Journal: PLoS ONE

Article Title: Mutational Spectrum of Semaphorin 3A and Semaphorin 3D Genes in Spanish Hirschsprung patients

doi: 10.1371/journal.pone.0054800

Figure Lengend Snippet: SEMA3A sequence variants detected in the current study.

Article Snippet: Sections were incubated at 4°C overnight with the primary antibodies anti- SEMA3A (1∶50 dilution, anti-rabbit, AntibodyBcn, Barcelona, Spain) and anti- SEMA3D (1∶5 dilution, anti-rabbit, Novus Biologicals, Cambridge, UK).

Techniques: Sequencing

Journal: PLoS ONE

Article Title: Mutational Spectrum of Semaphorin 3A and Semaphorin 3D Genes in Spanish Hirschsprung patients

doi: 10.1371/journal.pone.0054800

Figure Lengend Snippet: SEMA3A and SEMA3D missense variants detected in isolated HSCR patients.

Article Snippet: Sections were incubated at 4°C overnight with the primary antibodies anti- SEMA3A (1∶50 dilution, anti-rabbit, AntibodyBcn, Barcelona, Spain) and anti- SEMA3D (1∶5 dilution, anti-rabbit, Novus Biologicals, Cambridge, UK).

Techniques: Isolation, Mutagenesis

Journal: PLoS ONE

Article Title: Mutational Spectrum of Semaphorin 3A and Semaphorin 3D Genes in Spanish Hirschsprung patients

doi: 10.1371/journal.pone.0054800

Figure Lengend Snippet: The SEMA3A staining illustrated that the expression was present at smooth muscle (D, E, F) and submucous (G, H, I) layers, as well as in myenteric (J, K) and submucous plexuses (M, N) either in normal colon (A, D, G, J, M) and patients with A131T-3A (B, E, H, K, N) and S598G-3A mutations (C, F, I). The FFPE tissue block from patient with S598G had no all tissue layers. Scale bars: A–C = 200 µm and the rest of pictures = 10 µm.

Article Snippet: Sections were incubated at 4°C overnight with the primary antibodies anti- SEMA3A (1∶50 dilution, anti-rabbit, AntibodyBcn, Barcelona, Spain) and anti- SEMA3D (1∶5 dilution, anti-rabbit, Novus Biologicals, Cambridge, UK).

Techniques: Staining, Expressing, Blocking Assay

Journal: Biochemical and biophysical research communications

Article Title: Semaphorin 3A potentiates the profibrotic effects of transforming growth factor-β1 in the cornea

doi: 10.1016/j.bbrc.2019.10.107

Figure Lengend Snippet: A. Unoperated, cornea triple labelled for SEMA3A, α-SMA and DAPI. The epithelium is uppermost. Individual stains (monochrome) are shown in B-D. Keratocytes (arrowed) are faintly SEMA3A positive. Note total absence of α-SMA staining. E. Photograph of a central cat cornea 4 weeks after PRK, triple labelled as in A. Individual stains are shown in F-H. Arrows indicate α-SMA-negative cells that are strongly SEMA3A-positive. Note the zone of cellular hyper-density below the α-SMA-positive layer (demarcated with yellow line), where a significant proportion of cells [putative fibroblasts] are SEMA3A-positive.

Article Snippet: Sections in the ablation center (or the geometric center of unoperated corneas) were selected for immunohistochemistry; they were co-incubated with a primary mouse monoclonal anti-α-SMA (1:100; Thermo Fisher Scientific, Waltham, MA, USA) and a feline-specific, rabbit polyclonal anti-SEMA3A antibody (1:100; YenZym Antibodies LLC., South San Francisco, CA, USA) overnight at 4°C.

