Dynamic SPECT measurement of absolute myocardial blood flow in a porcine model.

UNLABELLED: Absolute myocardial blood flow (MBF) and myocardial flow reserve (MFR) provide incremental diagnostic and prognostic information over relative perfusion alone. Recent development of dedicated cardiac SPECT cameras with better sensitivity and temporal resolution make dynamic SPECT imaging more practical. In this study, we evaluate the measurement of MBF using a multipinhole dedicated cardiac SPECT camera in a pig model of rest and transient occlusion at stress using 3 common tracers: (201)Tl, (99m)Tc-tetrofosmin, and (99m)Tc-sestamibi. METHODS: Animals (n = 19) were injected at rest/stress with (99m)Tc radiotracers (370/1,100 MBq) or (201)Tl (37/110 MBq) with a 1-h delay between rest and dipyridamole stress. With each tracer, microspheres were injected simultaneously as the gold standard measurement for MBF. Dynamic images were obtained for 11 min starting with each injection. Residual resting activity was subtracted from stress data and images reconstructed with CT-based attenuation correction and energy window-based scatter correction. Dynamic images were processed with kinetic analysis software using a 1-tissue-compartment model to obtain the uptake rate constant K(1) as a function of microsphere MBF. RESULTS: Measured extraction fractions agree with those obtained previously using ex vivo techniques. Converting K(1) back to MBF using the measured extraction fractions produced accurate values and good correlations with microsphere MBF: r = 0.75-0.90 (P < 0.01 for all). The correlation in the MFR was between r = 0.57 and 0.94 (P < 0.01). CONCLUSION: Noninvasive measurement of absolute MBF with stationary dedicated cardiac SPECT is feasible using common perfusion tracers.

Biodistribution and radiodosimetry of a novel myocardial perfusion tracer 123I-CMICE-013 in healthy rats.

BACKGROUND: 123I-CMICE-013 is a novel radiotracer previously reported to have promising characteristics for single-photon emission computed tomography (SPECT) myocardial perfusion imaging. We evaluated the biokinetics and radiodosimetry of this rotenone-like 123I-labeled tracer in a microSPECT imaging-based study. METHODS: 37 to 111 MBq of 123I-CMICE-013 was synthesized and administered intravenously to 14 healthy rats. Images were acquired with a microSPECT/CT camera at various time intervals and reconstructed to allow activity quantification in the tissues of interest. Radiation dosage resulted from the injection of 123I-CMICE-013 was estimated base on the biodistribution data. Tissue uptake values from image analysis were verified by gamma-counting dissected organs ex vivo. RESULTS: The heart/stomach and heart/intestine uptake ratios peaked shortly after the injection of 123I-CMICE-013, meanwhile the heart/liver ratio reached 2 as early as at 23 min post-injection. Little activity was observed in the lung and overnight clearance was significant in most of the measured tissues. The radiation dosimetry analysis based on the time-activity curves provided an estimate of the effective human dose of 6.99E-03 mSv/MBq using ICRP 60 and 7.15E-03 mSv/MBq using ICRP 103, which is comparable to the popular myocardium perfusion imaging (MPI) agents such as 99mTc-tetrofosmin and 99mTc-sestamibi, as well as other 123I-based radiotracers. CONCLUSIONS: 123I-CMICE-013 demonstrated desirable characteristics in its biokinetic and radiodosimetric profiles, supporting its potential application as a novel myocardial perfusion imaging agent.

Characterization of the four isomers of (123)I-CMICE-013: a potential SPECT myocardial perfusion imaging agent.

