatment displayed significant improvement in the neuronal architecture and decrease of damage caused by rotenone in different brain regions. This appeared in the form of moderate protective efficacy with the persistence of loss in normal neuronal densities. However, combined treatment of rotenone-injected animals with chrysin and L-dopa was associated with more records of visible intact neurons and fewer records of abnormal degenerative changes in various brain neuronal areas This study concludes that chrysin may be a useful drug alone or with L-dopa for the treatment of rotenone-induced PD.

Cardiac c-Kit+ cells have a modest cardiogenic potential that could limit their efficacy in heart disease treatment. The present study was designed to augment the cardiogenic potential of cardiac c-Kit+ cells through class I histone deacetylase (HDAC) inhibition and evaluate their therapeutic potency in the chronic heart failure (CHF) animal model. Myocardial infarction (MI) was created by coronary artery occlusion in rats. c-Kit+ cells were treated with mocetinostat (MOCE), a specific class I HDAC inhibitor. At 3 weeks after MI, CHF animals were retrogradely infused with untreated (control) or MOCE-treated c-Kit+ cells (MOCE/c-Kit+ cells) and evaluated at 3 weeks after cell infusion. We found that class I HDAC inhibition in c-Kit+ cells elevated the level of acetylated histone H3 (AcH3) and increased AcH3 levels in the promoter regions of pluripotent and cardiac-specific genes. Epigenetic changes were accompanied by increased expression of cardiac-specific markers. Transplantation of CHF rats with either control or MOCE/c-Kit+ cells resulted in an improvement in cardiac function, retardation of CHF remodeling made evident by increased vascularization and scar size, and cardiomyocyte hypertrophy reduction. Compared with CHF infused with control cells, infusion of MOCE/c-Kit+ cells resulted in a further reduction in left ventricle end-diastolic pressure and total collagen and an increase in interleukin-6 expression. The low engraftment of infused cells suggests that paracrine effects might account for the beneficial effects of c-Kit+ cells in CHF. In conclusion, selective inhibition of class I HDACs induced expression of cardiac markers in c-Kit+ cells and partially augmented the efficacy of these cells for CHF repair. Significance The study has shown that selective class 1 histone deacetylase inhibition is sufficient to redirect c-Kit+ cells toward a cardiac fate. Epigenetically modified c-Kit+ cells improved contractile function and retarded remodeling of the congestive heart failure heart. This study provides new insights into the efficacy of cardiac c-Kit+ cells in the ischemic heart failure model.

Objectives Nitric oxide (NO) is a critical regulator of vascular tone and signal transduction in the cardiovascular system. NO is synthesized by three unique enzymes (nitric oxide synthases [NOS]): endothelial and neuronal NOS, both constitutively expressed, and inducible NOS (iNOS), which is induced by proinflammatory stimuli and subsequently produces a burst of NO. NO has been implicated as both an injurious and a beneficial mediator after cardiac arrest and resuscitation. A previous study in swine found that iNOS expression is absent in the myocardium prior to cardiac arrest and that it increases after 10 minutes of untreated ventricular fibrillation (VF), decreases somewhat during the early postresuscitation period, and then steadily increases up to 6 hours postresuscitation. Because this time course of iNOS expression mirrors that of postresuscitation myocardial dysfunction, this study was designed to test the hypothesis that selective inhibition of iNOS improves postresuscitation outcomes in swine. Methods Thirty-two domestic swine of either sex were randomly assigned to receive one of the following treatments 15 minutes after return of spontaneous circulation (ROSC): 1) NG-nitro-l-arginine methyl ester (l-NAME), a global NO inhibitor; 2) aminoguanidine (AG), a selective iNOS inhibitor; or 3) saline as control. After 10 minutes of untreated VF, swine received a standard resuscitation protocol. Twenty-four-hour survival, neurological status, left ventricular (LV) function, and hemodynamic measurements were obtained. Results Return of spontaneous circulation occurred in 28 of 32 animals (88%). Only successfully resuscitated animals were assigned to treatment groups and completed the study. There were no differences in survival or neurological outcomes between groups. There were also no differences in LV function or hemodynamic variables found between the control group and the AG group. Global inhibition of NOS with l-NAME post-ROSC increased aortic pressure and transiently decreased pulse pressure. Treatment with l-NAME also increased LV end diastolic pressure and decreased cardiac output within 30 minutes post-ROSC, which was sustained throughout the 4-hour measurements, compared to both the control and the AG groups. In addition, LV ejection fraction recovered to baseline measurements in both the control and AG groups, but failed to recover in the l-NAME group. Conclusions Global inhibition of NOS after cardiac arrest and resuscitation markedly worsens hemodynamic variables. Selective inhibition of iNOS after cardiac arrest and resuscitation does not prevent postresuscitation myocardial stunning. There were no significant differences in neurological outcome or survival between treatment groups.

Cardiovascular (CV) diseases are known to have a negative impact on the brain and neurocognition, and contribute to the development of vascular dementia and neurodegenerative diseases such as Alzheimer's disease (AD). Among CV diseases, congestive heart failure (CHF) after myocardial infarction (MI) is a condition where the ability of the left ventricle to eject blood to the circulation is impaired. As a consequence, CHF triggers inflammation and results in reduced cerebral blood flow which are considered among the risk factors for development of AD. However, biochemical alterations in the brain following MI and CHF remain unknown. To address this issue, we investigated microglia activation; levels of BACE1, the key rate-limiting enzyme involved in the pathogenesis of AD; and VEGF levels in the hippocampus and cortex following MI. We created MI by the ligation of the left anterior descending coronary artery in Sprague-Dawley male rats and collected brains either 3 days after MI (AMI) or 21 days after MI (CHF). We investigated microglia activation in AMI and CHF brains by immunohistochemistry and immunoblotting using macrophage/microglia marker Ionized calcium binding adaptor molecule 1 (Iba-1), and observed activated morphology of microglia in the cortex of rats in both AMI and CHF. We also showed the levels of BACE1 were increased in the cortex and hippocampus of CHF rats. To determine whether hypoxia occurs in the CHF brain, we assessed levels of VEGF in the hippocampus and cortex. Western blotting analysis showed up-regulation of VEGF in the hippocampus of CHF brains. These results suggest that neuroinflammation takes place secondary to myocardial infarction. In addition, CHF-induced hypoxia might play a role in the elevation of BACE1 and VEGF levels.

