Journal: Studies in Mycology

Article Title: Molecular systematics of the marine Dothideomycetes

doi: 10.3114/sim.2009.64.09

Figure Lengend Snippet: The list of species used in this study.

Article Snippet: Melanomma radicans , , , , ATCC 42522 , U43461 , U43479 , AY485625 , —.

Techniques: Algae

Journal: Cell Communication and Signaling

Article Title: NHERF2 is crucial in ERM phosphorylation in pulmonary endothelial cells

doi: 10.1186/1478-811x-11-99

Figure Lengend Snippet: Figure 1 EBP50 and NHERF2 have distinct interacting partners in EC. (A) EBP50 or NHERF2 was immunoprecipitated from lysates of BPAEC cells. Total cell lysate (input) and IP complexes were probed for ERM, EBP50 and NHERF2 specific antibodies. (B) Anti-c-myc antibody was utilized for immunoprecipitations form pCMV-myc ezrin (EZR), pCMV-myc radixin (RDX) or pCMV-myc moesin (MSN) transfected BPAEC cell lysates as described in Methods. Total cell lysates (input) and IP complexes were probed for c-myc tag, EBP50 and NHERF2. Ø AB: control of IP from BPAEC without the addition of antibody. Shown are representative data of at least 3 independent experiments.

Article Snippet: NHERF2 (SLC9A3R2) was silenced using 25 nM NHERF2 specific siRNA (sc-42522 from Santa Cruz or SI00068376, SI00068383, SI03075562 and SI03084977 from Qiagen) in complex with DharmaFECT-1 transfection reagent (Dharmacon) in serum-free medium.

Techniques: Immunoprecipitation, Transfection, Control

Journal: Cell Communication and Signaling

Article Title: NHERF2 is crucial in ERM phosphorylation in pulmonary endothelial cells

doi: 10.1186/1478-811x-11-99

Figure Lengend Snippet: Figure 2 Co-localization of phospho-ERM and NHERF2 during mitosis in BPAEC. Immunofluorescence staining of BPAEC was performed using anti-phospho-ERM (red) and anti-NHERF2 (green) primary antibodies. Phases of the cell cycle were identified using TO-PRO-3 Iodide staining. Scale bars: 10 μm. Shown are representative data of at least 3 independent experiments.

Article Snippet: NHERF2 (SLC9A3R2) was silenced using 25 nM NHERF2 specific siRNA (sc-42522 from Santa Cruz or SI00068376, SI00068383, SI03075562 and SI03084977 from Qiagen) in complex with DharmaFECT-1 transfection reagent (Dharmacon) in serum-free medium.

Techniques: Immunofluorescence, Staining

Journal: Cell Communication and Signaling

Article Title: NHERF2 is crucial in ERM phosphorylation in pulmonary endothelial cells

doi: 10.1186/1478-811x-11-99

Figure Lengend Snippet: Figure 3 Phospho-ERM co-immunoprecipitates with NHERF2 from mitotic EC. NHERF2 was immunoprecipitated from lysates of BPAEC without or with nocodazole (ND) treatment as described in Methods. Cell lysates (input) and IP complexes were probed for ERM, phospho-ERM and NHERF2. Shown are representative data of at least 3 independent experiments.

Article Snippet: NHERF2 (SLC9A3R2) was silenced using 25 nM NHERF2 specific siRNA (sc-42522 from Santa Cruz or SI00068376, SI00068383, SI03075562 and SI03084977 from Qiagen) in complex with DharmaFECT-1 transfection reagent (Dharmacon) in serum-free medium.

Techniques: Immunoprecipitation

Journal: Cell Communication and Signaling

Article Title: NHERF2 is crucial in ERM phosphorylation in pulmonary endothelial cells

doi: 10.1186/1478-811x-11-99

Figure Lengend Snippet: Figure 4 ERM cannot be phosphorylated in the absence of NHERF2. (A) Lysates of non transfected (ctr), non-siRNA or NHERF2 specific siRNA (sc-42522) treated cells were analyzed by Western blot using anti-EBP50, -NHERF2 and -actin antibodies. (B) Lysates of non transfected (ctr), non-siRNA or NHERF2 specific siRNA (sc-42522) treated cells without or with nocodazole (ND) challenge were analyzed by Western blot using antibodies against phospho-ERM, ERM, NHERF2 and actin as described in Methods. (C) NHERF2 or ROCK2 was immunoprecipitated from lysates of BPAEC. The IP complexes were probed for ROCK2 and NHERF2. (D) ROCK2 was immunoprecipitated from lysates of control, non-siRNA or NHERF2 specific siRNA treated cells. IP complexes were probed for ROCK2, ERM and NHERF2. Shown are representative data of at least 3 independent experiments.