UNLABELLED: Myocardial perfusion imaging (MPI) with single photon emission computed tomography (SPECT) is widely used in the assessment of coronary artery disease (CAD). We have developed (123)I-CMICE-013 based on rotenone, a mitochondrial complex I (MC-1) inhibitor, as a promising new MPI agent. Our synthesis results in a mixture of four species of (123)I-CMICE-013 A, B, C, D. In this study, we separated the four species and evaluated their biodistribution and imaging properties. The cold analogs (127)I-CMICE-013 A, B, C, D were isolated and characterized and their chemical structures proposed. METHODS: (123)I-CMICE-013 was synthesized by radiolabeling rotenone with Na(123)I in trifluoroacetic acid (TFA) with iodogen as the oxidizing agent at 60°C for 45min, and the four species were separated by RP-HPLC. The cold analogs (127)I-CMICE-013 A, B, C and D were isolated with a similar procedure and characterized by NMR and mass spectrometry. Biodistribution and microSPECT imaging studies were carried out on normal rats. RESULTS: We propose the mechanism of the rotenone iodination and the structures of the four species. First, I(+) forms an intermediate three-membered ring with 6' and 7' carbons. Second, the lone electron pair of the water molecule attacks the 6' or 7'-carbon, following by the formation of 6'-OH, and 7'-I bonds as in major products C and D, or 6'-I and 7'-OH bonds as in minor products A and B. The weaker 6'-I bond in the intermediate prompts the nucleophilic attachment of water at the favorable 6'-carbon to generate C and D. MicroSPECT images of (123)I-CMICE-013 A, B, C, D in rats showed clear visualization of myocardium and little interference from lung and liver. The imaging time activity curves and biodistribution data showed complex profiles for the four isomers, which is not expected from the structure activity relationship theory. CONCLUSION: (123/127)I-CMICE-013 A and B are constitutional isomers with C and D, while A and C are diastereomers of B and D, respectively. Overall, the biological characteristics of the four species are not correlated perfectly with their molecular structures.

Toxicological evaluation of a rotenone derivative in rodents for clinical myocardial perfusion imaging.

Myocardial perfusion scintigraphy is a valuable clinical tool for assessing coronary blood flow deficits in patients. We recently synthesized a new iodinated compound ((123)I-CMICE-013) based on rotenone and showed that it has excellent performance as a radiotracer for myocardial perfusion imaging. Here, we describe the cellular toxicity and subacute toxicity of CMICE-013 in rats. Cultured hepatocytes displayed sensitivity to rotenone but not CMICE-013 at equimolar concentrations. Following i.v. injection of CMICE-013 for 14 days, body weight, ambulation, behavior, grooming, guarding (abdominal, muscular), pale conjunctivae, and food intake were observed. Biochemical, hematological, and histopathological changes in tissues (heart, liver, kidney, spleen, lung, and brain) and echocardiography at pre- and post-dosing were also examined. All animals responded well to the daily injections of CMICE-013 and showed no mortality or adverse reactions with respect to the parameters above. Subacute i.v. injections at high- (5 µg/kg) and low (1 µg/kg)-dose levels did not result in any significant changes to either biochemical or hematological parameters and no detectable changes in histopathology compared to the vehicle or untreated animals. Echocardiographic analyses, including the measurements of cardiac function and anatomy (wall thickness, left atrial size, and left ventricular mass), were not different at pre- versus post-dose measures and were not different compared to the vehicle or untreated animals. Our observations in small animals reveal that CMICE-013 induces minimal toxicity when delivered intravenously for 14 days.

Synthesis and characterization of 123I-CMICE-013: a potential SPECT myocardial perfusion imaging agent.