This study was designed to determine if a viable biodegradable three-dimensional fibroblast construct (3DFC) patch implanted on the left ventricle after myocardial infarction (MI) improves left ventricular (LV) function and blood flow. We ligated the left coronary artery of adult male Sprague-Dawley rats and implanted the 3DFC at the time of the infarct. Three weeks after MI, the 3DFC improved LV systolic function by increasing ( p < 0.05) ejection fraction (37 ± 3% to 62 ± 5%), increasing regional systolic displacement of the infarcted wall (0.04 ± 0.02 to 0.11 ± 0.03 cm), and shifting the passive LV diastolic pressure volume relationship toward the pressure axis. The 3FDC improved LV remodeling by decreasing ( p < 0.05) LV end-systolic and end-diastolic diameters with no change in LV systolic pressure. The 3DFC did not change LV end-diastolic pressure (LV EDP; 25 ± 2 vs. 23 ± 2 mmHg) but the addition of captopril (2mg/L drinking water) lowered ( p < 0.05) LV EDP to 12.9 ± 2.5 mmHg and shifted the pressure–volume relationship toward the pressure axis and decreased ( p < 0.05) the LV operating end-diastolic volume from 0.49 ± 0.02 to 0.34 ± 0.03 ml. The 3DFC increased myocardial blood flow to the infarcted anterior wall after MI over threefold ( p < 0.05). This biodegradable 3DFC patch improves LV function and myocardial blood flow 3 weeks after MI. This is a potentially new approach to cell-based therapy for heart failure after MI.

Angiotensin II receptor blockade (ARB) increases vasorelaxation in heart failure by enhancing endothelial nitric oxide (NO). To determine the effects of valsartan on NO-mediated peripheral vascular function after myocardial infarction (MI), we treated adult male Sprague-Dawley rats immediately after MI with valsartan for 3 weeks (sham, n = 10; MI, n = 11) and 6 weeks (sham, n = 6; MI, n = 8). At both time points, valsartan lowered (P < 0.05) left ventricular (LV) systolic pressure (103 +/- 4 and 107 +/- 4 vs. 93 +/- 3 and 85 +/- 4 mm Hg, respectively) and LV end-diastolic pressure (25 +/- 1 and 25 +/- 2 to 13 +/- 2 and 18 +/- 3 mm Hg, respectively). Valsartan lowered (P < 0.05) LV dP/dt only at 6 weeks (4676 +/- 168 and 4503 +/- 232 vs. 4539 +/- 281 and 3372 +/- 417 mm Hg/sec); valsartan shortened (P < 0.05) the time constant of LV relaxation or tau only at 3 weeks (24.2 +/- 1.8 and 26.5 +/- 2.3 vs. 20.1 +/- 0.7 and 23.8 +/- 1.4 msec). At 6 weeks, the vasorelaxation response to acetycholine in aortic rings was decreased (P < 0.05) with MI and improved at acetycholine doses (10, 10, and 10; P < 0.06) with valsartan. Endothelial nitric oxide synthase (eNOS) protein was undetectable in aortic tissue from valsartan treated rats or from aortic tissue incubated with valsartan (2.5, 25, and 50 mg/mL). These data suggest that valsartan improves cardiac function after MI by modulating LV remodeling, decreasing LV end-diastolic pressure, and enhancing both LV diastolic and endothelial function. These effects are mediated, in part, by NO but upregulation of eNOS may not be required for improved systemic endothelial function in heart failure.

It is estimated that up to 128 million individuals might benefit from regenerative medicine therapy, or almost 1 in 3 individuals in the US. If accurate, the need to relieve suffering and reduce healthcare costs is an enormous motivator to rapidly bring stem cell therapies to the clinic. Unfortunately, embryonic stem (ES) cell therapies are limited at present by ethical and political constraints and, most importantly, by significant biologic hurdles. Thus, for the foreseeable future, the march of regenerative medicine to the clinic will depend on the development of non-ES cell therapies. At present, non-ES cells easily available in large numbers can be found in the bone marrow, adipose tissue and umbilical cord blood (CB). Each of these stem cells is being used to treat a variety of diseases. This review shows that CB contains multiple populations of pluripotent stem cells, and can be considered the best alternative to ES cells. CB stem cells are capable of giving rise to hematopoietic, epithelial, endothelial and neural tissues both in vitro and in vivo. Thus, CB stem cells are amenable to treat a wide variety of diseases including cardiovascular, ophthalmic, orthopedic, neurologic and endocrine diseases.

Background— This study was designed to determine the effects of pretreatment with an angiotensin receptor blocker on left ventricular (LV) function and remodeling during acute myocardial infarction (MI). Methods and Results— Sprague-Dawley rats were pretreated with candesartan (10 mg · kg −1 · d −1 ) for 2 weeks and studied at 1, 3, and 6 minutes after MI. Compared with untreated rats, pretreatment with candesartan lowered ( P P P P =0.04) and 70% ( P =0.002), and constitutive microtubulin increased 260% ( P =0.0005) and 111% ( P =0.003). Candesartan had no effect on LV tissue endothelial nitric oxide synthase levels but attenuated the increase in constitutive microtubulin by 77% ( P =0.004) and 37% ( P Conclusions— Pretreatment with candesartan before an acute MI improves global LV function, prevents LV dilation, and blunts the increase in constitutive microtubulin, with minimal effects on LV hemodynamics, regional function, or tissue endothelial nitric oxide synthase. Thus, candesartan given before an MI attenuates LV remodeling and alters the cytoskeleton matrix of the left ventricle.

This study was designed to determine whether tissue engineering could be used to reduce ventricular remodeling in a rat model of non-transmural, non-ST-elevation myocardial infarction.We grafted an acellular 3-dimensional (3D) collagen type 1 scaffold (solid porous foam) onto infarcted myocardium in rats. Three weeks after grafting, the scaffold was integrated into the myocardium and retarded cardiac remodeling by reducing left ventricular (LV) dilation. The LV inner and outer diameters, measured at the equator at zero LV pressure, decreased (p0.05) from 11,040 +/- 212 to 9,144 +/- 135 microm, and 13,469 +/- 187 to 11,673 +/- 104 microm (N = 12), after scaffold transplantation onto infarcted myocardium. The scaffold also shifted the LV pressure-volume curve to the left toward control and induced neo-angiogenesis (700 +/- 25 vs 75 +/- 11 neo-vessels/cm2, N = 5, p0.05). These vessels (75 +/- 11%) ranged in diameter from 25 to 100 microm and connected to the native coronary vasculature. Systemic treatment with granulocyte-colony stimulating factor (G-CSF), 50 microg/kg/day for 5 days immediately after myocardial injury, increased (p0.05) neo-vascular density from 700 +/- 25 to 978 +/- 57 neo-vessels/cm2.A 3D collagen type 1 scaffold grafted onto an injured myocardium induced neo-vessel formation and reduced LV remodeling. Treatment with G-CSF further increased the number of vessels in the myocardium, possibly due to mobilization of bone marrow cells.

