Kimberley J. Blackwood

In Vivo SPECT Quantification of Transplanted Cell Survival After Engraftment Using 111In-Tropolone in Infarcted Canine Myocardium

Current investigations of cell transplant therapies in damaged myocardium are limited by the inability to quantify cell transplant survival in vivo. We describe how the labeling of cells with 111In can be used to monitor transplanted cell viability in a canine infarction model. Methods: We experimentally determined the contribution of the 111In signal associated with transplanted cell (TC) death and radiolabel leakage to the measured SPECT signal when 111In-labeled cells were transplanted into the myocardium. Three groups of experiments were performed in dogs. Radiolabel leakage was derived by labeling canine myocardium in situ with free 111In-tropolone (n = 4). To understand the contribution of extracellular 111In (e.g., after cell death), we developed a debris impulse response function (DIRF) by injecting lysed 111In-labeled cells within reperfused (n = 3) and nonreperfused (n = 5) myocardial infarcts and within normal (n = 3) canine myocardium. To assess the application of the modeling derived from these experiments, 111In-labeled cells were transplanted into infarcted myocardium (n = 4; 3.1 × 107 ± 5.4 × 106 cells). Serial SPECT images were acquired after direct epicardial injection to determine the time-dependent radiolabel clearance. Clearance kinetics were used to correct for 111In associated with viable TCs. Results: 111In clearance followed a biphasic response and was modeled as a biexponential with a short (\batchmode \documentclass[fleqn,10pt,legalpaper]{article} \usepackage{amssymb} \usepackage{amsfonts} \usepackage{amsmath} \pagestyle{empty} \begin{document} \(\mathrm{T}_{1/2}^{\mathrm{s}}\) \end{document}) and long (\batchmode \documentclass[fleqn,10pt,legalpaper]{article} \usepackage{amssymb} \usepackage{amsfonts} \usepackage{amsmath} \pagestyle{empty} \begin{document} \(\mathrm{T}_{1/2}^{\mathrm{l}}\) \end{document}) biologic half-life. The \batchmode \documentclass[fleqn,10pt,legalpaper]{article} \usepackage{amssymb} \usepackage{amsfonts} \usepackage{amsmath} \pagestyle{empty} \begin{document} \(\mathrm{T}_{1/2}^{\mathrm{s}}\) \end{document} was not significantly different between experimental groups, suggesting that initial losses were due to transplantation methodology, whereas the \batchmode \documentclass[fleqn,10pt,legalpaper]{article} \usepackage{amssymb} \usepackage{amsfonts} \usepackage{amsmath} \pagestyle{empty} \begin{document} \(\mathrm{T}_{1/2}^{\mathrm{l}}\) \end{document} reflected the clearance of retained 111In. DIRF had an average \batchmode \documentclass[fleqn,10pt,legalpaper]{article} \usepackage{amssymb} \usepackage{amsfonts} \usepackage{amsmath} \pagestyle{empty} \begin{document} \(\mathrm{T}_{1/2}^{\mathrm{l}}\) \end{document} of 19.4 ± 4.1 h, and the \batchmode \documentclass[fleqn,10pt,legalpaper]{article} \usepackage{amssymb} \usepackage{amsfonts} \usepackage{amsmath} \pagestyle{empty} \begin{document} \(\mathrm{T}_{1/2}^{\mathrm{l}}\) \end{document} calculated from free 111In-tropolone injected in situ was 882.7 ± 242.8 h. The measured \batchmode \documentclass[fleqn,10pt,legalpaper]{article} \usepackage{amssymb} \usepackage{amsfonts} \usepackage{amsmath} \pagestyle{empty} \begin{document} \(\mathrm{T}_{1/2}^{\mathrm{l}}\) \end{document} for TCs was 74.3 h and was 71.2 h when corrections were applied. Conclusion: A new quantitative method to assess TC survival in myocardium using SPECT and 111In has been introduced. At the limits, method accuracy is improved if appropriate corrections are applied. In vivo 111In imaging most accurately describes cell viability half-life if \batchmode \documentclass[fleqn,10pt,legalpaper]{article} \usepackage{amssymb} \usepackage{amsfonts} \usepackage{amsmath} \pagestyle{empty} \begin{document} \(\mathrm{T}_{1/2}^{\mathrm{l}}\) \end{document} is between 20 h and 37 d.

Can the Inflammatory Response Be Evaluated Using 18F-FDG Within Zones of Microvascular Obstruction After Myocardial Infarction?