Article Snippet: NHERF2 (SLC9A3R2) was silenced using 25 nM NHERF2 specific siRNA (sc-42522 from Santa Cruz or SI00068376, SI00068383, SI03075562 and SI03084977 from Qiagen) in complex with DharmaFECT-1 transfection reagent (Dharmacon) in serum-free medium.

Techniques: Transfection, Western Blot, Immunoprecipitation, Control

Journal: Cell Communication and Signaling

Article Title: NHERF2 is crucial in ERM phosphorylation in pulmonary endothelial cells

doi: 10.1186/1478-811x-11-99

Figure Lengend Snippet: Figure 5 NHERF2 overexpression induces EC filopodia formation and spreading. (A) Lysates of BPAEC without transfection (ctr) or transfected with pCMV-HA wild type or mutant NHERF2 constructs were analyzed by Western blot using antibodies against phospho-ERM, ERM, and HA-tag. A representative Western blot is shown. Protein levels were quantified by densitometric analysis. Phospho-ERM protein levels were normalized against ERM protein levels. Bars represent mean ± SE. Significant changes, determined by Student’s t-test, are indicated by asterisks; **(P < 0.01), n = 6. (B) Left panel: Immunofluorescent staining of recombinant wild type or mutant NHERF2 overexpressing BPAEC was performed using anti-HA primary antibody (green). Actin microfilaments were stained with Texas Red conjugated phalloidin (red). Scale bars: 10 μm. Right panel: Wild type NHERF2 transfected BPAEC co-stained for phospho-ERM and HA-tag is shown. Arrows indicate co-localization of phosho-ERM and overexpressed NHERF2 on the merged image. (C): Non transfected (control), wild type (wt) and mutant (mu) NHERF2 transfected cells (left panel) or non-siRNA and NHERF2 specific siRNA treated cells (right panel) were plated onto 8W10E arrays. Spreading and attachment of cells were followed in time by ECIS measurement. Results are presented as means ± SD at least of four chambers for each sample.

Article Snippet: NHERF2 (SLC9A3R2) was silenced using 25 nM NHERF2 specific siRNA (sc-42522 from Santa Cruz or SI00068376, SI00068383, SI03075562 and SI03084977 from Qiagen) in complex with DharmaFECT-1 transfection reagent (Dharmacon) in serum-free medium.

Techniques: Over Expression, Transfection, Mutagenesis, Construct, Western Blot, Staining, Recombinant, Control

Journal: Cell Communication and Signaling

Article Title: NHERF2 is crucial in ERM phosphorylation in pulmonary endothelial cells

doi: 10.1186/1478-811x-11-99

Figure Lengend Snippet: Figure 6 NHERF2 is required for EC tube formation. (A) Control, non-siRNA or silencing RNA specific for NHERF2 transfected BPAEC were seeded on Matrigel-coated μ-Slide plates. F-actin staining was done at 5 h after seeding, and images were captured by confocal microscopy. Silencing of NHERF2 inhibits cord formation, resulting in the formation of cell aggregates in the Matrigel. (B) Western blot analysis of ERM and phospho-ERM in control, non-siRNA or NHERF2 specific silencing RNA transfected BPAEC grown in Matrigel and processed at 0, 2 and 4 h after seeding.

Article Snippet: NHERF2 (SLC9A3R2) was silenced using 25 nM NHERF2 specific siRNA (sc-42522 from Santa Cruz or SI00068376, SI00068383, SI03075562 and SI03084977 from Qiagen) in complex with DharmaFECT-1 transfection reagent (Dharmacon) in serum-free medium.

