Journal: Brain : a journal of neurology

Article Title: Connexin43 mimetic peptide reduces vascular leak and retinal ganglion cell death following retinal ischaemia.

doi: 10.1093/brain/awr338

Figure Lengend Snippet: Figure 3 Three hours of hypoxia and 6 h of reperfusion lead to significant endothelial cell death in vitro in rat brain microvascular endothelial cells (RBMVEC; A). Non-specific gap junction blocker carbenoxolone, non-specific hemichannel blocker LaCl3, and connexin43 (Cx43) mimetic peptide protected endothelial cells against hypoxic injury, with the number of viable cells significantly higher than no treatment, while scrambled peptide did not have any protective effects. The number of viable cells was expressed as percentage of the control without hypoxia. Hypoxia and reperfusion also lead to significant propidium iodide dye uptake into primary rat brain microvascular endothelial cells indicating open hemichannels (B). Carbenoxolone, LaCl3 and connexin43 mimetic peptide significantly prevented dye uptake compared to no treatment, indicating hemichannel closure, while scrambled peptide did not have any effect. Stars denote statistical significance when compared to the control group or compared between groups in brackets; P 5 0.05.

Article Snippet: Rat brain microvascular endothelial cells (R840K-05a, Cell Applications) were plated into 24-well plates (1 105 cells/well) in rat brain microvascular endothelial cell growth media (R819K-500, Cell Applications) and allowed to settle for 16 h. Medium was then removed and replaced with Dulbecco’s Modified Eagle’s Medium/F12 containing 0.5% foetal bovine serum and 1% glutamine.

Techniques: In Vitro, Control

Journal: International Journal of Nanomedicine

Article Title: Schisandrae fructus enhances myogenic differentiation and inhibits atrophy through protein synthesis in human myotubes

doi: 10.2147/IJN.S101299

Figure Lengend Snippet: SF-induced differentiation in HSkMCs. Notes: ( A ) SF was treated with various concentrations (0.1 μg/mL, 0.5 μg/mL, 1 μg/mL, 5 μg/mL, and 10 μg/mL) in growth medium. Two days after treatment, HSkMCs were counted. As a result, SF did not induce proliferation. ( B and C ) SF (0.1 μg/mL, 0.5 μg/mL, 1 μg/mL, 5 μg/mL, and 10 μg/mL) was treated with DM and changed with fresh DM every 2 days. HSkMCs were photographed three times every 2 days. For observing differentiation efficiency, fusion index and myotube area were analyzed. ( D ) Eight days after differentiation induction with SF, HSkMCs were photographed. A total of 0.5 μg/mL of SF significantly induced more differentiation. All data represented mean ± SEM (n=3). *Symbol indicates P <0.05 compared to control. Abbreviations: DM, differentiation medium; HSkMCs, human skeletal muscle cells; SF, Schisandrae fructus.

Article Snippet: Differentiation of HSkMCs was induced by HSkMC differentiation medium (DM) (C-23061; PromoCell, Heidelberg, Germany).

Techniques:

Journal: International Journal of Nanomedicine

Article Title: Schisandrae fructus enhances myogenic differentiation and inhibits atrophy through protein synthesis in human myotubes

doi: 10.2147/IJN.S101299

Figure Lengend Snippet: SF increased protein synthesis through mTOR/P70S6K and 4E-BP1 signaling. Notes: ( A ) SF (0.5 μg/mL) was treated with differentiation medium. HSkMCs’ differentiation was induced for 1 day, 2 days, and 4 days. Phosphorylation and expression of mTOR, P70S6K, and 4E-BP1 were observed for Western blot. GAPDH expression was analyzed to identify equal loading. ( B – D ) Levels of mTOR activation (phosphorylation) were normalized to the levels of GAPDH. Phosphorylation of P70S6K and 4E-BP1 was normalized to the levels of each total protein. All data represented mean ± SEM (n=3). *Symbol indicates P <0.05. Abbreviations: 4E-BP1, 4E-binding protein1; HSkMCs, human skeletal muscle cells; mTOR, mammalian target of rapamycin; p-4E-BP1, phosphorylated 4E-BP1; p-P70S6K, phosphorylated P70S6K; P70S6K, 70 kDa ribosomal protein S6 kinase; SF, Schisandrae fructus; SEM, standard error of mean; GAPDH, glyceraldehyde 3-phosphate dehydrogenase.

