Samuel M. Johnston

Geometric calibration for a dual tube/detector micro-CT system.

The authors describe a dual tube/detector micro-computed tomography (micro-CT) system that has the potential to improve temporal resolution and material contrast in small animal imaging studies. To realize this potential, it is necessary to precisely calibrate the geometry of a dual micro-CT system to allow the combination of projection data acquired with each individual tube/detector in a single reconstructed image. The authors present a geometric calibration technique that uses multiple projection images acquired with the two imaging chains while rotating a phantom containing a vertical array of regularly spaced metallic beads. The individual geometries of the imaging chains are estimated from the phantom projection images using analytical methods followed by a refinement procedure based on nonlinear optimization. The geometric parameters are used to create the cone beam projection matrices required by the reconstruction process for each imaging chain. Next, a transformation between the two projection matrices is found that allows the combination of projection data in a single reconstructed image. The authors describe this technique, test it with a series of computer simulations, and then apply it to data collected from their dual tube/detector micro-CT system. The results demonstrate that the proposed technique is accurate, robust, and produces images free of misalignment artifacts.

4D micro-CT for cardiac and perfusion applications with view under sampling*

Micro-CT is commonly used in preclinical studies to provide anatomical information. There is growing interest in obtaining functional measurements from 4D micro-CT. We report here strategies for 4D micro-CT with a focus on two applications: (i) cardiac imaging based on retrospective gating and (ii) pulmonary perfusion using multiple contrast injections/rotations paradigm. A dual source micro-CT system is used for image acquisition with a sampling rate of 20 projections per second. The cardiac micro-CT protocol involves the use of a liposomal blood pool contrast agent. Fast scanning of free breathing mice is achieved using retrospective gating. The ECG and respiratory signals are used to sort projections into ten cardiac phases. The pulmonary perfusion protocol uses a conventional contrast agent (Isovue 370) delivered by a micro-injector in four injections separated by 2 min intervals to allow for clearance. Each injection is synchronized with the rotation of the animal, and each of the four rotations is started with an angular offset of 22.5 from the starting angle of the previous rotation. Both cardiac and perfusion protocols result in an irregular angular distribution of projections that causes significant streaking artifacts in reconstructions when using traditional filtered backprojection (FBP) algorithms. The reconstruction involves the use of the point spread function of the micro-CT system for each time point, and the analysis of the distribution of the reconstructed data in the Fourier domain. This enables us to correct for angular inconsistencies via deconvolution and identify regions where data is missing. The missing regions are filled with data from a high quality but temporally averaged prior image reconstructed with all available projections. Simulations indicate that deconvolution successfully removes the streaking artifacts while preserving temporal information. 4D cardiac micro-CT in a mouse was performed with adequate image quality at isotropic voxel size of 88 µm and 10 ms temporal resolution. 4D pulmonary perfusion images were obtained in a mouse at 176 µm and 687 ms temporal resolution. Compared with FBP reconstruction, the streak reduction ratio is 70% and the contrast to noise ratio is 2.5 times greater in the deconvolved images. The radiation dose associated with the proposed methods is in the range of a typical micro-CT dose (0.17 Gy for the cardiac study and 0.21 Gy for the perfusion study). The low dose 4D micro-CT imaging presented here can be applied in high-throughput longitudinal studies in a wide range of applications, including drug safety and cardiopulmonary phenotyping.

Dual-energy micro-CT of the rodent lung

The purpose of this work is to investigate the use of dual-energy micro-computed tomography (CT) for the estimation of vascular, tissue, and air fractions in rodent lungs using a postreconstruction three material decomposition method. Using simulations, we have estimated the accuracy limits of the decomposition for realistic micro-CT noise levels. Next, we performed experiments involving ex vivo lung imaging in which intact rat lungs were carefully removed from the thorax, injected with an iodine-based contrast agent, and then inflated with different volumes of air (n = 2). Finally, we performed in vivo imaging studies in C57BL/6 mice (n = 5) using fast prospective respiratory gating in end inspiration and end expiration for three different levels of positive end expiratory pressure (PEEP). Before imaging, mice were injected with a liposomal blood pool contrast agent. The three-dimensional air, tissue, and blood fraction maps were computed and analyzed. The results indicate that separation and volume estimation of the three material components of the lungs are possible. The mean accuracy values for air, blood, and tissue were 93, 93, and 90%, respectively. The absolute accuracy in determining all fraction materials was 91.6%. The coefficient of variation was small (2.5%) indicating good repeatability. The minimum difference that we could detect in material fractions was 15%. As expected, an increase in PEEP levels for the living mouse resulted in statistically significant increases in air fractions at end expiration but no significant changes at end inspiration. Our method has applicability in preclinical pulmonary studies where changes in lung structure and gas volume as a result of lung injury, environmental exposures, or drug bioactivity would have important physiological implications.

