Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: Endothelial deletion of EPH receptor A4 alters single-cell profile and Tie2/Akap12 signaling to preserve blood–brain barrier integrity

doi: 10.1073/pnas.2204700120

Figure Lengend Snippet: Raw average gene expression values from all capillary-derived endothelial cells

Article Snippet: Rabbit anti-EphA4 (Proteintech), goat anti-Cd31 (BD Biosciences), rabbit anti-CD45 (Cell Signaling Technology), rat anti-CD68 (Invitrogen), rabbit anti-Akap12 (Alomone, Jerusalem), and goat anti-Tie2 (R&D Systems).

Techniques: Expressing, Activation Assay, Shear, Membrane, Control

Journal: Biomolecules

Article Title: Long Noncoding RNA DANCR Activates Wnt/β-Catenin Signaling through MiR-216a Inhibition in Non-Small Cell Lung Cancer

doi: 10.3390/biom10121646

Figure Lengend Snippet: LncRNA-DANCR regulates Wnt/β-catenin signaling. ( A ) β-Catenin activity in A549 cells cotransfected with TOPFlash luciferase reporter + DANCR shRNAs compared to cells transfected with TOPFlash luciferase reporter + scramble control ( n = 3, t test, *** p < 0.001, **** p < 0.0001); ( B ) fold change of β-catenin protein expression in A549 DANCR KD cells compared to scramble control ( n = 3, t test, * p < 0.05); ( C ) fold change of CMYC and AXIN2 gene expression in A549 DANCR KD cells compared to scramble control ( n = 3, t test, * p < 0.05, ** p < 0.01, *** p < 0.001); ( D ) fold change of c-myc and Axin2 protein expression in A549 DANCR KD cells compared to scramble control ( n = 3, t test, * p < 0.05, ** p < 0.01).

Article Snippet: The following antibodies were used from Santa Cruz Biotchnology, Inc. (Dallas, TX, USA): Sox2 (sc-365823), β-catenin (sc-7963), GAPDH (sc-32233); Cell Signaling: c-Myc (CST-5605), Axin2 (CST-5863); and Sigma: β-actin (A5441).

Techniques: Activity Assay, Luciferase, Transfection, Control, Expressing, Gene Expression

Journal: Biomolecules

Article Title: Long Noncoding RNA DANCR Activates Wnt/β-Catenin Signaling through MiR-216a Inhibition in Non-Small Cell Lung Cancer

doi: 10.3390/biom10121646

Figure Lengend Snippet: DANCR promotes Wnt/β-catenin signaling through miR-216a inhibition. ( A ) qRT-PCR measuring fold change of DANCR expression in A549 cells transfected with pBABE-DANCR compared to vector control ( n = 3; t test, ** p < 0.001); ( B ) fold change of β-catenin protein expression in A549 DANCR overexpressing cells ( n = 3, t test, ** p < 0.001); ( C ) (left) qRT-PCR measuring fold change of DANCR expression in A549 cells doubly transfected with vector + vector (EV + EV), vector + pBABE-miR-216a (EV + miR-216a), vector + pBABE-DANCR (EV + DANCR), or pBABE-DANCR + pBABE-miR-216a (DANCR + miR-216a); (right) qRT-PCR measuring fold change of miR-216a expression ( n = 4, t test, * p < 0.05, ** p < 0.001); ( D ) fold change of β-catenin protein expression in A549 cells doubly transfected with vector + vector (EV + EV), vector + pBABE-miR-216a (EV + miR-216a), vector + pBABE-DANCR (EV + DANCR), or pBABE-DANCR + pBABE-miR-216a (DANCR + miR-216a) ( n = 4, t test, * p < 0.05).

Article Snippet: The following antibodies were used from Santa Cruz Biotchnology, Inc. (Dallas, TX, USA): Sox2 (sc-365823), β-catenin (sc-7963), GAPDH (sc-32233); Cell Signaling: c-Myc (CST-5605), Axin2 (CST-5863); and Sigma: β-actin (A5441).

Techniques: Inhibition, Quantitative RT-PCR, Expressing, Transfection, Plasmid Preparation, Control

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Systemic lupus erythematosus immune complexes increase the expression of SLAM family members CD319 (CRACC) and CD229 (LY-9) on plasmacytoid dendritic cells and CD319 on CD56(dim) NK cells.

doi: 10.4049/jimmunol.1301022

Figure Lengend Snippet: FIGURE 5. pDCs lack the activating adaptor molecules but express inhibitory adaptor molecules of SLAM receptors. (A) IFN-a production by RNA-IC–stimulated pDCs after cross-linking of CD319, CD229, or BDCA-2 by plate-bound mAbs. IFN-a levels were determined in 20-h cell culture supernatants by an immunoassay and normalized to the IFN-a production in cultures without plate-bound mAb. The dashed line indicates 100%. Data (mean + SD) are from three donors from two independent experiments. Expression of EAT2 and SAP (B) and SHIP-1, SHP-1, SHP- 2, and CSK (C) in cell lysates from isolated pDCs or NK cells from two healthy donors was determined by Western blot. Expression of b-actin was used as control.