Article Snippet: Differentiation of HSkMCs was induced by HSkMC differentiation medium (DM) (C-23061; PromoCell, Heidelberg, Germany).

Techniques: Expressing, Western Blot, Activation Assay, Binding Assay

Journal: International Journal of Nanomedicine

Article Title: Schisandrae fructus enhances myogenic differentiation and inhibits atrophy through protein synthesis in human myotubes

doi: 10.2147/IJN.S101299

Figure Lengend Snippet: SF inhibited muscle atrophy through increasing protein synthesis. Notes: At 2 days of differentiation, atrophy of myotubes was induced by 100 μM DEX for 4 days, and DM was changed to fresh DM with SF every 2 days. ( A – C ) In the last 6 days after induction of differentiation, HSkMCs were photographed three times per group. For observing differentiation efficiency, fusion index and myotube area were analyzed. All data represented mean ± SEM (n=3). *Symbol indicates P <0.05 compared to control. # Symbol represents P <0.05 compared to DEX treatment alone. ( D ) Myotubes were fluorescence stained with anti-MYH (red) and DAPI (blue), which was observed as a marker of late differentiation. ( E and F ) Examples of representative Western blot were shown for MYH, p-P70S6K, P70S6K p-4E-BP1, 4E-BP1, GAPDH, and MuRF1. Levels of MYH, MuRF1, p-P70S6K, and p-4E-BP1 were normalized to the levels of GAPDH or total protein. Abbreviations: DEX, dexamethasone; DM, differentiation medium; 4E-BP1, 4E-binding protein1; HSkMCs, human skeletal muscle cells; MuRF1, muscle RING finger 1; MYH, myosin heavy chain; p-4E-BP1, phosphorylated eukaryotic translation initiation factor 4E-binding protein 1; p-P70S6K, phosphorylated P70S6K; P70S6K, 70 kDa ribosomal protein S6 kinase; SF, Schisandrae fructus; SEM, standard error of mean; GAPDH, glyceraldehyde 3-phosphate dehydrogenase.

Article Snippet: Differentiation of HSkMCs was induced by HSkMC differentiation medium (DM) (C-23061; PromoCell, Heidelberg, Germany).

Techniques: Fluorescence, Staining, Marker, Western Blot, Binding Assay

Journal: Clinical Epigenetics

Article Title: MicroRNA-101 is repressed by EZH2 and its restoration inhibits tumorigenic features in embryonal rhabdomyosarcoma

doi: 10.1186/s13148-015-0107-z

Figure Lengend Snippet: MiR-101 is directly targeted by EZH2 in RD cells. a , b Two independent ChIP assays on RD cells 72 h after EZH2 or CTR siRNA transfection showing the recruitment of EZH2 and histone H3 trimethylation on Lys27 (H3K27me3) levels on miR-101-2 promoter region and MCK regulatory regions. SMAD6 was the negative control gene. Rabbit IgG was used as a negative immunoprecipitation control. Histograms represent the percent of immunoprecipitated material relative to input DNA of the two independent experiments. c mRNA levels (RT-qPCR) of pri - miR-101-2 in RD cells 72 h after EZH2 siRNA treatment were normalized to GAPDH levels and expressed as fold increase over CTR siRNA. d Proposed model depicting the interplay between EZH2 and miR-101 in both normal myogenic differentiation ( left ) and eRMS ( right ). In muscle cells, when myogenesis is triggered, miR-101 is upregulated due to the lowering of EZH2 expression. Then, miR-101 directly inhibits EZH2 expression thus enforcing its own expression, driving late skeletal muscle differentiation. In eRMS, this circuit is dysregulated due to EZH2 over-expression, which leads to miR-101 down-regulation, thus maintaining the cells in an undifferentiated and proliferative state

Article Snippet: A human skeletal muscle differentiation model was obtained treating SkMC myoblasts for 14 days with a differentiating medium (DM) with appropriate supplements (C-23161 and C-39366, PromoCell GmbH, Heidelberg, Germany).

Techniques: Transfection, Negative Control, Immunoprecipitation, Quantitative RT-PCR, Expressing, Over Expression