Techniques: Control, Transfection, Staining, Confocal Microscopy, Western Blot

Journal: Cardiovascular Research

Article Title: Doxorubicin-induced cardiovascular toxicity: a longitudinal evaluation of functional and molecular markers

doi: 10.1093/cvr/cvad136

Figure Lengend Snippet: Proteomic analysis of aortic samples from DOX-treated mice at Weeks 2 and 6 and validation of two candidate biomarkers with western blotting. Plots of differentially expressed proteins at Weeks 2 and 6 ( A ). For A , horizontal grey line indicates significance threshold ( P < 0.05); up- and down-regulated proteins are indicated in blue and red, respectively; differentially expressed proteins are abbreviated (full names are listed in abbreviations list). Western blotting shows and corroborates higher levels of SERPINA3 in DOX-treated mice at Weeks 2 and 6, which persist at Week 9 ( B ). Western blotting also confirms higher levels of THBS1 in the DOX-treated group at Weeks 2 and 6 and shows a trend ( P = 0.052) towards increased THBS1 levels at Week 9 ( B ). Representative western blot images for SERPINA3 and THBS1 at Weeks 2, 6, and 9 ( C ). For C , ‘V’ and ‘D’ indicate lanes with vehicle and DOX aortic samples, respectively. D* on the blot at Week 9 contains a DOX sample for which no protein was detected and was excluded from analysis as such. For A , n = 8 in vehicle and DOX group for both Week 2 and Week 6. For B , vehicle group: n = 5, n = 6, and n = 6 at Weeks 2, 6, and 9, respectively; DOX group: n = 5, n = 6, and n = 5 at Weeks 2, 6, and 9, respectively. For B , Mann–Whitney U test for each time point. * and ** P < 0.05 and 0.01. Abbreviations for differentially expressed proteins in proteomics: SERPINA1E, α-1-antitrypsin; HMGN2, Non-histone chromosomal protein HMG-17; SERPINA3N, α-1-antichymotrypsin orthologue n; MT2, Metallothionein-2; MBL2, Mannose-binding protein C; MT1, Metallothionein-1; THBS1, Thrombospondin-1; FTL1, Ferritin light chain 1; COL14A1, Collagen α-1 (XIV) chain; LGALS3BP, Galectin-3-binding protein; RPL26, 60S ribosomal protein L26; IGHM, Immunoglobulin heavy constant µ; HAMP, Hepcidin; APOA2, Apolipoprotein A-II; HBA, Hemoglobin subunit α; HBB-B1, Hemoglobin subunit β-1; CA1, Carbonic anhydrase 1; KRT15, Keratin (type I cytoskeletal 15); SPTA1, Spectrin α; CA2; Carbonic anhydrase 2; RPL6, 60S ribosomal protein L6; VWA1, von Willebrand factor A domain-containing protein 1; RPL37A, 60S ribosomal protein L37a; CAPG, Macrophage-capping protein; RAD23A, UV excision repair protein RAD23 homolog A; IGHG1, Ig γ-1 chain C region (membrane-bound form); SLC4A1, Band 3 anion transport protein; ACLY, ATP-citrate synthase; ATPA1, Sodium/potassium-transporting ATPase subunit alpha-1.

Article Snippet: For validation of anti-THBS1 and anti-SERPINA3 primary antibodies, recombinant mouse THBS1 (catalogue number 7859-TH-050) and SERPINA3 (catalogue number 4709-PI-010) proteins were used as positive controls, which were purchased from Bio-Techne (Dublin, Ireland).

Techniques: Western Blot, MANN-WHITNEY, Binding Assay, Membrane

Journal: Cardiovascular Research

Article Title: Doxorubicin-induced cardiovascular toxicity: a longitudinal evaluation of functional and molecular markers

doi: 10.1093/cvr/cvad136

Figure Lengend Snippet: Quantification of SERPINA3 and THBS1 levels in plasma of control and AICT patients using ELISA. Schematic overview of the patient cohorts ( A ). AICT patients exhibited higher SERPINA3 and THBS1 levels compared with the control group ( B ). Both SERPINA3 and THBS1 showed an inverse Pearson correlation with LVEF ( C ). For B , data are presented as Tukey box plots with median (horizontal line) and the 25th and 75th percentile whiskers. For C , white and red dots represent control and AICT patients, respectively. n numbers are shown in A . For B , Mann–Whitney U test. * and **** P < 0.05 and 0.0001.

Article Snippet: For validation of anti-THBS1 and anti-SERPINA3 primary antibodies, recombinant mouse THBS1 (catalogue number 7859-TH-050) and SERPINA3 (catalogue number 4709-PI-010) proteins were used as positive controls, which were purchased from Bio-Techne (Dublin, Ireland).

Techniques: Enzyme-linked Immunosorbent Assay, MANN-WHITNEY