Brad Rice

Monitoring bacterial burden, inflammation and bone damage longitudinally using optical and μCT imaging in an orthopaedic implant infection in mice.

Background Recent advances in non-invasive optical, radiographic and μCT imaging provide an opportunity to monitor biological processes longitudinally in an anatomical context. One particularly relevant application for combining these modalities is to study orthopaedic implant infections. These infections are characterized by the formation of persistent bacterial biofilms on the implanted materials, causing inflammation, periprosthetic osteolysis, osteomyelitis, and bone damage, resulting in implant loosening and failure. Methodology/Principal Findings An orthopaedic implant infection model was used in which a titanium Kirshner-wire was surgically placed in femurs of LysEGFP mice, which possess EGFP-fluorescent neutrophils, and a bioluminescent S. aureus strain (Xen29; 1×103 CFUs) was inoculated in the knee joint before closure. In vivo bioluminescent, fluorescent, X-ray and μCT imaging were performed on various postoperative days. The bacterial bioluminescent signals of the S. aureus-infected mice peaked on day 19, before decreasing to a basal level of light, which remained measurable for the entire 48 day experiment. Neutrophil EGFP-fluorescent signals of the S. aureus-infected mice were statistically greater than uninfected mice on days 2 and 5, but afterwards the signals for both groups approached background levels of detection. To visualize the three-dimensional location of the bacterial infection and neutrophil infiltration, a diffuse optical tomography reconstruction algorithm was used to co-register the bioluminescent and fluorescent signals with μCT images. To quantify the anatomical bone changes on the μCT images, the outer bone volume of the distal femurs were measured using a semi-automated contour based segmentation process. The outer bone volume increased through day 48, indicating that bone damage continued during the implant infection. Conclusions/Significance Bioluminescent and fluorescent optical imaging was combined with X-ray and μCT imaging to provide noninvasive and longitudinal measurements of the dynamic changes in bacterial burden, neutrophil recruitment and bone damage in a mouse orthopaedic implant infection model.

Preclinical Validation of the Utility of BLZ-100 in Providing Fluorescence Contrast for Imaging Spontaneous Solid Tumors

There is a need in surgical oncology for contrast agents that can enable real-time intraoperative visualization of solid tumors that can enable complete resections while sparing normal surrounding tissues. The Tumor Paint agent BLZ-100 is a peptide-fluorophore conjugate that can specifically bind solid tumors and fluoresce in the near-infrared range, minimizing light scatter and signal attenuation. In this study, we provide a preclinical proof of concept for use of this imaging contrast agent as administered before surgery to dogs with a variety of naturally occurring spontaneous tumors. Imaging was performed on excised tissues as well as intraoperatively in a subset of cases. Actionable contrast was achieved between tumor tissue and surrounding normal tissues in adenocarcinomas, squamous cell carcinomas, mast cell tumors, and soft tissue sarcomas. Subcutaneous soft tissue sarcomas were labeled with the highest fluorescence intensity and greatest tumor-to-background signal ratio. Our results establish a foundation that rationalizes clinical studies in humans with soft tissue sarcoma, an indication with a notably high unmet need.

Adaptive row-action inverse solver for fast noise-robust three-dimensional reconstructions in bioluminescence tomography: theory and dual-modality optical/computed tomography in vivo studies

Abstract. A novel approach is presented for obtaining fast robust three-dimensional (3-D) reconstructions of bioluminescent reporters buried deep inside animal subjects from multispectral images of surface bioluminescent photon densities. The proposed method iteratively acts upon the equations relating the multispectral data to the luminescent distribution with high computational efficiency to provide robust 3-D reconstructions. Unlike existing algebraic reconstruction techniques, the proposed method is designed to use adaptive projections that iteratively guide the updates to the solution with improved speed and robustness. Contrary to least-squares reconstruction methods, the proposed technique does not require parameter selection or optimization for optimal performance. Additionally, optimized schemes for thresholding, sampling, and ordering of the bioluminescence tomographic data used by the proposed method are presented. The performance of the proposed approach in reconstructing the shape, volume, flux, and depth of luminescent inclusions is evaluated in a multitude of phantom-based and dual-modality in vivo studies in which calibrated sources are implanted in animal subjects and imaged in a dual-modality optical/computed tomography platform. Statistical analysis of the errors in the depth and flux of the reconstructed inclusions and the convergence time of the proposed method is used to demonstrate its unbiased performance, low error variance, and computational efficiency.

Improved Sensitivity by Applying Spectral Unmixing Prior to Fluorescent Tomography

Spectral unmixing technique was applied to the fluorescent tomography data acquired at multiple wavelengths to reduce the unwanted fluorophore signals and significantly improved the sensitivity and localization of the target fluorophore.

Small animal optical multispectral Cerenkov tomography

A novel pre-clinical imaging modality called Cerenkov luminescence imaging (CLI) has been recently introduced for small animals in vivo imaging. CLI is based on the detection of optical Cerenkov radiation generated by beta particles as they travel into the animal tissues with energy greater than Cerenkov threshold. The main goal of this work is the development of a novel optical image reconstruction method called multi spectral Cerenkov luminescence tomography (msCLT). The starting point of the msCLT reconstruction scheme is a set of 2D planar images acquired using several narrow bandpass filters. Because of the different tissues absorption at different wavelengths this provide distinctive information content that can be used for image reconstruction. More precisely the msCLT algorithm is based on a regularized iterative non-negative scheme in order to find the unknown source intensity solution, the theoretical Cerenkov emission spectrum was also included in the algorithm. In order to investigate the performances of the msCLT approach in vitro and in vivo imaging using 32P-ATP were performed by using the IVIS 200 (Caliper, a PerkinElmer company). A set of spatial resolution measurements were performed using a small capillary source placed between several slices of chicken breast at different depths. The spatial resolution obtained from the msCLT reconstructed images of the capillary showed that the FWHM is 1.5 mm for a source placed at 6 mm depth. In order to investigate the in vivo performances of the msCLT reconstruction method, a control nude mice injected with 10 MBq of 32P-ATP were imaged. Whole body MRI was acquired to provide an anatomical localization of the Cerenkov emission. msCLT reconstructed images co-registered with MRI images showed that the Cerenkov emission regions matches well with anatomical regions, such as the brain, heart and abdomen. These results were also confirmed by ex vivo imaging of organs such as intestine, brain, heart and ribs.