Journal: Oncotarget

Article Title: Axon guidance molecule semaphorin3A is a novel tumor suppressor in head and neck squamous cell carcinoma

doi: 10.18632/oncotarget.6831

Figure Lengend Snippet: ( A ) Immunohistochemistry (IHC) staining for SEMA3A in normal oral epithelium. SEMA3A is highly expressed in normal oral epithelium. ( B ) IHC staining for SEMA3A in HNSCC specimens. SEMA3A is absent or reduced from HNSCC specimens. ( C ) Specimens without incubation with polyclonal antibody served as a negative control. (Scale bar: 100 μm). ( D ) Kaplan-Meier overall survival (OS) curves for 100 patients with HNSCC, according to SEMA3A expression level.

Article Snippet: 48 hours later, culture medium were collected to measure the amount of SEMA3A using the human SEMA3A ELISA kit (Elabscience Biotechnology), according to the manufacturer's instructions.

Techniques: Immunohistochemistry, Incubation, Negative Control, Expressing

Journal: Oncotarget

Article Title: Axon guidance molecule semaphorin3A is a novel tumor suppressor in head and neck squamous cell carcinoma

doi: 10.18632/oncotarget.6831

Figure Lengend Snippet: Correlation of SEMA3A expression and the clinical-pathological parameters of HNSCC specimens

Article Snippet: 48 hours later, culture medium were collected to measure the amount of SEMA3A using the human SEMA3A ELISA kit (Elabscience Biotechnology), according to the manufacturer's instructions.

Techniques: Expressing

Journal: Oncotarget

Article Title: Axon guidance molecule semaphorin3A is a novel tumor suppressor in head and neck squamous cell carcinoma

doi: 10.18632/oncotarget.6831

Figure Lengend Snippet: Univariate and multivariate cox regression analysis of clinical characteristics and SEMA3A expression

Article Snippet: 48 hours later, culture medium were collected to measure the amount of SEMA3A using the human SEMA3A ELISA kit (Elabscience Biotechnology), according to the manufacturer's instructions.

Techniques: Expressing

Journal: Oncotarget

Article Title: Axon guidance molecule semaphorin3A is a novel tumor suppressor in head and neck squamous cell carcinoma

doi: 10.18632/oncotarget.6831

Figure Lengend Snippet: ( A ) SEMA3A expression in HNSCC cell lines HN4, HN6, HN13, CAL27, SCC9 and SCC25 assessed by Western blot analysis (up), and quantification of the protein expression (down). ( B ) SEMA3A expression of CAL27, HN6 and SCC25 cells infected with SEMA3A adenovirus on protein (up) and gene (down) levels, detected by Western blot and real-time RT-PCR. ( C ) Images of colonies of SEMA3A-transduced cells (Ad-SEMA3A-CAL27, Ad-SEMA3A-HN6) and control vector (Ad-Con-CAL27, Ad-Con-HN6) stained with crystal violet (up), and the quantification of cell colonies (down). Each data point represents the mean ± SD of data from 3 independent trials. (** P < 0.01) ( D ) Transfection efficiency of SCC9, HN4 and HN13 cells determined by Western blot (up) and real-time RT-PCR (down), after 48 hours’ SEMA3A-siRNA transfection. ( E ) Images of colonies of SEMA3A-siRNA-transfected cells (Si-SEMA3A-SCC9, Si-SEMA3A-HN4) and negative control (Scramble) cells (Si-Scramble-SCC9, Si-Scramble-HN4) stained with crystal violet (left), and the quantification of cell colonies (right). ( F ) Graphs of growth curves of Ad-SEMA3A-cells (grey lines), Ad-Con-cells (black lines) and control cells (dotted lines), as CCK-8 assays carried out over 5 days. ( G, H ) Images of protein expression of CDKs (2, 4, 6), cyclins (E1, D1, D3) and inhibitors of CDKs (P21, P27) in SEMA3A-over-expressed cells (G) or SEMA3A-siRNA-transfected cells (H). ( I ) Flow cytometric analysis of cell-cycle changes in Ad-SEMA3A/Con-CAL27 and Ad-SEMA3A/Con-HN6 cells.

