Tony S. Ma

Tumor necrosis factor-alpha gene and protein expression in adult feline myocardium after endotoxin administration.

TNF alpha mRNA and protein biosynthesis were examined in the adult feline heart after stimulation with endotoxin. When freshly isolated hearts were stimulated with endotoxin in vitro, de novo TNF alpha mRNA expression occurred within 30 min, and TNF alpha protein production was detected within 60-75 min; however, TNF alpha mRNA and protein production were not detected in diluent-treated hearts. Immunohistochemical studies localized TNF alpha to endothelial cells, smooth muscle cells, and cardiac myocytes in the endotoxin-treated hearts, whereas TNF alpha immunostaining was absent in the diluent-treated hearts. To determine whether the cardiac myocyte was a source for TNF alpha production, two studies were performed. First, in situ hybridization studies, using highly specific biotinylated probes, demonstrated TNF alpha mRNA in cardiac myocytes from endotoxin-stimulated hearts; in contrast, TNF alpha mRNA was not expressed in myocytes from diluent-treated hearts. Second, TNF alpha protein production was observed when cultured cardiac myocytes were stimulated with endotoxin, whereas TNF alpha protein production was not detected in the diluent-treated cells. The functional significance of the intramyocardial production of TNF alpha was determined by examining cell motion in isolated cardiac myocytes treated with superfusates from endotoxin- and diluent-stimulated hearts. These studies showed that cell motion was depressed in myocytes treated with superfusates from the endotoxin-treated hearts, but was normal with the superfusates from the diluent-treated hearts; moreover, the negative inotropic effects of the superfusates from the endotoxin-treated hearts could be abrogated completely by pretreatment with an anti-TNF alpha antibody. Finally, endotoxin stimulation was also shown to result in the intramyocardial production of TNF alpha mRNA and protein in vivo. Thus, this study shows for the first time that the adult mammalian myocardium synthesizes biologically active TNF alpha.

Determination of the catalytic site of creatine kinase by site-directed mutagenesis

Site-directed mutagenesis was used to alter the amino-acid residues at the presumed catalytic site Cys-283 and ATP binding site Asp-340 of human creatine kinase B cDNA. In addition, a highly conserved arginine residue, Arg-292, was also mutated. Transfection of 0.1 to 1 microgram of recombinant plasmid into COS cells produced increasing creatine kinase activity in the cell lysate. The expression of mutant Cys283-Tyr and Cys283-Ser resulted in complete abolition of homodimer BB isoform enzymatic activity without alteration of the capacity for dimerization. Expression of mutants Arg292-His, Arg292-Leu, and Arg292-Gln produced non-functional homodimers, whereas expression of mutant Arg292-Lys produced a homodimer with enzymatic activity that was 42% of the enzymatic activity of the wild type. Expression of the Asp340-Glu mutant creatine kinase did not alter enzyme activity as compared to the wild type. Following heterodimerization, there was inhibition of the normal subunit by the mutant subunit, for both the BB and the MB dimer. The results showed residues Cys-283 and Arg-292 are essential for enzyme catalysis. The best fit model for the dimer is one in which there is close apposition of the two catalytic sites. The interaction of the individual subunits during dimerization provides a molecular approach for dominant negative modulation of the creatine kinase isozyme system in future genetic manipulative experiments.

Impact of Alpha 1-Adrenergic Antagonist Use for Benign Prostatic Hypertrophy on Outcomes in Patients With Heart Failure