Summary Background For prolonged VF, perfusion of the myocardium by pre-shock chest compressions can improve myocardial readiness for successful defibrillation. Characteristics of the VF waveform correlate with the duration of VF when there is no structural heart disease. A "smart" automated external defibrillator (AED) could therefore analyze the VF waveform, determine if VF has been prolonged, and then direct rescuers to either deliver a shock first or chest compressions first. We hypothesized that ischemic heart failure might alter the waveform content of ventricular fibrillation compared with normal hearts, complicating the determination of VF duration. Methods Myocardial infarction was induced by ligating the proximal left coronary artery. Six weeks later, VF was then induced in 10 rats with myocardial infarction and heart failure (MI-CHF) and 9 control rats. Waveforms were analyzed for total signal amplitude, median frequency, dominant frequency and bandwidth (the frequency interval containing 50% of the total amplitude about the median frequency). Results All of these VF waveform characteristics were altered substantially in MI-CHF rats compared to normal controls. In particular, MI-CHF rats had decreased signal amplitude early in VF ( p =0.02), a broader bandwidth ( p =0.001) and different frequency characteristics over time ( p Conclusions VF waveforms vary over time in a typical manner among rats with and without ischemic heart failure. However, the time-course and waveform characteristics of ventricular fibrillation are altered in rats with myocardial infarctions and ischemic heart failure compared to normal controls. These findings have important implications regarding the use of waveform analyses to determine the duration of VF.

This review briefly highlights the major mechanisms of left ventricular (LV) remodeling following myocardial infarction in heart failure and presents novel strategies to limit infarct size and retard the progression of LV remodeling. This is important since interventions that have been shown to improve mortality in patients with heart failure such as angiotensin converting enzyme inhibitor and beta-blockers had a favorable impact on the LV remodeling process. Although several other animal models of heart failure are available, we chose the rat infarct model because of its accurate reflection of human pathophysiology. It is clear from this review that the understanding of the mechanisms responsible for the regression of ventricular remodeling is still unclear. Further investigations in this area will certainly lead to the development of new therapy for heart failure.

Background Interstitial fibrosis and fibrotic scar formation contribute to cardiac remodeling and loss of cardiac function in myocardial infarction (MI) and heart failure. Recent studies showed that histone deacetylase (HDAC) inhibitors retard fibrosis formation in acute MI settings. However, it is unknown whether HDAC inhibition can reverse cardiac fibrosis in ischemic heart failure. In addition, specific HDAC isoforms involved in cardiac fibrosis and myofibroblast activation are not well defined. Thus, the purpose of this study is to determine the effects of selective class I HDAC inhibition on cardiac fibroblasts activation and cardiac fibrosis in a congestive heart failure (CHF) model secondary to MI. Methods MI was created by left anterior descending (LAD) coronary artery occlusion. Class I HDACs were selectively inhibited via Mocetinostat in CD90+ fibroblasts isolated from atrial and ventricular heart tissue in vitro. In vivo, Class I HDACs were inhibited in 3 weeks post MI rats by injecting Mocetinostat for the duration of 3 weeks. Cardiac function and heart tissue were analyzed at 6 weeks post MI. Results In sham hearts, HDAC1 and HDAC2 displayed differential expression patterns where HDAC1 mainly expressed in cardiac fibroblast and HDAC2 in cardiomyocytes. On the other hand, we showed that HDAC1 and 2 were upregulated in CHF hearts, and were found to co-localize with CD90+ cardiac fibroblasts. In vivo treatment of CHF animals with Mocetinostat improved left ventricle end diastolic pressure and dp/dt max and decreased the total collagen amount. In vitro treatment of CD90+ cells with Mocetinostat reversed myofibroblast phenotype as indicated by a decrease in α-Smooth muscle actin (α-SMA), Collagen III, and Matrix metalloproteinase-2 (MMP2). Furthermore, Mocetinostat increased E-cadherin, induced β-catenin localization to the membrane, and reduced Akt/GSK3β signaling in atrial cardiac fibroblasts. In addition, Mocetinostat treatment of atrial CD90+ cells upregulated cleaved-Caspase3 and activated the p53/p21 axis. Conclusions Taken together, our results demonstrate upregulation of HDAC1 and 2 in CHF. In addition, HDAC inhibition reverses interstitial fibrosis in CHF. Possible anti-fibrotic actions of HDAC inhibition include reversal of myofibroblast activation and induction of cell cycle arrest/apoptosis.

The objective of this study was to determine how maturation and aging affects beta (beta)-adrenergic receptor (AR) control of arterial vasorelaxation. Left ventricular (LV) hemodynamics and arterial vasorelaxation in thoracic artery segments were studied in Brown Norway, Fisher 344 cross rats at 6 weeks, 6 months, and 23 months of age. We defined changes in maturation as occurring between 6 weeks and 6 months of age and changes in aging as occurring between 6 months and 23 months of age. With maturation, isoproterenol resulted in a downward shift in heart rate and an upward shift in both LV dP/dt and peripheral vascular resistance responses. Similar changes were noted with aging except for the downward shift in LV dP/dt isoproterenol response. There was a dose-dependent increase in arterial vasorelaxation in response to isoproterenol in all age groups, but the 6-week-old animals had a 5-fold (P

After myocardial infarction (MI), nitric oxide (NO)-mediated vasorelaxation is attenuated in both conduit and resistance arteries. To determine if the attenuated vasorelaxation after MI is due to downregulation of eNOS protein, pharmacological, immunoblotting, and gene transfer of eNOS were performed in rats 3 weeks after MI. Gene transfer was accomplished using a "first-generation" serotype 5, replication-deficient, adenoviral vector (1.2 x 10(9) pfus) containing eNOS cDNA in the hindlimb vasculature for 30 min. Five days after infection, overexpression of eNOS protein was confirmed by immunohistochemical staining and immunoblotting. Recombinant gene expression was localized primarily to the vascular endothelial cells. After MI, eNOS protein level decreased (3.3 +/- 0.9 vs 2.1 +/- 0.8 intensity units/microg protein, n=6, P

Angiotensin II type 1 (AT1) receptor blockade attenuates myocardial fibrosis after myocardial infarction (MI). However, whether inhibition of fibrosis by AT1 receptor blockade influences myocardial stiffness and contractility is unknown. We measured left ventricular (LV) hemodynamics, papillary muscle function, and myocardial stiffness and fibrosis in rats randomized to losartan or placebo 1 day after MI and treated subsequently for 8 wk. Losartan decreased LV and right ventricular weights as well as mean aortic and LV systolic pressures in sham and MI rats. LV end-diastolic pressure increased after MI and was decreased with losartan. Maximal developed tension and peak rate of tension rise and decline were decreased in MI vs. sham rats. Interstitial fibrosis developed after MI and was prevented in losartan-treated MI rats. The development of abnormal myocardial stiffness after MI was prevented by losartan. After MI, AT1 receptor blockade prevents an abnormal increase in myocardial collagen content. This effect was associated with a normalization of passive myocardial stiffness.