Inflammation that occurs after acute myocardial infarction plays a pivotal role in healing by facilitating the creation of a supportive scar. 18F-FDG, which is taken up avidly by macrophages, has been proposed as a marker of cell-based inflammation. However, its reliability as an accurate indicator of inflammation has not been established, particularly in the early postinfarction period when regional myocardial perfusion is often severely compromised. Methods: Nine adult dogs underwent left anterior descending coronary occlusion with or without reperfusion. Animals were imaged between 7 and 21 d after infarction with PET/MR imaging after bolus injection of gadolinium-diethylenetriaminepentaacetic acid (DTPA), bolus injection of 18F-FDG, bolus injection of 99Tc-DTPA to simulate the distribution of gadolinium-DTPA (which represents its partition coefficient in well-perfused tissue), and injection of 111In-labeled white blood cells 24 h earlier. After sacrifice, myocardial tissue concentrations of 18F, 111In, and 99Tc were determined in a well counter. Linear regression analysis evaluated the relationships between the concentrations of 111In and 18F and the dependence of the ratio of 111In/18F to the apparent distribution volume of 99mTc-DTPA. Results: In 7 of 9 animals, 111In increased as 18F increased with the other 2 animals, showing weak negative slopes. With respect to the dependence of 111In/18F with partition coefficient, 4 animals showed no dependence and 4 showed a weak positive slope, with 1 animal showing a negative slope. Further, in regions of extensive microvascular obstruction, 18F significantly underestimated the extent of the presence of 111In. Conclusion: In the early post–myocardial infarction period, 18F-FDG PET imaging after a single bolus administration may underestimate the extent and degree of inflammation within regions of microvascular obstruction.

Hybrid SPECT/cardiac-gated first-pass perfusion CT: locating transplanted cells relative to infarcted myocardial targets

PURPOSE A challenge with cardiac cell therapy is determining the location of cells relative to infarct tissue. As cells are viable following ¹¹¹In-labeling, and first-pass CT imaging can identify regions of myocardial infarction, we evaluated the feasibility of a SPECT/CT system to localize cells relative to infarcted myocardium in a canine model. METHODS Ten canines underwent surgical ligation of the left-anterior-descending artery and endothelial progenitor cells labeled with ¹¹¹In-tropolone were transplanted endocardially or epicardially. SPECT/CT was performed on day of transplantation, 4 and 10 days post-transplantation. For each imaging session first-pass perfusion CT was performed to delineate the area of reduced perfusion. SPECT and first-pass CT images were fused and evaluated. Contrast-to-noise ratios (CNR) were calculated for ¹¹¹In-SPECT images to evaluate cell detection. RESULTS The zone of reduced perfusion was well delineated on first-pass perfusion CT in all canines. The ¹¹¹In signal was visualized within this zone in all cases. Analysis of the CNRs suggests that cells may be followed for 11 effective half-lives using the images from first-pass perfusion CT to provide the anatomic landmarks. CONCLUSION In the setting of an acute myocardial infarction SPECT/[first-pass perfusion CT] is an effective hybrid platform for the localization of cells in relation to the area of reduced blood flow.

Imaging of post-infarction myocardial inflammation with hybrid FDG PET/MR: feasibilty and preliminary findings in a canine model

Background An understanding of inflammation following myocardial infarction may be of importance in the development of novel therapeutics to limit the development of heart failure following myocardial injury. However, the quantification of inflammation in this setting continues to be challenging. Hybrid FDG PET/MR may offer a non-invasive in vivo solution through intrinsic registration of these complementary modalities. This study sought to validate its use in a canine model of myocardial infarction (MI).

Comparison of 111In Leakage from Labeled Endocardial and Epicardial Cells: Impact on Modeling Viability of Cells to Be Transplanted into Myocardium

Introduction. Previously we proposed a cellular imaging technique to determine the surviving fraction of transplanted cells in vivo. Epicardial kinetics using Indium-111 determined the Debris Impulse Response Function (DIRF) and leakage coefficient parameters. Convolution-based modeling which corrected for these signal contributions indicated that 111In activity was quantitative of cell viability with half-lives within 20 hrs to 37 days. We determine if the 37-day upper limit remains valid for endocardial injections by comparing previous epicardial cell leakage parameter estimates to those for endocardial cells. Methods. Normal canine myocardium was injected (111In-tropolone) epicardially (9 injections) or endocardially (10 injections). Continuous whole body and SPECT scans for 5 hours were acquired with three weekly follow-up imaging sessions up to 20–26 days. Time-activity curves evaluated each injection type. Results. The epicardial and endocardial kinetics were not significantly different (Epi: 1286 ± 253; Endo: 1567 ± 470 hours P = .62). Conclusion. The original epicardial estimate of leakage kinetics has been validated for use in endocardial injections.