Lung perfusion imaging in small animals using 4D micro-CT at heartbeat temporal resolution

PURPOSE Quantitative in vivo imaging of lung perfusion in rodents can provide critical information for preclinical studies. However, the combined challenges of high temporal and spatial resolution have made routine quantitative perfusion imaging difficult in small animals. The purpose of this work is to demonstrate 4D micro-CT for perfusion imaging in rodents at heartbeat temporal resolution and isotropic spatial resolution. METHODS We have recently developed a dual tube/detector micro-CT scanner that is well suited to capture first pass kinetics of a bolus of contrast agent used to compute perfusion information. Our approach is based on the paradigm that similar time density curves can be reproduced in a number of consecutive, small volume injections of iodinated contrast agent at a series of different angles. This reproducibility is ensured by the high-level integration of the imaging components of our system with a microinjector, a mechanical ventilator, and monitoring applications. Sampling is controlled through a biological pulse sequence implemented in LABVIEW. Image reconstruction is based on a simultaneous algebraic reconstruction technique implemented on a graphic processor unit. The capabilities of 4D micro-CT imaging are demonstrated in studies on lung perfusion in rats. RESULTS We report 4D micro-CT imaging in the rat lung with a heartbeat temporal resolution (approximately 150 ms) and isotropic 3D reconstruction with a voxel size of 88 microm based on sampling using 16 injections of 50 microL each. The total volume of contrast agent injected during the experiments (0.8 mL) was less than 10% of the total blood volume in a rat. This volume was not injected in a single bolus, but in multiple injections separated by at least 2 min interval to allow for clearance and adaptation. We assessed the reproducibility of the time density curves with multiple injections and found that these are very similar. The average time density curves for the first eight and last eight injections are slightly different, i.e., for the last eight injections, both the maximum of the average time density curves and its area under the curve are decreased by 3.8% and 7.2%, respectively, relative to the average time density curves based on the first eight injections. The radiation dose associated with our 4D micro-CT imaging is 0.16 Gy and is therefore in the range of a typical micro-CT dose. CONCLUSIONS 4D micro-CT-based perfusion imaging demonstrated here has immediate application in a wide range of preclinical studies such as tumor perfusion, angiogenesis, and renal function. Although our imaging system is in many ways unique, we believe that our approach based on the multiple injection paradigm can be used with the newly developed flat-panel slip-ring-based micro-CT to increase their temporal resolution in dynamic perfusion studies.

Temporal and spectral imaging with micro-CT.

PURPOSE Micro-CT is widely used for small animal imaging in preclinical studies of cardiopulmonary disease, but further development is needed to improve spatial resolution, temporal resolution, and material contrast. We present a technique for visualizing the changing distribution of iodine in the cardiac cycle with dual source micro-CT. METHODS The approach entails a retrospectively gated dual energy scan with optimized filters and voltages, and a series of computational operations to reconstruct the data. Projection interpolation and five-dimensional bilateral filtration (three spatial dimensions + time + energy) are used to reduce noise and artifacts associated with retrospective gating. We reconstruct separate volumes corresponding to different cardiac phases and apply a linear transformation to decompose these volumes into components representing concentrations of water and iodine. Since the resulting material images are still compromised by noise, we improve their quality in an iterative process that minimizes the discrepancy between the original acquired projections and the projections predicted by the reconstructed volumes. The values in the voxels of each of the reconstructed volumes represent the coefficients of linear combinations of basis functions over time and energy. We have implemented the reconstruction algorithm on a graphics processing unit (GPU) with CUDA. We tested the utility of the technique in simulations and applied the technique in an in vivo scan of a C57BL∕6 mouse injected with blood pool contrast agent at a dose of 0.01 ml∕g body weight. Postreconstruction, at each cardiac phase in the iodine images, we segmented the left ventricle and computed its volume. Using the maximum and minimum volumes in the left ventricle, we calculated the stroke volume, the ejection fraction, and the cardiac output. RESULTS Our proposed method produces five-dimensional volumetric images that distinguish different materials at different points in time, and can be used to segment regions containing iodinated blood and compute measures of cardiac function. CONCLUSIONS We believe this combined spectral and temporal imaging technique will be useful for future studies of cardiopulmonary disease in small animals.

