Analyzing red blood cell-deformability distributions.

BACKGROUND: Red blood cells (RBCs) must deform to pass the smallest capillaries of the microcirculation. Available techniques for measuring RBC deformability often provide an indication of the mean deformability. The latter may be decreased either by a slight overall deformability reduction or by the presence of a small fraction of rigid cells. A distinction between these two cases can be made with a RBC-deformability distribution (RBC-DD). METHODS: This paper explores RBC-DDs of healthy individuals and of cells with anomalous mechanical properties (sickle cell disease, dialysis patients, elliptocytosis, and cultivated malaria tropica). The distributions were measured with an automated rheoscope, which uses advanced image analysis techniques to obtain the deformability index of a large number of individual cells subjected to simple shear flow. RESULTS: The RBC-DD of healthy volunteers is close to a normal distribution. In the investigated patients, distributions were markedly different and yielded significant changes in the mean, in the standard deviation, or in both. The presence of hypodeformable and hyperdeformable cell fractions can qualitatively and quantitatively be assessed from the deformability distribution (DD). In elliptocytosis, cells orient differently with respect to the streamlines, compared to normal cells. This causes the DD to be biased. CONCLUSIONS: The RBC-DD is a powerful representation to establish subpopulations with anomalous deformability. Fractions of hypodeformable and hyperdeformable cells and the standard deviation of the DD are new and excellent quantitative parameters to assess alterations in RBC deformability.

Conversion of left ventricular endocardial positions from patient-independent co-ordinates into biplane fluoroscopic projections.

Electrocardiographic body surface mapping is used clinically to guide catheter ablation of cardiac arrhythmias by providing an estimate of the site of origin of an arrhythmia. The localisation methods used in our group produce results in left-ventricular cylinder co-ordinates (LVCCs), which are patient-independent but hard to interpret during catheterisation in the electrophysiology laboratory. It is preferable to provide these results as three-dimensional (3D) co-ordinates which can be presented as projections in the biplane fluoroscopic views that are used routinely to monitor the catheter position. Investigations were carried out into how well LVCCs can be converted into fluoroscopic projections with the limited anatomical data available in contemporary clinical practice. Endocardial surfaces from magnetic resonance imaging (MRI) scans of 24 healthy volunteers were used to create an appropriate model of the left-ventricular endocardial wall. Methods for estimation of model parameters from biplane fluoroscopic images were evaluated using simulated biplane data created from these surfaces. In addition, the conversion method was evaluated, using 107 catheter positions obtained from eight patients, by computing LVCCs from biplane fluoroscopic images and reconstructing the 3D positions using the model. The median 3D distance between reconstructed positions and measured positions was 4.3mm.

ECG of the "newborn" mouse (Mus domesticus) with specific reference to comparative AV transmission.

INTRODUCTION: The objective of this study was to record the ECG of the smallest living mammal to extend the domain of data for comparative AV nodal electrophysiologic purposes. These data are needed to establish the relationship between the PR interval and heart size in mammalian species of all sizes. METHODS AND RESULTS: In recently born mice (age 1.5 to 8 weeks) weighing between 2.5 and 10 g and with estimated heart weights between 15 and 60 mg, ECGs, using bipolar limb leads, were recorded during general anesthesia. The PR interval, representing AV transmission time was about 40 msec, which is quite long for hearts of this size. On the basis of detailed analysis of the data, we postulate the presence of a fixed delay or discontinuous propagation in the AV node not only in newborn mice, but in mammals of all sizes. CONCLUSION: AV transmission times obtained in mammals (including humans) cannot be explained on the basis of generally accepted, classic AV conduction theories. The acceptance of the presence of a fixed delay in the AV node may ultimately be of value to better understand AV node function during sinus rhythm and supraventricular arrhythmias.

Dynamic behavior of DNA replication domains.

