Temporal sequence of splenic dysfunction in sickle cell disease.

Sickle cell patients develop splenic dysfunction early in the course of their disease as shown by failure to visualize the organ on technetium-99m colloid scintigraphy. However, preliminary studies from our center have shown that, when the spleen is not demonstrable on colloid uptake, it may be visualized on technetium-99m heat-denatured RBC scintigraphy. With time, however, the spleen can no longer be visualized with both tests in many patients. We have studied 46 patients aged 2 to 16 years, which included 36 SS, 7 Sbeta(0) thal, and 3 SD. Eighteen (39.1%) had normal splenic colloid uptake (Group 1), 15 (32.6%) had partial uptake (Group 2), and 13 (28.3%) had absent uptake (Group 3). When the patients in Group 1 were compared to those in the two other groups, there was no significant difference in the mean age and Hb F values. However, the mean Hb of 10.2 g/dl in Group 1 was significantly higher than the value of 9.0 g/dl in the other two groups. In addition, 60% of the SS patients with normal uptake and 40% of those with partial or absent uptake had co-existing alpha-thal trait; the difference in this proportion is significant (chi(2) = 85, P < 0.0001). Heat-denatured RBC scintigraphy was carried out on five patients in Group 2, and the spleen was visible in all, while of 12 children in Group 3, the spleen was visible only in 4 patients. This study demonstrates that the phagocytic function of the spleen, which is tested by colloid uptake, is the first to be lost while the filtration function, tested by denatured RBC uptake, persists for much longer. This finding may have significant implications for the clinical symptomatology and therapeutic strategies of sickle cell disease.

Acute 4-aminopyridine seizures increase the regional cerebral blood flow in the thalamus and neocortex, but not in the entire allocortex of the mouse brain.

Systemic injections of 4-aminopyridine precipitate epileptiform generalized seizures characterized mainly by shivering of the body, tail movements and tonic-clonic convulsions in rats and mice. However, only few details are known as concerns which brain regions are possibly affected and stimulated by the compound. The aim of the present study was to investigate the changes in regional cerebral blood flow in mice by using the lipophilic compound technetium-99m-hexamethyl-propyleneamineoxime (99mTc-HMPAO). Whilst the uptake of 99mTc-HMPAO was increased significantly in the neocortex and thalamus following the induction of acute 4-aminopyridine seizures, no such changes were observed in the allocortex of the mice. The increases in uptake in the neocortex and thalamus were completely prevented by carbamazepine (which abolished the symptoms of the seizure, too). The primary involvement of the neocortex and thalamus points to the importance of thalamocortical circuits in the precipitation and maintenance of experimental 4-aminopyridine convulsions.

Assessment of skeletal muscle damage in experimental animal using In-111 antimyosin.

Skeletal muscle injury induced in the thigh muscles of New Zealand rabbits was investigated using radionuclide imaging technique after intravenous injection of In-111 antimyosin. Images were obtained for 3 weeks beginning from the first day of injury. The injury was visualized as early as 3 h after injection of the tracer. The mean uptake ratio of the injury to the normal muscle was 2.8 +/- 0.62 (p < 0.001) on the third to fifth day, decreasing with healing to 1.40 +/- 0.15 by the end of the study (3 weeks). The mean uptake ratio of the more severely crushed left to less severely crushed right side was 1.36 +/- 0.17 (p < 0.01) on the third day. The results of this study show that the radionuclide imaging technique using In-111 antimyosin is useful for the assessment of skeletal muscle damage and also for monitoring the progress of healing. This technique can be applied clinically to evaluate skeletal muscle damage in problematic fractures.

The use of radiopharmaceuticals as an effective toxicologic technique for studying nephrotoxicity of drugs: cyclosporine-A.