Article Snippet: 48 hours later, culture medium were collected to measure the amount of SEMA3A using the human SEMA3A ELISA kit (Elabscience Biotechnology), according to the manufacturer's instructions.

Techniques: Expressing, Western Blot, Infection, Quantitative RT-PCR, Control, Plasmid Preparation, Staining, Transfection, Negative Control, CCK-8 Assay

Journal: Oncotarget

Article Title: Axon guidance molecule semaphorin3A is a novel tumor suppressor in head and neck squamous cell carcinoma

doi: 10.18632/oncotarget.6831

Figure Lengend Snippet: ( A ) A gain of apoptosis-like phenotype of SEMA3A-transfected cells on 48 and 96 hours after transfection. Red arrowheads indicated apoptotic cells. Scale bar: 100 μm. ( B ) Flow cytometric analysis of apoptosis in Ad-SEMA3A/Con-CAL27/HN6. Cells were stained with Annexin V-allophycocyanin (APC) and 7-aminoactinomycin D (7-AAD), followed by FACS (fluorescence-activated cell sorting) analysis (up). Apoptosis was determined by FACS analysis (early apoptotic death cells in lower right plot quadrants and late apoptotic death cells in upper right plot quadrants) and plotted (down). As showed, apoptosis rate in Ad-SEMA3A cells was significantly higher than controls (** P < 0.01, t -test). ( C ) Protein changes of caspase (caspase-3, -5, -7) induced by SEMA3A overexpression and depletion were measured by western blotting. Representative images of WB are shown. ( D ) CAL27 cells were treated with rhSEMA3A protein for 48 hours of different concentrations. As the concentration increased, more cells adopted an apoptosis-like phenotype. The remaining cells were counted and plotted. Error bars represent the SD (standard deviation) from three separate experiments. *or**, statistically significant difference compared with the control at p < 0.05 or p < 0.01, respectively.

Article Snippet: 48 hours later, culture medium were collected to measure the amount of SEMA3A using the human SEMA3A ELISA kit (Elabscience Biotechnology), according to the manufacturer's instructions.

Techniques: Transfection, Staining, Fluorescence, FACS, Over Expression, Western Blot, Concentration Assay, Standard Deviation, Control

Journal: Oncotarget

Article Title: Axon guidance molecule semaphorin3A is a novel tumor suppressor in head and neck squamous cell carcinoma

doi: 10.18632/oncotarget.6831

Figure Lengend Snippet: ( A ) HNSCC xenograft tumor models were established. Images of mice with xenograft tumors were showed 7 weeks after adenovirus injection. Red arrow heads indicated xenograft tumors. ( B ) An illustration of scheme of CAL27 cells injection and adenovirus administration. ( C ) Tumor growth curves of nude mice in Ad-control group (Group 1, n = 5) and Ad-SEMA3A group (Group 2, n = 5). ( D ) Subcutaneous tumors isolated from nude mice 7 weeks after adenovirus injection.

Article Snippet: 48 hours later, culture medium were collected to measure the amount of SEMA3A using the human SEMA3A ELISA kit (Elabscience Biotechnology), according to the manufacturer's instructions.

Techniques: Injection, Control, Isolation

Journal: Oncotarget

Article Title: Axon guidance molecule semaphorin3A is a novel tumor suppressor in head and neck squamous cell carcinoma

doi: 10.18632/oncotarget.6831

Figure Lengend Snippet: ( A ) Cellular morphological changes were shown 48 hours after SEMA3A depletion and rhSEMA3A protein (100 ng/ml) treatment. ( B ) Western blot analysis were used to assess the expression of epithelial (E-cadherin and β-catenin) and mesenchymal (N-cadherin and Vimentin) markers inAd-SEMA3A/Con cells (CAL27, SCC25) and Si-SEMA3A/Scramble cells (HN4, SCC9). ( C ) The differences in the migration ability between the Ad-SEMA3A-CAL27/HN6 and Ad-Con-CAL27/HN6 cells, were measured using the wound-healing assay. ( D ) A transwell assay was employed to analyze the cell invasion ability.