Abstract —To examine arterial mechanical changes during aging, pressure-radius and axial force–radius curves were measured in vivo in carotid arteries from 6- and 23-month-old Brown Norway X Fischer 344 rats. Incremental passive circumferential stiffness (measured at 50, 100, and 200 mm Hg) was higher ( P 2 ×10 3 , respectively). Incremental passive axial stiffness was increased ( P 2 ×10 3 , respectively). Active incremental circumferential arterial stiffness at 100 and 200 mm Hg was increased ( P P P P

This study was designed to determine if adenoviral-mediated delivery of a transgene encoding the beta 2-adrenergic receptor (beta 2-AR) to the carotid arterial wall could result in alterations in in vivo vascular function. De-endothelialized rat carotid arteries were infused in vivo with 0.1 mg/ml elastase and adenovirus [6 x 10(9) plaque forming units (PFU)] containing either the marker gene beta-galactosidase (Adeno-beta-gal), DNA encoding the human beta 2-AR (Adeno-beta 2-AR), or no transgene. This low concentration of elastase increased the water permeability (5.2 +/- 0.6 v 1.9 +/- 0.4 x 10(-8) cm/s/mmHg, n = 4, P < 0.0001) without affecting either the vasomotor responsiveness or the morphology of the arterial wall. A transfection efficiency of 73% was achieved with Adeno-beta-gal (n = 3). beta-gal expression was associated with infrequent appearance of T and B lymphocytes, or neutrophil infiltration. Five days after infection with Adeno-beta 2-AR, the total beta-AR density increased six-fold (67.8 +/- 3.4 v 397.0 +/- 155.5 fmol/mg protein, n = 5, P < 0.01); isoproterenol-induced vasorelaxation at transmural pressures from 10-110/mmHg increased (P < 0.01) compared to arteries exposed to control virus (empty adenovirus), n = 4; and isoproterenol-stimulated cAMP production was increased by 65% (n = 5). Thus, adenoviral-mediated delivery of beta 2-ARs into large artery walls results in enhanced beta-AR-mediated vasorelaxation via augmentation in cAMP levels in vascular smooth muscle cells.

— Adaptations of the aging left ventricle (LV) to hemodynamic overload are functionally and structurally distinct from those of the young organism. This study describes the influence of aging on LV hemodynamics and remodeling late after myocardial infarction (MI) in Fischer 344 Brown Norway rats. In sham rats at 23 mo, LV weight, myocyte cross-sectional area (CSA), and myocardial fibrosis were increased, whereas LV dP/d t, LV relaxation, and maximal LV systolic function declined with respect to younger rats (7, 12, and 18 mo of age). Isometric myocardial function was evaluated in papillary muscles of 12- and 23-mo-old sham rats. Myocardial systolic function was decreased in older rats. To determine how aging affects LV function and remodeling after MI, rats were infarcted at 7 and 18 mo of age and were studied 5 mo later. Infarct size was similar in each group. Right ventricular weight, LV end-diastolic pressure, and volume index were increased, whereas LV dP/d t, peak cardiac index, and peak developed LV pressure declined after MI. However, there were no significant differences between young and older rats in any variable of LV systolic function or remodeling after MI. Myocyte CSA increased in younger rats after MI but was unchanged in 23-mo-old rats. After MI, myocardial fibrosis was significantly increased from baseline only in younger rats. The negative interaction of aging and MI on myocyte hypertrophy and fibrosis was highly significant. The findings indicate that baseline LV and myocardial function decline with age. In the aging rat after MI, despite limited compensatory hypertrophy and more advanced baseline myocardial fibrosis, the long-term functional and structural adaptations to MI are similar to those of the mature adult heart.

Background Progenitor cells isolated from cardiac explant–derived cells improve cardiac function after myocardial infarction (MI). To fully realize the therapeutic potential of these cells, it is essential to develop a safe and efficient delivery method. Therefore, the objective of this study was to determine the efficacy of our newly developed approach to retrograde coronary vein (RCV) infusion of cardiac c-Kit + cells in a small-animal model of congestive heart failure (CHF). Methods Sprague-Dawley rats underwent experimental MI. After 21 days, cardiac explant–derived c-Kit + cells were delivered to both sham and CHF animals using RCV delivery. Vehicle-treated (serum-free medium) sham and CHF animals were used as controls. Cardiac function and heart tissues were evaluated 21 days post-transplantation. Results RCV-delivered cells were retained in infarcted hearts for at least 21 days after transplantation. At 21 days post-RCV infusion, the majority of transplanted c-Kit + /GFP + cells were localized in the left ventricle. Compared with vehicle-treated CHF animals, RCV-treated rats showed a significant improvement in cardiac function. Furthermore, RCV-treated rats exhibited an increase in capillary density, a decrease in total heart collagen, and a reduction in both infarct size and cardiomyocyte hypertrophy when compared with vehicle-treated CHF rats. Conclusions Our study showed that the RCV infusion approach is an efficient technique for targeted cell delivery to the infarcted myocardium. Cardiac c-Kit + cells, delivered using RCV infusion ameliorated progression of heart failure, improved cardiac function and retarded myocardial remodeling in heart failure rats.

Varying strategies are currently being evaluated to develop tissue-engineered constructs for the treatment of ischemic heart disease. This study examines an angiogenic and biodegradable cardiac construct seeded with neonatal cardiomyocytes for the treatment of chronic heart failure (CHF).We evaluated a neonatal cardiomyocyte (NCM)-seeded 3-dimensional fibroblast construct (3DFC) in vitro for the presence of functional gap junctions and the potential of the NCM-3DFC to restore left ventricular (LV) function in an in vivo rat model of CHF at 3 weeks after permanent left coronary artery ligation.The NCM-3DFC demonstrated extensive cell-to-cell connectivity after dye injection. At 5 days in culture, the patch contracted spontaneously in a rhythmic and directional fashion at 43 ± 3 beats/min, with a mean displacement of 1.3 ± 0.3 mm and contraction velocity of 0.8 ± 0.2 mm/sec. The seeded patch could be electrically paced at nearly physiologic rates (270 ± 30 beats/min) while maintaining coordinated, directional contractions. Three weeks after implantation, the NCM-3DFC improved LV function by increasing (p0.05) ejection fraction 26%, cardiac index 33%, dP/dt(+) 25%, dP/dt(-) 23%, and peak developed pressure 30%, while decreasing (p0.05) LV end diastolic pressure 38% and the time constant of relaxation (Tau) 16%. At 18 weeks after implantation, the NCM-3DFC improved LV function by increasing (p0.05) ejection fraction 54%, mean arterial pressure 20%, dP/dt(+) 16%, dP/dt(-) 34%, and peak developed pressure 39%.This study demonstrates that a multicellular, electromechanically organized cardiomyocyte scaffold, constructed in vitro by seeding NCM onto 3DFC, can improve LV function long-term when implanted in rats with CHF.