The concept of altered biologic behavior of administered radiopharmaceuticals is used routinely in clinical nuclear medicine to increase the sensitivity of diagnosis, monitor the efficacy of chemotherapeutic drugs and radiation treatment, and determine injury caused by a drug whose effect has exceeded its therapeutic value. In this study, cyclosporine-A (CsA) an immunosuppressant drug known to cause nephrotoxicity due to tubular impairment and Tc-99m MAG-3, a renal imaging radiopharmaceutical secreted by the tubules have been used in animal models to establish a method for investigating the nephrotoxicity of drugs. New Zealand rabbits and Wistar rats were used. The rabbits and rats were treated with 30 mg/kg of CsA for 4 and 28 consecutive days respectively. Plasma creatinine and urea were measured and renogram studies were performed in the rabbits prior to and on 1, 4, 8, 11 and 15 days after treatment with CsA. For the renogram, the rabbits were given an intravenous bolus injection of 44.4 MBq (1.5 mCi) of Tc-99m MAG-3. The Tmax, T1/2, TTHM and uptake slope of the Tc-99m MAG-3 were calculated. Each rat was injected intravenously with 185 MBq (5 mCi) of Tc-99m MAG-3, killed 3 min later, the kidneys removed and 20 mm frozen sections made. Autoradiograms were generated from the frozen sections. Creatinine and urea levels were also measured in the rats. There was no consistent difference in creatinine and urea levels between control and CsA treated rabbits and rats. However, for the rabbit, on day 1 or 4 after treatment, there was significant increase in the values of Tmax, T1/2, TTHM and uptake slope between the control and CsA treated animals, indicating intrarenal vasoconstriction and delayed transit of Tc-99m MAG-3 from the parenchyma to the collecting system. This delay is dramatically shown in the tissue autoradiograms of the rats. The results are consistent with reported nephrotoxicity of CsA using other techniques. The results of this study, therefore, indicate that the concept of altered biologic behavior of Tc-99m MAG-3 can be used effectively as a toxicologic method for studying nephrotoxicity of drugs as exemplified by CsA.

Placental binding and transfer of radiopharmaceuticals: technetium-99m d, 1-HMPAO.

UNLABELLED: Placental binding and transfer of 99mTc d, 1-hexamethyl propyleneamine oxime (HMPAO) was studied in vitro using human placenta and pregnant guinea pigs. MATERIALS AND METHODS: Five pieces of human placenta were incubated in 50 ml Earle's solution containing 1.85 MBq 99mTc d, 1-HMPAO. The percent binding of the tracer to the placenta per 1 ml standard solution was calculated. Pregnant guinea pigs representing first, second and third trimesters were each injected with 74 MBq 99mTc d, 1-HMPAO through the jugular or femoral vein after sedation was induced with pentabarbital sodium. Static images were obtained, the guinea pigs were killed, and the fetuses were removed, weighed and imaged separately. The placentas, maternal and fetal brains, lungs, livers and kidneys also were removed, and the radioactivity was assayed in a dose calibrator for each organ. The percent radioactivity in each organ was calculated. RESULTS: The binding of 99mTc d, 1-HMPAO to human placenta ranged from 2.95% +/- 1.5% to 5.82% +/- 0.3% per 1 ml standard solution. Both the binding of 99mTc d, 1-HMPAO to guinea pig placenta and its transfer to the fetus increased with gestational age. The percent binding ranged from 0.09% +/- 0.06% to 0.43% +/- 0.05%, whereas that of transfer to the fetus ranged from 0.05% +/- 0.03% to 2.19% +/- 0.64%. Of the amount transfered to the fetus, the order of accumulation in the fetal organs was liver > blood >> brain > lungs > kidneys > heart. CONCLUSION: Technetium-99m d, 1-HMPAO binds to the placenta, and a minimal amount crosses the placental barrier and is transfered into the fetal circulation, mostly in the liver but a measurable amount is found in brain tissue.

Modified preparation and rapid quality control test for technetium-99m-tetrofosmin.