Like many nuclear processes, DNA replication takes place in distinct domains that are scattered throughout the S-phase nucleus. Recently we have developed a fluorescent double-labeling procedure that allows us to visualize nascent DNA simultaneously with "newborn" DNA that had replicated earlier in the same nucleus during the same S-phase. Using this procedure we have shown that all DNA in a replication domain is replicated within 1 h (Manders et al., 1992, J. Cell Sci. 103, 857-862). Here we extend these studies by analyzing the behavior of replication domains on a time scale of less than 1 h. We have carried out a series of double-labeling experiments in which we varied the time interval between nascent DNA and newborn DNA from 0 to 60 min. Subsequently, we determined from the confocal, 3D images the spatial position of replicated DNA domains and identified pairs of nearest neighbor domains containing newborn and nascent DNA, respectively. The distance between the centers of the two domains in a pair gradually increases. Accurate measurements show that domains containing nascent DNA and domains containing newborn DNA gradually separate from each other at a rate that is on the order of 0.5 micron/h. This indicates that either newly synthesized DNA moves away from sites of replication activity or the replication machinery is moving itself. This rate is essentially the same during early and late S-phase.

Largest contour segmentation: a tool for the localization of spots in confocal images.

An accurate determination of the 3-D positions of multiple spots in images obtained by confocal microscopy is essential for the investigation of the spatial distribution of specific components or processes in biological specimens. The position of the centroid, as an estimator for the position of a spot, can be calculated on the basis of all voxels that belong to the domain of the spot. For this calculation a domain that defines which voxels belong to the spot must be delimited. To create a boundary for a domain we developed a 3-D image segmentation procedure: the largest contour segmentation (LCS). This procedure is based on an iterative region-growing procedure around each local maximum of intensity. By means of this procedure the position of each spot was determined accurately and automatically. Qualities of the procedure were evaluated by means of simulated test-images as well as 3-D images of real biological specimens.

Effect of mains interference on time coherent averaging.

The paper deals with the influence of mains interference on time coherent averaging using an internal trigger. The general distribution of the time jitter is derived and applied to a simplified signal. The authors prove that no amount of averaging can do away with this type of interference. Finally, the deformation this type of jitter has on a step signal is computed.

Method for the computation of an accurate zero reference for ECG signals.

When the instantaneous magnitude of an electrical signal is to be determined, a reference has to be known against which the signal value should be measured. In electrocardiology, this reference is commonly derived from the isoelectrical period in the ECG. However, this period is not a clearly defined part of the ECG, and its use can be problematic. The signal level obtained from it is often not sufficiently unambiguous and in the case of a high heart rate this period is too short to be used or not present at all. In the paper a method is developed to compute a reference value, independent of the isoelectrical period and of heart rate. The method is derived from a method published before, which we developed for the separation of the surface His-Purkinje signal from the P-wave in the case of overlap. In the present application of the method, two successive heartbeats are separated in an analogous manner, revealing the signal level at zero heart activity. In applications described in the paper, this signal level is used as zero reference in nonlinear techniques. The method proved to work well and accurately.

A method for the measurement of field placement errors in digital portal images.

Correct placement of radiation fields relative to patient anatomy is essential in radiotherapy in order to minimise serious side effects to reduce the probability of recurrence of the tumour. One way to determine patient setup accuracy is to analyse portal images obtained in the therapy beam distal to the patient. A field placement analysis (FPA) method has been developed for detailed evaluation of patient setup by comparing positions of corresponding radiation field edges in digitised simulator and portal images. A simulator image is matched to a portal image using similar anatomical landmarks in both images and mapping these landmarks against each other applying a least squares minimisation approach. Discrepancies between the simulator field edge (reference) and a portal field edge are determined by comparing the distances between the central axis of the beam and corresponding edge segments and the angles of these segments with a reference line. Uncertainties in these distances and angles are to a large extent determined by the magnification, rotation and translation procedure. Uncertainties due to the FPA method itself are of about 1.0 mm and 0.5 degrees in portal images of head and neck fields. These FPA uncertainties are in general smaller than the variations due to patient setups. Matching of simulator and portal images of lateral pelvic fields revealed larger uncertainties: 1.7 mm and 1.1 degrees. Setup variations in this kind of pelvic radiation field are usually also larger, and therefore meaningful results can be obtained with the new FPA method.

Determinants of the occurrence of vortex rings in the left ventricle during diastole.

