Journal: Life

Article Title: Modulation of Oxidative and ER Stress Pathways by the ADAM17 Inhibitor GW280264X in LPS-Induced Acute Liver Injury

doi: 10.3390/life15121877

Figure Lengend Snippet: Effects of GW280264X on redox regulatory proteins in liver tissue. ( A ) Kelch-like ECH-associated protein 1 (Keap1), ( B ) nuclear factor erythroid 2 2-related factor 2 (Nrf2), and ( C ) glutathione (GSH) levels. Data are presented as mean ± SEM ( n = 5). * p < 0.05 vs. 0.9% NaCl, # p < 0.05 vs. LPS. Black squares represent individual data points for each animal.

Article Snippet: Commercial ELISA kits were used to quantify liver tissue levels of KEAP1 (Cat. No. E2198Mo, BT-Laboratory, Shanghai, China), CHOP (Cat. No. E2718Mo, BT-Laboratory), 4-HNE (Cat. No. E1293Mo, BT-Laboratory), MDA (Cat. No. E0625Mo, BT-Laboratory), NRF2 (Cat. No. E1367Mo, BT-Laboratory), TNF-α (Cat. No. E-EL-M3063, Elabscience, Houston, TX, USA), GRP78 (Cat. No. E-EL-M2696, Elabscience, Houston, TX, USA), glutathione (GSH) colorimetric assay kit (Cat No. E-EL-0026, Elabscience, Houston, TX, USA) and ATF6 (Cat. No. ELK4479, Elk Biotechnology, Wuhan, China).

Techniques:

Journal: Journal of Cell Communication and Signaling

Article Title: Unveiling the role of Jagged2 in hypoxic pulmonary arterial hypertension: A NOX2‐mediated pathway

doi: 10.1002/ccs3.70032

Figure Lengend Snippet: Effects of NADPH oxidase 2 (NOX2) overexpression on proliferation and migration of pulmonary artery smooth muscle cells (PASMCs) under hypoxic conditions. (A) Expression of Jag2 and NOX2 in PASMCs at various time points under hypoxic treatment detected by immunofluorescence. (B) mRNA levels of Jag2 and NOX2 in PASMCs at various time points under hypoxic treatment detected by qRT‐PCR. (C) Cell proliferation levels were measured by the CCK‐8 assay. (D) Cell proliferation levels assessed by EdU staining, scale bar = 50 μm. (E) Protein expression of Jag2, NOX2, proliferating cell nuclear antigen (PCNA), and survivin in different groups was detected by the western blot. (F) Cell migration levels were assessed using a Transwell assay. (G) Protein expression of nuclear factor erythroid 2–related factor 2 (Nrf2) and SOD2 in different groups detected by the western blot; levels of superoxide dismutase (SOD), malondialdehyde (MDA), total glutathione (GSH), reduced GSH, and oxidized GSH measured by assay kits; reactive oxygen species (ROS) levels assessed by 2',7'‐dichlorodihydrofluorescein diacetate (DCFH‐DA) staining. (H) Protein expression of caspase‐3, cleaved caspase‐3, B‐cell lymphoma 2 (Bcl2), and Bcl‐2‐associated X protein (BAX) in different groups detected by the western blot. (I) Apoptosis levels in cells of different groups were assessed by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay, scale bar = 50 μm. (J) Apoptosis levels in cells of different groups were measured by flow cytometry (FCM). * indicates p < 0.05 compared to the hypoxia + siJag2 group, ** indicates p < 0.01 compared to the hypoxia + siJag2 group; all cell experiments were performed in triplicate.

Article Snippet: The membrane was blocked with 5% nonfat dry milk at room temperature for 1 h. The PVDF membrane was incubated overnight at 4°C with primary antibodies diluted in tris‐buffered saline with tween 20 (TBST): Jag2 (bs‐4244R, Bioss), NOX2 (19013‐1‐AP, Proteintech), nuclear factor erythroid 2‐related factor 2 (Nrf2) (16396‐1‐AP, Proteintech), proliferating cell nuclear antigen (PCNA) (10205‐2‐AP, Proteintech), survivin (10508‐1‐AP, Proteintech), SOD2 (24127‐1‐AP, Proteintech), cleaved caspase‐3 (68773‐1‐lg, Proteintech), B‐cell lymphoma 2 (68103‐1‐lg, Proteintech), Bcl‐2‐associated X protein (BAX) (50599‐2‐lg, Proteintech), α‐SMA (14395‐1‐AP, Proteintech), vimentin (10366‐1‐AP, Proteintech), CD31 (28083‐1‐AP, Proteintech), VE‐cadherin (A25003, Abclonal), and β‐actin (81115‐1‐RR, Proteintech).

