Matthew Sullivan

Significant improvement of heart function by cotransplantation of human mesenchymal stem cells and fetal cardiomyocytes in postinfarcted pigs

BACKGROUND Viable cardiomyocytes after myocardial infarction (MI) are unable to repair the necrotic myocardium due to their limited capability of regeneration. The present study investigated whether intramyocardial transplantation of human mesenchymal stem cells (hMSCs) or cotransplantation of hMSCs plus human fetal cardiomyocytes (hFCs; 1:1) reconstituted impaired myocardium and improved cardiac function in MI pigs. METHODS AND RESULTS Cultured hMSCs were transfected with green fluorescent protein (GFP). Six weeks after MI induction and cell transplantation, cardiac function was significantly improved in MI pigs transplanted with hMSCs alone. However, the improvement was even markedly greater in MI pigs cotransplanted with hMSCs plus hFCs. Histological examination demonstrated that transplantation of hMSCs alone or hMSCs plus hFCs formed GFP-positive engrafts in infarcted myocardium. In addition, immunostaining for cardiac alpha-myosin heavy chain and troponin I showed positive stains in infarcted regions transplanted with hMSCs alone or hMSCs plus hFCs. CONCLUSIONS Our data demonstrate that transplantation of hMSCs alone improved cardiac function in MI pigs with a markedly greater improvement from cotransplantation of hMSCs plus hFCs. This improvement might result from myocardial regeneration and angiogenesis in injured hearts by engrafted cells.

Electrocardiographic findings in mdx mice: A cardiac phenotype of Duchenne muscular dystrophy

The mdx mouse is a model of Duchenne muscular dystrophy (DMD). As many DMD patients die of cardiac failure, we investigated whether mdx mice exhibited clinically relevant cardiac phenotypes. We applied a recently developed method for noninvasively recording electrocardiograms (ECGs) to study male mdx mice (n = 15) and control mice (n = 15). The mdx mice had significant tachycardia and decreased heart rate variability, consistent with observations in DMD patients. Heart rate was nearly 15% faster in mdx mice than control mice (P < 0.05). The rate‐corrected QT interval duration and PR interval were shorter in mdx compared to control mice (P < 0.05). The muscarinic antagonist atropine significantly increased heart rate and decreased PR interval in C57 mice. In contrast, atropine significantly decreased heart rate and increased PR interval in all mdx mice. Pharmacological autonomic blockade and baroreflex sensitivity testing demonstrated an imbalance in autonomic nervous system modulation of heart rate, with decreased parasympathetic activity and increased sympathetic activity in mdx mice. Baseline ECGs and contrary responses to muscarinic blockade by atropine in mice deficient in neuronal nitric oxide synthase (nNOS) suggest that the autonomic dysfunction in mdx mice may be independent of decreased myocardial nNOS. These electrocardiographic findings in dystrophin‐deficient mice may provide new bases for diagnosing, understanding, and treating DMD patients.

Induction of Cardiac Cytochrome P450 in Cocaine-Treated Mice

Cytochrome P450 (P450) is a ubiquitous family of enzymes responsible for the metabolism of a wide variety of drugs and their metabolites, including cocaine. To investigate the effects of cocaine on myocardial injuries and cardiac P450 expression, BALB/c mice were injected daily intraperitoneally with cocaine (30 mg/kg) or cocaine plus pretreatment of P450 inhibitors for 14 days. Tumor necrosis factor-α (TNF-α) content and creatine phosphokinase (CPK) activity in mice hearts and serums were significantly increased after long-term treatment with cocaine. Pretreatment with the P450 inhibitor, cimetidine (Cime, 50 mg/kg) or metyrapone (Mety, 40 mg/kg) abolished or significantly attenuated the effects of cocaine on TNF-α and CPK activity. Western blot analysis shows that mouse cardiac tissues express the P450 isoforms CYP1A1, CYP1A2, and CYP2J2. The protein levels normalized with cyclophilin A were 1.20 ± 0.07, 0.67 ± 0.03, and 1.48 ± 0.01 for CYP1A1, CYP1A2, and CYP 2J2, respectively. After cocaine administration, CYP2J2 increased by 43.6% and CYP1A1 increased by 108.5%, but CYP1A2 was not significantly altered. However, the cytochrome P450 inhibitors Cime and Mety suppressed the cocaine-induced increase in CYP1A1 and CYP2J2 expression. Moreover, application of Cime or Mety alone did not alter the level of cardiac TNF-α or the expression of P450. Our results demonstrate that long-term exposure to cocaine causes an increase in cardiac CYP1A1 and CYP2J2 concentration. We speculate that induction of P450 isoforms may cause cardiac injury due to cocaine metabolites locally catalyzed by P450 or the increase in P450 expression itself.