Background Cardiac c‐Kit+ cells isolated from cardiac explant‐derived cells modestly improve cardiac functions after myocardial infarction; however, their full potential has not yet been realized. For instance, the majority of potential candidates for cell therapy suffer from chronic heart failure ( CHF ), and it is unclear how this disease affects the explant‐derived progenitor cells. Therefore, the objective of this study was to determine the effect of CHF on the number and phenotype of cardiac explant c‐Kit+ progenitors and elucidate mechanisms of their regulation. Methods and Results Myocardial infarction was created by left anterior descending coronary artery occlusion. Sham‐operated animals were used as a control group. CHF ‐developed infarcted animals were selected on the basis of left ventricle end‐diastolic pressure ≥20 mm Hg and scar size ≥30%. Here, we found that CHF atrial explants produced less c‐Kit+ cells than sham explants. CHF ‐derived c‐Kit+ cells exhibited upregulated transforming growth factor–β ( TGF ‐β) signaling, increased level of epithelial to mesenchymal transition markers, and diminished expression of pluripotency markers compared with shams. We show that intervention with TGF ‐β signaling by inhibiting TGF ‐β receptor type I or Smad 2/3 using small‐molecule inhibitors improved c‐Kit+ cell yield, attenuated epithelial to mesenchymal transition markers, stimulated the pluripotency marker Nanog, and improved efficiency of c‐Kit+ cell differentiation toward cardiomyocyte‐like cells in vitro. Conclusions Taken together, our findings suggest that TGF ‐β inhibition positively modulates c‐Kit+ cell phenotype and function in vitro, and this strategy may be considered in optimizing cardiac progenitor function and cell expansion protocols for clinical application.

A pilot clinical study using autologous c-Kit+ cells showed improvement in cardiac functions in congestive heart failure (CHF), however, it is unclear if c-Kit+ cells isolated from CHF hearts are equally as potent as cells from controls. To test the potency of CHF c-Kit+ cells, myocardial infarction (MI) was created by permanent ligation of the left anterior descending coronary artery. Six weeks after MI, animals with left ventricle end-diastolic pressure (LVEDP) ≥20 mmHg and scar size ≥30% of left ventricle (LV) were designated as CHF rats. We found that CHF atrial explants generated less c-Kit+ cells compared to shams (15.7% vs. 11% sham vs. CHF). CHF c-Kit+ cells exhibited elevated levels of epicardial to mesenchymal transition markers, including Snail (2.5 fold) and Pai1 (3 fold), while the expression level of epithelial marker, E-cadherin was 3 fold lower in CHF c-Kit+ cells. Moreover, CHF c-Kit+ cells exhibited reduced gene expressions of pluripotency markers; 2.1 fold decrease in Nanog and 4.5 fold decrease in Sox 2 compared to sham cells. To evaluate the potency of the c-Kit+ cells, 1 x 10 6 cells isolated from CHFs or shams were delivered to 3 weeks post-MI CHF hearts. Cells were pre-labeled with GFP to enable their tracing in vivo and delivered to the infarcted myocardium via left coronary vein by a retrograde coronary sinus cell infusion (RCI). RCI delivery resulted in a cell distribution of LV (30%), right atrium (30%) and right ventricle (20%), while only 10% of cells were found in a left atrium. Three weeks after cells delivery, rats transplanted with sham c-Kit+ cells showed improved LVEDP (29.4 ± 6 vs. 11.7 ± 3.5 mmHg, CHF vs. CHF+ sham c-Kit+ cells) and a rise in peak rate of pressure (dPdt max) (3988 ± 520 vs. 5333 ± 597 mmHg/s). In contrast, no functional improvement was detected in rats transplanted with CHF c-Kit+ cells. Histological analysis demonstrated that transplanted c-Kit+/GFP+ cells were mostly incorporated into blood vessels and co-localized with endothelial marker vWf, and α-smooth muscle actin. Our results showed that left coronary vein is an efficient route for c-Kit+ cell delivery and that c-Kit+ cells isolated from CHF rats are less potent when transplanted in chronic heart failure rat model compared to those isolated from control.

Cardiac progenitor cells are generated from atria explants; however the cellular origin and the mechanisms of cell outgrowth are unclear. Using transgenic tamoxifen-induced Willms tumor 1 (Wt1)-Cre/ERT and Cre-activated GFP reporter mice, we found approximately 40% of explant-derived cells and 74% of explant-derived c-Kit+ cells originated from the epicardium. In atria from sham hearts, Wt1+ cells were located in a thin epicardial layer, while c-Kit+ cells were primarily found within both the sub-epicardium and the myocardium, albeit at low frequency. No overlap between c-Kit+ and Wt1+ cells was observed, suggesting that epicardial Wt1+ cells do not express c-Kit marker in vivo, but more likely the c-Kit marker was acquired in culture. Compared with 4 days in culture, at day 21 we observed 7 folds increase in Snail gene expression; 32% increase in α-smooth muscle actin (SMA) marker, and 30% decrease in E-cadherin marker, suggesting that the explant-derived cells underwent epithelial to mesenchymal transition (EMT) in vitro. Cell outgrowths released TGF-β (1036.4 ± 1.18 pm/ml) and exhibited active TGF-β signaling, which might triggered the EMT. Compared to shams, CHF cell outgrowths exhibited elevated levels of EMT markers, SMA (49% vs. 34%) and Snail (2 folds), and reduced level of Wt1 (11% vs. 22%). In addition, CHF cell outgrowths had two folds increase in Pai1 gene expression, a direct target of TGF-β signaling. In c-Kit+ cells derived from CHF explants, Nanog gene expression was 4 folds lower and Sox 2 was 2 folds lower compared with cells from shams. Suppression of EMT in cell outgrowth increased the percentage of c-Kit+ and Wt1+ cells by 17%, and 15%, respectively. Also suppression of EMT in c-Kit+ cells resulted in 4 folds increase in Nanog and 3 fold increase in Sox2 gene expressions. Our results showed that CHF may further exuberates EMT while diminishes the re-activation of pluripotency genes. Thus, EMT modulation in CHF is a possible strategy to regulate both the yield and the pluripotency of cardiac-explant-derived progenitor cells.

Myofibroblast activation after myocardial infarction (MI) is one of the major mechanisms of myocardial fibrosis and subsequent heart failure. Elucidating the mechanisms of myofibroblast activation will provide a strategy to treat heart failure. Histone Deacetylases (HDAC) were reported to regulate myofibroblast differentiation and extracellular matrix deposition in idiopathic pulmonary fibrosis. Furthermore, HDAC inhibitors were shown to retards myocardial remodeling and improve cardiac function after MI. Therefore, here we examined whether HDAC1/2 are up-regulated after MI, and identify which myocardial cell type is predominately expressing HDAC. MI was created in rats by ligation of the left coronary artery. First, we assessed expression of HDAC 1 and 2 following MI in heart sections. Both HDACs uniformly expressed in myocardium in right ventricular (LV) and left ventricular (LV) in sham hearts. On the other hand, we observed up-regulation of both HDAC1 and 2 in the infarcted LV following MI. In addition, HDAC1 and 2 were co-localized with fibroblast marker Vimentin in the infarcted area. Next, we performed western blot analysis to investigate protein levels of HDAC1 and HDAC2 in right ventricular (RV), septum (S) and LV (infarcted and remote area). We found up-regulation of HDAC1 and HDAC2 only in the infarcted area compared to sham rats. Altogether these results suggest HDAC1 and 2 play a role in fibrosis following MI.

