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Culturing Mouse Cardiac Valves in the Miniature Tissue Culture System

Heart valve disease is a major burden in the Western world and no effective treatment is available. This is mainly due to a lack of knowledge of the ...

New Jersey Institute of Technology

It is well established that the aging heart exhibits left ventricular (LV) diastolic dysfunction and changes in mechanical properties, which are thought to be due to alterations in the extracellular matrix. We tested the hypothesis that the mechanical properties of cardiac myocytes significantly change with aging, which could contribute to the global changes in LV diastolic dysfunction. We used atomic force microscopy (AFM), which determines cellular mechanical property changes at nanoscale resolution in myocytes, from young (4 mo) and old (30 mo) male Fischer 344 x Brown Norway F1 hybrid rats. A measure of stiffness, i.e., apparent elastic modulus, was determined by analyzing the relationship between AFM indentation force and depth with the classical infinitesimal strain theory and by modeling the AFM probe as a blunted conical indenter. This is the first study to demonstrate a significant increase (P < 0.01) in the apparent elastic modulus of single, aging cardiac myocytes (from 35.1 +/- 0.7, n = 53, to 42.5 +/- 1.0 kPa, n = 58), supporting the novel concept that the mechanism mediating LV diastolic dysfunction in aging hearts resides, in part, at the level of the myocyte.

Aging increases stiffness of cardiac myocytes measured by atomic force microscopy nanoindentation.

The objective of this study was to test the hypothesis that the mechanism mediating left ventricular (LV) dysfunction in the aging rat heart involves, in part, changes in cardiac cytoskeletal components. Our results show that there were no significant differences in heart rate, LV pressure, or LV diameter between conscious, instrumented young [5.9 +/- 0.3 mo (n = 9)] and old rats [30.6 +/- 0.1 mo (n = 10)]. However, the first derivative of LV pressure (LV dP/dt) was reduced (8,309 +/- 790 vs. 11,106 +/- 555 mmHg/s, P < 0.05) and isovolumic relaxation time (tau) was increased (8.7 +/- 0.7 vs. 6.3 +/- 0.6 ms, P < 0.05) in old vs. young rats, respectively. The differences in baseline LV function in young and old rats, which were modest, were accentuated after beta-adrenergic receptor stimulation with dobutamine (20 mug/kg), which increased LV dP/dt by 170 +/- 9% in young rats, significantly more (P < 0.05) than observed in old rats (115 +/- 5%). Volume loading in anesthetized rats demonstrated significantly impaired LV compliance in old rats, as measured by the LV end-diastolic pressure and dimension relationship. In old rat hearts, there was a significant (P < 0.05) increase in the percentage of LV collagen (2.4 +/- 0.2 vs. 1.3 +/- 0.2%), alpha-tubulin (92%), and beta-tubulin (2.3-fold), whereas intact desmin decreased by 51%. Thus the cardiomyopathy of aging in old, conscious rats may be due not only to increases in collagen but also to alterations in cytoskeletal proteins.

Cardiac dysfunction in aging conscious rats: altered cardiac cytoskeletal proteins as a potential mechanism.

Respiratory assist using an intravenous catheter may be a potential treatment for patients suffering from acute or acute-on-chronic lung failure. The objective of this study was to evaluate a novel respiratory catheter that uses an impeller within the fiber bundle to enhance gas exchange efficiency, thus requiring a smaller fiber bundle and insertional size (25 Fr) and permitting simple percutaneous insertion. Bench testing of gas exchange in deionized water was used to evaluate eight impeller designs. The three best performing impeller designs were evaluated in acute studies in four calves (122 + or - 10 kg). Gas exchange increased significantly with increasing impeller rotation rate. The degree of enhancement varied with impeller geometry. The maximum gas exchange efficiency (exchange per unit surface area) for the catheter with the best performing impeller was 529 + or - 20 ml CO(2)/min/m(2) and 513 + or - 21 ml CO(2)/min/m(2) for bench and animal studies, respectively, at a rotation rate of 20,000 rpm. Absolute CO(2) exchange was 37 and 36 ml CO(2)/min, respectively. Active mixing by rotating impellers produced 70% higher gas exchange efficiency than pulsating balloon catheters. The sensitivity of gas exchange to impeller design suggests that further improvements can be made by computational fluid dynamics-based optimization of the impeller.

Evaluation of a respiratory assist catheter that uses an impeller within a hollow fiber membrane bundle.

Valvular heart disease is a leading cause of morbidity and mortality in adults but little is known about the underlying etiology. A better understanding of the genetic and hemodynamic mechanisms involved in growth and remodeling of heart valves during physiological and pathological conditions is needed for a better understanding of valvular heart disease. Here, we report the design of a miniature tissue culture system (MTCS) that allows the culture of mitral valves from perinatal to adult mice. The design of the MTCS is novel in that fine positioning and cannulation can be conducted with hearts of different sizes (perinatal to adult). Perfusion of the heart and hence, culture of the mitral valve in its natural position, occurs in a hydraulically sealed culture bath environment. Using the MTCS, we successfully cultured the mitral valve of adult mouse hearts for 3 days. Histological analysis indicated that the cultured valves remained viable and their extracellular matrix organization was similar to age-matched native valves. Gene expression could also be modified in cultured valves by perfusion with medium containing beta-galactosidase-expressing adenovirus. Thus, the MTCS is a new tool to study the genetic and hemodynamic mechanisms underlying the three-dimensional organization of the heart valves, which could provide insights in the pathology of valvular heart disease and be used in animal models for the development of tissue-engineered heart valves.

Samuel C. Lieber

Department of Engineering Technology

New Jersey Institute of Technology

Culturing Mouse Cardiac Valves in the Miniature Tissue Culture System