Quantitative blood flow measurements in the small animal cardiopulmonary system using digital subtraction angiography

PURPOSE The use of preclinical rodent models of disease continues to grow because these models help elucidate pathogenic mechanisms and provide robust test beds for drug development. Among the major anatomic and physiologic indicators of disease progression and genetic or drug modification of responses are measurements of blood vessel caliber and flow. Moreover, cardiopulmonary blood flow is a critical indicator of gas exchange. Current methods of measuring cardiopulmonary blood flow suffer from some or all of the following limitations--they produce relative values, are limited to global measurements, do not provide vasculature visualization, are not able to measure acute changes, are invasive, or require euthanasia. METHODS In this study, high-spatial and high-temporal resolution x-ray digital subtraction angiography (DSA) was used to obtain vasculature visualization, quantitative blood flow in absolute metrics (ml/min instead of arbitrary units or velocity), and relative blood volume dynamics from discrete regions of interest on a pixel-by-pixel basis (100 x 100 microm2). RESULTS A series of calibrations linked the DSA flow measurements to standard physiological measurement using thermodilution and Ficks method for cardiac output (CO), which in eight anesthetized Fischer-344 rats was found to be 37.0 +/- 5.1 ml/min. Phantom experiments were conducted to calibrate the radiographic density to vessel thickness, allowing a link of DSA cardiac output measurements to cardiopulmonary blood flow measurements in discrete regions of interest. The scaling factor linking relative DSA cardiac output measurements to the Ficks absolute measurements was found to be 18.90 x CODSA = COFick. CONCLUSIONS This calibrated DSA approach allows repeated simultaneous visualization of vasculature and measurement of blood flow dynamics on a regional level in the living rat.

Dual-energy micro-CT imaging for differentiation of iodine- and gold-based nanoparticles

Spectral CT imaging is expected to play a major role in the diagnostic arena as it provides material decomposition on an elemental basis. One fascinating possibility is the ability to discriminate multiple contrast agents targeting different biological sites. We investigate the feasibility of dual energy micro-CT for discrimination of iodine (I) and gold (Au) contrast agents when simultaneously present in the body. Simulations and experiments were performed to measure the CT enhancement for I and Au over a range of voltages from 40-to-150 kVp using a dual source micro-CT system. The selected voltages for dual energy micro-CT imaging of Au and I were 40 kVp and 80 kVp. On a massconcentration basis, the relative average enhancement of Au to I was 2.75 at 40 kVp and 1.58 at 80 kVp. We have demonstrated the method in a preclinical model of colon cancer to differentiate vascular architecture and extravasation. The concentration maps of Au and I allow quantitative measure of the bio-distribution of both agents. In conclusion, dual energy micro-CT can be used to discriminate probes containing I and Au with immediate impact in pre-clinical research.

Assessing the Radiation Response of Lung Cancer with Different Gene Mutations Using Genetically Engineered Mice