Embryonic stem cells attenuate viral myocarditis in murine model.

We used mice to test our hypothesis that in response to viral invasion, stem cells may migrate into the heart and attenuate the effect of viral myocarditis. Male BALB/c mice were divided into three groups: mouse embryonic stem (ES) cell control, encephalomyocarditis virus (EMCV), and EMCV + ES cells. After administration of ES cells via tail vein, mice were immediately inoculated with EMCV. Mice were sacrificed at different days after EMCV inoculation. Mortality was recorded. Inflammatory cell infiltration and necrosis (major pathological changes of viral myocarditis) were evaluated by hematoxylin-eosin staining. ES cell migration and differentiation were identified by immunofluorescence. The survival rate in the EMCV + ES cell group (80%) was significantly increased (p < 0.05) over the EMCV-alone group (64%). Also, the incidence of inflammatory cell infiltration and myocardial lesions was lower in the EMCV + ES cell mice. Furthermore, the result of green fluorescent protein (GFP) and a-actinin analysis indicated that ES cells migrated into the heart and differentiated into myocytes after virus inoculation. In conclusion, ES cells significantly increased the survival of viral myocarditis mice and also decreased the necrosis and infiltration of inflammatory cells. These results demonstrated the ability of stem cells to mitigate the effects of viral infection on the heart and illustrated their potential therapeutic application to other mammalian species, including humans.

Overexpression of Na+/Ca2+ exchanger gene attenuates postinfarction myocardial dysfunction

We monitored myocardial function in postinfarcted wild-type (WT) and transgenic (TG) mouse hearts with overexpression of the cardiac Na(+)/Ca(2+) exchanger. Five weeks after infarction, cardiac function was better maintained in TG than WT mice [left ventricular (LV) systolic pressure: WT, 41 +/- 2; TG, 58 +/- 3 mmHg; P < 0.05; maximum rising rate of LV pressure (+dP/dt(max)): WT, 3,750 +/- 346; TG, 5,075 +/- 334 mmHg/s; P < 0.05]. The isometric contractile response to beta-adrenergic stimulation was greater in papillary muscles from TG than WT mice (WT, 13.2 +/- 0.9; TG, 16.3 +/- 1.0 mN/mm(2) at 10(-4) M isoproterenol). The sarcoplasmic reticulum (SR) Ca(2+) content investigated by rapid cooling contractures in papillary muscles was greater in TG than WT mouse hearts. We conclude that myocardial function is better preserved in TG mice 5 wk after infarction, which results from enhanced SR Ca(2+) content via overexpression of the Na(+)/Ca(2+) exchanger.

A microfluidic vibrating wire viscometer for operation at high pressure and high temperature

This article discusses a microfluidic vibrating wire viscometer with an internal volume of a few microliters. Accuracy of order ±10% for viscosities ranging from 0.1 to 100 cP is demonstrated for temperatures (10 °C-175 °C) and pressures (10-24,000 psi) suitable for oilfield use by prior calibration in air and toluene. Comparison between multiple data sets indicates that a large fraction of the discrepancy between literature values is systematic, indicating that future refinements may be possible with better interpretation. Confinement effects are evaluated and are not found to play a significant role, which is surprising since the ratio (6.6) of the fluid channel width to the wire diameter is very low.

