Mattheus G. Lötter

Kinetics, Distribution and Sites of Destruction of 111Indium‐labelled Human Platelets

The survival, tissue distribution and fate of 111In‐oxine labelled autologous platelets in six normal humans were studied with serial blood sampling, scintillation camera and computer‐assisted imaging, whole body profile scanning, and rectilinear scanning. 111In‐platelets recovery in the circulation was 72±16% and survival was 216±17 h. Platelet survival curves fitted a linear function best. Initially platelets pooled rapidly in the spleen as a single exponential function, and at 90 min 26% of the injected 111In was located in this organ. Early hepatic uptake was also significant and at 90 min constituted 16% of total body 111In‐activity. As labelled platelets disappeared from the circulation there was a threefold increase of radioactivity in the liver to reach 39% of whole body activity at 216 h. Radioactivity also increased significantly in the spleen (33±3% at 216 h). There was significant residual radioactivity in the thoracic and lower abdominal regions at 216 h, suggesting that platelets are also sequestered in the bone marrow. Radioactivity in the lower limbs almost disappeared with time (0±7% at 216 h), indicating that utilization of platelets in the peripheral vasculature is not marked in normal subjects.

Chelator effect on ion diffusion in ferrous-sulfate-doped gelatin gel dosimeters as analyzed by MRI.

Ferrous-sulfate-doped gelatin gel dosimeters are useful tools for the measurement of three-dimensional absorbed radiation dose distributions. The diffusion of ferric ions through these gels causes degradation with time of the dose distribution image. It would be useful to reduce ferric ion diffusion without decreasing gel sensitivity. The amount of ferric ion diffusion is a function of the time delay after radiation, the gel temperature, and the gel concentration. These effects can be quantified by measuring the ferric ion diffusion coefficient. Determination of the diffusion coefficient by irradiating the lower section of a cylinder of gel, which was then imaged repeatedly over time with a clinical magnetic resonance imager, is described. Analysis of the edge spread function formed at each of several times after irradiation by drawing a profile over the imaged junction between the irradiated and unirradiated halves of the cylinder, gave estimates of the variance of the edge spread function. These variances were used to obtain an estimate of the ferric ion diffusion coefficient for the gel. A method of reducing ferric ion diffusion by adding a chelator and the cross linkage agent formaldehyde is suggested. The chelators investigated were 1,10 phenanthroline, xylenol orange, and bathophenanthroline disulfonic acid. These reduced diffusion to varying extents, and influenced the gel sensitivity. The diffusion coefficient in gels containing xylenol orange was found to be 0.44 mm2h-1. The gel sensitivity was 0.0093 s-1Gy-1. This compared with a diffusion coefficient of 0.82 mm2h-1 for the base line gel that did not contain formaldehyde or chelators. The sensitivity of this base line gel was 0.0129 s-1Gy-1. The addition of xylenol orange produced the most improved gel dosimeter of the gels studied. This gel had a decreased ferric ion diffusion coefficient and a decreased sensitivity. It was still sensitive enough to be useful.

A computer program in compiled basic for the IBM personal computer to calculate the mean platelet survival time with the multiple-hit and weighted mean methods

We developed an easy-to-operate computer program for the IBM personal computer to calculate, display and store in a database platelet kinetic data determined by analysis of the rate of clearance of radiolabeled blood platelets from the circulation. This was done by curve fitting using the weighted mean method and multiple-hit model. These models are complementary and calculating the mean platelet survival time with both is recommended. Improvement of the weighted mean method was investigated. The optimized weighting and fitting the exponential function with the Marquardt non-linear least squares method improved the weighted mean method. The weighted mean and multiple-hit models fit the survival curve data equally well. The calculation of the mean platelet survival time with the weighted mean method was very fast. The duration of calculation with the multiple-hit model could take up to 2 minutes. Calculation of the mean platelet survival time using both models has the advantage that conditions when calculation of the mean platelet survival time would be invalid, can be detected. The computer program will promote the valid comparison of results obtained at different institutions.

