Imaging the Alternatively Spliced D Domain of Tenascin C in a Preclinical Model of Inflammatory Bowel Disease.

PURPOSE: To image colon-expressed alternatively spliced D domain of tenascin C in preclinical colitis models using near infrared (NIR)-labeled targeted molecular imaging agents. PROCEDURES: A human IgG1 with nanomolar binding affinity specific to the alternatively spliced D domain of tenascin C was generated. Immunohistochemistry identified disease-specific expression of this extracellular matrix protein in the colon of mice given dextran sulfate sodium in the drinking water. The antibody reagent was labeled with the NIR fluorophore IRDye 800CW via amine chemistry and intravenously dosed to evaluate in vivo targeting specificity. Increasing doses of imaging agent were given to estimate the saturating dose. RESULTS: The NIR-labeled proteins successfully targeted colonic lesions in a murine model of colitis. Co-administration of a molar excess competing unlabeled dose reduced normalized uptake in diseased colon by > 70%. Near infrared ex vivo images of colon resected from diseased animals showed saturation at doses exceeding 1 nmol and was confirmed with additional quantitative ex vivo biodistribution. Cellular-level specificity and protein stability were assessed via microscopy. CONCLUSIONS: Our imaging data suggest the alternatively spliced D domain of tenascin C is a promising target for delivery-based applications in inflammatory bowel diseases.

Quantitative in vivo micro-computed tomography for monitoring disease activity and treatment response in a collagen-induced arthritis mouse model.

A painful, chronic condition, Rheumatoid Arthritis, is marked by bone erosion and soft tissue swelling at the joint. As treatments are investigated in pre-clinical models, characterizing disease progression is integral to assessing treatment efficacy. Here, in vivo and ex vivo micro-computed tomography (µCT) are used in parallel with traditional caliper score measurement to quantify physiological changes in the tarsal region in a murine, collagen-induced arthritis model. In vivo imaging methods, which are validated here through comparison to ex vivo and caliper methods, afford longitudinal analysis of both bone and soft tissue through a single image acquisition. This method removes the subjectivity of swelling quantification which is inherently associated with traditional caliper measurements. Histopathology offers an additional assessment of bone erosion and inflammation by providing a microscopic characterization of disease activity. In comparison to untreated animals, daily prednisolone (glucocorticoid) treatment is shown to restore bone volume, as reflected through in vivo and ex vivo µCT images, as well as histopathology. Prednisolone-associated reduction in inflammation is shown through in vivo µCT soft tissue volume measurements, paw caliper measurements, and histopathology. The findings reported here provide a comprehensive validation of in vivo µCT with a sensitivity that enables characterization of pre-clinical disease assessment in response to treatment in a murine, collagen-induced arthritis model.

Near infrared readouts offer sensitive and rapid assessments of intestinal permeability and disease severity in inflammatory bowel disease models.

Intestinal permeability and neutrophil activity are closely linked to inflammatory bowel disease (IBD) pathophysiology. Here we discuss two techniques for assessing permeability and neutrophil activity in mouse IBD models using near infrared (NIR) detection. To address the limitation of visible light readouts-namely high background-IRDye 800CW was used to enable rapid, non-terminal measurements of intestinal permeability. The increased sensitivity of NIR readouts for colon permeability is shown using dextran sulfate sodium (DSS) and anti-CD40 murine colitis models in response to interleukin-22 immunoglobulin Fc (IL22Fc) fusion protein and anti-p40 monoclonal antibody treatments, respectively. In addition to enhanced permeability, elevated levels of neutrophil elastase (NE) have been reported in inflamed colonic mucosal tissue. Activatable NIR fluorescent probes have been extensively used for disease activity evaluation in oncologic animal models, and we demonstrate their translatability using a NE-activatable reagent to evaluate inflammation in DSS mice. Confocal laser endomicroscopy (CLE) and tissue imaging allow visualization of spatial NE activity throughout diseased colon as well as changes in disease severity from IL22Fc treatment. Our findings with the 800CW dye and the NE probe highlight the ease of their implementation in preclinical IBD research.

Distinct spatiotemporal pattern of CNS lesions revealed by USPIO-enhanced MRI in MOG-induced EAE rats implicates the involvement of spino-olivocerebellar pathways.

USPIO-enhanced MRI allows non-invasive visualization of mononuclear cell infiltration into CNS lesions in MS and EAE. Herein, we show a distinct spatiotemporal pattern of CNS lesions that reveals the involvement of spino-olivocerebellar pathways in MOG-induced EAE rats using USPIO-enhanced MRI. Specifically, lesions of the inferior olives were observed primarily in the acute phase whereas lesions of cerebellum or spinal cord/brainstem were observed during the relapse phase. Further, behavioral deficits observed from these animals are consistent with the functional role of spino-olivocerebellar pathways in coordination and movement. Collectively, our results provide new insights into the pathophysiology of this animal model of MS.