Experimental Observation of Inertia-Dominated Squeeze Film Damping in Liquid

We have studied the phenomenon of squeeze film damping in a liquid with a microfabricated vibrating plate oscillating in its fundamental mode with out-of plane motion. It is paramount that this phenomenon be understood so that proper choices can be made in terms of sensor design and packaging. The influences of plate-wall distance h, effective plate radius R, and fluid viscosity and density on squeeze film damping have been studied. We experimentally observe that the drag force is inertia dominated and scales as 1/h 3 even when the plate is far away from the wall, a surprising but understandable result for a Microfluidic device where the ratio of h to the viscous penetration depth is result for a microfluidic device where the ratio of h to the viscous penetration depth is large. We observe as well that the drag force scales as R 3 , which is inconsistent with squeeze film damping in the lubrication limit. These two cubic power laws arise due to the role of inertia in the high frequency limit.

Metoprolol attenuates postischemic depressed myocardial function in papillary muscles isolated from normal and postinfarction rat hearts

The present study was designed to test the hypothesis that metoprolol treatment may enhance tolerance to ischemia in normal and postinfarction rat myocardium. Myocardial infarction was induced by permanent ligation of the left coronary artery in adult rats. Animals were divided into sham-operated and infarction groups with or without metoprolol treatment. Metoprolol treatment (60 mg/kg/day via gastric gavage) was started on the second day after surgery and continued until sacrifice at 6 weeks after myocardial infarction. Isometric force and intracellular Ca(2+) ([Ca(2+)](i)) transients were simultaneously recorded in isolated left ventricular papillary muscles. Ischemia was simulated by immersing the muscles into fluorocarbon with hypoxia. Metoprolol treatment induced a significant improvement of isometric force and ameliorated diastolic [Ca(2+)](i) overload in postinfarction rat myocardium at baseline. Metoprolol treatment also reduced diastolic [Ca(2+)](i), ameliorated the depression of developed tension during ischemia, and enhanced recovery of postischemic depressed myocardial function in sham-operated and postinfarction rat papillary muscles. Protein levels of the sarcoplasmic reticulum Ca(2+) ATPase of left ventricles and postischemic papillary muscles from metoprolol-treated rats were higher than those in placebo-treated animals. We concluded, therefore, that metoprolol treatment produced appreciable improvement of intracellular Ca(2+) handling during ischemia-reoxygenation cycles, and enhanced recovery of postischemic depressed myocardial function in both normal and postinfarction rat myocardium.

Genistein attenuates postischemic depressed myocardial function by increasing myofilament Ca2+ sensitivity in rat myocardium.

The present study investigated whether genistein, a broad-spectrum tyrosine kinase inhibitor, could increase the myofilament Ca2+ sensitivity and partially reverse postischemic depressed myocardial function. Left ventricular papillary muscles were isolated from adult Wistar rats and loaded with the Ca2+ indicator, aequorin. The use of fluorocarbon immersion with hypoxia simulated a model of ischemia. Myofilament responsiveness to Ca2+ was evaluated from force-[Ca2+], relationship recorded during tetani in papillary muscles. Protein levels of troponin I (Tnl) were measured in postischemic papillary muscles with the Western blot technique. Isometric contraction was depressed during the period of ischemia and remained low after 60 min of reoxygenation without a corresponding significant change of peak [Ca2+], in the control group (n = 7). In contrast, the depression of isometric contraction was ameliorated during ischemia in muscle preparations in the presence of genistein (2 μM; n = 8), and postischemic depressed myocardial contractility partially recovered after a 60-min reperfusion. The myofilament Ca2+ responsiveness was significantly increased in papillary muscles in the presence of genistein. Protein levels of Tnl were reduced in postischemic papillary muscles, whereas genistein partially restored decreased protein levels of Tnl. Our results reveal that genistein produces an effective attenuation of postischemic depressed myocardial function and improves myofibrillar Ca2+ responsiveness in rat myocardium.