OBJECTIVE: The objectives of this study were to: modify the preparation of 99mTc-tetrofosmin by using twice the amount of 99mTcO4- recommended by the manufacturer; evaluate the use of miniaturized rapid paper chromatography (MRPC) for quality control (QC) testing; and determine the in vitro stability of the modified preparation using MRPC. METHODS: Two preparations of 99mTc-tetrofosmin were made: one with 4.4-8.8 GBq (120-240 mCi) and the other with 13.9-17.6 GBq (380-480 mCi) 99mTcO4-, referred to as regular and modified preparations, respectively. Routine QC tests were performed using MRPC and instant thin-layer chromatography/silica-gel (ITLC/SG) systems. The preparations were injected into 58 patients. Planar and SPECT images of stress and rest studies were obtained. The technical quality of the SPECT images was graded visually by four observers. Heart-to-lung and heart-to-background ratios were calculated from the planar images. RESULTS: The QC testing procedure took 4.18 +/- 0.15 min with MRPC and 54 +/- 5.3 min with ITLC/SG systems. The percent labeling efficiency, as determined by both techniques, ranged from 95.6 +/- 1.6 to 97.2% +/- 0.8%. Both preparations were stable up to 6 hr after reconstitution. There was no difference between the cardiac-to-lung and cardiac-to-background ratios of the two preparations. CONCLUSION: The results indicate that MRPC is a faster and effective chromatographic technique for routine QC testing of 99mTc-tetrofosmin. Doubling the amount of 99mTcO4- used in preparing 99mTc-tetrofosmin did not affect its in vitro stability, its efficacious use in patients or the technical quality of the images.

Experimental studies on multiorgan toxicity of cyclosporine-A in rats using a radiopharmaceutical and comparison with histopathological findings.

Iodine-125 HIPDM was evaluated as a screening agent for studying multiorgan toxicity due to Cyclosporine-A (CsA) and the results were compared to histopathological findings of the tissues. The rats were injected subcutaneously with 50 mg/kg (body weight) of CsA or with equal volume of the vehicle, Cremophor-EL, for 7 consecutive days. A dose of 10 microCi of I-125 HIPDM was injected intravenously at the end of the treatment period. The results indicated that there was a significant increase in the uptake of I-125 HIPDM in the kidney, liver, heart and blood compared to control rats (P < 0.05). However, there was a significant decrease in the uptake of I-125 HIPDM in the spleen compared to control animals (P < 0.001). The lung and brain of CsA treated rats showed no change in the uptake of I-125 HIPDM when compared to control rats. The change in the uptake of I-125 HIPDM in these organs was assumed to indicate tissue response to the toxic effects of CsA. The radiopharmaceutical results were comparable with the histopathological findings in which the organs showed varying degrees of tissue degeneration. It is concluded that the lipophilic radiopharmaceutical, I-125 HIPDM, can be used as an effective screening agent to study multiorgan toxicity due to CsA.

The effect of cyclosporine-A on labeling blood cells with radiopharmaceuticals.

The effect of cyclosporine-A (CsA) on the labeling efficiencies of red blood cells with reduced 99mTcO4-; leukocytes and platelets with 111In oxine was studied. Blood was used from rats treated with CsA (30 mg/kg body weight) for 28 consecutive days and from control rats. For 99mTc labeling of RBCs, blood was obtained from individual rats and in vitro labeling technique was used. For leukocyte and platelet labeling, blood was pooled from 5 rats either treated with CsA or control. Leukocytes/platelets were labeled with 111In oxine using routine techniques. The labeling efficiency for 99mTc RBCs was 83.42 +/- 0.83% (CsA treated) and 84.85 +/- 0.62% (control); 111In-oxine leukocytes was 38.5 +/- 1.75% (CsA treated) and 42.5 +/- 3.53% (control); and for 111In-oxine platelets, it was 74.0 +/- 2.5% (CsA treated) and 78.0 +/- 1.41% (control). Comparison of the results indicate that there is no difference between the percent labeling efficiencies of 99mTc RBCs, 111In-oxine leukocytes, and 111In-oxine platelets for CsA-treated and control rats. Hence, CsA does not interfere with the labeling process of blood cells with radiopharmaceuticals.