Previous clinical trials have shown that alpha(1)-adrenergic antagonists are not effective in subjects with heart failure (HF) and might increase HF rates when used for hypertension. However, alpha(1)-adrenergic antagonists may be prescribed to subjects with HF who have symptomatic benign prostatic hyperplasia. We sought to determine any association between alpha(1)-adrenergic antagonist use, commonly prescribed for benign prostatic hyperplasia, and the clinical outcomes of subjects with HF receiving contemporary therapy. An existing database of 388 subjects with decompensated HF admissions from 2002 to 2004 at the Veterans Affairs Hospital was analyzed according to the use of alpha(1)-adrenergic antagonists at discharge. Covariate-adjusted Cox proportional hazard models were used to examine any association with future admissions for decompensated HF and total mortality. Alpha-1-adrenergic antagonist therapy was prescribed in 25% of our HF population, predominantly for benign prostatic hyperplasia, and was not associated with significant increases in the combined risk of all-cause mortality and rehospitalization for HF (hazard ratio 1.24, 95% confidence interval 0.93 to 1.65, p = 0.14), HF hospitalization (hazard ratio 1.20, 95% confidence interval 0.85 to 1.70, p = 0.31), or all-cause mortality (hazard ratio 1.10, 95% confidence interval 0.78 to 1.56, p = 0.57). In patients not receiving beta-blocker therapy, alpha(1)-adrenergic antagonist therapy was significantly associated with increased HF hospitalizations (hazard ratio 1.94, 95% confidence interval 1.14 to 3.32, p = 0.015). In conclusion, in patients with chronic HF, the use of alpha(1)-adrenergic antagonists was significantly associated with more HF hospitalizations when prescribed without concomitant beta blockade. Thus, background beta-blocker therapy appears to be protective against the potential harmful effects of alpha(1)-adrenergic antagonist therapy in patients with HF.

Modeling left ventricular diastolic dysfunction: classification and key indicators

BackgroundMathematical modeling can be employed to overcome the practical difficulty of isolating the mechanisms responsible for clinical heart failure in the setting of normal left ventricular ejection fraction (HFNEF). In a human cardiovascular respiratory system (H-CRS) model we introduce three cases of left ventricular diastolic dysfunction (LVDD): (1) impaired left ventricular active relaxation (IR-type); (2) increased passive stiffness (restrictive or R-type); and (3) the combination of both (pseudo-normal or PN-type), to produce HFNEF. The effects of increasing systolic contractility are also considered. Model results showing ensuing heart failure and mechanisms involved are reported.MethodsWe employ our previously described H-CRS model with modified pulmonary compliances to better mimic normal pulmonary blood distribution. IR-type is modeled by changing the activation function of the left ventricle (LV), and R-type by increasing diastolic stiffness of the LV wall and septum. A 5th-order Cash-Karp Runge-Kutta numerical integration method solves the model differential equations.ResultsIR-type and R-type decrease LV stroke volume, cardiac output, ejection fraction (EF), and mean systemic arterial pressure. Heart rate, pulmonary pressures, pulmonary volumes, and pulmonary and systemic arterial-venous O2 and CO2 differences increase. IR-type decreases, but R-type increases the mitral E/A ratio. PN-type produces the well-described, pseudo-normal mitral inflow pattern. All three types of LVDD reduce right ventricular (RV) and LV EF, but the latter remains normal or near normal. Simulations show reduced EF is partly restored by an accompanying increase in systolic stiffness, a compensatory mechanism that may lead clinicians to miss the presence of HF if they only consider LVEF and other indices of LV function. Simulations using the H-CRS model indicate that changes in RV function might well be diagnostic. This study also highlights the importance of septal mechanics in LVDD.ConclusionThe model demonstrates that abnormal LV diastolic performance alone can result in decreased LV and RV systolic performance, not previously appreciated, and contribute to the clinical syndrome of HF. Furthermore, alterations of RV diastolic performance are present and may be a hallmark of LV diastolic parameter changes that can be used for better clinical recognition of LV diastolic heart disease.