Sci—Fri AM: Imaging — 09: Hybrid SPECT and First‐Pass Perfusion CT: Application in Cell Localization

Purpose: A challenge with cardiac cell therapy is determining the location of cells relative to infarct tissue. As cells are viable following 111In‐labeling, and first‐pass CTimaging can identify regions of myocardial infarction, we evaluated the feasibility of a SPECT/CT system to localize and track cells relative to infarcted myocardium in a canine model. Methods: To determine accuracy of SPECT/CT registration, images were acquired of capillary tubes filled with CT and SPECT contrast agents. Accuracy was measured by comparing locations of tube centroids in SPECT and CT. Ten canines underwent surgical ligation of the left‐anterior‐descending artery and endothelial progenitor cells labeled with 111In‐tropolone were transplanted endocardially or epicardially. SPECT/CT was performed on day of transplantation, 4, and 10 days post‐transplantation. For each imaging session first‐pass perfusion CT was performed to delineate the infarct zone. SPECT and first‐pass CTimages were fused and evaluated. Delayed‐enhanced MRI was performed to validate CT infarct localization. Contrast‐to‐noise ratios (CNR) were calculated for 111In‐SPECT images to evaluate cell detection. Results: Phantom SPECT/CT registration accuracy was approximately 1mm. The infarct zone was well delineated on first‐pass perfusion CT in all canines and correlated well with MRI findings. 111In signal was visualized within the infarct zone in all cases. Analysis of the CNRs suggests that cells can be followed for 11 effective half‐lives provided the location of the cells can be inferred by first‐pass CT.Conclusion: SPECT/[First‐Pass Perfusion CT] is an effective hybrid platform for the localization and tracking of stem cells in relation to infarct tissue.

Sci‐Fri PM Imaging‐11: Evaluation of SPECT/CT for Regenerative Medicine Applications: Tracking Transplanted Cells in a Large Animal Model

Introduction: The purpose of this study was to characterize the performance of dual‐radionuclide SPECT/CT in quantitative tasks associated with tracking transplanted cellsin vivo. Previous studies identified matters of hardware design, whereas we focus on biological variables impacting performance such as non‐specific uptake of radiolabels, and radiolabel dilution with cell colony growth.Methods: Using experimental SPECT/CT data, a canine digital phantom was developed of in vitro 111 In ‐radiolabeled stem cells, transfected with a reporter gene, transplanted into canine infarcted myocardium, and interrogated post‐transplantation using a peripherally‐injected 131 I ‐radiolabeled reporter probe. Single‐ and dual‐head SPECT/CT acquisitions were simulated. Simulations included physical effects of radionuclide‐specific attenuation, scatter, resolution, and dual‐radionuclide crosstalk. CT data was used to simulate photon attenuation. Performance was characterized using an estimation task, incorporating the statistical properties of SPECTimaging, where the precision of parameter estimates ( 111 In and 131 I radiolabel quantity, cell colony size and location, and background) was tracked as the phantom evolved to simulate 111 In ‐label efflux, cell colony growth, and improved reporter probe specificity. Results: In vitro pre‐labeling of transplanted cells with 111 In improved precision of parameter estimates via a priori size and location information. Precision of radiolabel quantity estimates improved with cell colony growth, despite 111 In radiolabel dilution; size and location parameters were influenced little. Precision of radiolabel quantity estimates improved with reduced 131 I reporter probe non‐specific uptake. Conclusion: The performance of SPECT/CT in cell tracking is influenced strongly by biological variables. These should be considered when planning experiments or developing SPECT/CT technology for cell tracking.

Sci‐Fri AM General‐07: Dual Isotope SPECT to Track Transplanted Cells in Canine Myocardium Using Molecular Imaging

Introduction: Non‐invasive cell tracking techniques that provide repetitive and functional assessments of transplanted cells will be an asset in cell‐based therapies. Specifically, reporter gene (RG) expression may be used to signal certain molecular events as they occur in vivo. Objective: To compare the in vivo kinetics of a radiolabeled reporter probe (RP) specifically targeted by RG expression to a non‐specific radiolabel (NSL) in canine bone marrow cells (BMC). Cells were co‐labeled in vitro, transplanted into normal canine myocardium, and tracked using dual‐isotope SPECT. Methods: Following canine bone marrow harvest (n=1), BMCs were isolated, transfected with RG, and grown in culture for 4 weeks. Transfected cells (5.4×106) were then incubated with RP and NSL followed by direct injection into the left canine ventricle. The recipient of transplanted cells was also the donor. Forty 1‐hr SPECTimages were acquired to obtain washout kinetics of RP and NSL. Time activity curves (TAC) were generated from SPECT region of interest analysis. After sacrifice, ex vivo measurements of RP and NSL activity remaining in the myocardium were made with a high purity germanium gamma‐ray well counter. Results: Decay‐corrected TACs demonstrated faster washout kinetics for RP compared to NSL with biological half‐lives of 19 and 39 hours, respectively. This suggests that RP loss is greater than that attributed to cell death alone and demonstrates differences in label stability. Ex vivotissue analysis confirmed these results. Conclusion: Multispectral SPECT shows potential for monitoring RG stability and expression to follow cell viability and function non‐invasively in large animals.