Techniques: Over Expression, Migration, Expressing, Immunofluorescence, Quantitative RT-PCR, CCK-8 Assay, Staining, Western Blot, Transwell Assay, Measured Assay, TUNEL Assay, Flow Cytometry

Journal: Journal of Cell Communication and Signaling

Article Title: Unveiling the role of Jagged2 in hypoxic pulmonary arterial hypertension: A NOX2‐mediated pathway

doi: 10.1002/ccs3.70032

Figure Lengend Snippet: Mechanistic study of Jag2 promoting pulmonary artery smooth muscle cell (PASMC) proliferation and migration under hypoxic conditions through the NADPH oxidase 2 (NOX2)/reactive oxygen species (ROS) pathway. (A) CCK‐8 assay to measure cell proliferation levels in each group; (B, C) EdU staining to measure cell proliferation levels in each group (B), with panel C showing the bar graph statistical results of panel B, scale bar = 50 μm; (D) western blot analysis of Jag2, NOX2, proliferating cell nuclear antigen (PCNA), and survivin protein expression in each group; (E) Transwell assay to measure migration levels in each group; (F) western blot analysis of nuclear factor erythroid 2–related factor 2 (Nrf2) and SOD2 protein expression in each group; (G) kit assays to measure superoxide dismutase (SOD) and malondialdehyde (MDA) levels in each group; (H) kit assays to measure total glutathione (GSH), reduced GSH, and oxidized GSH levels in each group; (I) 2',7'‐dichlorodihydrofluorescein diacetate (DCFH‐DA) staining to measure ROS levels in each group; (J) western blot analysis of caspase‐3, cleaved caspase‐3, B‐cell lymphoma 2 (Bcl2), and Bcl‐2‐associated X protein (BAX) expression in each group; (K) terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay to measure apoptosis levels in each group, scale bar = 50 μm; (L) flow cytometry (FCM) to measure apoptosis levels in each group. * p < 0.05 compared to the normoxia group, ** p < 0.01 compared to the normoxia group, # p < 0.05 compared to the hypoxia + siNC group, ## p < 0.01 compared to the hypoxia + siNC group. All experiments were repeated three times.

Article Snippet: The membrane was blocked with 5% nonfat dry milk at room temperature for 1 h. The PVDF membrane was incubated overnight at 4°C with primary antibodies diluted in tris‐buffered saline with tween 20 (TBST): Jag2 (bs‐4244R, Bioss), NOX2 (19013‐1‐AP, Proteintech), nuclear factor erythroid 2‐related factor 2 (Nrf2) (16396‐1‐AP, Proteintech), proliferating cell nuclear antigen (PCNA) (10205‐2‐AP, Proteintech), survivin (10508‐1‐AP, Proteintech), SOD2 (24127‐1‐AP, Proteintech), cleaved caspase‐3 (68773‐1‐lg, Proteintech), B‐cell lymphoma 2 (68103‐1‐lg, Proteintech), Bcl‐2‐associated X protein (BAX) (50599‐2‐lg, Proteintech), α‐SMA (14395‐1‐AP, Proteintech), vimentin (10366‐1‐AP, Proteintech), CD31 (28083‐1‐AP, Proteintech), VE‐cadherin (A25003, Abclonal), and β‐actin (81115‐1‐RR, Proteintech).

Techniques: Migration, CCK-8 Assay, Staining, Western Blot, Expressing, Transwell Assay, TUNEL Assay, Flow Cytometry

Journal: Journal of Cell Communication and Signaling

Article Title: Unveiling the role of Jagged2 in hypoxic pulmonary arterial hypertension: A NOX2‐mediated pathway

doi: 10.1002/ccs3.70032

Figure Lengend Snippet: Effects of the Jag2/NADPH oxidase 2 (NOX2) pathway on inflammation, oxidative stress, and apoptosis in a hypoxic pulmonary arterial hypertension (PAH) rat model. (A) Immunohistochemistry detection of CD68 expression in the lungs of different groups of rats, scale bar = 50 μm; (B) ELISA detection of tumor necrosis factor‐alpha (TNF‐α) and IL‐6 levels in serum and lung tissue; (C) measurement of superoxide dismutase (SOD) and malondialdehyde (MDA) levels in the serum of different rat groups; (D) western blot detection of nuclear factor erythroid 2–related factor 2 (Nrf2) and SOD2 expression in lung tissue; (E) DHE staining for reactive oxygen species (ROS) levels in lung tissue, scale bar = 50 μm; (F) terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) detection of apoptosis levels in lung tissue, scale bar = 20 μm. * p < 0.05 compared to the control group, ** p < 0.01 compared to the control group, # p < 0.05 compared to the model + AAV‐shNC group, ## p < 0.01 compared to the model + AAV‐shNC group, N = 8.

Article Snippet: The membrane was blocked with 5% nonfat dry milk at room temperature for 1 h. The PVDF membrane was incubated overnight at 4°C with primary antibodies diluted in tris‐buffered saline with tween 20 (TBST): Jag2 (bs‐4244R, Bioss), NOX2 (19013‐1‐AP, Proteintech), nuclear factor erythroid 2‐related factor 2 (Nrf2) (16396‐1‐AP, Proteintech), proliferating cell nuclear antigen (PCNA) (10205‐2‐AP, Proteintech), survivin (10508‐1‐AP, Proteintech), SOD2 (24127‐1‐AP, Proteintech), cleaved caspase‐3 (68773‐1‐lg, Proteintech), B‐cell lymphoma 2 (68103‐1‐lg, Proteintech), Bcl‐2‐associated X protein (BAX) (50599‐2‐lg, Proteintech), α‐SMA (14395‐1‐AP, Proteintech), vimentin (10366‐1‐AP, Proteintech), CD31 (28083‐1‐AP, Proteintech), VE‐cadherin (A25003, Abclonal), and β‐actin (81115‐1‐RR, Proteintech).

Techniques: Immunohistochemistry, Expressing, Enzyme-linked Immunosorbent Assay, Western Blot, Staining, TUNEL Assay, Control