Journal: Oncoimmunology

Article Title: Formyl peptide receptor 1 suppresses gastric cancer angiogenesis and growth by exploiting inflammation resolution pathways

doi: 10.1080/2162402X.2017.1293213

Figure Lengend Snippet: ALOXs and GPR32 involvement in GC angiogenic response. (A) Tumor growth curves of AGS shCTR, shALOX5 (three clones) and shALOX15 (three clones) xenografts in immunodeficient mice. *p < .05 compared to shCTR xenografts. (B) Representative images and quantification (five fields/sample) of the proliferation index (Ki-67), vessel density (CD31), and apoptotic rate (Cleaved Caspase 3), assessed by immunohistochemistry, of shCTR, shALOX5, and shALOX15 cell xenografts harvested 28 d post-inoculation. *p < .05 compared to shCTR xenografts. (C) ALOX15 and ALOX5 mRNA expression levels of 295 patients affected by gastric adenocarcinoma stratified for disease-free and overall survival status. *p < .05 between the two groups. (D) Tumor growth curves of AGS shCTR and shGPR32 xenografts (average of three clones) cells in immunodeficient mice *p < .05 compared to shCTR xenografts. (E) Representative images of the vessel density (CD31), assessed by immunohistochemistry, of shCTR and shGPR32 AGS cell xenografts harvested 28 d post-inoculation.

Article Snippet: Xenografts in mice Each group of 10 mice (4-week-old female CD1 nu/nu mice, Charles River, Wilmington, MA) was inoculated subcutaneously with shCTR, shFPR1, shALOX5, shALOX15, shGPR32 AGS cells (1×10 7 cells/mouse).

Techniques: Clone Assay, Immunohistochemistry, Expressing

Journal: Oncoimmunology

Article Title: Formyl peptide receptor 1 suppresses gastric cancer angiogenesis and growth by exploiting inflammation resolution pathways

doi: 10.1080/2162402X.2017.1293213

Figure Lengend Snippet: ALOXs and GPR32 are required for FPR1 tumor suppressor role. (A) VEGF-B, -C, and CXCL1 mRNA synthesis was inhibited in shCTR, but not in shALOX5 or shALOX15 AGS cells upon fMLF (10−9 M) treatment. Data are represented as mean ± SD of three independent experiments. *p < .05 vs. the relative untreated cells (dotted line). (B) Pre-treatment of AGS cells with NDGA (ALOX inhibitor) reverted the ability of fMLF to inhibit the mRNA synthesis of pro-angiogenic molecules. Data are represented as mean ± SD of three independent experiments. *p < .05 compared to fMLF treated cells. (C) In AGS cells, fMLF induced ALOX5 and ALOX15A mRNA overexpression and concomitant VEGF-B and CXCL1 mRNA downregulation. These effects were reverted by a neutralizing GPR32 antibody. An isotype-matched antibody was used as a control. Data are represented as mean ± SD of three independent experiments. *p < .05 compared to untreated cells (dotted line). §p < .05 compared to isotype-matched control. (D) fMLF significantly induced ALOX5, ALOX15A, and ALOX15B mRNA levels and significantly reduced the mRNA levels of pro-angiogenic mediators in AGS shCTR, but not in shGPR32 cells. Data are represented as mean ± SD of three independent experiments. *p < .05 compared to shCTR cells. (E) A neutralizing GPR32 antibody inhibited fMLF-induced ALOX5 and ALOX15A mRNA overexpression and concomitant VEGF-B and CXCL1 mRNA downregulation in AGS shFPR2 cells. An isotype-matched antibody was used as a control. Data are represented as mean ± SD of three independent experiments. *p < .05 compared to untreated cells (dotted line). §p < .05 compared to isotype-matched control. (F) Activation kinetics of AKT, MAPK, JNK, p38, STAT3, and SRC in AGS shCTR, AGS shFPR1, MKN45 pcDNA, MKN45 FPR1 cells assessed by western blot for their phosphorylated forms. Two STAT3 inhibitors (5-15 DPP and FLLL31) reverted the anti-angiogenic effects of RvD1 in AGS shFPR1 cells. Data are represented as mean ± SD of three independent experiments. *p < .05 compared to untreated cells (NT). §p < .05 compared to RvD1 treated cells.

Article Snippet: Xenografts in mice Each group of 10 mice (4-week-old female CD1 nu/nu mice, Charles River, Wilmington, MA) was inoculated subcutaneously with shCTR, shFPR1, shALOX5, shALOX15, shGPR32 AGS cells (1×10 7 cells/mouse).

Techniques: Over Expression, Control, Activation Assay, Western Blot

Journal: Journal of Biological Macromolecules

Article Title: Generation of ribonuclease H2 A subunit (RH2A)-knockout HEK293 cells and analysis of the ribonucleotide content of their genomic DNA

doi: 10.14533/jbm.24.33

Figure Lengend Snippet: Fig. 3. Microscopic image and growth curve of HEK293 WT and RH2A-KO cells.

Article Snippet: After separation, the proteins were transferred by electroblotting onto a polyvinylidene difluoride (PVDF) membrane Sequi- BlotTM PVDF (BioRad, Hercules, CA) in 25 mM Tris-HCl buffer (pH 8.3), 192 mM glycine, 20% v/v methanol at 25 V for 50 min. After blotting, the membrane was washed with 50 mM Tris-HCl buffer (pH 8.3), 138 mM NaCl, 2.7 mM KCl, 0.05% Tween 20 (TBS-T), blocked with TBS-T containing 2% w/v skim milk, and incubated with mouse anti human RH2A polyclonal antibody, Anti RNASEH2A (Proteintech, Rosemont, IL, 1:1000 in TBS-T containing 1% w/v skim milk).