Article Snippet: Rabbit polyclonal Abs to Ewing’s sarcoma-activated transcript 2 (EAT2), SHIP-1, SHP-2, CSK (Proteintech Group, Chicago, IL), and a rabbit mAb to SHP-1 (EPR5519; Epitomics, Burlingame, CA), followed by HRP-conjugated goat anti-rabbit IgG (H+L) (Invitrogen), were used to detect the indicated signaling molecules.

Techniques: Cell Culture, Expressing, Isolation, Western Blot, Control

Journal: Oncology reports

Article Title: SRPX and HMCN1 regulate cancer‑associated fibroblasts to promote the invasiveness of ovarian carcinoma.

doi: 10.3892/or.2019.7379

Figure Lengend Snippet: Figure 3. Knockdown of SRPX and HMCN1 in WS1 cells significantly reduced TOV21G cell invasion through the RhoA signaling pathway. Representative multi- ples of changes in comparisons of reverse‑transcription‑quantitative polymerase chain reaction results for (A) SRPX and (B) HMCN1 (***P<0.001). (C) Images of western blot analysis of the expression of SRPX, HMCN1, ROCK, RhoA, cdc42 and MLC in WS1 parental, mock‑transduced, SRPX‑downregulated cells (SRPX‑1 and SRPX‑2) and HMCN1‑downregulated cells (HMCN1‑1 and HMCN1‑2). Similar results were obtained from at least three independent experi- ments. Western blot analysis of HMCN1 was not performed due to no commercially available antibody. SRPX, sushi repeat‑containing protein, X; HMCN1, hemicentin 1; P, parental (normal WS1) cells; Z, mock‑transduced; RhoA, Ras homology family member A; ROCK, Rho kinase; cdc42, cell division cycle 42; MLC, myosin II regulatory light chain; ns, not significant.

Article Snippet: Protein (30 μg) from each well was dissolved in loading sample buffer under reducing conditions, separated by 12% SDS‐PAGE, and transferred to 0.2‐μm nitrocellulose membranes for immunolabelling with the following antibodies: Primary antibodies against SRPX (1:4,000, cat. co. 5473, Cell Signaling Technology, Inc.), RhoA (1:1,000, cat. co. 2117, Cell Signaling Technology, Inc.), ROCK (1:1,000, cat. co. 4035, Cell Signaling Technology, Inc.), cell division cycle 42 (cdc42) (1:1,000, cat. co. 2466, Cell Signaling Technology, Inc.), MLC (1:500, cat. co. 21157, Cell Signaling Technology, Inc.) and β‐actin (1:10,000, cat. co. MA5‐15739, Thermo Fisher Scientific, Inc.) were purchased from Novus Biologicals and incubated at 4 ̊C overnight with β‐actin as a loading control.

Techniques: Knockdown, Polymerase Chain Reaction, Western Blot, Expressing

Journal: Oncology reports

Article Title: SRPX and HMCN1 regulate cancer‑associated fibroblasts to promote the invasiveness of ovarian carcinoma.

doi: 10.3892/or.2019.7379

Figure Lengend Snippet: Figure 4. Overexpression of SRPX in WS1 fibroblasts induces TOV21G cell invasion. Representative (A) changes of SRPX and HMCN1 detected by reverse‑transcription‑quantitative polymerase chain reaction analysis. (B) images of the western blots of the expression of SRPX, ROCK, RhoA and MLC in WS1 parental, mock‑transduced and SRPX‑overexpressing fibroblasts. (C) Overexpression of SRPX in WS1 fibroblasts increased TOV21G cell invasion abili- ties (***P<0.001). The experiment was repeated at least three times. SRPX, sushi repeat‑containing protein, X; HMCN1, hemicentin 1; P, parental (normal WS1) cells; RhoA, Ras homology family member A; ROCK, Rho kinase; cdc42, cell division cycle 42; MLC, myosin II regulatory light chain; ns, not significant.

Article Snippet: Protein (30 μg) from each well was dissolved in loading sample buffer under reducing conditions, separated by 12% SDS‐PAGE, and transferred to 0.2‐μm nitrocellulose membranes for immunolabelling with the following antibodies: Primary antibodies against SRPX (1:4,000, cat. co. 5473, Cell Signaling Technology, Inc.), RhoA (1:1,000, cat. co. 2117, Cell Signaling Technology, Inc.), ROCK (1:1,000, cat. co. 4035, Cell Signaling Technology, Inc.), cell division cycle 42 (cdc42) (1:1,000, cat. co. 2466, Cell Signaling Technology, Inc.), MLC (1:500, cat. co. 21157, Cell Signaling Technology, Inc.) and β‐actin (1:10,000, cat. co. MA5‐15739, Thermo Fisher Scientific, Inc.) were purchased from Novus Biologicals and incubated at 4 ̊C overnight with β‐actin as a loading control.

Techniques: Over Expression, Polymerase Chain Reaction, Western Blot, Expressing