We and others have shown that transplantation of explant-derived cells (EDCs) obtained from cardiac biopsies improved cardiac function after myocardial infarction. The current study was designed to examine the molecular mechanisms regulating expression of pluripotent genes in these cells. Toward this end, EDCs were separated based on expression of c-Kit antigen after 21 days in culture. We found that Notch signaling was activated mainly in c-Kit+ cells, but to a lesser degree in c-Kit- cells. In addition, we found that the pluripotency markers Sca-1, Nanog and Sox2 were mainly expressed in c-Kit+ cells. Also, in c-Kit+ cells, forced activation of Notch signaling via over-expression of Notch intracellular domain (NICD) induced cellular and molecular changes typical of epithelial-mesenchymal transition (EMT), evident by a decrease in VE cadherin and increases in N-cadherin, MMP, and ICAM. . Suppression of Notch signaling was associated with c-Kit+ cells exhibition of an epithelial/endothelial morphology. This observation is further supported by the increase in Nanog, Sca1, Sox2, VEGFR2 gene expressions after Notch suppression. In contrast, overexpression of NICD resulted in down-regulation of pluripotency gene expressions. Furthermore, suppression of Notch signaling was coincided with stabilization of β-catenin and accumulation of phosphorylated glycogen synthase kinase 3 beta (pGSK3β) suggestive of a crosstalk between Notch and canonical Wnt pathway. Thus, Notch regulates expression of pluripotency genes and mesenchymal transition of c-Kit+ cardiac EDCs.

Background Progenitor cell therapy is emerging as a novel treatment for heart failure. However the molecular mechanisms regulating the generation of cardiac progenitor cells is not fully understood. We hypothesized that cardiac progenitor cells are generated from cardiac explant via a process similar to epithelial to mesenchymal transition (EMT). Methods/Findings Explant-derived cells were generated from partially digested atrial tissue. After 21 days in culture, c-Kit+ cells were isolated from cell outgrowth. The majority of explant-originated c-Kit+ cells expressed the epicardial marker Wt1. Cardiac cell outgrowth exhibits a temporal up-regulation of EMT-markers. Notch stimulation augmented, while Notch inhibition suppressed, mesenchymal transition in both c-Kit+ and c-Kit- cells. In c-Kit+ cells, Notch stimulation reduced, while Notch inhibition up-regulated pluripotency marker expressions such as Nanog and Sox2. Notch induction was associated with degradation of β-catenin in c-Kit- cells. In contrast, Notch inhibition resulted in β-catenin accumulation, acquisition of epitheloid morphology, and up-regulation of Wnt target genes in c-Kit- cells. Conclusion Our study suggests that Notch-mediated reversible EMT process is a mechanism that regulates explant-derived c-Kit+ and c-Kit- cells.

This study examines a viable biodegradable three-dimensional fibroblast construct (3DFC) in a model of chronic heart failure. The viable fibroblasts, cultured on a vicryl mesh, secrete growth factors that stimulate angiogenesis.We ligated the left coronary artery of male Sprague-Dawley rats, implanted the 3DFC 3 weeks after myocardial infarction and obtained end point data 3 weeks later, that is, 6 weeks after myocardial infarction.Implanting the 3DFC increases (p0.05) myocardial blood flow twofold, microvessel formation (0.02±0.01 vs. 0.07±0.03 vessels/μm2), and ventricular wall thickness (0.53±0.02 to 1.02±0.17mm). The 3DFC shifts the passive pressure volume loop toward the pressure axis but does not alter left ventricular (LV) ejection fraction, systolic displacement, LV end-diastolic pressure/dimension, or LV cavity area. The 3DFC stimulates selected cytokine activation with a decrease in the proinflammatory cascade and increased total protein content stimulated by strained 3DFC in vitro.The 3DFC functions as a cell delivery device providing matrix support for resident cell survival and integration into the heart. The imbedded fibroblasts of the 3DFC release a complex blend of cardioactive cytokines promoting increases in microvessel density and anterior wall blood flow but does not improve ejection fraction or alter LV remodeling.

Cell-based therapy for myocardial infarction (MI) holds great promise; however, the ideal cell type and delivery system have not been established. Obstacles in the field are the massive cell death after direct injection and the small percentage of surviving cells differentiating into cardiomyocytes. To overcome these challenges we designed a novel study to deliver cardiac progenitor cells as a cell sheet.Cell sheets composed of rat or human cardiac progenitor cells (cardiospheres), and cardiac stromal cells were transplanted onto the infarcted myocardium after coronary artery ligation in rats. Three weeks later, transplanted cells survived, proliferated, and differentiated into cardiomyocytes (14.6 +/- 4.7%). Cell sheet transplantation suppressed cardiac wall thinning and increased capillary density (194 +/- 20 vs. 97 +/- 24 per mm(2), P0.05) compared with the untreated MI. Cell migration from the sheet was observed along the necrotic trails within the infarcted area. The migrated cells were located in the vicinity of stromal-derived factor (SDF-1) released from the injured myocardium, and about 20% of these cells expressed CXCR4, suggesting that the SDF-1/CXCR4 axis plays, at least, a role in cell migration. Transplantation of cell sheets resulted in a preservation of cardiac contractile function after MI, as was shown by a greater ejection fraction and lower left ventricular end diastolic pressure compared with untreated MI.The scaffold-free cardiosphere-derived cell sheet approach seeks to efficiently deliver cells and increase cell survival. These transplanted cells effectively rescue myocardium function after infarction by promoting not only neovascularization but also inducing a significant level of cardiomyogenesis.

Myocardial infarction (MI) and stroke are the first and third leading causes of death in the U.S.A. accounting for more than 1 in 3 deaths per annum. Despite interventional and pharmaceutical advances, the number of people diagnosed with heart disease is on the rise. Therefore, new clinical strategies are needed. Cell-based therapy holds great promise for treatment of these diseases and is currently under extensive preclinical as well as clinical trials. The source and types of stem cells for these clinical applications are questions of great interest. Human umbilical cord blood (hUCB) appears to be a logical candidate as a source of cells. hUCB is readily available, and presents little ethical challenges. Stem cells derived from hUCB are multipotent and immunologically naive. Here is a critical literature review of the beneficial effects of hUCB cell therapy in preclinical trials.