Influence of Platelet Membrane Sialic Acid and Platelet Associated IgG on Ageing and Sequestration of Blood Platelets in Baboons

Platelets were isolated from blood of baboons and treated with neuraminidase to remove platelet membrane sialic acid, a process which artificially ages the platelets. The platelets were then labelled with 111In and their mean life span, in vivo distribution and sites of sequestration were measured. The effect of removal of sialic acid on the attachment of immunoglobulin to platelets were investigated and related to the sequestration of the platelets by the spleen, liver, and bone marrow. Removal of sialic acid by neuraminidase did not affect the aggregation of platelets by agonists in vitro, nor their sites of sequestration. The removal of 0.51 (median, range 0.01 to 2.10) nmol sialic acid/10(8) platelets shortened their life span by 75 h (median, range 0 to 132) h (n = 19, p < 0.001), and there was an exponential correlation between the shortening of the mean platelet life span and the amount of sialic acid removed. The increase in platelet-associated IgG was 0.112 (median, range 0.007 to 0.309) fg/platelet (n = 25, p < 0.001) after 0.79 (median, range 0.00 to 6.70) nmol sialic acid/10(8) platelets was removed (p < 0.001). There was an exponential correlation between the shortening of mean platelet life span after the removal of sialic acid and the increase in platelet-associated IgG. The results suggest that platelet membrane sialic acid influences ageing of circulating platelets, and that the loss of sialic acid may have exposed a senescent cell antigen that binds IgG on the platelet membrane.(ABSTRACT TRUNCATED AT 250 WORDS)

An improved method for the quantification of the in vivo kinetics of a representative population of 111In-labelled human platelets

The recommended and commonly used methods for the isolation of platelets from whole blood do not harvest a representative platelet population. There is evidence that these methods may result in the loss of a functionally more active platelet subpopulation. We describe a method whereby a completely representative population of platelets was isolated from the whole blood of 28 normal human volunteers by repeated washing of platelets from the red-cell layer. The harvesting efficiency was 98.3%±2.8%. The platelets were labelled with 111In-oxine in a saline milieu with a labelling efficiency of 86.4%±6.8%. The disappearance of reinjected labelled autologous platelets from the circulation was almost linear, and the mean platelet survival was estimated to be 224±23 h. At equilibrium, 61%±12% of the labelled platelets were recovered from the circulation. The in vivo distribution at equilibrium and the sites of sequestration of the senescent labelled platelets were determined by geometric-mean whole-body quantification in six of the volunteers. This improved method permits accurate quantification of organ 111In radioactivity. Following reinjection, the labelled platelets pooled in the spleen and the accumulated activity can be presented by a single exponential function. At equilibrium, 31.1%±6.1% and 9.6%±1.2% of the platelets were in the spleen and liver, respectively. Splenic and hepatic radioactivity increased significantly with time, and at the end of the platelet life span, 35.6%±9.7% and 28.7%±8.3% of the labelled platelets were sequestrated in these organs, respectively. The 30.3%±7.8% remaining platelets were probably sequestrated mainly in the reticuloendothelial component of the bone marrow and other tissues. These techniques of platelet labelling and measurement of the in vivo distribution of 111In-labelled platelets are relatively simply and accurate methods for the study of platelet kinetics in man.

Radiation dose from human platelets labelled with indium 111

The biological distribution of 111In-labelled platelets in normal subjects was determined by whole-body counting and scintillation-camera computer-assisted imaging. Using these data, organ radiation dose was quantitated. The highest radiation dose of 7.4 mGy/MBq (27.4 rad/mCi) was received by the spleen and 0.97 mGy/MBq (3.6 rad/mCi) by the liver. While body radiation dose was 0.25 mGy/MBq (0.9 rad/mCi). The gonad radiation dose of males was 0.14 mGy/MBq (0.5 rad/mCi) and that of females 0.22 mGy/MBq (0.8 rad/mCi). These estimates indicate that radiation doses received from 8.6 MBq of 111In-labelled platelets are well within acceptable limits, and that 111In is a safe labelling agent for the study of platelet kinetics.

An evaluation of four methods of 111In planar image quantification

The accurate quantification of the in vivo distribution of 111In labeled platelets, other cells, and proteins with a scintillation camera is important in clinical and experimental medicine. Planar techniques of image quantification were therefore evaluated with the aim of improving on the accuracy, and simplifying the techniques currently in use. The attenuation of the 172- and 247-keV photons of 111In, singly and in combination, was determined for varying diameter flat sources (3.4 to 16.9 cm). The influence of region of interest (ROI) selection on the shape of the attenuation curves was also determined for five different ROIs. Defining the attenuation curves mathematically generated parameters of fit for three approaches to in vivo quantification, namely: a single exponential geometric mean approach that takes into account source size, depth-dependent, and depth-independent buildup factor approaches to account for the contribution of scatter. The accuracy of these techniques was ascertained and compared to the classical geometric mean method. This was done in a waxen phantom of a human thorax with a hollow liver and spleen. The results indicated that the depth-independent buildup factor is the best method; the error for quantification in the spleen was 0.8% +/- 2.2%. The classical geometric mean approach gave a corresponding error of 43.3% +/- 3.4%. Since the attenuation of the two energies of 111In differ, their ratio changes with depth. This phenomenon was investigated with the goal of determining whether the depth of an object can be estimated from one set of planar images. This was not successful.(ABSTRACT TRUNCATED AT 250 WORDS)

Kinetics, in vivo redistribution and sites of sequestration of indium-111-labelled platelets in giant platelet syndromes.

