Cardioprotective effect of nicorandil on isoproterenol induced cardiomyopathy in the Mdx mouse model

Abstract Background Duchenne muscular dystrophy (DMD) associated cardiomyopathy is a major cause of morbidity and mortality. In an in vitro DMD cardiomyocyte model, nicorandil reversed stress-induced cell injury through multiple pathways implicated in DMD. We aimed to test the efficacy of nicorandil on the progression of cardiomyopathy in mdx mice following a 10-day treatment protocol. Methods A subset of mdx mice was subjected to low-dose isoproterenol injections over 5 days to induce a cardiac phenotype and treated with vehicle or nicorandil for 10 days. Baseline and day 10 echocardiograms were obtained to assess cardiac function. At 10 days, cardiac tissue was harvested for further analysis, which included histologic analysis and assessment of oxidative stress. Paired student’s t test was used for in group comparison, and ANOVA was used for multiple group comparisons. Results Compared to vehicle treated mice, isoproterenol decreased ejection fraction and fractional shortening on echocardiogram. Nicorandil prevented isoproterenol induced cardiac dysfunction. Isoproterenol increased cardiac fibrosis, which nicorandil prevented. Isoproterenol increased gene expression of NADPH oxidase, which decreased to baseline with nicorandil treatment. Superoxide dismutase 2 protein expression increased in those treated with nicorandil, and xanthine oxidase activity decreased in mice treated with nicorandil during isoproterenol stress compared to all other groups. Conclusions In conclusion, nicorandil is cardioprotective in mdx mice and warrants continued investigation as a therapy for DMD associated cardiomyopathy

Host interleukin 6 production regulates inflammation but not tryptophan metabolism in the brain during murine GVHD

Graft-versus-host disease (GVHD) induces pathological damage in peripheral target organs leading to well-characterized, organ-specific clinical manifestations. Patients with GVHD, however, can also have behavioral alterations that affect overall cognitive function, but the extent to which GVHD alters inflammatory and biochemical pathways in the brain remain poorly understood. In the current study, we employed complementary murine GVHD models to demonstrate that alloreactive donor T cells accumulate in the brain and affect a proinflammatory cytokine milieu that is associated with specific behavioral abnormalities. Host IL-6 was identified as a pivotal cytokine mediator, as was host indoleamine 2,3-dioxygenase (IDO-1), which was upregulated in GVHD in an IL-6-dependent manner in microglial cells and was accompanied by dysregulated tryptophan metabolism in the dorsal raphe nucleus and prefrontal cortex. Blockade of the IL-6 signaling pathway significantly reduced donor T cell accumulation, inflammatory cytokine gene expression, and host microglial cell expansion, but did not reverse GVHD-induced tryptophan metabolite dysregulation. Thus, these results indicate that inhibition of IL-6 signaling attenuates neuroinflammation, but does not reverse all of the metabolic abnormalities in the brain during GVHD, which may also have implications for the treatment of neurotoxicity occurring after other T cell-based immune therapies with IL-6-directed approaches

The nucleotide prodrug CERC‐913 improves mtDNA content in primary hepatocytes from DGUOK‐deficient rats

Loss‐of‐function mutations in the deoxyguanosine kinase (DGUOK) gene result in a mitochondrial DNA (mtDNA) depletion syndrome. DGUOK plays an important role in converting deoxyribonucleosides to deoxyribonucleoside monophosphates via the salvage pathway for mtDNA synthesis. DGUOK deficiency manifests predominantly in the liver; the most common cause of death is liver failure within the first year of life and no therapeutic options are currently available. in vitro supplementation with deoxyguanosine or deoxyguanosine monophosphate (dGMP) were reported to rescue mtDNA depletion in DGUOK‐deficient, patient‐derived fibroblasts and myoblasts. CERC‐913, a novel ProTide prodrug of dGMP, was designed to bypass defective DGUOK while improving permeability and stability relative to nucleoside monophosphates. To evaluate CERC‐913 for its ability to rescue mtDNA depletion, we developed a primary hepatocyte culture model using liver tissue from DGUOK‐deficient rats. DGUOK knockout rat hepatocyte cultures exhibit severely reduced mtDNA copy number (~10%) relative to wild type by qPCR and mtDNA content remains stable for up to 8 days in culture. CERC‐913 increased mtDNA content in DGUOK‐deficient hepatocytes up to 2.4‐fold after 4 days of treatment in a dose‐dependent fashion, which was significantly more effective than dGMP at similar concentrations. These early results suggest primary hepatocyte culture is a useful model for the study of mtDNA depletion syndromes and that CERC‐913 treatment can improve mtDNA content in this model

Cannabinoid CB2 receptors in the mouse brain: relevance for Alzheimer’s disease

Abstract Background Because of their low levels of expression and the inadequacy of current research tools, CB2 cannabinoid receptors (CB2R) have been difficult to study, particularly in the brain. This receptor is especially relevant in the context of neuroinflammation, so novel tools are needed to unveil its pathophysiological role(s). Methods We have generated a transgenic mouse model in which the expression of enhanced green fluorescent protein (EGFP) is under the control of the cnr2 gene promoter through the insertion of an Internal Ribosomal Entry Site followed by the EGFP coding region immediately 3′ of the cnr2 gene and crossed these mice with mice expressing five familial Alzheimer’s disease (AD) mutations (5xFAD). Results Expression of EGFP in control mice was below the level of detection in all regions of the central nervous system (CNS) that we examined. CB2R-dependent-EGFP expression was detected in the CNS of 3-month-old AD mice in areas of intense inflammation and amyloid deposition; expression was coincident with the appearance of plaques in the cortex, hippocampus, brain stem, and thalamus. The expression of EGFP increased as a function of plaque formation and subsequent microgliosis and was restricted to microglial cells located in close proximity to neuritic plaques. AD mice with CB2R deletion exhibited decreased neuritic plaques with no changes in IL1β expression. Conclusions Using a novel reporter mouse line, we found no evidence for CB2R expression in the healthy CNS but clear up-regulation in the context of amyloid-triggered neuroinflammation. Data from CB2R null mice indicate that they play a complex role in the response to plaque formation

Additional file 1: of Cannabinoid CB2 receptors in the mouse brain: relevance for Alzheimer’s disease

CB2 Western blots. Test of different CB2 primary antibodies in spleen samples (with high CB2 expression levels in normal conditions) harvested from CB2EGFP mice (lines 1, 2, and 3) and CB2KO mice (lines 4, 5, and 6). GFP and beta-actin immunodetection was employed as internal controls. (PPTX 6471 kb

Additional file 3: of Cannabinoid CB2 receptors in the mouse brain: relevance for Alzheimer’s disease

Table S1. Statistical analysis of the data provided in the manuscript. (DOCX 17 kb

Additional file 2: of Cannabinoid CB2 receptors in the mouse brain: relevance for Alzheimer’s disease

Figure S1. Sequences of the primers employed in the present studies. (DOCX 13 kb