(Circulation. 1997;95:1577-1584.)
© 1997 American Heart Association, Inc.
Articles |
Very Early Noninvasive Detection of Acute Experimental Nonreperfused Myocardial Infarction With 99mTc-Labeled Glucarate
Presented for the Young Investigator Award of the Cardiovascular Council in the 41st Annual Meeting of the Society of Nuclear Medicine, June 5, 1994, Orlando, Fla.
From the Center for Drug Targeting and Analysis (J.N., A.P., B.C.L., B.-A.K.), Northeastern University, Boston, Mass; Cardiac Unit (J.N., B.-A.K.), Massachusetts General Hospital and Harvard Medical School, Boston, Mass; and Molecular Targeting Technologies Inc (K.-Y.P., B.-A.K.), Malvern, Pa.
Correspondence to Ban-An Khaw, PhD, George D. Behrakis Professor of Pharmaceutical Sciences, Center for Drug Targeting and Analysis, 205 Mugar, Northeastern University, Boston, MA 02115.
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Abstract |
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Background 99mTc glucarate has recently been reported to be an infarct-avid agent. The feasibility of imaging with 99mTc glucarate was evaluated for the early diagnosis of nonreperfused and reperfused myocardial infarction and compared with localization of simultaneously administered 111In anti-myosin.
Methods and Results Four groups of six rabbits each were studied. The left anterior descending coronary artery (LAD) was kept persistently occluded (n=6) or reperfused after 40 minutes (n=6) in rabbits. After confirmation of LAD occlusion by 201Tl scintigraphy, a mixture of 99mTc glucarate (15.7±1.6 mCi) and 111In anti-myosin (0.53±0.03 mCi) was administered intravenously. Another group of rabbits (n=6) with 5 or 15 minutes of LAD occlusion were used to assess the affinity of 99mTc glucarate for the ischemic myocardium. The remaining 6 rabbits with reperfused myocardial infarction were used for the assessment of subcellular localization of 99mTc glucarate. 99mTc glucarate cleared rapidly from circulation (elimination t1/2, 36 minutes). Infarcts were visualized within 10 minutes in reperfused and within 30 minutes in nonreperfused coronary territories after intravenous administration. 111In anti-myosin delineated reperfused infarcts within 1 to 3 hours, but no uptake was seen in persistently occluded rabbits. 99mTc glucarate uptake in reperfused and nonreperfused infarct centers was 28 and 12 times greater, respectively, than that in normal myocardium (P=.0001). A direct correlation between glucarate and anti-myosin localization (r=.60 for nonreperfused; 0.76 for reperfused; P<.0001) was observed. Ischemic hearts showed no glucarate uptake. Subcellularly, 99mTc glucarate localized predominantly in the nuclear fraction of the infarct, with lesser extents in the mitochondrial and cytoplasmic fractions.
Conclusions Noninvasive imaging of myocardial infarcts with 99mTc glucarate is possible within minutes in persistently occluded or reperfused myocardial infarcts. Early detectability results from the rapid blood clearance and high avidity of glucarate for the acutely necrotic myocardial tissue.
Key Words: myocardial infarction • imaging • antibodies • coronary disease • myosin • reperfusion
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Introduction |
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The diagnosis of acute myocardial infarction is made on the basis of typical chest pain, the evolution of ECG changes, and the pattern of serum cardiac enzyme release in the majority of patients.1 In several instances these findings are less helpful, such as in the differentiation of acute infarction from prolonged severe ischemia, infarction in the presence of conduction abnormalities, or infarct recurrence in the ECG territory of previous infarction.1 2 In such cases, the confirmation of diagnosis requires other noninvasive diagnostic aids. The additional diagnostic aid would be of greatest value if the occurrence of acute myocardial infarction could be established early to allow thrombolytic therapy.1 The time window amenable to thrombolysis is short.3 4 5 If a hot spot–imaging agent were to fulfill this role, it should target the infarct zone rapidly despite the persistently occluded status of the coronary artery, clear rapidly from the circulation, and possess high avidity for the necrotic myocardial tissue to attain sufficient target-to-background ratios.