Influence of alpha-thalassemia trait on spleen function in sickle cell anemia patients with high HbF.

Spleen function was studied in a group of 20 Kuwaiti SS patients (aged 2-12 years), using 99mTc-labeled tin colloid scintigraphy. They were screened for the alpha-thalassemia determinants which are prevalent in the Arabian Peninsula [-alpha (3.7 kb) deletion, alpha2-globin gene polyadenylation signal (AATAAA => AATAAG) mutation, and 5' IVS-I splice junction pentanucleotide (GAGGTGAGG => GAGG) deletion] with a combination of polymerase chain reaction and allele-specific oligonucleotide (ASO) hybridization techniques. The patients were divided into three groups depending on the result of their colloid uptake. Group I consisted of 7 patients (35.0%) with normally visualized spleens, Group II consisted of 5 (25.0%) with partial visualization, and in Group III there were 8 (40.0%) in whom the spleen was not visualized at all. The significant distinguishing features among those in Groups I and III were mean corpuscular volumes (MCVs) of 74.1 +/- 5.1 and 90.1 +/- 6.6 fl (P<0.0001) and mean corpuscular hemoglobins (MCHs) of 22.4 +/- 2.7 and 27.5 +/- 4.0 pg (P<0.05), respectively. The overall frequency of alpha-thalassemia determinants in the study was 35.0%; however, the frequencies in Groups I, II, and III were 57.1, 30.0, and 18.8%, respectively. alpha-Thalassemia trait, therefore, appears to be associated with normal splenic function in these patients.

A miniaturized rapid paper chromatographic procedure for quality control of technetium-99m sestamibi.

A miniaturized rapid paper chromatographic technique (MRPC) for quality control of a technetium-99m sestamibi preparation was developed and compared with the manufacturer's and Hung et al. techniques. The MRPC system involves the use of 6.0x0.5cm Whatman 3MM paper strip developed in ethyl acetate. The procedure was completed within 3min while that of the manufacturer and Hung techniques took 30-35 and 4min respectively. The Rf range of 99mTc-sestamibi using MRPC was 0.55-0.75 while that of the other two techniques was 0.9-1.0. The results indicate that MRPC can be used to separate 99mTc-sestamibi from any 99mTc contaminant that migrates with the solvent front. The MRPC is a fast and effective chromatographic technique for routine quality control testing of 99mTc-sestamibi preparation.

Is cyclosporine toxic to the heart?

Cardiotoxicity caused by cyclosporine was studied in experimental rats by the uptake of radiopharmaceuticals (technetium 99m-labeled pyrophosphate and indium 111-labeled antimyosin) and histologic examination of heart tissues. A dose of 50 mg/kg (body weight) of cyclosporine and an equal volume of vehicle (cremophor-EL) were injected into the rats subcutaneously for 7 or 11 consecutive days. A statistically significant difference (p < 0.05) was noted between the uptake of the radiopharmaceuticals in the hearts of cyclosporine-treated rats compared to the control rats. For 99mTc pyrophosphate, the cardiac uptake ratios of cyclosporine-treated rats to control rats were 2.13 and 4.08 for 7-day and 11-day treatment periods, respectively. For 111In antimyosin, the ratios were greater than 2 for both 7-day and 11-day treatment periods. Histologically, vacuoles were found in single or focal groups of myocytes with interstitial edema in the hearts of cyclosporine-treated rats compared to the control rats. The results of both the uptake of the radiopharmaceuticals and the histologic evidence indicate cell injury in the hearts of cyclosporine-treated rats. Cyclosporine therefore seems to be toxic to the heart tissue.

New technique for assessing muscle damage after trauma.