Article Snippet: 48 hours later, culture medium were collected to measure the amount of SEMA3A using the human SEMA3A ELISA kit (Elabscience Biotechnology), according to the manufacturer's instructions.

Techniques: Western Blot, Expressing, Migration, Wound Healing Assay, Transwell Assay

Journal: Oncotarget

Article Title: Axon guidance molecule semaphorin3A is a novel tumor suppressor in head and neck squamous cell carcinoma

doi: 10.18632/oncotarget.6831

Figure Lengend Snippet: ( A ) Real time RT-PCR was used to detect the expression of classic EMT-inducers Zeb (Zeb1, Zeb2) and Snail (SNAI1, SNAI2) protein in Ad-SEMA3A/Con-CAL27 cells. ( B ) Western blotting was used to detect the expression of SNAI2 protein in Ad-SEMA3A/Con cells (CAL27, SCC25) and Si-SEMA3A/Con cells (HN4, SCC9). ( C ) Western blotting was performed to assess the expression of NF-κB pathway-related proteins in cytoplasmic and nuclear extracts from Ad-SEMA3A/Con-HN6 and Si-SEMA3A/Scramble-SCC9 cells. β-actin and histone H1 were employed as the positive controls for cytoplasmic and nuclear proteins, respectively. Semi-quantitative analysis of changes in protein expression as determined by scanning of the immunoreactive bands. ( D ) A glycosylated form of NRP1 was detected by Western blot analysis.

Article Snippet: 48 hours later, culture medium were collected to measure the amount of SEMA3A using the human SEMA3A ELISA kit (Elabscience Biotechnology), according to the manufacturer's instructions.

Techniques: Quantitative RT-PCR, Expressing, Western Blot

Journal: Oncotarget

Article Title: Axon guidance molecule semaphorin3A is a novel tumor suppressor in head and neck squamous cell carcinoma

doi: 10.18632/oncotarget.6831

Figure Lengend Snippet: Expression of SEMA3A in normal oral epithelium and HNSCC

Article Snippet: 48 hours later, culture medium were collected to measure the amount of SEMA3A using the human SEMA3A ELISA kit (Elabscience Biotechnology), according to the manufacturer's instructions.

Techniques: Expressing

Journal: Cell & Bioscience

Article Title: An increase in Semaphorin 3A biases the axonal direction and induces an aberrant dendritic arborization in an in vitro model of human neural progenitor differentiation

doi: 10.1186/s13578-022-00916-1

Figure Lengend Snippet: Sema 3A activates the microglia proinflammatory M1 phenotype 48 h after transfection. A Representative immunofluorescence pictures of the percentage of microglia cells transfected with Sema 3A-GFP (Sema 3A) or GFP empty vector (GFP). As control, we used non-transfected microglia cells (Ctrl). Scale bar: 55 µm. 10 × objective. B ELISA assay on media from Sema 3A, GFP and Ctrl microglia 48 h after transfection. Sema 3A levels were expressed as ng/ml and normalized on the number of alive cells (mean ± SEM from 10 different fields) for each experimental point. Data are the mean ± SEM of three independent experiments performed in triplicate. ****P < 0.0001 vs Sema 3A media. C Extent of cell survival obtained by counting the number of DAPI positive nuclei before and after Sema 3A transfection as well as in GFP and Ctrl microglia. Data are the mean ± SEM of three independent experiments in quadruplicate. One-way ANOVA followed by Tukey’s test for multiple comparisons. ****P < 0.0001 vs Sema 3A. D Quantitative analysis of TMEM 119, Iba1, CD86, CD68, and E iNOS and TNFα positive cells. MFI was performed on the entire slide using Zeiss Celldiscoverer7. Values are normalized on the number of DAPI positive cells for each slide and expressed as % of Ctrl. Data are the mean ± SEM of three independent experiments performed in quadruplicate. One-way ANOVA followed by Tukey’s test for multiple comparisons. *P < 0.05; **P < 0.01; ***P < 0.001. Representative images of iNOS (red) and TNFα (red) and Sema 3A or GFP (green) staining are reported in F . Scale bar: 10 µm. 63 × objective