Using a human cardiovascular-respiratory model to characterize cardiac tamponade and pulsus paradoxus

BackgroundCardiac tamponade is a condition whereby fluid accumulation in the pericardial sac surrounding the heart causes elevation and equilibration of pericardial and cardiac chamber pressures, reduced cardiac output, changes in hemodynamics, partial chamber collapse, pulsus paradoxus, and arterio-venous acid-base disparity. Our large-scale model of the human cardiovascular-respiratory system (H-CRS) is employed to study mechanisms underlying cardiac tamponade and pulsus paradoxus. The model integrates hemodynamics, whole-body gas exchange, and autonomic nervous system control to simulate pressure, volume, and blood flow.MethodsWe integrate a new pericardial model into our previously developed H-CRS model based on a fit to patient pressure data. Virtual experiments are designed to simulate pericardial effusion and study mechanisms of pulsus paradoxus, focusing particularly on the role of the interventricular septum. Model differential equations programmed in C are solved using a 5th-order Runge-Kutta numerical integration scheme. MATLAB is employed for waveform analysis.ResultsThe H-CRS model simulates hemodynamic and respiratory changes associated with tamponade clinically. Our model predicts effects of effusion-generated pericardial constraint on chamber and septal mechanics, such as altered right atrial filling, delayed leftward septal motion, and prolonged left ventricular pre-ejection period, causing atrioventricular interaction and ventricular desynchronization. We demonstrate pericardial constraint to markedly accentuate normal ventricular interactions associated with respiratory effort, which we show to be the distinct mechanisms of pulsus paradoxus, namely, series and parallel ventricular interaction. Series ventricular interaction represents respiratory variation in right ventricular stroke volume carried over to the left ventricle via the pulmonary vasculature, whereas parallel interaction (via the septum and pericardium) is a result of competition for fixed filling space. We find that simulating active septal contraction is important in modeling ventricular interaction. The model predicts increased arterio-venous CO2 due to hypoperfusion, and we explore implications of respiratory pattern in tamponade.ConclusionOur modeling study of cardiac tamponade dissects the roles played by septal motion, atrioventricular and right-left ventricular interactions, pulmonary blood pooling, and the depth of respiration. The study fully describes the physiological basis of pulsus paradoxus. Our detailed analysis provides biophysically-based insights helpful for future experimental and clinical study of cardiac tamponade and related pericardial diseases.

Serial Alu sequence transposition interrupting a human B creatine kinase pseudogene

We have isolated, sequenced, and characterized a single-copy B creatine kinase pseudogene. The chromosomal assignment of this gene is 16p13 and a unique sequence probe from this locus detects EcoRI restriction fragment length polymorphisms of 7.8 and 5.4 kb. In 26 unrelated individuals, the frequencies for the 7.8- and 5.4-kb B creatine kinase pseudogene alleles were calculated to be 17.3 and 82.7%, respectively. The B creatine kinase pseudogene is interrupted by a 904-bp DNA insertion composed of three Alu repeat sequences in tandem flanked by an 18-bp direct repeat, derived from the pseudogene sequence. Nucleotide sequence analysis of the Alu elements suggests that the Alu sequences were incorporated into this locus in three separate integration events. Several complex clustered Alu repeat sequences without defined integration borders have been previously identified at different genomic loci. This is the first evidence that complex tandem Alu elements can integrate in an apparently serial manner in the human genome and supports the contention that Alu repeats integrate nonrandomly into the human genome.

Modeling study of the failing heart and its interaction with an implantable rotary blood pump

The effectiveness of clinical diagnosis and treatment of heart failure is a direct function of clinical signs that can be measured in a patient within cost and safety constraints. Large-scale mathematical modeling can be a key tool in revealing important, measurable clinical signs of heart failure, furthering medical understanding and development of treatment. In the first part of this study we have created two models of left heart failure — diastolic and systolic, using our human cardiovascular-respiratory system (H-CRS) model, and we present a comparison of the two types with emphasis on novel and differentiating clinical signs, such as tricuspid flow and septal motion. In the event of compromised left ventricular performance, mechanical left ventricular assist devices (LVAD) are often implanted to augment or completely replace the pumping action of the left ventricle (LV). One such type is the implantable rotary blood pump (iRBP). Several design issues related to the iRBP are difficult to study experimentally due to procedure complexity and limitations in animal models of heart failure [2]. Therefore, modeling has become a key tool in iRBP development. In the second part of this study, we have introduced an iRBP model based on [1]–[2] in the systolic failing heart to study the interactions. We consider optimal motor settings for different levels of LV assistance, the effects of the iRBP on the right heart, septum, and pulmonary circulation. Our model results align with those reported in [1]–[2]. Improvement in cardiac output, pulmonary congestion, and heart work are seen with the iRBP. We observe lowered septal assistance to RV and LV ejection with increasing pump speeds, elevating right ventricular (RV) work, reducing LVET, and causing ventricular mechanical dyssynchrony in ejection. These results suggest right heart compromise via the septums reduced role with the introduction of an iRBP. This work emphasizes the critical role of modeling in heart failure and treatment studies.