UNLABELLED: Coronary artery disease (CAD) is a major cause of death in Canada and the United States. Single photon emission computed tomography (SPECT) myocardial perfusion imaging (MPI) is a useful diagnostic test in the management of patients with CAD. The widely used SPECT MPI agents, (99m)Tc sestamibi and (99m)Tc tetrofosmin, exhibit less than ideal pharmacokinetic properties with decreasing uptake with higher flows. (123)I has a similar energy as (99m)Tc, an ideal half life, and is readily available from cyclotrons. The objective of this study was to develop an (123)I labeled MPI agent based on rotenone, a mitochondrial complex I inhibitor, as an alternative to currently available SPECT MPI agents. METHODS: (123)I-CMICE-013 was synthesized by radiolabeling rotenone with (123)I in trifluoroacetic acid (TFA) with iodogen as the oxidizing agent at 60 °C for 45 min, followed by RP-HPLC purification. The product was formulated in 5% EtOH in 10 mM NaOAc pH 6.5. The inactive analog (127)I-CMICE-013 was isolated and characterized by NMR and mass spectrometry, and the structure determined. Micro-SPECT imaging studies were carried out in normal and infarcted rats. Biodistribution studies were performed in normal rats at 2 h (n=6) and 24 h (n=8) post injection (p.i.). RESULTS: (123)I-CMICE-013 was isolated with >95% radiochemical purity and high specific activity (14.8-111 GBq/µmol; 400-3000 mCi/µmol). Structural analysis showed that rotenone was iodinated at 7'-position, with removal of the 6',7'-double bond, and addition of a hydroxy group at 6'-position. MicroSPECT images in normal rats demonstrated homogeneous and sustained myocardial uptake with minimal interference from lung and liver. Absent myocardial perfusion was clearly identified in rats with permanent left coronary artery ligation and ischemia-reperfusion injury. In vivo biodistribution studies in normal rats at 2 h p.i. showed significant myocardial uptake (2.01±0.48%ID/g) and high heart to liver (2.98±0.93), heart to lung (4.11±1.04) and heart to blood (8.37±3.97) ratios. At 24 h p.i., the majority of (123)I-CMICE-013 was cleared from tissues, and a significant amount of tracer was found in the thyroid, indicating in vivo deiodination of the tracer. CONCLUSION: (123)I-CMICE-013 is a promising new radiotracer for SPECT MPI with high myocardial uptake, very good target to background ratios and favorable biodistribution characteristics.

Intravascular and extravascular microvessel formation in chronic total occlusions a micro-CT imaging study.

OBJECTIVES: The purpose of this study was to characterize the 3-dimensional structure of intravascular and extravascular microvessels during chronic total occlusion (CTO) maturation in a rabbit model. BACKGROUND: Intravascular microchannels are an important component of a CTO and may predict guidewire crossability. However, temporal changes in the structure and geographic localization of these microvessels are poorly understood. METHODS: A total of 39 occlusions were created in a rabbit femoral artery thrombin model. Animals were sacrificed at 2, 6, 12, and 24 weeks (n > or =8 occlusions per time point). The arteries were filled with a low viscosity radio-opaque polymer compound (Microfil) at 150 mm Hg pressure. Samples were scanned in a micro-computed tomography system to obtain high-resolution volumetric images. Analysis was performed in an image processing package that allowed for labeling of multiple materials. RESULTS: Two distinct types of microvessels were observed: circumferentially oriented "extravascular" and longitudinally oriented "intravascular" microvessels. Extravascular microvessels were evident along the entire CTO length and maximal at the 2-week time point. There was a gradual and progressive reduction in extravascular microvessels over time, with very minimal microvessels evident beyond 12 weeks. In contrast, intravascular microvessel formation was delayed, with peak vascular volume at 6 weeks, followed by modest reductions at later time points. Intravascular microvessel formation was more prominent in the body compared with that in the proximal and distal ends of the CTO. Sharply angulated connections between the intravascular and extravascular microvessels were present at all time points, but most prominent at 6 weeks. At later time points, the individual intravascular microvessels became finer and more tortuous, although the continuity of these microvessels remained constant beyond 2 weeks. CONCLUSIONS: Differences are present in the temporal and geographic patterns of intravascular and extravascular microvessel formation during CTO maturation.

Natural history of experimental arterial chronic total occlusions.

OBJECTIVES: We sought to perform the first systematic study of the natural history of chronic total arterial occlusions (CTOs) in an experimental model. BACKGROUND: Angioplasty of CTOs has low success rates. The structural and perfusion changes during CTO maturation, which may adversely affect angioplasty outcome, have not been systematically studied. METHODS: Occlusions were created in 63 rabbit femoral arteries by thrombin injection. Histology, contrast-enhanced magnetic resonance imaging, relative blood volume (RBV) index, and micro-computed tomography imaging were analyzed at 2, 6, 12, and 18 to 24 weeks. RESULTS: Early changes were characterized by an acute inflammatory response and negative arterial remodeling, with >70% reduction of arterial cross-sectional area (CSA) from 2 to 6 weeks. Intraluminal neovascularization of the CTO occurred with a 2-fold increase in total (media + intima) microvessel CSA from 2 to 6 weeks (0.014 +/- 0.002 mm2 to 0.023 +/- 0.005 mm2, p = 0.0008) and a 3-fold increase in RBV index (5.1 +/- 1.9% to 16.9 +/- 2.7%, p = 0.0008). However at later time periods, there were significant reductions in both RBV (3.5 +/- 1.1%, p < 0.0001) and total microvessel CSA (0.017 +/- 0.002 mm2, p = 0.011). Micro-computed tomography imaging demonstrated a corkscrew-like recanalization channel at the proximal end at 6 weeks that regressed at later time points. These vascular changes were accompanied by a marked decrease in proteoglycans and accumulation of a collagen-enriched extracellular matrix, particularly at the entrance ("proximal fibrous cap"). CONCLUSIONS: This study is the first to systematically analyze compositional changes occurring during CTO maturation, which may underlie angioplasty failure. Negative remodeling, regression of intraluminal channels, and CTO perfusion, together with the accumulation of dense collagen, may represent important targets for novel therapeutic interventions.