We report a new technique which is a useful objective method for assessing the extent and severity of traumatic muscle damage using scintigraphic imaging. Radionuclide indium-111 antimyosin was used in 13 patients with trauma. The increased uptake was confined to the injured muscles and the intensity was related to the severity of damage.

Effect of hyperthermia on somatosensory evoked potentials in the anaesthetized rat.

In the rat heat stroke model, established by heating to a climatic chamber temperature of 42 degrees C, the brain temperature was found to be consistently lower than the rectal temperature, suggesting efficient brain cooling mechanisms in the rat. In response to heating, with increasing brain temperature, the latencies of the somatosensory evoked potentials (SEPs) showed an initial decrease followed by an increase (inflection point). Studies were done on rats heated up to, before, or after the inflection point and then cooled. Reversibility with cooling of functional and structural changes induced by heat was evaluated by analysis of SEPs, survival time, brain blood perfusion and histopathology. The evidence from these studies demonstrated that the brain temperature at which the inflection in wave P2 latency occurred was critical, beyond which hyperthermia produced irreversible changes in the SEP, shorter survival time, relative reduction in brain blood perfusion and evidence of brain histopathological damage. The suggestion that endorphins may mediate brain dysfunction in hyperthermia was investigated. In rats heated and then cooled after wave P2 latency inflection naloxone, the endorphin antagonist, was injected (10 mg/kg, intravenously) just prior to the inflection. It produced reversibility of SEP changes as well as longer survival time (P less than 0.001) compared to saline-treated rats.

Forskolin impregnation of small calibre PTFE grafts lowers early platelet graft sequestration and improves patency in a sheep model.

Synthetic vascular grafts have a thrombogenic surface which plays a role in graft failure. Systemic pharmacologic interventions have been used to lower platelet sequestration onto the graft surface but are associated with side effects. In this communication we describe the results of a new therapeutic principle of applying forskolin, a powerful cyclic adenosine monophosphate stimulator (cAMP) to the inner surface of PTFE vascular grafts. The grafts were evaluated with Indium-III-oxine labelled platelets and by graft patency on 3 consecutive days after implantation at 1 month and 3 months. Forskolin significantly lowered early platelet sequestration onto the treated graft surface when compared with controls. Graft potency at 1 and 3 months was also significantly higher in the forskolin treated grafts.

Direct platelet effect of low molecular weight dextran in small calibre PTFE grafts.

The thrombogenicity of synthetic vascular grafts is a major factor in occlusion of grafts when they are used to bypass small calibre arteries. In this paper, the effect of low molecular weight dextran (LMWD, Dextran-40) on graft surface-platelet interaction was studied using Indium-III-oxine labelled platelets. It was found that LMWD significantly reduced platelet deposition onto graft surfaces (P less than 0.001). Dextran had a direct antiplatelet effect independent of plasma volume expansion as dextran-soaked grafts significantly reduced platelet deposition when compared to systemic dextran administration (P less than 0.001). We therefore conclude that LMWD has a direct antiplatelet effect which is beneficial in reducing platelet deposition on synthetic PTFE grafts which may improve the early patency of such grafts.

Use of indium-111-labeled transferrin to study plasma extravasation during endotoxin shock and the effects of the beta-2 agonist terbutaline.

Endotoxemia and sepsis are common causes of respiratory distress (ARDS), which is characterized by increased pulmonary vascular permeability to plasma proteins resulting in "noncardiac pulmonary edema." The aim of this series was to study the effects of the beta-2 receptor agonist, terbutaline, on plasma extravasation in multiple organs, in sheep exposed to endotoxin shock. A double isotope technique was used and the radioactivity was recorded in different organs (lungs, liver, spleen, kidneys, intestine) by a computerized gamma camera. Tc-99m-labeled erythrocytes were used as a marker for intravascular volume and In-111m-labeled transferrin for tracing extravascular plasma leakage. An organ-transferrin index (organ-TI) was calculated for each organ which corrects for changes in blood distribution. Fourteen sheep were anesthetized and ventilated. After stabilization (t = 0) all animals received E. coli endotoxin 10 micrograms/kg by IV infusion during 30 min. At t = 30, seven animals (group T) received IV infusion of terbutaline, 20 micrograms/kg/hr, during 4 h, while the other seven received normal saline and served as controls (group E). The endotoxin infusion caused an immediate and significant increase in the transferrin index in the lungs and in the liver in both groups. The transferrin index continued to rise in the control group towards the end of the experiment (t = 240), while in group T it reached a maximum 60 min after endotoxin. Four hours after endotoxin the transferrin index was significantly higher in the controls than in the terbutaline treated group, both in the lungs and in the liver (P less than 0.01). No significant changes were recorded in the kidneys or over the intestine.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of cyclosporin A on bile flow in experimental animals using technetium-99m EHIDA.