This study employs classical inviscid fluid dynamics theory to investigate whether LV diastolic inflow volume and the size of the LV play a role in vortex ring formation. Fluid injection across an orifice into a large container results in the generation of a vortex ring having a constant size and speed. Relations between the vortex size and speed and the injection were obtained by applying conservation laws regarding kinetic energy, impulse and vorticity; the initial state was computed using a bolus injection model, and the final state by using the Kelvin vortex model. An important parameter in the equations is the relative injection length, i.e., the ratio of the length of the injected bolus and the radius of the orifice (L/R). Its estimated highest value in man, L/R = 15, produces a rather thick vortex ring (relative thickness 0.77). Comparable results following from the Hill vortex model convinced us that the Kelvin vortex model can be applied in the whole range of injection lengths in the human left ventricle. In an in vitro model it is shown experimentally that vortex rings can be generated for L/R in the range from 2 to 16. The measured traveling speed of the vortex ring is in fair agreement with the theory, as well as the ring radius for large injections. A vortex ring located in a narrow channel cannot reach its proper traveling speed. The method of images is used to estimate the speed reduction of vortex rings within a cylinder. It turns out that propagation of vortex rings is possible when the ratio of orifice to cylinder radius is less than about 0.5.(ABSTRACT TRUNCATED AT 250 WORDS)

Deformation-corrected computer-aided three-dimensional reconstruction of immunohistochemically stained organs: application to the rat heart during early organogenesis.

The application of a computer-assisted, three-dimensional reconstruction procedure for serial sections to embryonic rat hearts during the period of cardiac looping and compartmentalization is described. The procedure relies on immunohistochemical staining for the introduction of selective contrast and on episcopic and diascopic images of each of the sections for alignment and correction of compression due to sectioning. Episcopic (reference) images are taken from the embedding block just before the cutting of a slice and are still aligned and undeformed. Diascopic images are taken from the sections after immunohistochemical processing and, hence, contain selective contrast but are deformed and no longer aligned. The three-dimensional images are visualized as shaded voxel models. This approach allowed the unequivocal delineation of the developing myocardium and the inspection of its changing architecture both from the outside and from within. Furthermore, it allowed a quantification of myocardial volume. Because standardized and hence comparable views of three different stages were generated, changes in the shape of the cardiac loop, the atria, and the ventricles as well as changes in the position of the atrioventricular canal and interventricular foramen could be accurately described. Characteristic changes in the position of both the right ventricle and the atrioventricular canal that are essential for the formation of a correctly functioning four-chambered heart could be observed. These changes in shape occur while the myocardial size increases dramatically.

Circadian rhythm of proteinuria: consequences of the use of urinary protein:creatinine ratios.

Proteinuria in patients with glomerular disease has a circadian rhythm, but for creatinine such a rhythm is either absent or of low amplitude. We found in 18 of 23 admitted patients (group I) and in seven outpatients (group II) a marked circadian rhythm of the protein: creatinine ratio. Estimates of 24-h proteinuria were obtained by multiplying the protein:creatinine ratio of 3-h urine samples with 24-h creatinine excretion, calculated from age, sex and bodyweight. Estimated proteinuria could be as low as 19% or as high as 349% of actually measured 24-h proteinuria; the mean SD was 23%. The best estimate was obtained with the 06.00-09.00 hours urine samples. The estimates correlated better with actually measured 24-h proteinuria than the protein: creatinine ratio per se correlated with the 24-h proteinuria. Day-to-day variation of proteinuria estimates from samples taken at the same time of the day was of similar magnitude as day-to-day variation of actual 24-h proteinuria. We conclude that the usefulness of the protein: creatinine ratio of a random urine sample for estimation of proteinuria is limited, because of the circadian rhythm of proteinuria. However, samples collected at a fixed time of the day are an acceptable alternative for 24-h urine collections in the clinical follow-up of individual patients.

Singular value decomposition for identification of parameters in multicompartment analysis.

Standard singular value decomposition is used in a practical approach to parameter identification in multicompartment analysis; a solution can be found even in not well-defined situations. The method is demonstrated on a simple 2-compartment glucose model and applied to an extended 7-compartment model.