Severe hemodilutional anemia increases cerebral tissue injury following acute neurotrauma.

Anemia may worsen neurological outcomes following traumatic brain injury (TBI) by undefined mechanisms. We hypothesized that hemodilutional anemia accentuates hypoxic cerebral injury following TBI. Anesthetized rats underwent unilateral TBI or sham injury (n > or = 7). Target hemoglobin concentrations between 50 and 70 g/l were achieved by exchanging 40-50% of the blood volume (1:1) with pentastarch. The effect of TBI, anemia, and TBI-anemia was assessed by measuring brain tissue oxygen tension (Pbr(O(2))), regional cerebral blood flow (rCBF), jugular venous oxygen saturation (Sjv(O(2))), cerebral contusion area, and nuclear staining for programmed cell death. Baseline postinjury Pbr(O(2)) values in the TBI and TBI-anemia groups (9.3 +/- 1.3 and 11.3 +/- 4.1 Torr, respectively) were lower than the uninjured controls (18.2 +/- 5.2 Torr, P < 0.05 for both). Hemodilution caused a further reduction in Pbr(O(2)) in the TBI-anemia group relative to the TBI group without anemia (7.8 +/- 2.7 vs. 14.8 +/- 3.9 Torr, P < 0.05). The rCBF remained stable after TBI and increased comparably after hemodilution in both anemia and TBI-anemia groups. The Sjv(O(2)) was elevated after TBI (87.4 +/- 8.9%, P < 0.05) and increased further following hemodilution (95.0 +/- 1.6%, P < 0.05). Cerebral contusion area and nuclear counts for programmed cell death were increased following TBI-anemia (4.1 +/- 3.0 mm(2) and 686 +/- 192, respectively) relative to TBI alone (1.3 +/- 0.3 mm(2) and 404 +/- 133, respectively, P < 0.05 for both). Hemodilutional anemia reduced cerebral Pbr(O(2)) and oxygen extraction and increased cell death following TBI. These results support our hypothesis that acute anemia accentuated hypoxic cerebral injury after neurotrauma.

Hypoxia alters the expression of inhibitor of apoptosis proteins after brain trauma in the mouse.

Hypoxia worsens brain injury following trauma, but the mechanisms remain unclear. The purpose of this study was to determine the effect of traumatic brain injury (TBI) and secondary hypoxia (9% oxygen) on apoptosis-related protein expression, cell death, and behavior. Using a murine weight-drop model, TBI led to an early (6 h) increase followed by a later (24 h) decrease in neuronal apoptosis inhibitor protein (NAIP) expression in the olfactory and motor cortex; in contrast, TBI led to a sustained (6 h to 7 days) increase in NAIP in the striatum. The peak increase in the expression of NAIP (6-12 h) following TBI alone was delayed (1-7 days) when hypoxia was added to TBI. Hypoxia following TBI further depleted other apoptosis inhibitor proteins (IAPs) and activated caspases, as well as increased contusion size and worsened cell death. Hypoxia added to TBI also increased motor and feeding activity on days 2 and 4 compared to TBI alone. Hypoxia without TBI had no effect on the expression of IAPs or cell death. These findings show that IAPs have a potential role in the increased vulnerability of brain cells to hypoxia following TBI, and have implications for configuring future therapies for TBI.