Herpes Zoster and Its Cardiovascular Complications in the Elderly – Another Look at a Dormant Virus

Herpes zoster (shingles) is a reactivation of latent Varicella-zoster virus (VZV). We present a case of pleuropericarditis simulating acute myocardial infarction and another with complete heart block in the setting of acute/recent VZV reactivation. These cases are consistent with a modified concept: (1) the VZV dormancy is comprised of multiple foci of infections in the sensory and autonomic ganglia, and (2) the VZV reactivation could involve co-incident activations of two or more loci. Recognition of this possibility of cardiovascular complications of VZV should be helpful in the clinical management of the elderly, in the differential diagnosis of chest pain, ST elevation, and heart block etiology in the setting of acute or recent VZV reactivation.

Creatine Phosphate Shuttle Pathway in Tissues with Dynamic Energy Demand

This chapter discusses the creatine phosphate shuttle pathway in tissues with dynamic energy demand. All cells use adenosine tri phosphate (ATP) as the immediate energy source. Cellular ATP is generated by oxidative metabolism and by glycolysis. Many specialized cells in an organism require fast and constant energy utilization to maintain specialized cellular function. It is evident that many cells with constant and rapid energy demand have a compartmentalized CK system. New recombinant deoxyribonucliec acid (DNA) methodology such as “site-directed mutagenesis” facilitates the dissection of the creatine kinase (CK) molecule and demonstrates the molecular mechanism through which the CKs catalyze the enzymatic reaction as well as delineate the various features of the enzyme molecule which targets it to regions of diverse cellular ATPases. Future technologies such as “gene targeting,” “transgenic models,” and “dominant negative modulation” provide the opportunity to alter the cellular CK level at selected tissue sites and at selected developmental or experimental time points. Understanding the contribution of the creatine phosphate shuttle pathway in cellular homeostasis under conditions of stress, such as decreased energy supply or increased energy demand provides new insights and approaches for the management of diverse disease states.

Cardiovascular Interactions Tutorial: An Update

The goal of the CVI (Cardiovascular Interaction) project was to develop a computerized tutorial to allow learners (users) to investigate the integration and control of the cardiovascular system. Recent publications in Advances in Physiology Education are concluding that new methods for teaching physiology are more effective than old ones if they get students actively involved [1]. Rothe’s models and the Lab Book can be used either for classroom discussions or for self instruction. Both ways of using his models can get students actively involved. This goal has not changed; however, it was necessary to update the implementation to continue to attain this goal. The original five-compartment model, discussed in references [2, 3], was a two-part implementation consisting of a Lab Book in the form of a tutorial as a Microsoft Word document that drives the cardiovascular simulation model. The six-compartment model had a similar structure. Both models were implemented in Visual Basic. Due to changes in the Windows operating system, the interactive tutorial is no longer functional. The models, however, are fully functional and can stand alone. A recent effort was undertaken to reimplement the cardiovascular simulation models so they can be run on the current operating systems. (The Lab Book can still be followed, and the learner still gets the benefit of that process.) While updating the implementation of the models, it was discovered that information on the six-compartment model had never appeared in the literature. The focus of what follows is the six-compartment model.