The use of magnetic resonance imaging (MRI) for evaluating joint injuries is often considered superior to radiography due to the capacity of MRI for visualizing both soft and hard tissues. While longitudinal studies regarding cartilage repair have been undertaken on patients and in larger animal models, a method has yet to be developed for mouse cartilage to be repeatedly and non-invasively evaluated over time. The aim of this pilot study was to investigate if morphological changes following a focal cartilage injury in mice could be measured by 9.4 T magnetic resonance imaging. Focal cartilage defects were induced in the left knee of 4-6 weeks old C57BL/6 and MRL/MpJ mice. At endpoints 0, 2, and 4 weeks post-injury, legs were dissected out and imaged ex vivo. The defect could be detected by MRI immediately after injury, appearing as a hyperintense focal point and with size similar to that of the surgical tool used. Defects were visible in both strains up to 4 weeks post-injury, although signal intensity decreased over time. One C57BL/6 in particular, displayed extensive fibrosis in the patellar tendon at 4 weeks as assessed by histology, while the MR images of the same animal displayed a clear, structural distinction between the patella and the new tissue growth. Overall, our results suggest that MRI could be used for longitudinal studies in murine cartilage injury models to evaluate certain characteristics of repair not detectable through histology. (C) 2015 Elsevier Inc. All rights reserved.

Purpose: Articular cartilage is often thought to have low/no repair capability, however, the presence of mesenchymal progenitor cells in the synovial membrane and synovial fluid, suggest the possibility that some type of endogenous repair mechanism might exist in vivo. A main drawback in studying cartilage repair, is that few model systems have demonstrated ‘validated’ articular cartilage repair/regeneration in vivo. There are, however, a number of studies documenting cartilage regeneration in mice, specifically those from the MRL/MpJ ‘superhealer’ strain. Differences in cartilage repair have been observed between MRL/MpJ and C57BL/6 mice at 6 and 12 weeks after full thickness cartilage defects were induced. In this study, we sought to use magnetic resonance imaging (MRI) to characterize the earlier stages of the repair process following a full thickness cartilage defect in an attempt to identify at what specific point the healing processes diverge between the two mice strains. Methods: Two groups of MRL/MpJ and C57BL/6 male mice (5 weeks of age; n = 6 per time point) were used in the study. A custom made depth controlled needle (26 gauge) was used to introduce a full thickness cartilage defect into the femoral groove of the left knee. At 4 and 6 weeks post-surgery, the respective groups were euthanized and the legs were dissected out and fixed in 10% NBF. Using a 9.4 T magnet and a quadrature coil, samples underwent ex vivo magnetic resonance imaging using a RARE sequence (Repetition time = 2000 ms; Echo time = 7.6 ms; FOV =1.92; Matrix = 256). After imaging, samples were decalcified using 10% EDTA, embedded in paraffin, and sectioned at 7 um thickness. Sections were then stained with Safranin-O and immunohistochemistry (IHC) was performed using the anti-mouse Ly-6A/E (also known as Stem Cell Antigen- 1; Sca-1). Results: MRI scans at 4 weeks showed higher signal intensity at the cartilage compared to those scanned at 6 weeks (Fig1). There were no observable differences between strains at each time point. Safranin-O staining of the C57 sections at the 4 weeks, showed lower proteoglycan content than the MRL at the same time point. Defects were observable in C57 mice at all time points, but were not observed in MRL mice (Fig2). However, tissues within the MRL defect were not identical with the surrounding tissues, in specific regards to proteoglycan content, matrix structure, and chondrocyte orientation. Also of interest, it was observed that Sca-1 positive cells were enriched within the C57 defect, but were not found within the MRL defect (Fig2). Conclusion: In this study, we sought to characterize the early stages of endogenous cartilage repair that occurs in MRL/MpJ mice using both a non-invasive imaging technique (MRI) and tradition histological methods. Interestingly, MRI scans revealed that at 4 weeks post injury, there were no observable differences between C57 and MRL mice, though the Safranin-O stains did show increased cellularity in MRL samples at the defect compared to C57. Furthermore, the scarcity of Sca-1+ cells in MRL samples was surprising as the Safranin-o stains had revealed the superior healing and increased cellularity of the defect compared to the C57’s. A possible explanation could be that the recruitment of Sca-1 cells in the MRL's occurs much earlier than the time points used in this study. This would explain the lack of Sca-1 cells as well as the increased cellularity and structural organization at the defects of the MRL's compared to the C57’s. Future efforts will be focused on earlier time points than what was used in the present study.Figure 24 weeks post injury. Safranin-O stain of a) 057 and b) MRL with the respective Sca-1 IHC on the right.View Large Image Figure ViewerDownload Hi-res image Download (PPT)