Summary Aim Increases in serum cytokines have been reported after successful resuscitation from prolonged ventricular fibrillation (VF). Pro-inflammatory cytokines can stimulate inducible nitric oxide synthase (iNOS) to produce excessive levels of nitric oxide (NO). High levels of both myocardial inflammatory cytokines and nitric oxide levels can depress myocardial contractile function. We hypothesized that myocardial pro-inflammatory cytokines and iNOS activity would increase following successful resuscitation from prolonged ventricular fibrillation cardiac arrest, and that such increases would parallel the development of post-resuscitation myocardial dysfunction. Methods Ventricular fibrillation cardiac arrest was induced in seven domestic swine (25 ± 5 kg). After 10 min of untreated VF, the animals were defibrillated and resuscitated. Left ventricular (LV) systolic and diastolic function measurements, serum samples (arterial and coronary sinus) for IL-8 cytokine quantification, and LV myocardial biopsies were collected before, during, and after resuscitation. Quantification of myocardial endothelial (eNOS) and inducible (iNOS) nitric oxide synthase protein levels were determined using immunoblot analyses and protein localization was examined using immunohistochemistry. Results Post-resuscitation LV systolic and diastolic functions were depressed while increases in both coronary sinus IL-8 levels and myocardial iNOS activity were found. Compared to pre-arrest baseline, levels of iNOS protein increased during VF ( p ≤ 0.05) and continued to increase throughout the post-resuscitation study period of 6 h ( p ≤ 0.05). Conclusions Myocardial inflammatory cytokines and iNOS activity increase during and after prolonged cardiac arrest and successful resuscitation. These increases correspond to the well described decrease in LV function post-resuscitation.

The heart does not mend itself after infarction. However, stem cells may revolutionize heart disease treatment. A vast and growing body of evidence indicates that cell-based strategies have promising therapeutic potential. Recent clinical and pre-clinical studies demonstrate varying degrees of improvement in cardiac function using different adult stem cell types such as bone marrow (BM)-derived progenitor cells and skeletal myoblasts. However, the efficacy of cell therapy after myocardial infarction (MI) is inconclusive and the cellular source with the highest potential for regeneration is unclear. Clinically, BM and skeletal muscle are the most commonly used sources of autologous stem cells. One major pitfall of using autologous stem cells is that the number of functional cells is generally depleted in the elderly and chroni- cally ill. Therefore, there is an urgent need for a new source of adult stem cells. Human umbilical cord blood (CB) is a candidate and appears to have several key advantages. CB is a viable and practical source of progenitor cells. The cells are naive and what's more, CB contains a higher number of immature stem/progenitor cells than BM. We review recent clinical experience with adult stem cells and explore the potential of CB as a source of cells for cardiac repair following MI. We conclude that there is a conspicuous absence of clinical studies utilizing CB-derived cells and there is a pressing need for large randomized double-blinded clinical trials to assess the overall efficacy of cell-based ther- apy.

Altering the renin-angiotensin aldosterone system improve mortality in heart failure (HF) in part through an improvement in nitric oxide (NO)-mediated endothelial function. This study examined if spironolactone affects endothelial nitric oxide synthase (eNOS) and NO-mediated vasorelaxation in HF.Rats with HF after coronary artery ligation were treated with spironolactone for 4 weeks. Rats with HF had a decrease (P.05) in left ventricular (LV) systolic pressure (130 +/- 7 versus 118 +/- 6 mm Hg) and LV pressure with respect to time (9,122 +/- 876 versus 4,500 +/- 1971 mm Hg/second) with an increase in LV end-diastolic pressure (4 +/- 2 versus 23 +/- 8 mm Hg). Spironolactone did not affect hemodynamics but it improved (P.05) endothelial-dependent vasorelaxation at more than 10(-8) M acetylcholine that was abolished with N(G)-monomethyl-L-arginine. The eNOS levels were decreased (P.05) in the LV and the aorta; spironolactone restored LV and aortic eNOs levels to normal.Spironolactone prevents the decrease in eNOS in the LV and aorta and improves NO-dependent vasorelaxation, suggesting that one potential mechanism of spironolactone is an improvement in vasoreactivity mediated though an increase in NO.

Published guidelines recommend continuing β-adrenergic receptor blockade in patients undergoing stress testing. We evaluated the role of pharmacological versus exercise stress testing in achieving target heart rate (THR) among patients on β-adrenergic blockade. We compared data from 140 patients who underwent dobutamine stress echo (DSE) and 143 patients who underwent exercise treadmill testing (ETT). In both groups, β-adrenergic blocker was continued at the time of stress testing. Overall, patients undergoing DSE achieved THR more frequently than ETT. With β-adrenergic blockade, DSE patients met THR more frequently than ETT patients (p < 0.001). Without β-adrenergic blockade, there was no difference between either modality in achieving THR. In both DSE and ETT patients, absence of β-adrenergic blockade increased the odds of achieving THR [odds ratio (OR): 2.46, p = 0.042 and OR: 7.44, p < 0.001, respectively]. Atropine use with DSE increased the odds of achieving THR (OR: 3.76, p = 0.006). In conclusion, pharmacological stress testing appears to be superior to exercise stress testing in achieving THR among patients on β-adrenergic blockade.

This study was designed to determine if the thyroid hormone analog 3,5 diiodothyropropionic acid (DITPA), now in clinical trials for heart failure, alters endothelial function after myocardial infarction (MI). Three weeks after MI, adult Sprague-Dawley rats were randomly assigned to DITPA (375 microg/100 g subcutaneous) or no treatment of 3 weeks. In MI rats, left ventricular (LV) end-diastolic pressure and LV dP/dt decreased (P < 0.05). DITPA did not change MAP (87 +/- 10 versus 90 +/- 7 mm Hg) or LV end-diastolic pressure (23 +/- 3 versus 19 +/- 9 mm Hg) but did lower (P < 0.05) LV dP/dt (4,633 +/- 797 versus 3,650 +/- 1,236 mm Hg/s). In aortic segments from MI rats, DITPA enhanced the acetylcholine dependent vasorelaxation (59 +/- 11% at 10(-4) M, P < 0.05) and isoproterenol induced vasorelaxation (57 +/- 13% at 10(-4) M, P < 0.05). The increases in vasorelaxation were blocked with l-NAME and restored with L-arginine. Treatment with DITPA increased (P < 0.05) eNOS protein content in aortic tissue from sham rats (3.8 +/- 2.8 to 44.5 +/- 7.1 integrated intensity units (II)/microg) and in MI rats (5.3 +/- 3.4 to 28.3 +/- 8.9 II/microg). In endothelial cells, 24 hours' treatment with DITPA (10 microM) increased (P < 0.01) eNOS protein expression from 22.1 +/- 4.8 to 52.7 +/- 16.8 II/microg protein and DITPA (20 microM) increased eNOS to 49.1+/- 15.2 II/microg protein. The thyroid analog DITPA enhances endothelial nitric oxide and beta-adrenergic-mediated vasorelaxation by increasing nitric oxide in the vasculature.