Two currently used infarct-avid agents (99mTc pyrophosphate6 7 8 and 111In anti-myosin9 10 11 ) have not allowed the hyperacute localization and visualization of myocardial infarction. Recently, 99mTc glucarate, a six-carbon dicarboxylic acid,12 has been shown to concentrate in regions of myocardial necrosis.13 14 15 99mTc glucarate localization in experimental acute reperfused myocardial infarct in a canine model was shown to occur on the day of the infarct.14 It was also demonstrated that glucarate uptake diminished significantly by 48 to 72 hours after the acute event.15 The present study was undertaken to evaluate the role of glucarate scintigraphy in the detection of acute nonreperfused myocardial infarction and to compare it with its uptake in reperfused rabbit infarction as well as in severely ischemic myocardial tissues. Glucarate uptake was also compared with anti-myosin antibody–delineated myocardial necrosis. Ex vivo assessment of the myocardial tissues was undertaken to identify the site of subcellular localization of 99mTc glucarate in the infarcted myocardium.
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Methods |
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99mTc glucarate uptake in the ischemic and necrotic myocardium was evaluated in 24 New Zealand White rabbits (weight, 2.5 to 3.0 kg). Acute experimental myocardial infarcts were induced by the occlusion of the left anterior descending coronary artery (LAD) in 12 rabbits for noninvasive
scintigraphy; in the
first group of 6 animals, the LAD territory was kept persistently
occluded, and in the second group of 6 animals, the LAD territory was
reperfused. The noninvasive imaging studies were performed in a third
group of 6 animals after the induction of severe myocardial
ischemia. In the remaining 6 rabbits of the fourth group, acute
reperfused myocardial infarcts were produced, and the hearts were
excised and immediately used for the assessment of subcellular
localization of 99mTc glucarate activity in the infarcted
myocardial tissue. All animals received care in compliance with the
principles of laboratory animal care formulated by the National Society
of Medical Research and the Guidelines for the Care and Use of
Laboratory Animals prepared by the National Academy of Science
(NIH publication No. 85-23, revised 1985).
Experimental Acute Myocardial Infarction
All rabbits were anesthetized with a mixture of ketamine and
xylazine (0.5 to 0.6 mg/kg, 10:1 vol/vol mixture of 100 mg/mL each).
Surgical tracheostomy was performed, and ventilation was maintained
with a Harvard rodent positive-pressure respirator. Anesthesia was
maintained on sodium pentobarbital (1 to 2 mL/h, 6.5 mg/mL). The heart
was exposed through parasternal thoracotomy, and the pericardium was
removed. The region of the LAD was identified between the aortic root
and the left auricle, and a monofilament suture was placed at the site.
The LAD was occluded by tightening the snare created by passing the
suture through a polyethylene tubing. For nonreperfused infarcts, the
LAD was kept persistently occluded, and for reperfused myocardial
infarcts, the snare was removed from the LAD after 40 minutes of
occlusion (Fig 1
).
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Experimental Myocardial Ischemia
In 6 rabbits, the LAD was ligated as described above. The
coronary artery was reperfused after 5 minutes in 3 rabbits and after
15 minutes in the remaining 3 animals. 201Tl was
administered during the LAD occlusion to ascertain sufficient
myocardial region of ischemia.
99mTc Glucarate Preparation
Glucarate kits were provided by Molecular Targeting
Technology, Inc. Each kit consists of a lyophilized mixture of sodium
glucarate (12.5 mg) and stannous chloride (50 µg). To each kit, 30 to
40 mCi of generator-eluted
99mTcO4- was added and the
reaction was allowed to incubate at room temperature for 30 minutes.
Quality control was then performed using 3-mm paper (Whatmann)
chromatography, developed with 60:40 acetonitrile/water.16
TcO2 colloids remained at the origin, labeled glucarate
moved with an RF of 0.3, and free pertechnetate
was recovered at the solvent front. Quality control was also assessed
by thin-layer chromatography with Bakerflex (Gelman Sciences)
SiO2 strips. Chromatograms developed in saline showed
glucarate activity near the solvent front with the colloid remaining at
the origin. On the other hand, the chromatograms developed in methyl
ethyl ketone provided glucarate activity at the origin and free
pertechnetate at the solvent front. Every kit labeled and analyzed as
above showed labeling efficiency of >98%.
111In Anti-Myosin Antibody
The anti-myosin Fab–DTPA was provided by Centocor. The
molar ratio of DTPA to anti-myosin Fab was 1:1. Monoclonal anti-myosin
R11D10 was generated by hybridization of immune murine spleen cells
with SP2/OA murine myeloma cells and purified subsequently according to
the methods previously described.17 Bicyclic anhydride of
DTPA prepared according to the method of Hnatowich et al18
was used to modify anti-myosin Fab.