Techniques:

Journal: Journal of Biological Macromolecules

Article Title: Generation of ribonuclease H2 A subunit (RH2A)-knockout HEK293 cells and analysis of the ribonucleotide content of their genomic DNA

doi: 10.14533/jbm.24.33

Figure Lengend Snippet: Fig. 5. Expression of RH2A.

Article Snippet: After separation, the proteins were transferred by electroblotting onto a polyvinylidene difluoride (PVDF) membrane Sequi- BlotTM PVDF (BioRad, Hercules, CA) in 25 mM Tris-HCl buffer (pH 8.3), 192 mM glycine, 20% v/v methanol at 25 V for 50 min. After blotting, the membrane was washed with 50 mM Tris-HCl buffer (pH 8.3), 138 mM NaCl, 2.7 mM KCl, 0.05% Tween 20 (TBS-T), blocked with TBS-T containing 2% w/v skim milk, and incubated with mouse anti human RH2A polyclonal antibody, Anti RNASEH2A (Proteintech, Rosemont, IL, 1:1000 in TBS-T containing 1% w/v skim milk).

Techniques: Expressing

Journal: Journal of Biological Macromolecules

Article Title: Generation of ribonuclease H2 A subunit (RH2A)-knockout HEK293 cells and analysis of the ribonucleotide content of their genomic DNA

doi: 10.14533/jbm.24.33

Figure Lengend Snippet: Fig. 6. Expression of RH2A variants with AGS- causing mutation in RH2A-KO cells.

Article Snippet: After separation, the proteins were transferred by electroblotting onto a polyvinylidene difluoride (PVDF) membrane Sequi- BlotTM PVDF (BioRad, Hercules, CA) in 25 mM Tris-HCl buffer (pH 8.3), 192 mM glycine, 20% v/v methanol at 25 V for 50 min. After blotting, the membrane was washed with 50 mM Tris-HCl buffer (pH 8.3), 138 mM NaCl, 2.7 mM KCl, 0.05% Tween 20 (TBS-T), blocked with TBS-T containing 2% w/v skim milk, and incubated with mouse anti human RH2A polyclonal antibody, Anti RNASEH2A (Proteintech, Rosemont, IL, 1:1000 in TBS-T containing 1% w/v skim milk).

Techniques: Expressing, Mutagenesis

Journal: Journal of Diabetes Research

Article Title: The Involvement of Notch1-RBP-Jk/Msx2 Signaling Pathway in Aortic Calcification of Diabetic Nephropathy Rats

doi: 10.1155/2017/8968523

Figure Lengend Snippet: The mRNA expressions of Notch1 (a), RBP-Jk (b), Msx2 (c), and Jagged1 (d) in aortic tissues. Data presented as mean ± standard deviation (SD). ∗ p < 0.05, ∗∗ p < 0.01, and ∗∗∗ p < 0.001 versus Nor group; Δ p > 0.05, # p < 0.05, ## p < 0.01, and ### p < 0.001. Nor group: normal controls; DN + VDN group: diabetic nephropathy rats with vitamin D3/nicotine-induced vascular calcification.

Article Snippet: The following primary antibodies were used for immunohistochemical analysis: monoclonal rabbit anti-rat RBP-Jk, Msx2, and N1-ICD (Shanghai Bioworld Tech Inc., China); monoclonal rabbit anti-rat Jagged1 and Runx2 (Beijing BioTech Inc., China); monoclonal rabbit anti-rat Notch1 (Wuhan Proteintech Group Inc., China); and monoclonal mouse anti-rat α -SMA (Wuhan Boster Bio-Tech., Ltd, China).

Techniques: Standard Deviation

Journal: Journal of Diabetes Research

Article Title: The Involvement of Notch1-RBP-Jk/Msx2 Signaling Pathway in Aortic Calcification of Diabetic Nephropathy Rats

doi: 10.1155/2017/8968523

Figure Lengend Snippet: Detection of Notch1, RBP-Jk, Msx2, Jagged1, and N1-ICD levels by immunohistochemical analysis. The representative images and quantitative analysis of Notch1 (a-b), RBP-Jk (c-d), Msx2 (e-f), Jagged1 (g-h), and N1-ICD (i-j) in aortic tissues. The left panel of histological images: Nor group; right panel: DN + VDN group at 8, 12 , and 16 weeks, respectively. Data presented as mean ± standard deviation (SD). ∗ p < 0.05, ∗∗ p < 0.01, and ∗∗∗ p < 0.001 versus Nor group; Δ p > 0.05, # p < 0.05, ## p < 0.01, and ### p < 0.001. Nor group: normal controls; DN + VDN group: diabetic nephropathy rats with vitamin D3/nicotine-induced vascular calcification.

Article Snippet: The following primary antibodies were used for immunohistochemical analysis: monoclonal rabbit anti-rat RBP-Jk, Msx2, and N1-ICD (Shanghai Bioworld Tech Inc., China); monoclonal rabbit anti-rat Jagged1 and Runx2 (Beijing BioTech Inc., China); monoclonal rabbit anti-rat Notch1 (Wuhan Proteintech Group Inc., China); and monoclonal mouse anti-rat α -SMA (Wuhan Boster Bio-Tech., Ltd, China).

Techniques: Immunohistochemical staining, Standard Deviation