Construction of adenoviral vectors.

Recombinant adenovirus vectors have proven to be useful tools in facilitating gene transfer. Construction of such vectors requires a knowledge of the adenovirus genome structure and its life cycle. A commonly used recombinant adenovirus involves deletion of the E1 region; such a recombinant is traditionally produced by overlap recombination after cotransfection of 293 cells with a plasmid shuttle vector and a large right-end restriction fragment of viral DNA. The shuttle vector contains a cassette for a transgene placed in region E1 and flanking sequences from adenovirus for recombination. Normally, a high background of parental virus results because of the difficulty in separating right-end restriction fragment length DNA from uncut DNA. This paper describes a negative selection based on the traditional cotransfection method using viral DNA from an E1-deleted adenoviral recombinant that expresses green fluorescent protein (GFP). In situ fluorescent microscopy is used to distinguish the recombinant plaques (white or nonfluorescent) from the parental virus plaques (green or fluorescent). In addition, this system allows for the detection of contaminating parental virus at later stages when production lots of the recombinant vector are being made.

The left ventricular ejection effect.

A recently developed model of the left ventricle, based on experimental data, has been shown to exhibit the main features of the heart's ability to pump. Two special cases during blood ejection, termed pressure deactivation and hyperactivation, were identified. This study proposes an 'ejection effect' correction to the model that addresses deactivation, hyperactivation and adjusts the shape of the computed ventricular ejection curve in late systole. Also, a new approach based on new animal experiments is proposed to identify the ejection effect mechanism(s).

Human circulatory system model based on Frank's mechanism.

A new analytical model of the left ventricle as a pump, developed from isolated canine experiments, was adapted to describe each of the four heart chambers in a complete human circulatory system model. Each chamber is embodied as a volume and time dependent isovolumic pressure source, after Otto Frank's classic experiments. Analytical results show that a small set of equations is sufficient to describe the main features of the heart as a pump, including isovolumic and ejecting beats for a wide range of ventricular and circulation conditions. This model allows interactive teaching of cardiovascular system dynamics. Computed results demonstrate that, with additional experiments, a quantitative description of the human circulation for research purposes may emerge from this approach.

The changing view of the heart through the centuries.

In human perception, the heart was not always part of the blood circulating system. It was later included as a suction pump until Harvey argued that the heart is actually a compression pump, the central organ of the circulation, and the only organ responsible for the motion of blood. Considered initially as an autonomous pump, the heart gradually became viewed as subservient to the needs of the peripheral organs it perfuses. Constant properties assigned to the heart had to be replaced, one after another, by adjustable parameters. Even the adequacy of the heart as the sole pump has been doubted, an issue that resurfaces today.

A perspective on myocardial contractility.

Myocardial contractile properties form the cornerstone of the heart's ability to pump blood. Efforts have been made to characterize these properties via classic elasticity theory concepts, which can lead to spurious results, as demonstrated by experiments measuring intramyocardial pressure. Two ways out of these difficulties are identified. One is to start at the cellular level, the other at the chamber level. The latter allows separation of ventricle (source) and arterial (load) effects on measured pressure and flow, distinct from previous definitions of ventricular contractility which tended to lump the two.

Automated in-vivo measurement of quasi-static lung compliance in the rat.

Instrumentation to automate quasi-static lung compliance measurement in the rat was developed and values obtained with it were compared with manual measurements by a trained technician. Designed to be used during mechanical ventilation, this system interrupts ventilation to inflate and deflate the lungs and measures lung transmural pressure and volume as functions of time. Animal experiments demonstrated that the automated system is capable of generating correctly shaped volume-pressure curves. These curves yielded reproducible lung compliance values that compare favorably with those obtained by the manual method. No statistically significant difference was observed comparing the two methods when evaluating either inter- or intra-animal variation. This automated system thereby obviates the need for highly trained personnel to perform the test.

Thresholds for premature ventricular contractions in frog hearts exposed to lithotripter fields.

Piezoelectrically generated lithotripter shocks were shown to produce premature ventricular contractions of the frog heart. Anesthetized grass frogs, Rana pipiens, were studied following implantation of an aortic catheter and EKG leads. The most sensitive phase of the heart cycle for the generation of premature ventricular contractions with lithotripter shocks at 30 MPa peak pressure was found to be the T-P segment. During this phase of the heart cycle, the minimum peak-positive pressure shock wave necessary to produce a premature ventricular contraction in a frog heart was between 5 MPa and 10 MPa.