111In chloride (0.6 to 0.8 mCi) in 0.5 mol/L citrate,
pH 5.5, was used to label 100 µg of DTPA/anti-myosin
Fab.19 The reaction mixture was allowed to incubate at
room temperature for 
30 minutes. Antibody-bound 111In
was separated from free 111In with the use of Sephadex G-25
(Sigma Chemical Co) column chromatography (10-mL column). Peak tubes in
the void volume containing the radiolabeled antibody were pooled and
used within 1 hour of radiolabeling. An average of 95% of total
activity was recovered in the peak tubes containing the radiolabeled
antibody.
Imaging Protocol and Biodistribution Studies
The myocardial area at risk in all animals was first confirmed
with the use of 201Tl perfusion scintigraphy. Subsequently,
99mTc glucarate was administered to assess its uptake in
the necrotic myocardium and was compared with the uptake of
simultaneously administered 111In anti-myosin Fab.
In vivo planar imaging was performed with an Ohio-Nuclear
camera (Technicare Corp) equipped with a 3-mm pinhole collimator.
201Tl (0.67±0.08 mCi) was injected intravenously 30
minutes after reperfusion or the corresponding time point in the
persistently occluded group of animals (Fig 1
). Left lateral planar
images were obtained for 10 to 15 minutes after injection. Immediately
afterward, a mixture of 99mTc glucarate (15.7±1.6 mCi) and
111In-labeled anti-myosin Fab (0.53±0.03 mCi) was injected
intravenously. Sequential 1-minute images were acquired for the
first 60 minutes and then every 15 minutes. 201Tl imaging
was performed with a centerline setting of 80 keV, 140 keV for
99mTc, and 247-keV photopeak for 111In. A 15%
window was used for all radioisotopes. Blood samples were withdrawn
from the femoral artery at 1 to 5, 10, 15, 20, 30, 45, 60, 75, 90, 105,
120, 150, and 180 minutes after intravenous administration of the
mixture of radiolabeled glucarate and antibody. All animals were killed
at 3 hours with an overdose of pentobarbital. Ex vivo imaging of the
entire heart was performed; then, the hearts were cut into slices, and
images of the slices were made. Slices with faint or no uptake on
ex vivo imaging were also subjected to macroautoradiography.
For histochemical delineation of the myocardial infarcts, the
slices were incubated with nitroblue tetrazolium for 30 minutes at
37°C. Nitroblue tetrazolium has provided more reproducible staining
for macroscopic delineation of rabbit myocardial infarcts than did
triphenyl tetrazolium chloride. The outline of the unstained pale
infarcted region of the slices was traced onto transparent acetate
sheets or photographed. Each slice was then divided into eight pieces:
201Tl, 99mTc, and 111In uptakes in
the pieces of the heart were determined with scintillation counting
(LKB Compugamma 1282; Pharmacia LKB Nuclear, Inc). Biodistribution in
the kidneys, liver, spleen, and skeletal muscles was also performed,
using the automatic spill-correction settings for the three isotopes.
However, thallium counting data were not used for comparative analysis
because 201Tl counts at the end of the experiment may not
be representative of its initial distribution, especially after
postmortem tissue fixation.
Intracellular Distribution of 99mTc Glucarate
To determine the intracellular distribution of 99mTc
glucarate in the infarcted and normal myocardium, 6 additional rabbits
with reperfused myocardial infarction were studied. Localization of
99mTc glucarate was allowed to occur for 1 hour (n=3) or 3
hours (n=3) after intravenous administration. 201Tl and
111In anti-myosin were not used in these animals. The
animals were killed as previously described. The infarcted myocardial
region was identified through imaging of the slices and confirmed
through staining with nitroblue tetrazolium. The infarcted myocardial
and normal posterior myocardial areas were dissected, washed, weighed,
and counted in a scintillation counter. Then, the myocardial
tissues were homogenized as described previously.20 21 The
nuclei were separated (pelleted) from the mitochondria and cytoplasm
(supernatant) through sucrose (0.25 mol/L) gradient centrifugation
(800g for 5 minutes). After decanting, the supernatant was
recentrifuged at 10 000g for 10 minutes to separate the
mitochondria (pellet) from the cytoplasm, contractile proteins, and
other cellular organelles (supernatant). Each fraction was then counted
in a counter to determine the subcellular distribution of
99mTc activity. To confirm the efficiency of fractionation,
aliquots of nuclear, mitochondrial, and cytosolic fractions were
stained with Wright's stain, ethidium bromide, and rhodamine 123.
