Journal: Genes & Cancer

Article Title: Erythropoietin Receptor Signaling through STAT3 Is Required for Glioma Stem Cell Maintenance

doi: 10.1177/1947601909356352

Figure Lengend Snippet: Targeting erythropoietin receptor (EPOR) expression in glioma stem cells (GSCs) reduces cell growth associated with decreased survival and neurosphere formation capacity. (A) Western blotting demonstrated successful knockdown of EPOR expression using lentivirus expressing two different EPOR-targeted short-hairpin RNA (shRNA) constructs (EPOR shRNA1 and EPOR shRNA2) in comparison to a nontargeting shRNA (NT) in T3691 and T3359 GSCs. (B) EPOR knockdown with two distinct shRNAs decreased the growth of T3691 GSCs in comparison to nontargeting control. *P < 0.001, one-way analysis of variance (ANOVA), n = 3. (C) Targeting EPOR significantly inhibited GSC proliferation. [3H]thymidine incorporation was decreased in T3691 GSCs infected with either one of two different EPOR shRNAs in comparison to a nontargeting control shRNA. *P < 0.001, one-way ANOVA, n = 5. (D) Loss of EPOR expression significantly increased apoptosis. Targeting EPOR with two different shRNAs increased relative caspase 3/7 activity (Promega, Madison, WI) in comparison to nontargeting control shRNA in T3691 GSCs. *P < 0.001, one-way ANOVA, n = 3. (E) Representative images demonstrating reduced neurosphere formation in T3359 and T3691 GSCs with two different EPOR targeting shRNA constructs (EPOR shRNA1 and EPOR shRNA2) in comparison to nontargeting (NT) control. (F) Targeting EPOR inhibits neurosphere formation capacity in primary passages. Increasing numbers of T3359 GSCs infected with the indicated shRNAs were plated per well and the percentage of wells demonstrating neurospheres determined after 4 days (n = 12). (G) Targeting EPOR inhibits neurosphere formation in sequential passages. Ten T3359 GSCs were plated in the presence of 1 µg/mL recombinant human soluble EPO receptor or bovine serum albumin as the control. For the primary passage, the percentage of wells with neurospheres and the number of neurospheres per well were quantified after 7 days. For the secondary passage, neurospheres formed in the primary passage were harvested and dissociated and the experiment repeated. Representative images of neurospheres formed in primary and secondary passages are shown. *P < 0.05, Student’s t test, n = 5.

Article Snippet: For the serial neurosphere formation assay with soluble EPOR treatment, 10 T3359 or D456MG glioma stem cells were plated into each well of 96-well plates with 200 μL completed neurobasal medium with either 1 μg/mL recombinant human soluble EPO receptor (sEPOR, R&D Systems) or equal amount of bovine serum albumin (BSA; GIBCO, Carlsbad, CA) as the control.

Techniques: Expressing, Western Blot, Knockdown, shRNA, Construct, Comparison, Control, Infection, Activity Assay, Recombinant

Journal: Molecular Medicine Reports

Article Title: Gaseous signalling molecule SO 2 via Hippo-MST pathway to improve myocardial fibrosis of diabetic rats

doi: 10.3892/mmr.2017.7714

Figure Lengend Snippet: SO 2 improves myocardial fibrosis in diabetic rats. (A) Morphological changes in myocardium assessed by Masson staining. Images were acquired at ×400 magnification. Expression levels of (B) MMP9, (C) MMP24 and (D) TIMP1 in each group. Date are expressed as mean ± standard deviation (n=3). *P<0.05 vs. control group; # P<0.05 vs. STZ group. SO 2 , sulfur dioxide; MMP, matrix metalloproteinase; TIMP, tissue inhibitor of metalloproteinase; STZ, streptozotocin; HDX, L-Aspartic acid β-hydroxamate.

Article Snippet: The antibodies for matrix metalloproteinase (MMP)9, MMP24, tissue inhibitor of metalloproteinase (TIMP)1 and GAPDH were purchased from Wuhan Boster Biological Technology, Ltd. (Wuhan, China).

Techniques: Staining, Expressing, Standard Deviation, Control

Journal:

Article Title: The glucocorticoid receptor is required for stress erythropoiesis

doi:

Figure Lengend Snippet: Proliferation of murine ebls in vitro requires ligand-activated GR. Fetal livers were isolated on day 14.5 from wild-type (wt, ●) and GRnull/null (□) littermate embryos. Cells were suspended and cultivated in media containing Dex, SCF, and Epo. (A) Cells were counted at daily intervals, and cumulative cell numbers were determined. (B) Aliquots from the cultures were cytocentrifuged onto slides on day 6 and processed for histological staining. Images were taken using a CCD camera and processed with Adobe Photoshop software. Hemoglobin positive cells appear grey to black (Wessely et al. 1997). Note the large ebls in wild-type cultures. This contrasts with the mostly small, highly hemoglobinized reticulocytes/erythrocytes or dead cells and granulocytes occurring in GRnull/null cultures.

Article Snippet: For evaluation of CFU-E, MethoCult 3236 was supplemented with human recombinant Epo (5 units; CILAG AG, Schaffhausen, CH), 100 ng/ml murine SCF (R&D), IGF-I (40 ng/ml; Sigma), and LDL (40 μg/ml; Sigma), and small, compact colonies were counted on day 2.

Techniques: In Vitro, Isolation, Staining, Software