Journal: Evidence-based Complementary and Alternative Medicine : eCAM

Article Title: Quercetin Improves Mitochondrial Function and Inflammation in H 2 O 2 -Induced Oxidative Stress Damage in the Gastric Mucosal Epithelial Cell by Regulating the PI3K/AKT Signaling Pathway

doi: 10.1155/2021/1386078

Figure Lengend Snippet: Que alleviates H 2 O 2 -induced oxidative stress, apoptosis, and inflammatory in GES-1 cells. (a) CCK-8 was performed to measure the viability of GES-1 cells. Flow cytometry was performed to evaluate the proportion of (b) cells with increase in ROS and (c) apoptotic cells. (d) Western blot was performed to detect the apoptosis-related proteins Bax, Bcl-2, and cleaved caspase 3 expression in GES-1 cells. qRT-PCR was performed to detect mRNA expression of (e) IL-10 and (f) TNF- α in GES-1 cells. Data presets as mean ± SD, n − = 3, ∗ P < 0.05 vs. control group, # P < 0.05 vs. model group.

Article Snippet: After blocking with 5% skim milk, the membranes were cultured for 1 h with primary antibodies against BCL-2 associated X protein (BaX) (1 : 1000, Bioswamp, PAB30040), B-cell lymphoma-2 (Bcl-2) (1 : 1000, Bioswamp, PAB33482), cleaved caspase 3 (1 : 1000, Abcam, Ab2302), density-regulated protein 1 (DRP1) (1 : 1000, Bioswamp, PAB33409) phosphorylation (p)-DRP1 (1 : 1000, Abcam, ab193216), mitochondrial profusion protein Mitofusins (Mfn2) (1 : 1000, Bioswamp, PAB41825), occludin (1 : 1000, Bioswamp, PAB33418), claudin-1 (1 : 1000, Bioswamp, PAB33267), zona occludens 1(ZO-1) (1 : 1000, Bioswamp, PAB36669), phosphatidylinositol-3 kinase (PI3K) (1 : 1000, Bioswamp, PAB30084), p-PI3K (1 : 1000, Abcam, Ab182651), AKT (1 : 1000, Bioswamp, PAB34089), p-AKT (1 : 1000, Abcam, Ab38449), and GAPDH (1 : 1000, Housekeeping, Bioswamp, PAB36269), followed by 1 h of incubation with goat anti-rabbit IgG secondary antibody (1 : 20000, Bioswamp, SAB43714).

Techniques: CCK-8 Assay, Flow Cytometry, Western Blot, Expressing, Quantitative RT-PCR, Control

Journal: Frontiers in Cell and Developmental Biology

Article Title: LncRNA FEZF1-AS1 Promotes Multi-Drug Resistance of Gastric Cancer Cells via Upregulating ATG5

doi: 10.3389/fcell.2021.749129

Figure Lengend Snippet: FEZF1-AS1 was upregulated in chemo-resistant GC tissues and knockdown of FEZF1-AS1 improved chemo-sensitivity of GC cells. (A) qRT-PCR on the level of FEZF1-AS1 in 27 pairs of specimens from GC patients with poor response to chemotherapy were 3.5-fold increase compared with patients with good response. (B) CCK8 assay on the viability of SGC-7901 and SGC-7901/CDDP cells treated with 10–80 μg/ml of CDDP for 24 h. (C) qRT-PCR on the level of FEZF1-AS1 in SGC-7901 and SGC-7901/CDDP cells. (D) CCK8 assay on the viability of SGC-7901 and SGC-7901/5-FU cells treated with 5–25 μg/ml of 5-FU for 24 h. (E) qRT-PCR on the level of FEZF1-AS1 in SGC-7901 and SGC-7901/5-FU cells. (F) Representative images and quantitation of flow cytometric analysis of apoptotic SGC-7901/CDDP cells with or without the treatment of FEZF1-AS1 siRNA after 24 h incubation with CDDP (40 mg/ml). (G) Western blot detected apoptotic proteins, BCL2, BAX, and Cleaved caspase3 in SGC-7901/CDDP cells with or without the treatment of FEZF1-AS1 siRNA after 24 h incubation with CDDP (40 μg/ml). (H) Representative images and quantitation of flow cytometric analysis of apoptotic SGC-7901/5-FU cells with or without the treatment of FEZF1-AS1 siRNA after 24 h incubation with 5-FU (20 μg/ml). (I) Western blot detected apoptotic proteins, BCL2, BAX, and Cleaved caspase3 in SGC-7901/5-FU cells with or without the treatment of FEZF1-AS1 siRNA after 24 h incubation with 5-FU (20 μg/ml). The protein expression of MDR1 and MRP1 in SGC-7901/CDDP (J) and SGC-7901/5-FU (L) cells detected by Western blot, FEZF1-AS1 siRNA decreased the protein levels of MDR1 and MRP1 in both cells. The knockdown efficiency of FEZF1-AS1 in SGC-7901/CDDP (K) and SGC-7901/5-FU (M) cells detected by qRT-PCR. Each experiment was performed in triplicate. Data are mean ± SD. * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001.

Article Snippet: Membranes were blocked with 5% non-fat milk in TBST for 1 h at room temperature and incubated overnight at 4°C with the first antibodies against drug resistance-associated proteins MDR1 (1:2000, PAB30805, Bioswamp), MRP1 (1:1000, PAB33537, Bioswamp), ATG5 (1:1000, 10181-2-AP, Proteintech), β-actin (1:1000, 66009-1-Ig, Proteintech), Ubiquitin (1:1000, 3936, CST), LC3B (1:1000, 3836, CST), BCL2 (1:1000, 12789-1-AP, Proteintech), BAX (1:1000, 50599-2-Ig, Proteintech) and Cleaved caspase3 (1:1000, 9661, CST).

Techniques: Knockdown, Quantitative RT-PCR, CCK-8 Assay, Quantitation Assay, Incubation, Western Blot, Expressing