Statistical Analysis
All statistical analyses were performed with the use of
Statgraphics 5.0 (Manugistics). The scintillation counts were
calculated as percent of the total injected dose per gram and as
absolute tracer uptake per gram of tissue or blood. All values were
expressed as mean±SEM. On the basis of nitroblue tetrazolium staining
of the slices, areas of the infarct center and border zone as well as
the uninvolved myocardial regions were identified. From 6 nonreperfused
animals of the first group, 28 myocardial pieces represented the
infarct center and 29 and 23 pieces represented the infarct periphery
and normal myocardial tissues, respectively. Similarly, in 6 reperfused
rabbits of the second group, 27, 24, and 30 myocardial pieces were
obtained from the infarct center, infarct periphery, and normal
myocardial regions, respectively. Mean percent injected doses per gram
of 99mTc and 111In in these zones were
compared. A three-way ANOVA was used to determine the effects of the
perfusion status of the coronary artery (reperfused or persistently
occluded), location of the infarct (infarct center, infarct border, and
normal myocardium), and the radiopharmaceutical (99mTc
glucarate or 111In anti-myosin). A Newman-Keuls
multiple-range test was used to determine the statistical significance
of the difference of radiopharmaceutical uptake in the three myocardial
regions. Subsequently, a total of 340 myocardial pieces were used for
the analysis of correlation: 169 reperfused and 171 nonreperfused. For
this purpose, infarct-to-normal ratios were obtained for all
radionuclides, and the correlation between their distributions was
obtained. Simple linear regression routine was used for the correlation
of anti-myosin and glucarate uptake in the myocardial tissues from
reperfused and nonreperfused infarcts. Because 201Tl is
expected to redistribute over the course of the experiment and
postmortem fixation, 201Tl distribution data were not used
for analysis.
Similar to the tissue samples, percent total injected dose per gram of blood was determined from the serial blood samples. One-minute postinjection activity was considered to be 100%, and percent residual activity was calculated for the subsequent samples. Blood clearance characteristics and half-time estimations of 99mTc glucarate and 111In anti-myosin Fab were determined from a two-compartment model using a PCNONLIN 3.0 nonlinear least-squares estimation program (Statistical Consultations, Inc).
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Results |
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Imaging Acute Myocardial Infarction With 99mTc Glucarate
99mTc glucarate was observed to clear rapidly from the blood, which facilitated very early visualization of the infarcts. 99mTc glucarate uptake could be seen in the area of myocardial necrosis within 10 minutes after intravenous administration in reperfused infarcts, with unequivocal visualization within 30 minutes (Fig 2
). 99mTc glucarate
delineated myocardial hot spot activity correlated with the zone of
thallium defect (Fig 3). Antimyosin-delineated infarct
was visualizable at 1 to 3 hours after intravenous injection. At 3
hours (the time at which the animals were killed), glucarate- and
anti-myosin–delineated regions of myocardial infarction were similar.
However, target-to-background ratios as well as the absolute
radiotracer uptake of 99mTc glucarate in the infarcts were
significantly greater than those of 111In anti-myosin.
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99mTc glucarate concentration was less in rabbits
with persistently occluded coronary arteries. Although, infarcts could
be visualized earlier than 30 minutes, unequivocal visualization was
possible only at 40 to 60 minutes after intravenous administration of
the radiopharmaceutical (Fig 4). Delineation of the
infarct with the use of 99mTc glucarate closely
corresponded to the regions of initial thallium defects. Anti-myosin
antibody localization in nonreperfused necrotic zones did not reach
sufficiently high target-to-background ratio to enable in vivo
visualization, even at 3 hours after intravenous injection.
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No glucarate uptake was observed in 6 animals with myocardial
ischemia on the basis of in vivo imaging or ex
vivo imaging of the explanted heart or of the slices or
in the macroautoradiographs of the slices (Fig 5).