Techniques: Staining

Journal: Biochemical and biophysical research communications

Article Title: Semaphorin 3A potentiates the profibrotic effects of transforming growth factor-β1 in the cornea

doi: 10.1016/j.bbrc.2019.10.107

Figure Lengend Snippet: Representative Western blot showing similar, relative SEMA3A expression levels in whole cell lysates and concentrated supernatants (S/N) of cat corneal fibroblasts (F) and myofibroblasts (M). Expression of α-SMA was used as an indicator of myofibroblast differentiation in the cell lysates.

Article Snippet: Sections in the ablation center (or the geometric center of unoperated corneas) were selected for immunohistochemistry; they were co-incubated with a primary mouse monoclonal anti-α-SMA (1:100; Thermo Fisher Scientific, Waltham, MA, USA) and a feline-specific, rabbit polyclonal anti-SEMA3A antibody (1:100; YenZym Antibodies LLC., South San Francisco, CA, USA) overnight at 4°C.

Techniques: Western Blot, Expressing

Journal: Biochemical and biophysical research communications

Article Title: Semaphorin 3A potentiates the profibrotic effects of transforming growth factor-β1 in the cornea

doi: 10.1016/j.bbrc.2019.10.107

Figure Lengend Snippet: A. Representative Western blot showing protein levels for COL1, t-FN, EDA-FN and α-SMA in cells treated with recombinant human SEMA3A with/without TGF-β1. Adding SEMA3A potentiated TGF-β1-induced increases in EDA-FN, t-FN, COL1 and α-SMA relative to TGF-β1 alone. B. Plot of relative densitometry data for EDA-FN, t-FN, COL1 and α-SMA with respect to β-actin. Data are means±SEM over 4 experiments. **ANOVA relative to baseline, p<0.0005. *ANOVA between 2 treatment groups, p<0.05. C. Representative Western blot showing SEMA3A levels in cells transfected with Sema3A siRNA or control (non-targeting) siRNA. D. Representative Western blot showing relative levels of COL1, t-FN, EDA-FN and α-SMA in fibroblasts transfected with control siRNA or Sema3A siRNA and cultured with/without TGF-β1. E. Plot of relative densitometry data for EDA-FN, t-FN, COL1 and α-SMA with respect to β-actin. Data are means±SEM over 3 experiments. *ANOVA relative to baseline, p<0.05. ns ANOVA between 2 treatment groups, p=0.227.

Article Snippet: Sections in the ablation center (or the geometric center of unoperated corneas) were selected for immunohistochemistry; they were co-incubated with a primary mouse monoclonal anti-α-SMA (1:100; Thermo Fisher Scientific, Waltham, MA, USA) and a feline-specific, rabbit polyclonal anti-SEMA3A antibody (1:100; YenZym Antibodies LLC., South San Francisco, CA, USA) overnight at 4°C.

Techniques: Western Blot, Recombinant, Transfection, Control, Cell Culture

Journal: Biochemical and biophysical research communications

Article Title: Semaphorin 3A potentiates the profibrotic effects of transforming growth factor-β1 in the cornea

doi: 10.1016/j.bbrc.2019.10.107

Figure Lengend Snippet: A. Phase contrast images (top row) show a change from fibroblastic to myofibroblastic morphologies in both cells transfected with control and Sema3A siRNA, but α-SMA staining (green in bottom row) is decreased in Sema3A siRNA-treated cells. B. Phalloidin staining of different regions on the same plates as in A shows persistence of F-actin fibers and clear morphological characteristics of larger myofibroblasts in the two plates treated with TGF-β1, even when α-SMA expression was decreased, as in the Sema3A siRNA+TGF-β1 condition.

Article Snippet: Sections in the ablation center (or the geometric center of unoperated corneas) were selected for immunohistochemistry; they were co-incubated with a primary mouse monoclonal anti-α-SMA (1:100; Thermo Fisher Scientific, Waltham, MA, USA) and a feline-specific, rabbit polyclonal anti-SEMA3A antibody (1:100; YenZym Antibodies LLC., South San Francisco, CA, USA) overnight at 4°C.

Techniques: Transfection, Control, Staining, Expressing