A reporter mouse for optical imaging of inflammation in mdx muscles

BACKGROUND: Duchenne muscular dystrophy (DMD) is due to mutations in the gene coding for human DMD; DMD is characterized by progressive muscle degeneration, inflammation, fat accumulation, and fibrosis. The mdx mouse model of DMD lacks dystrophin protein and undergoes a predictable disease course. While this model has been a valuable resource for pre-clinical studies aiming to test therapeutic compounds, its utility is compromised by a lack of reliable biochemical tools to quantifiably assay muscle disease. Additionally, there are few non-invasive assays available to researchers for measuring early indicators of disease progression in mdx mice. METHODS: Mdx mice were crossed to knock-in mice expressing luciferase from the Cox2 promoter. These reporter mice (Cox2(FLuc/+)DMD(−/−)) were created to serve as a tool for researchers to evaluate muscle inflammation. Luciferase expression was assayed by immunohistochemistry to insure that it correlated with muscle lesions. The luciferase signal was quantified by optical imaging and luciferase assays to verify that the signal correlated with muscle damage. As proof of principle, Cox2(FLuc/+)DMD(−/−) mice were also treated with prednisolone to validate that a reduction in luciferase signal correlated with prednisone treatment. RESULTS: In this investigation, a novel reporter mouse (Cox2(FLuc/+)DMD(−/−) mice) was created and validated for non-invasive quantification of muscle inflammation in vivo. In this dystrophic mouse, luciferase is expressed from cyclooxygenase 2 (Cox2) expressing cells and bioluminescence is detected by optical imaging. Bioluminescence is significantly enhanced in damaged muscle of exercised Cox2(FLuc/+)DMD(−/−) mice compared to non-exercised Cox2(FLuc/+)DMD(+/+) mice. Moreover, the Cox2 bioluminescent signal is reduced in Cox2(FLuc/+)DMD(−/−) mice in response to a course of steroid treatment. Reduction in bioluminescence is detectable prior to measurable therapy-elicited improvements in muscle strength, as assessed by traditional means. Biochemical assay of luciferase provides a second means to quantify muscle inflammation. CONCLUSIONS: The Cox2(FLuc/+)DMD(−/−) mouse is a novel tool to evaluate the therapeutic benefits of drugs intended to target inflammatory aspects of dystrophic pathology. This mouse model will be a useful adjunct to traditional outcome measures in assessing potential therapeutic compounds

Chromatin very small angle neutron scattering: further evidence for a 30 nm diameter super coil in dilute solutions.

Intact chromatin, chromatin minus histone H1, and nuclease digestion fragments have been studied by very small angle neutron scattering. The results are not consistent with a straight chain of nucleosomes and require the presence of a higher order coiling in monovalent salt solutions. The data are interpretable by a structure having a cross section radius of gryration of 8.5 +/- 1 nm, which suggests an outer diameter for a coil of nucleosomes of 27 +/- 3 nm

Chronic effects of the strobilurin fungicide azoxystrobin in the leaf shredder Gammarus fossarum (Crustacea; Amphipoda) via two effect pathways

Fungicides pose a risk for crustacean leaf shredders serving as key-stone species for leaf litter breakdown in detritus-based stream ecosystems. However, little is known about the impact of strobilurin fungicides on shredders, even though they are presumed to be the most hazardous fungicide class for aquafauna. Therefore, we assessed the impact of the strobilurin azoxystrobin (AZO) on the survival, energy processing (leaf consumption and feces production), somatic growth (growth rate and molting activity), and energy reserves (neutral lipid fatty and amino acids) of the amphipod crustacean Gammarus fossarum via waterborne exposure and food quality mediated (through the impact of leaf colonizing aquatic microorganisms) and thus indirect effects using 2 x 2-factorial experiments over 24 days. In a first bioassay with 30 mu g AZO/L, waterborne exposure substantially reduced survival, energy processing and affected molting activity of gammarids, while no effects were observed via the dietary pathway. Furthermore, a negative growth rate (indicating a body mass loss in gammarids) was induced by waterborne exposure, which cannot be explained by a loss in neutral lipid fatty and amino acids. These energy reserves were increased indicating a disruption of the energy metabolism in G. fossarum caused by AZO. Contrary to the first bioassay, no waterborne AZO effects were observed during a second experiment with 15 mu g AZO/L. However, an altered energy processing was determined in gammarids fed with leaves microbially colonized in the presence of AZO, which was probably caused by fungicide-induced effects on the microbial decomposition efficiency ultimately resulting in a lower food quality. The results of the present study show that diet-related strobilurin effects can occur at concentrations below those inducing waterborne toxicity. However, the latter seems to be more relevant at higher fungicide concentrations

Chronic effects of the strobilurin fungicide azoxystrobin in the leaf shredder Gammarus fossarum (Crustacea; Amphipoda) via two effect pathways

Fungicides pose a risk for crustacean leaf shredders serving as key-stone species for leaf litter breakdown in detritus-based stream ecosystems. However, little is known about the impact of strobilurin fungicides on shredders, even though they are presumed to be the most hazardous fungicide class for aquafauna. Therefore, we assessed the impact of the strobilurin azoxystrobin (AZO) on the survival, energy processing (leaf consumption and feces production), somatic growth (growth rate and molting activity), and energy reserves (neutral lipid fatty and amino acids) of the amphipod crustacean Gammarus fossarum via waterborne exposure and food quality mediated (through the impact of leaf colonizing aquatic microorganisms) and thus indirect effects using 2 x 2-factorial experiments over 24 days. In a first bioassay with 30 mu g AZO/L, waterborne exposure substantially reduced survival, energy processing and affected molting activity of gammarids, while no effects were observed via the dietary pathway. Furthermore, a negative growth rate (indicating a body mass loss in gammarids) was induced by waterborne exposure, which cannot be explained by a loss in neutral lipid fatty and amino acids. These energy reserves were increased indicating a disruption of the energy metabolism in G. fossarum caused by AZO. Contrary to the first bioassay, no waterborne AZO effects were observed during a second experiment with 15 mu g AZO/L. However, an altered energy processing was determined in gammarids fed with leaves microbially colonized in the presence of AZO, which was probably caused by fungicide-induced effects on the microbial decomposition efficiency ultimately resulting in a lower food quality. The results of the present study show that diet-related strobilurin effects can occur at concentrations below those inducing waterborne toxicity. However, the latter seems to be more relevant at higher fungicide concentrations

Lebetin peptides, a potent platelet aggregation inhibitors: chemical synthesis, biological activity and structure activity relationships

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