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Quantitative Glucarate Uptake in Reperfused and Nonreperfused
Infarcts
Delineation of the infarct by nitroblue tetrazolium staining
corresponded to the images of 99mTc glucarate in the
myocardial slices (Fig 6). Tissue samples from the
center of the infarct, periphery of the infarct, and the normal
posterior myocardium remote from the infarcted tissues were assessed
for the distribution of 99mTc and 111In
activities. In the reperfused animals, 99mTc glucarate
uptake in the center and the border zone of the infarct was
0.108±0.0121% and 0.024±0.005%, respectively (Table 1 and Fig 7a). Mean infarct-to-normal ratio was
28:1 in the infarct center and 6.3:1 in the border zone
(P=.0001). In the corresponding tissue samples,
111In anti-myosin uptake values were 1.131±0.154% and
0.251±0.04%, respectively (Fig 7b). The mean infarct-to-normal
myocardium ratio of anti-myosin uptake was 14:1 in the infarct center
and 3:1 in the infarct border at 3 hours (P=.0001).
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In persistently occluded animals, glucarate uptake was
0.064±0.005% in the infarct center, 0.041±0.005% in the infarct
border, and 0.0055±0.0005% in normally perfused myocardium (Fig 7a).
Target-to-background ratios of glucarate were 12- and 7.4-fold higher
in the infarct center and border zones, respectively. 111In
anti-myosin uptake in the infarcted myocardium was, however, only
approximately twofold higher than that in normal myocardium. The mean
percent dose per gram of 111In anti-myosin uptake in the
infarct center, infarct border, and normal myocardium was
0.148±0.020%, 0.130±0.012%, and 0.066±0.004%, respectively (Fig 7b).
Absolute uptake of both of the radiopharmaceuticals (glucarate and
anti-myosin) in persistently occluded infarct centers was lower than
the corresponding values in the reperfused infarcts (Table 1).
Visualization of the necrotic zones was possible relatively early in
reperfused infarcts due to higher tracer localization. Although
absolute glucarate uptake in nonreperfused animals was sufficient for
noninvasive imaging relatively early, anti-myosin uptake did not
achieve sufficiently high contrast to enable visualization or
differentiation from the blood pool activity, even at 3 hours.
Correlation of Glucarate Uptake With 111In
Anti-Myosin
A direct correlation between glucarate and anti-myosin
uptake was observed in both reperfused
(y=0.004+0.071x; r=.76,
P<.0001; Fig 8a) and nonreperfused
(y=-0.0003+0.25x; r=.60,
P<.0001; Fig 8b) myocardial infarcts. Despite a significant
correlation between anti-myosin and 99mTc glucarate in the
two models, the slopes of the correlation were different. In reperfused
infarcts, uptake of each isotope was intense. On the other hand, in
nonreperfused infarcts, uptake of only glucarate was intense, whereas a
disproportionately lower anti-myosin uptake (Table 1) resulted in a
significant shift of the slope in the direction of glucarate uptake
(Fig 8b). Despite this shift, a significant direct correlation existed
between the two radiopharmaceuticals.
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Blood Clearance of Glucarate
99mTc glucarate cleared rapidly from the circulation
and demonstrated a biexponential clearance characteristic. The
elimination t1/2 was 36 minutes with a
t1/2
of 6 minutes and a
t1/2ß of 133 minutes (Fig 9).
Anti-myosin blood clearance also demonstrated a biexponential
characteristic, with an elimination half-life of 150 minutes
(t1/2=20 minutes and
t1/2ß=186 minutes).
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Subcellular Uptake of 99mTc Glucarate
In the 6 rabbits included in the analysis of the subcellular
distribution of 99mTc glucarate, 3 rabbits were killed at 1
hour, and the remaining 3 were killed at 3 hours after glucarate
administration (Table 2). The mean percent injected dose
per gram uptake of glucarate in the infarcted myocardium at 1 hour was
0.146±0.004%, which increased to 0.177±0.011% by 3 hours. Normal
myocardial activity at 1 hour (0.016±0.004%) decreased by 2.7-fold at
3 hours to 0.006±0.000%, indicating that glucarate is not retained by
the normal myocardium.
