Hesperidin modulates dextran sulfate sodium‐induced ulcerative colitis in rats: Targeting sphingosine kinase‐1‐ sphingosine 1 phosphate signaling pathway,mitochondrial biogenesis,inflammation, and apoptosis

Ulcerative colitis (UC) is a chronic gastrointestinal disorder interfering with life quality. A total of 60 male Wistar rats were divided into four equal groups: Control (group I), hesperidin only (group II), UC untreated (group III), and UC treated with hesperidin (group IV). Hesperidin had modulatory effects on UC pathogenesis, which might be through alleviating colonic sphingosine phosphate phosphatase 2 messenger RNA expression and sphingosine kinase‐1 levels, thus suppressing the subsequent downstream inflammatory and apoptotic cascades represented by decreased macrophage inflammatory protein‐1α and enhancement of B‐cell lymphoma 2 immunohistochemistry expression. Also, it improved mitochondrial biogenesis by increasing the peroxisome proliferator‐activated receptor‐gamma‐coactivator 1‐α level. It successfully restored redox potential as evidenced by marked alleviations of the nitric oxide and peroxynitrite levels, increasing total antioxidant capacity, and activating the superoxide dismutase enzyme. Also, hesperidin alleviated the UC disease activity index and improved the histopathological picture. These findings may offer a new therapeutic strategy for UC treatment.     

P2RX7 Purinoceptor: A Therapeutic Target for Ameliorating the Symptoms of Duchenne Muscular Dystrophy

Background

Duchenne muscular dystrophy (DMD) is the most common inherited muscle disease, leading to severe disability and death in young men. Death is caused by the progressive degeneration of striated muscles aggravated by sterile inflammation. The pleiotropic effects of the mutant gene also include cognitive and behavioral impairments and low bone density.Current interventions in DMD are palliative only as no treatment improves the long-term outcome. Therefore, approaches with a translational potential should be investigated, and key abnormalities downstream from the absence of the DMD product, dystrophin, appear to be strong therapeutic targets. We and others have demonstrated that DMD mutations alter ATP signaling and have identified P2RX7 purinoceptor up-regulation as being responsible for the death of muscles in the mdx mouse model of DMD and human DMD lymphoblasts. Moreover, the ATP–P2RX7 axis, being a crucial activator of innate immune responses, can contribute to DMD pathology by stimulating chronic inflammation. We investigated whether ablation of P2RX7 attenuates the DMD model mouse phenotype to assess receptor suitability as a therapeutic target.

Methods and Findings

Using a combination of molecular, histological, and biochemical methods and behavioral analyses in vivo we demonstrate, to our knowledge for the first time, that genetic ablation of P2RX7 in the DMD model mouse produces a widespread functional attenuation of both muscle and non-muscle symptoms. In dystrophic muscles at 4 wk there was an evident recovery in key functional and molecular parameters such as improved muscle structure (minimum Feret diameter, p < 0.001), increased muscle strength in vitro (p < 0.001) and in vivo (p = 0.012), and pro-fibrotic molecular signatures. Serum creatine kinase (CK) levels were lower (p = 0.025), and reduced cognitive impairment (p = 0.006) and bone structure alterations (p < 0.001) were also apparent. Reduction of inflammation and fibrosis persisted at 20 mo in leg (p = 0.038), diaphragm (p = 0.042), and heart muscles (p < 0.001). We show that the amelioration of symptoms was proportional to the extent of receptor depletion and that improvements were observed following administration of two P2RX7 antagonists (CK, p = 0.030 and p = 0.050) without any detectable side effects. However, approaches successful in animal models still need to be proved effective in clinical practice.

Conclusions

These results are, to our knowledge, the first to establish that a single treatment can improve muscle function both short and long term and also correct cognitive impairment and bone loss in DMD model mice. The wide-ranging improvements reflect the convergence of P2RX7 ablation on multiple disease mechanisms affecting skeletal and cardiac muscles, inflammatory cells, brain, and bone. Given the impact of P2RX7 blockade in the DMD mouse model, this receptor is an attractive target for translational research: existing drugs with established safety records could potentially be repurposed for treatment of this lethal disease.     

RNA-Seq is not required to determine stable reference genes for qPCR normalization

Assessment of differential gene expression by qPCR is heavily influenced by the choice of reference genes. Although numerous statistical approaches have been proposed to determine the best reference genes, they can give rise to conflicting results depending on experimental conditions. Hence, recent studies propose the use of RNA-Seq to identify stable genes followed by the application of different statistical approaches to determine the best set of reference genes for qPCR data normalization. In this study, however, we demonstrate that the statistical approach to determine the best reference genes from commonly used conventional candidates is more important than the preselection of ‘stable’ candidates from RNA-Seq data. Using a qPCR data normalization workflow that we have previously established; we show that qPCR data normalization using conventional reference genes render the same results as stable reference genes selected from RNA-Seq data. We validated these observations in two distinct cross-sectional experimental conditions involving human iPSC derived microglial cells and mouse sciatic nerves. These results taken together show that given a robust statistical approach for reference gene selection, stable genes selected from RNA-Seq data do not offer any significant advantage over commonly used reference genes for normalizing qPCR assays.     

Synthesis and biological evaluation of halogenated phenoxychalcones and their corresponding pyrazolines as cytotoxic agents in human breast cancer

Novel halogenated phenoxychalcones 2a–f and their corresponding N-acetylpyrazolines 3a–f were synthesised and evaluated for their anticancer activities against breast cancer cell line (MCF-7) and normal breast cell line (MCF-10a), compared with staurosporine. All compounds showed moderate to good cytotoxic activity when compared to control. Compound 2c was the most active, with IC₅₀ = 1.52 µM and selectivity index = 15.24. Also, chalcone 2f showed significant cytotoxic activity with IC₅₀ = 1.87 µM and selectivity index = 11.03. Compound 2c decreased both total mitogen activated protein kinase (p38α MAPK) and phosphorylated enzyme in MCF-7 cells, suggesting its ability to decrease cell proliferation and survival. It also showed the ability to induce ROS in MCF-7 treated cells. Compound 2c exhibited apoptotic behaviour in MCF-7 cells due to cell accumulation in G2/M phase and elevation in late apoptosis 57.78-fold more than control. Docking studies showed that compounds 2c and 2f interact with p38alpha MAPK active sites.     

Mechanical stretch regulates the expression of specific miRNA in extracellular vesicles released from lung epithelial cells

The underlying mechanism of normal lung organogenesis is not well understood. An increasing number of studies are demonstrating that extracellular vesicles (EVs) play critical roles in organ development by delivering microRNAs (miRNA) to neighboring and distant cells. miRNAs are important for fetal lung growth; however, the role of miRNA–EVs (miRNAs packaged inside the EVs) during fetal lung development is unexplored. The aim of this study was to examine the expression of miRNA–EVs in MLE-12, a murine lung epithelial cell line subjected to mechanical stretch in vitro with the long-term goal to investigate their potential role in the fetal lung development. Both cyclic and continuous mechanical stretch regulate miRNA differentially in EVs released from MLE-12 and intracellularly, demonstrating that mechanical signals regulate the expression of miRNA–EVs in lung epithelial cells. These results provide a proof-of-concept for the potential role that miRNA–EVs could play in the development of fetal lung.     

HIRA directly targets the enhancers of selected cardiac transcription factors during in vitro differentiation of mouse embryonic stem cells

Molecular Biology Reports - HIRA is a histone chaperone known to modulate&nbsp;gene expression through the deposition of H3.3. Conditional knockout of Hira in embryonic mouse hearts leads to...