A decrease in cerebral glucose metabolic uptake is an early and characteristic sign of Alzheimer’s disease (AD). Streptozotocin (STZ) is a bacterial toxin which damages insulin-producing cells and insulin receptors. Intracerebroventricular (icv) application of STZ in rats has been found to chronically decrease cerebral glucose uptake and produce other effects that bear a resemblance to several other molecular and pathological features of AD. In the present experiments in vivo 1H MR Spectroscopy with short echo time (3 ms) was used to non-invasively obtain a neurochemical profile of rat brains, 3 weeks and 2 months after double icv injections of STZ or vehicle. Seventeen metabolites were quantified from 27 μL tissue volume which included hippocampus and a part of cerebral cortex, using the LCModel and unsuppressed water signal as an internal reference. Three weeks after icv STZ several metabolites were significantly decreased, the most prominent changes noted in glycerophosphocholine and phosphocholine (−38 ± 5%), glutathione (−37 ± 4%), taurine (−30 ± 19%), glutamate (−26 ± 14%), phosphocreatine (−23 ± 15%) and N-acetylaspartate (−16 ± 6%). On the contrary, the concentration of N-acetylaspartylglutamate was found significantly increased (+38 ± 18%). After 2 months some of these changes were even more pronounced. We conclude that in vivo 1H MRS of rat brain following icv STZ injections provides a new input into a better understanding of the critical dependency of neural function and structure on brain glucose consumption, and may be of relevance in further studies of AD pathomechanism.

We have developed a magnetic resonance molecular imaging method using a novel iron-oxide contrast agent targeted towards P-selectin - MNP-PBP (magnetic nanoparticle-P-selectin binding peptide) - to image endothelial activation following cerebral ischemia/reperfusion. MNP-PBP consists of approximately 1000 PBP ligands (primary sequence: GSIQPRPQIHNDGDFEEIPEEYLQ GGSSLVSVLDLEPLDAAWL) conjugated to a 50 nm diameter aminated dextran iron oxide particle. In vitro P- and E-selectin binding was assessed by competition ELISA. Transient focal cerebral ischemia was induced in male C57/BL 6 mice followed by contrast injection (MNP-PBP; MNP-NH2; Feridex; MNP-PBP-FITC) at 24 h after reperfusion and T(2) magnetic resonance imaging at 9.4 T was performed. Infarction and microvasculature accumulation of contrast agent was assessed in coronal brain sections. MNP-PBP attenuated antibody binding to P-selectin by 34.8 +/- 1.7%. P-selectin was preferentially increased in the infarct hemisphere and MNP-PBP-FITC accumulation in the infarct hemisphere microvasculature was observed. Compared with the nontargeted iron oxide agents MNP-NH2 and Feridex, MNP-PBP showed a significantly greater T(2) effect within the infarction. MR imaging of P-selectin expression with a targeted iron oxide nanoparticle contrast agent may reveal early endothelial activation in stroke and other neuroinflammatory states.

B. Blasiak, B. Tomanek, T. Foniok, D. Kirk, D. Rushforth, R. MacKenzie, A. Abulrob, U. Iqbal, D. Stanimirovic, X. Lung, and P. Forsyth Department of Clinical Neurosciences, University of Calgary, Calgary, Alberta, Canada, Institute of Nuclear Physics, Polish Academy of Sciences, Krakow, malopolskie, Poland, Institute of Biodiagnostics(West), National Research Council of Canada, Calgary, Alberta, Canada, Institute of Biological Sciences, National Research Council of Canada, Ottawa, Ontario, Canada, Faculty of Medicine, University of Ottawa, Ottawa, Ontario, Canada