The effect of cyclosporin A (CyA) on bile flow was studied in experimental rabbits by radionuclide imaging and measurement of bilirubin levels. The rabbits were dosed intravenously with 15 mg/kg CyA for 4 consecutive days. Each rabbit served as its own control. The rabbits were injected IV with technetium (Tc)-99m EHIDA, and dynamic images were obtained prior to and 1, 4, 8, and 15 days after CyA treatment. There was no difference in half-times of blood clearance in control and CyA-treated rabbits. There were differences in the half-times of liver curve and bilirubin measurements between control and 4-day treated rabbits. By the 15th day after CyA treatment both the radionuclide findings and bilirubin levels became normal. The results suggest that CyA causes intrahepatic cholestasis and demonstrate evidence of reversibility of CyA's toxic effect on bile flow after treatment is discontinued.

The use of 125I-HIPDM for studying tissue response due to toxic effects of cyclosporin-A in rats.

125I-HIPDM was used to study the response of various tissues in cyclosporin-A, CyA, treated and control rats. The rats were given 50 mg/kg of CyA for 7 consecutive days. The liver, kidney and heart showed significant increase while the spleen had a pronounced decrease in the uptake of 125I-HIPDM in CyA treated compared to control rats. This difference in the uptake of 125I-HIPDM between CyA treated and control rats is assumed to be the tissue response to toxic effects of CyA. The results indicate that CyA is toxic to liver, kidney, spleen and probably heart. There was no difference in the uptake of 125I-HIPDM in the lung and brain of CyA treated and control rats. This lack of difference is assumed to indicate that CyA does not adversely affect the lung and brain.

Leucocyte sequestration in endotoxemia and the effect of low-molecular-weight dextran.

Leucocyte sequestration in various organs during endotoxin-induced shock in sheep was studied using leucocytes labelled with indium 111 oxine. A moderate dose of Escherichia coli endotoxin (10 micrograms/kg body weight) was slowly infused intravenously in 16 sheep, 9 of which subsequently received a continuous i.v. infusion of low-molecular-weight dextran (LMWD) given at an infusion rate of 15 ml/h over 4 h, starting 30 min after administration of the endotoxin. By that time, signs of acute lung injury had developed, thus mimicking a clinical situation. The remaining animals were untreated and served as controls. A marked increase in lung, liver and kidney leucocyte sequestration, together with a sharp, corresponding drop in splenic activity and leucocyte count in peripheral blood, occurred shortly after the endotoxin infusion in both groups. However, after 90 min there was a significantly lower leucocyte activity in the lungs, liver and kidneys of LMWD-treated animals as compared with controls. Less marked hemodynamic and respiratory alterations were also observed in animals treated with LMWD. The present study confirms previous reports that significant leucocyte sequestration in the lungs occurs early during endotoxemia. Furthermore, we found that leucocyte sequestration also occurs in the liver and kidneys, which could explain the development of multi-organ failure, frequently described in clinical sepsis. Even after injury to organs, LMWD infusion seems to be beneficial by significantly lowering leucocyte sequestration and could therefore be justified as an addition to the arsenal of interventions used in the treatment of endotoxemia.