Purpose: Non-small cell lung cancers (NSCLC) are a heterogeneous group of carcinomas harboring a variety of different gene mutations. We have utilized two distinct genetically engineered mouse models of human NSCLC (adenocarcinoma) to investigate how genetic factors within tumor parenchymal cells influence the in vivo tumor growth delay after one or two fractions of radiation therapy (RT). Materials and Methods: Primary lung adenocarcinomas were generated in vivo in mice by intranasal delivery of an adenovirus expressing Cre-recombinase. Lung cancers expressed oncogenic KrasG12D and were also deficient in one of two tumor suppressor genes: p53 or Ink4a/ARF. Mice received no radiation treatment or whole lung irradiation in a single fraction (11.6 Gy) or in two 7.3 Gy fractions (14.6 Gy total) separated by 24 h. In each case, the biologically effective dose (BED) equaled 25 Gy10. Response to RT was assessed by micro-CT 2 weeks after treatment. Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) and immunohistochemical staining were performed to assess the integrity of the p53 pathway, the G1 cell-cycle checkpoint, and apoptosis. Results: Tumor growth rates prior to RT were similar for the two genetic variants of lung adenocarcinoma. Lung cancers with wild-type (WT) p53 (LSL-Kras; Ink4a/ARFFL/FL mice) responded better to two daily fractions of 7.3 Gy compared to a single fraction of 11.6 Gy (P = 0.002). There was no statistically significant difference in the response of lung cancers deficient in p53 (LSL-Kras; p53FL/FL mice) to a single fraction (11.6 Gy) compared to 7.3 Gy × 2 (P = 0.23). Expression of the p53 target genes p21 and PUMA were higher and bromodeoxyuridine uptake was lower after RT in tumors with WT p53. Conclusion: Using an in vivo model of malignant lung cancer in mice, we demonstrate that the response of primary lung cancers to one or two fractions of RT can be influenced by specific gene mutations.

4D Micro-CT using Fast Prospective Gating

Micro-CT is currently used in preclinical studies to provide anatomical information. But, there is also significant interest in using this technology to obtain functional information. We report here a new sampling strategy for 4D micro-CT for functional cardiac and pulmonary imaging. Rapid scanning of free-breathing mice is achieved with fast prospective gating (FPG) implemented on a field programmable gate array. The method entails on-the-fly computation of delays from the R peaks of the ECG signals or the peaks of the respiratory signals for the triggering pulses. Projection images are acquired for all cardiac or respiratory phases at each angle before rotating to the next angle. FPG can deliver the faster scan time of retrospective gating (RG) with the regular angular distribution of conventional prospective gating for cardiac or respiratory gating. Simultaneous cardio-respiratory gating is also possible with FPG in a hybrid retrospective/prospective approach. We have performed phantom experiments to validate the new sampling protocol and compared the results from FPG and RG in cardiac imaging of a mouse. Additionally, we have evaluated the utility of incorporating respiratory information in 4D cardiac micro-CT studies with FPG. A dual-source micro-CT system was used for image acquisition with pulsed x-ray exposures (80 kVp, 100 mA, 10 ms). The cardiac micro-CT protocol involves the use of a liposomal blood pool contrast agent containing 123 mg I ml(-1) delivered via a tail vein catheter in a dose of 0.01 ml g(-1) body weight. The phantom experiment demonstrates that FPG can distinguish the successive phases of phantom motion with minimal motion blur, and the animal study demonstrates that respiratory FPG can distinguish inspiration and expiration. 4D cardiac micro-CT imaging with FPG provides image quality superior to RG at an isotropic voxel size of 88 μm and 10 ms temporal resolution. The acquisition time for either sampling approach is less than 5 min. The radiation dose associated with the proposed method is in the range of a typical micro-CT dose (256 mGy for the cardiac study). Ignoring respiration does not significantly affect anatomic information in cardiac studies. FPG can deliver short scan times with low-dose 4D micro-CT imaging without sacrificing image quality. FPG can be applied in high-throughput longitudinal studies in a wide range of applications, including drug safety and cardiopulmonary phenotyping.

GPU-based iterative reconstruction with total variation minimization for micro-CT

Dynamic imaging with micro-CT often produces poorly-distributed sets of projections, and reconstructions of this data with filtered backprojection algorithms (FBP) may be affected by artifacts. Iterative reconstruction algorithms and total variation (TV) denoising are promising alternatives to FBP, but may require running times that are frustratingly long. This obstacle can be overcome by implementing reconstruction algorithms on graphics processing units (GPU). This paper presents an implementation of a family of iterative reconstruction algorithms with TV denoising on a GPU, and a series of tests to optimize and compare the ability of different algorithms to reduce artifacts. The mathematical and computational details of the implementation are explored. The performance, measured by the accuracy of the reconstruction versus the running time, is assessed in simulations with a virtual phantom and in an in vivo scan of a mouse. We conclude that the simultaneous algebraic reconstruction technique with TV minimization (SART-TV) is a time-effective reconstruction algorithm for producing reconstructions with fewer artifacts than FBP.