Hemodynamic fluctuations and baroreflex sensitivity in humans: a beat-to-beat model.

A beat-to-beat model of the cardiovascular system is developed to study the spontaneous short-term variability in arterial blood pressure (BP) and heart rate (HR) data from humans at rest. The model consists of a set of difference equations representing the following mechanisms: 1) control of HR and peripheral resistance by the baroreflex, 2) Windkessel properties of the systemic arterial tree, 3) contractile properties of the myocardium (Starling's law and restitution), and 4) mechanical effects of respiration on BP. The model is tested by comparing power spectra and cross spectra of simulated data from the model with spectra of actual data from resting subjects. To make spectra from simulated data and from actual data tally, it must be assumed that respiratory sinus arrhythmia at rest is caused by the conversion of respiratory BP variability into HR variability by the fast, vagally mediated baroreflex. The so-called 10-s rhythm in HR and BP appears as a resonance phenomenon due to the delay in the sympathetic control loop of the baroreflex. The simulated response of the model to an imposed increase of BP is shown to correspond with the BP and HR response in patients after administration of a BP-increasing drug, such as phenylephrine. It is concluded that the model correctly describes a number of important features of the cardiovascular system. Mathematical properties of the difference-equation model are discussed.

Toward computerized morphometric facilities: a review of 58 software packages for computer-aided three-dimensional reconstruction, quantification, and picture generation from parallel serial sections.

This review gives an inventory of 58 computer-aided three-dimensional reconstruction applications in the domain of biomedical research. It is devoted to the formulation of a set of recommendations thought to be necessary for improved performance of software packages in this field. These recommendations can be used to select packages and to guide future developments of existing reconstruction systems. The survey is restricted to three-dimensional reconstructions based upon a series of parallel sections of an object. Subjects treated are programming languages, resolution and sampling, input preparation, realignment, local deformation of slices, numerical quantifications, topological complexity, internal representation, display complexity (hidden surfaces, shading, smoothing), structure extraction, descriptive elements, database, data compression, time efficiency of systems and algorithms, hardware configuration, input devices, input media, interactive aids, display devices, and output devices. Information for this survey comes from articles that appeared between 1965 and 1985.

A new method for microdensitometer slit length correction of radiographic noise power spectra.

When the power spectrum of radiographic noise is estimated from scans obtained with a microdensitometer equipped with a rectangular slit, a bias is present due to the finite length of this slit. A method to correct for this bias was developed and was tested on both a simulated and a measured spectrum. No bias remains with our method, in situations where a conventional method shows a significantly biased result. Good agreement was found with the synthesized slit method, described by Sandrik and Wagner.

Simple assessment of the efficacy of peritoneal transport in continuous ambulatory peritoneal dialysis patients.

The efficacy of peritoneal transport was assessed in 13 permeability studies in 11 continuous ambulatory peritoneal dialysis (CAPD) patients. During each study the in situ intraperitoneal volume was measured as well as the dialysate and plasma concentrations of various solutes with a molecular weight range from 60 to 5,500. As clearance estimations are unsuitable for the purpose of permeability studies, mass transfer area coefficients were used. By applying a simple mathematical model assuming first-order kinetics, these coefficients were calculated for urea, lactate, creatinine, glucose, kanamycin, and inulin. The accuracy of the calculations is indicated by their r values. After pooling these correlation coefficients, the mean approached 1.00 for all solutes with high confidence limits, indicating the usefulness of the model. A further simplification was tested using only an initial- and end-dialysate sample and two blood samples, without the measurement of the in situ intraperitoneal volume. Except for inulin the results of this simplification correlated well with the results described above. The reproducibility of the simplified mass transfer area coefficient calculations was investigated on 15 occasions in 3 other CAPD patients. The coefficients of variation of low molecular weight solutes varied between 15 and 20%. It is concluded that mass transfer area coefficient estimations using the latter method can be performed in any CAPD patient and probably yield sufficient information to establish the efficacy of the membrane transport mechanism during clinical follow-up.