Article Snippet: Sema 3A protein levels in media from microglia overexpressing Sema 3A (Sema 3A media) or GFP (GFP media) or non-transfected (Ctrl media) were assessed using Elisa kit (ELISA, Cusabio, Houston, TX, USA #CSB-E15913h), according to the manufacturer instructions.

Techniques: Transfection, Immunofluorescence, Plasmid Preparation, Control, Enzyme-linked Immunosorbent Assay, Staining

Journal: Biomedicines

Article Title: Can Repetitive Transcranial Magnetic Stimulation (rTMS) Promote Neurogenesis and Axonogenesis in Subacute Human Ischemic Stroke?

doi: 10.3390/biomedicines12030670

Figure Lengend Snippet: Neurogenesis-related molecular biomarker plasma levels at different time points by real, sham, and healthy control groups.

Article Snippet: Total plasma BDNF (MyBioSource, San Diego, CA, USA; catalog #: MBS824804), Ntn-1 (MyBioSource; catalog #: MBS700176), and Sema3A (MyBioSource; catalog #: MBS059778) concentrations were determined by specific enzyme-linked immunosorbent assay (ELISA), in accordance with the manufacturer’s protocol.

Techniques: Biomarker Discovery, Clinical Proteomics, Control

Journal: Biomedicines

Article Title: Can Repetitive Transcranial Magnetic Stimulation (rTMS) Promote Neurogenesis and Axonogenesis in Subacute Human Ischemic Stroke?

doi: 10.3390/biomedicines12030670

Figure Lengend Snippet: Levels of neurogenesis-related molecular biomarkers at T14 by real and sham groups. ( A ) Netrin-1. ( B ) miR-25. ( C ) miR-93. ( D ) miR-106b. ( E ) BDNF. ( F ). Semaphorin 3A. Boxes are drawn from the first quartile to the third quartile, horizontal lines are medians, vertical lines indicate maximum and minimum. Asterisks in ( B , C ) and the dot in ( C ) represent outliers. p values < 0.05 indicates statistically significant differences.

Article Snippet: Total plasma BDNF (MyBioSource, San Diego, CA, USA; catalog #: MBS824804), Ntn-1 (MyBioSource; catalog #: MBS700176), and Sema3A (MyBioSource; catalog #: MBS059778) concentrations were determined by specific enzyme-linked immunosorbent assay (ELISA), in accordance with the manufacturer’s protocol.

Techniques:

Journal: Biomedicines

Article Title: Can Repetitive Transcranial Magnetic Stimulation (rTMS) Promote Neurogenesis and Axonogenesis in Subacute Human Ischemic Stroke?

doi: 10.3390/biomedicines12030670

Figure Lengend Snippet: Temporal profile of median neurogenesis-related molecular biomarker levels across the different time points by real vs. sham groups ( A ) Netrin-1. ( B ) BDNF. ( C ) miR-25. ( D ) miR-93. ( E ) miR-106b. ( F ) Semaphorin 3A. Data on miRs are presented as 2-dCt where Ct is cycle threshold.

Article Snippet: Total plasma BDNF (MyBioSource, San Diego, CA, USA; catalog #: MBS824804), Ntn-1 (MyBioSource; catalog #: MBS700176), and Sema3A (MyBioSource; catalog #: MBS059778) concentrations were determined by specific enzyme-linked immunosorbent assay (ELISA), in accordance with the manufacturer’s protocol.

Techniques: Biomarker Discovery