Six patients with giant platelet syndrome were examined: four with Bernard‐Soulier syndrome (two were asplenic); one with hereditary thrombopathic thrombocytopenia; and one with May‐Hegglin anomaly. Autologous platelets were labelled with In‐111‐oxine and in vivo redistribution and sites of sequestration measured with quantitative imaging. In Bernard‐Soulier syndrome platelet survival was normal or moderately shortened; platelet turnover was decreased only in the two patients with thrombocytopenia. In the patients with thrombopathia or May‐Hegglin anomaly, platelet survival and turnover was moderately decreased. In those patients with normal‐sized spleens, the mean splenic platelet pool consisted of 35.5% of the platelet mass, i.e. normal. The intrasplenic transmit time of the megathrombocytes was prolonged. Splenic blood flow was within normal limits. There was a marked accumulation of platelets in the liver at equilibrium: 15.5‐58.8% of whole body radioactivity (normal 9.6±1.2%). This finding is unexplained. The final sites of sequestration of platelets were mainly in the liver and spleen, similar to that seen in normal subjects. We conclude that there is no inverse relationship between cell size and splenic platelet transit time. Platelet size therefore does not determine the size of the splenic platelet pool. The size of the platelets also does not seem to affect the sites of sequestration at the end of their life span.

The effects of different correction techniques on absolute volume determination with SPECT using a threshold edge detection method.

Quantitation of planar radionuclide images is hampered by structures containing radioactivity which overlie or underlie the organ of interest. The introduction of single photon emission computerized tomography (SPECT) overcame this problem to a large extent and enhanced the contrast of the images. Attenuation of photons, however, degrades the resultant SPECT images and correction methods for photon absorption and scatter were subsequently proposed. The different correction methods have variable effects on the reconstructed images. If threshold techniques are used to quantitate organ volume, i.e., combining pixels with the same percentage of the maximum pixel count in the volume, the selected threshold values which give the most accurate volume determination, will be affected by the specific correction method used. In this study, the effect of various SPECT image correction methods on threshold was investigated. A thorax phantom containing volumes ranging from 30 to 1200 ml was used. Threshold values varying from 45.6% (210 ml without any correction) to 23.7% (1200 ml with a combination of scatter subtraction and attenuation correction) were used to produce correct quantitation when different methods were investigated. A negative correlation was found between threshold and volume. This reduction in threshold was most prominent when scatter and attenuation correction were combined. This study shows that correction methods for attenuation of photons influence the threshold value for volume quantitation and the use of a constant threshold value could lead to underestimation of larger volumes.

The influence of the ‘collection injury’ on the survival and distribution of Indium- 111 -labelled canine platelets

Summary. The extent of the ‘collection injury’ sustained by platelets during labelling with In‐lll‐oxine was investigated in three matched pairs of beagle dogs. The influence of the procedure on the survival, kinetics, in vivo distribution and fate of the labelled platelets was determined by serial blood sampling and quantitative computerized scintillation camera studies. Injured labelled platelets were removed in the matched dog acting, as a biological filter. The survival, distribution and fate of the ‘filtered’ and ‘unfiltered’ platelets were compared. The mean platelet lifespan of the ‘filtered’ and ‘unfiltered’ platelets did not differ significantly, but the shape of the survival curve of the filtered platelets fitted a linear function more closely than that of the unfiltered platelets. Radioactivity in the different organs and regions was serially quantitated and expressed as a percentage of whole body radioactivity. Splenic and hepatic radioactivity of filtered and unfiltered platelets did not differ significantly at equilibrium or at the end of platelet lifespan. It is concluded that the currently employed isolation and labelling techniques for platelets are suitable for quantitative in vivo studies with a computerized scintillation camera system. The shape of platelet survival curves should, however, be interpreted with some caution as it may be influenced by these procedures.