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The subcellular distribution of glucarate in the infarcted myocardium demonstrated that approximately three fourths of the radioactivity was recovered in the nuclear fraction (73% to 76%). The remaining activity was equally distributed in the mitochondrial (10% to 14%) and cytoplasmic (12% to 14%) fractions. Glucarate uptake in the nuclear fraction in the infarcted myocardial region was 11-fold higher than that in the normal myocardium at 1 hour, which increased to 45:1 by 3 hours. The corresponding infarct-to-normal ratios were 21 and 36 for the mitochondrial fractions and 4 and 12 for the cytoplasmic fractions, respectively.
General Biodistribution of 99mTc Glucarate
99mTc glucarate showed an early accumulation in
the kidneys, with 0.7% to 0.8% injected dose per gram, and was the
major organ of excretion of the radiopharmaceutical (Fig 10). All other organs, including the liver and the
blood compartment, had <0.05% injected dose per gram. On the other
hand, circulating anti-myosin Fab activity was significantly high at 3
hours after intravenous administration (>0.3% injected dose per
gram). Maximum sequestration of anti-myosin Fab was also observed in
the kidneys (0.3%). In addition, the liver demonstrated high
radiolabeled antibody accumulation (0.1%) because it is an organ of
protein catabolism. Other organs demonstrated low levels of anti-myosin
Fab uptake.
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Discussion |
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The need for rapid detection of acute myocardial infarction has prompted the development of several infarct-avid imaging techniques. 99mTc pyrophosphate and 111In anti-myosin have most commonly been used for this purpose.6 7 8 9 10 11 Pyrophosphate imaging predominantly targets the sequestered calcium in the infarcted or severely ischemic mitochondria. The process of mitochondrial calcium sequestration takes
48 hours6 7 8 ;
99mTc pyrophosphate, therefore, is most effective in
delineating myocardial infarcts at this time. Due to the small size of
the pyrophosphate molecule, its localization in reperfused and
nonreperfused infarct is equally effective. Pyrophosphate uptake, which
occurs in the infarcted myocardium up to 10 days after the acute event,
provides a window on the age of the infarct.6
111In anti-myosin, on the other hand, targets the
intracellular cardiac myosin, which is exposed to the extracellular
milieu subsequent to the ischemic disruption of the sarcolemma.
Although the interaction of anti-myosin antibody with the homologous
antigen in the infarcted myocardium occurs almost instantly, it
requires 12 to 48 hours for the blood pool activity in humans to
decrease below the target activity to enable visualization of the areas
of myocardial infarction.9 Furthermore, because of the
slower blood clearance of a larger molecule, anti-myosin antibody
localizes more intensely in reperfused infarcts.22 Uptake
of anti-myosin in the infarcts continues up to 6 weeks after the
acute event and decreases precipitously thereafter.23
Anti-myosin antibody therefore does not have the capability to
differentiate an acute from a subacute event. However, the exclusive
specificity of anti-myosin antibody for the necrotic myocardium allows
precise quantification of the infarct size.
Although both of these infarct-avid imaging agents, pyrophosphate and
anti-myosin, are highly effective for the delineation of acute
myocardial infarction, they do not allow localization of the infarcted
myocardium in a time window that is amenable for thrombolytic
intervention. Identification of the infarct beyond this time limit may
not be of significant therapeutic advantage.3 4 5 In the
present study, glucarate uptake could be visualized with the use of
imaging within 30 to 60 minutes in infarcts with a persistently
occluded coronary artery. 99mTc glucarate uptake was almost
12-fold higher in the infarct center than in the normal myocardium and
7-fold higher in the infarct periphery in persistently occluded
rabbit infarcts. In the corresponding persistently occluded infarcts,
anti-myosin could achieve only a 2:1 ratio in the infarct center or
periphery at 3 hours after administration of the radiopharmaceutical.
Development of high target-to-normal glucarate uptake ratios appeared
to be a direct consequence of the rapid clearance from the circulation
and its avidity for the necrotic myocardium.
