Bioengineering probiotic bacterium contrast agents for monitoring of inflammation using multispectral optoacoustic tomography - PROJECT SUMMARY Inflammatory diseases include a vast array of disorders including, arthritis, autoimmune diseases, hepatitis, and inflammatory bowel disease (IBD) among others. The ability to image inflammation in vivo is critical to improve the understanding of the pathophysiology underlying various disease etiologies. Current technologies to image IBD suffer from using radiation, lack of specificity, high cost, or invasive procedures that carry risk from both sedation or anesthesia and colonic perforation. Therefore, there is an urgent need to develop a noninvasive imaging modality that accurately identify early inflammatory changes, that can be used longitudinally to monitor the status of disease. With the use of a newly emerging imaging technology, multispectral optoacoustic tomography (MSOT), MSOT has potential for detecting inflammation and has the capability to detect multiple contrast agents simultaneously. The development of contrast agents detectable for this modality has largely explored nanomaterials, dye-labeled peptides, or dye-labeled antibodies. Although synthetic contrast agents for molecular imaging have shown great efficacy on disease diagnosis in preclinical trials, many of these synthetic agents have limitations such as low biocompatibility, material-associated toxicity, poor clearance, and high cost of pilot scale production. To overcome these limitations, the objective is to develop the first probiotic bacterial- based contrast agent to monitor inflammation using MSOT. Probiotics comprise primarily lactic acid bacteria, Lactococcus lactis (L. lactis) and are designated as generally regarded as safe organisms by FDA. L. lactis is well established as an efficient expression system for recombinant proteins and is being intensively investigated for delivery of antigens and cytokines at the mucosal level. It was demonstrated that the blue color generated from L. lactis-Blue (LlB) is detectable by MSOT. The goal is to bioengineer a stimuli-responsive L. lactis-Blue and dual label it with Zn-DPA-CF750 to result in a bacteria-based dual labeled contrast agent to monitor inflammation, using colitis as an inflammatory model, by MSOT. The hypothesis is that endogenous hemoglobin alone with MSOT will be insufficient to monitor chronic inflammatory diseases, thus, the proposed single or dual bacterial-based contrast agent, will target inflammation and provide additional molecular information to facilitate monitoring of inflammatory diseases. This hypothesis will be tested as follow: 1) Engineer and characterize LIB as a biologically-based contrast detectable using MSOT, 2) Construct and characterize L. lactis-750 and LIB-750 as single and dual bacterial-based contrast agents detectable using MSOT, and 3) Determine the potential of LIB, L-750, and LIB-750 as biological contrast agents to identify areas of inflammation using MSOT in a mouse model of colitis. Collectively, this proposal represents one of the first developments of a bacterial-based optoacoustic detectable contrast agent to further enhance the detection of inflammation associated with colitis.