Coronary artery disease (CAD) remains the leading cause of death in the Western world. The high impact of its main sequelae, acute myocardial infarction and congestive heart failure (CHF), on the quality of life of patients and the cost of health care drives the search for new therapies. The recent finding that stem cells contribute to neovascularization and possibly improve cardiac function after myocardial infarction makes stem cell therapy the most highly active research area in cardiology. Although the concept of stem cell therapy may revolutionize heart failure treatment, several obstacles need to be addressed. To name a few: 1) Which patient population should be considered for stem cell therapy? 2) What type of stem cell should be used? 3) What is the best route for cell delivery? 4) What is the optimum number of cells that should be used to achieve functional effects? 5) Is stem cell therapy safer and more effective than conventional therapies? The published studies vary significantly in design, making it difficult to draw conclusions on the efficacy of this treatment. For example, different models of ischemia, species of donors and recipients, techniques of cell delivery, cell types, cell numbers and timing of the experiments have been used. However, these studies highlight the landmark concept that stem cell therapy may play a major role in treating cardiovascular diseases in the near future. It should be noted that stem cell therapy is not limited to the treatment of ischemic cardiac disease. Non-ischemic cardiomyopathy, peripheral vascular disease, and aging may be treated by stem cells. Stem cells could be used as vehicle for gene therapy and eliminate the use of viral vectors. Finally, stem cell therapy may be combined with pharmacological, surgical, and interventional therapy to improve outcome. Here we attempt a systematic overview of the science of stem cells and their effects when transplanted into ischemic myocardium.

To determine whether angiotensin receptor blockade decreases vascular tone in heart failure by improving endothelial-dependent vasorelaxation and increasing nitric oxide (NO) bioavailability, we treated infarcted adult male Sprague-Dawley rats with candesartan for 7 days or 8 weeks (10 mg/kg/day in drinking water). Candesartan, at both time points, lowered left ventricular (LV) systolic pressure (P < 0.05) (122 +/- 22 versus 74 +/- 16 and 73 +/- 10 mm Hg) and LV dP/dt (5914 +/- 1294 versus 2857 +/- 1672 versus 3175 +/- 769 mm Hg/s), but lowered LV end-diastolic pressure only at 8 weeks (16.9 +/- 9.7 versus 11.2 +/- 5.7 versus 6.9 +/- 5.3 mm Hg). The vasorelaxation response to acetylcholine (ACh) in thoracic aortic segments was decreased with infarction (P < 0.05), remained unchanged with 1 week of candesartan, but increased 84 and 86% at 10-4 and 10-5 M ACh (P < 0.05) at 8 weeks. The enhanced candesartan-induced vasorelaxation at 8 weeks was abolished with NG-nitro-l-arginine methyl ester (200 microM). In bovine pulmonary endothelial cells, 20 microM candesartan increased endothelial nitric-oxide synthase (eNOS) protein levels (P < 0.05) (28.9 +/- 2.6 versus 16.1 +/- 3.7 intensity units/microg of protein); the increased eNOS was abolished by a specific angiotensin subtype 2 (AT2) receptor antagonist, PD 123319. These data suggest that AT1 receptor blockade enhances vasorelaxation in heart failure by increasing NO bioavailability, in part via an AT2 receptor-mediated up-regulation of eNOS protein.

The efficacy of amiodarone used in combination with a pacemaker cardioverter defibrillator (PCD) to decrease episodes of ventricular tachycardia and subsequent PCD shocks is not clear. We examined a retrospective registry of 82 patients with PCD implantation to define the efficacy of amiodarone treatment. We compared patients treated with amiodarone (for 24 consecutive months without interruption) versus no amiodarone. In patients treated with amiodarone there was a 3-fold increase (p = 0.02) in PCD shocks; in patients not on beta-blockers, amiodarone resulted in a 6-fold increase (p < 0.05) in PCD shocks. Patients with a left ventricular ejection fraction >30% on amiodarone and patients

Background: Intramedullary rods and external fixators are reported to be more widely used than plating in osteosynthesis of femoral shaft fractures in children. In our institution plating is the routine, and we are reporting our experience in a retrospective study. Methods: Two groups of patients were treated by AO plating for femoral shaft fractures. The first included 36 polytraumatized children and the second involved 10 cases of old malunited fractures. Average follow-up for the patients was 5 years. Results: All children achieved union after one operation except one. There were no major complications except one case of mechanical failure. Insignificant limb length inequality occurred in only six children. Conclusion: We believed that plating of femoral shaft fractures is a reasonable option in treating children with fresh and neglected fractures.

Somatic gene therapy of vascular diseases is a promising new field in modern medicine. Recent advancements in gene transfer technology have greatly evolved our understanding of the pathophysiologic role of candidate disease genes. With this knowledge, the expression of selective gene products provides the means to test the therapeutic use of gene therapy in a multitude of medical conditions. In addition, with the completion of genome sequencing programs, gene transfer can be used also to study the biologic function of novel genes in vivo. Novel genes are delivered to targeted tissue via several different vehicles. These vectors include adenoviruses, retroviruses, plasmids, plasmid/liposomes, and oligonucleotides. However, each one of these vectors has inherent limitations. Further investigations into developing delivery systems that not only allow for efficient, targeted gene transfer, but also are stable and nonimmunogenic, will optimize the clinical application of gene therapy in vascular diseases. This review further discusses the available mode of gene delivery and examines six major areas in vascular gene therapy, namely prevention of restenosis, thrombosis, hypertension, atherosclerosis, peripheral vascular disease in congestive heart failure, and ischemia. Although we highlight some of the recent advances in the use of gene therapy in treating vascular disease discovered primarily during the past two years, many excellent studies published during that period are not included in this review due to space limitations. The following is a selective review of practical uses of gene transfer therapy in vascular diseases. This review primarily covers work performed in the last 2 years. For earlier work, the reader may refer to several excellent review articles. For instance, Belalcazer et al. (6) reviewed general aspects of somatic gene therapy and the different vehicles used for the delivery of therapeutic genes. Gene therapy in restenosis and stimulation of angiogenesis in the cardiac muscle are discussed in reviews by several investigators (13,26,57,74,83). In another review, Meyerson et al. (43) discuss advances in gene therapy for vascular proliferative disorders and chronic peripheral and cardiac ischemia.

Congestive heart failure (HF) is characterized by inadequate nitric oxide (NO) production in the vasculature. Because NO is degraded by oxygen radicals, we hypothesized that NO is degraded faster in HF from inadequate peripheral arterial antioxidant reserves. HF was induced in male Sprague-Dawley rats by left coronary artery ligation. Vascular endothelial function was evaluated by measuring the NO-mediated vasorelaxation response to acetylcholine (ACh; 10−9–10−4M) in excised aortas. This was repeated with the free radical generator pyrogallol (20 μM) and again with pyrogallol and superoxide dismutase (SOD; 60 U/ml). Aortic and myocardial SOD activity was also determined. ACh-induced vasorelaxation was reduced in HF ( n = 9) compared with normal control rats ( n = 11; P < 0.001). Pyrogallol further reduced vasorelaxation in HF: 74 ± 11% at 10−4M ACh versus 58 ± 10% in normal control rats ( P < 0.004). There was a trend ( P = 0.06) toward reduced SOD activity in HF aortas. In conclusion, altered NO-dependent vasorelaxation in HF is in part due to excessive degradation of NO and is likely related to reduced vascular SOD activity.