Predominant localization of 99mTc glucarate radioactivity
occurred in the nuclear fraction, followed by the mitochondria and
cytoplasm. Uptake in the infarcted tissues increased over time after
intravenous administration of the radiolabeled glucarate. On the other
hand, glucarate uptake decreased consistently in noninfarcted
myocardium, indicating that glucarate is not sequestered by the normal
myocardial tissues (Table 2). Our studies have further indicated a high
likelihood of an ionic interaction between negatively charged glucarate
and positively charged histones as well as the possible in situ
radiolabeling of the phosphates of the DNA by transchelation of
99mTc from glucarate to the phosphates. Mitochondrial and
cytosolic uptake of glucarate is different from that reported for
99mTc gluconate, a monocarboxylic sugar.24 25
99mTc gluconate activity was predominantly recovered in the
mitochondrial fraction; chromatographic subfractionation through
high-performance liquid chromatography suggested targeting of
mitochondrial protein cytochrome AA3.25
Gluconate uptake in disintegrating nuclei has not been
investigated.24 25
The regions delineated by the use of 99mTc glucarate and 111In anti-myosin were essentially similar in the present study as well as in our previously reported study of a canine model of reperfused myocardial infarction.26 Despite the significant agreement in the trends of uptake of the two radiopharmaceuticals in the infarcted and normal myocardium, glucarate demonstrated distinctly superior uptake characteristics in nonreperfused myocardium. This property of glucarate imaging may be due to the small size of the glucarate, which may offer an advantage of very early visualization of nonreperfused infarcts, thereby allowing early thrombolytic intervention in acute myocardial infarcts. The lack of glucarate uptake in the ischemic myocardium and the identical pattern of uptake in the infarcted and normal myocardium, similar to those offered by anti-myosin antibody, suggest that the loss of sarcolemmal integrity is a prerequisite for intense glucarate localization. Previous reports of glucarate localization in myocardial infarction support the concept that glucarate uptake is exclusively observed in the infarcted tissue and that there is no appreciable uptake in the ischemic zones. Orlandi et al14 reported that no glucarate uptake occurred in the canine ischemic myocardium produced by 15 minutes of coronary artery occlusion. Similarly, glucarate uptake in an isolated-perfused rat heart model was observed in the infarcted myocardium produced by 90 minutes of no perfusate flow; no glucarate uptake was observed in an ischemic tissue of 15 minutes of no flow.27 Yaoita et al13 compared glucarate uptake with 3H-deoxyglucose in a rabbit infarct model. They demonstrated a discordance in the uptake of the two radiopharmaceuticals; 3H-deoxyglucose was mainly observed in the severely ischemic zone, but predominant glucarate uptake was observed in the infarcted region. It is likely that severely ischemic tissue in their model represented histomorphologically leaky membranes. Some degree of glucarate uptake by ischemic tissue may be accounted for by the possible increase in the utilization of the sugar transport system in the hypoxic states or the existence of an admixture of predominantly ischemic myocardium and some infarcted myocardium. In vitro experiments have indeed showed glucarate uptake in the renal LLC-PK1 cells to be inhibited by fructose.13
The precise duration of glucarate positivity in the infarcted myocardium remains to be determined. In 10-day-old canine experimental infarcts, no glucarate uptake occurred.14 In rodents, glucarate uptake was seen only in 1- and 2-day-old infarcts; no uptake was obtained by the third day.15 If the early glucarate uptake is substantiated in clinical studies, glucarate imaging may complement the 2- to 10-day time window of 99mTc pyrophosphate for the precise determination of the age of the infarct. Furthermore, it may also be possible to reevaluate patients soon after the initial imaging for assessment of the final infarct size after thrombolysis in acute myocardial infarction.
Conclusions
Rapid blood clearance coupled with a strong avidity of
99mTc glucarate for the necrotic myocardium enabled
generation of high target-to-background ratios early after experimental
irreversible myocardial injury. Glucarate uptake occurred equally
effectively in reperfused and nonreperfused myocardial infarct models.
Although the percent injected dose per gram tracer localization is less
with 99mTc glucarate than with 111In-labeled
anti-myosin (due to the rapid blood clearance), target-to-background
ratios as well as the absolute radioisotope uptake were significantly
greater with 99mTc glucarate than with 111In
anti-myosin Fab. The present study suggests that high-contrast images
can be obtained within minutes after the onset of injury. If early
glucarate uptake in myocardial infarction is confirmed in clinical
studies, it may not only help direct the use of thrombolytic therapy in
patients presenting with equivocal diagnosis but also allow
differentiation of acute from recent infarcts.
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Acknowledgments |
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This work was partially supported by Small Business Innovative Research (SBIR) grant NHLBI-1R43-HL-54410-01 and by Molecular Targeting Technology, Inc (Malvern, Pa).
Received July 26, 1996; revision received November 7, 1996; accepted November 14, 1996.
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