Journal: Translational Psychiatry

Article Title: Carboxypeptidase E conditional knockout mice exhibit learning and memory deficits and neurodegeneration

doi: 10.1038/s41398-023-02429-y

Figure Lengend Snippet: a – e Western blot analysis of p -TrkB, BDNF, p -mTOR, mTOR, p -AKT, and AKT levels in the hippocampus. f Immunofluorescence of CPE, MAP2, DCX, and GFAP; and the relative fluorescent intensities of g CPE in Sub, h MAP2 in Sub, i MAP2 in hilius, j DCX in DG, k GFAP in DG of WT, CPE flox/− , and CPE flox/flox mice at 100× and 400× (square in the panel). n = 6; * P < 0.05 and ** P < 0.01 compared with WT; values are mean ± SEM.

Article Snippet: After blocking with 5% nonfat milk, the membrane was blotted with antibodies against mouse anti-β-actin antibody (1:1000, CST, 4967S), mouse anti-CPE antibody (1:1000; BD bioscience, 610758), mouse anti-BDNF antibody (1:600, Abcam, UK, ab108319), rabbit anti-p-TrkB antibody (1:1000; Boster, Wuhan, China, BM4437), rabbit anti-AKT (1:2000, Cell Signaling Technology, 4691S), and anti-p-AKT (1:2000, Cell Signaling Technology, 23430S) overnight at 4 °C.

Techniques: Western Blot, Immunofluorescence

Journal: Experimental and Therapeutic Medicine

Article Title: Cucurbitacin B induces apoptosis in colorectal cells through reactive oxygen species generation and endoplasmic reticulum stress pathways

doi: 10.3892/etm.2023.12183

Figure Lengend Snippet: CuB induces ER stress in CRC cells. After CRC cells were treated with CuB (5 µM) for 48 h, the expression levels of ER stress-related proteins was evaluated by western blotting. (A and C) The expression of ERS-related proteins in different groups of HT-29 and SW620 cells. (B and D) Quantification of expression levels of ERS-related proteins. Data are presented as the mean ± SD of three independent experiments. * P<0.05, ** P<0.01, *** P<0.001 vs. control. ATF4, activating transcription factor 4; CRC, colorectal cancer; CuB, cucurbitacin B; eIF2α, eukaryotic translation initiation factor 2α; GRP78, glucose regulated protein 78; p-, phosphorylated; PERK, protein kinase R-like ER kinase; XBP1-s, X-box binding protein 1.

Article Snippet: Primary antibodies against the following proteins were used: GAPDH (1:10,000; 36 KDa; cat. no. 10494-1-AP; Wuhan Sanying Biotechnology); Bax (1:5,000; 21 kDa; cat. no. GR3275117-D; Abcam) and Bcl2 (1:2,000; 26 kDa; cat. no. GR3325129-9; Abcam); glucose regulated protein 78 (GRP78; 1:1,000; 78kDa; cat. no. 11587-1-AP; Wuhan Sanying Biotechnology), CHOP (27 kDa; cat. no. 44266; 1:1,000; GeneTex, Inc.), activating transcription factor 4 (ATF4; 42 kDa; cat. no. 39568; 1:1,000; GeneTex, Inc.) and X-box binding protein 1 splicing form (XBP1-s; 55 kDa; cat. no. 44286; 1:1,000; GeneTex, Inc.); IRE1 (1:1,000; 130 kDa; cat. no. 3294S; Cell Signaling Technology, Inc.); phosphorylated (p)-protein kinase R-like ER kinase (PERK; 1:1,000; 125 kDa; cat. no. 6N04; SAB Biotherapeutics, Inc.); PERK (1;1,000; 115 kDa; cat. no. 5683S; Cell Signaling Technology, Inc.); p-eukaryotic translation initiation factor 2α (eIF2α; 1:1,000; 32 kDa; cat. no. AP0692; ABclonal Biotech Co., Ltd.) and eIF2α (1:1,000; 38 kDa; cat. no. 42774; GeneTex, Inc.).

Techniques: Expressing, Western Blot, Binding Assay

Journal: Experimental and Therapeutic Medicine

Article Title: Cucurbitacin B induces apoptosis in colorectal cells through reactive oxygen species generation and endoplasmic reticulum stress pathways

doi: 10.3892/etm.2023.12183

Figure Lengend Snippet: CuB mediates ROS production in CRC cells and induces ERS. (A and B) After CRC cells were treated with CuB (0-5 µM) for 48 h, the dichlorodihydrofluorescein diacetate fluorescent probe was used to evaluate the amount of ROS produced. Scale bars, 200 µm. CRC cells were pretreated with NAC (5 mM) for 1 h before adding CuB (final concentration of 5 µM). After 48 h of treatment, the expression of ERS-related proteins was evaluated by western blotting. (C and E) The expression of ERS-related proteins in different groups of HT-29 and SW620 cells. (D and F) Quantification of expression levels of ERS-related proteins. Data are presented as the mean ± SD of three independent experiments. * P<0.05, ** P<0.01, *** P<0.001 vs. control or as indicated. CRC, colorectal cancer; CuB, cucurbitacin B; eIF2α, eukaryotic translation initiation factor 2α; ERS, endoplasmic reticulum stress; GRP78, glucose regulatory protein 78; NAC, N-acetylcysteine; p-, phosphorylated; PERK, protein kinase R-like ER kinase; ROS, reactive oxygen species; XBP1-s, X-box binding protein 1.

Article Snippet: Primary antibodies against the following proteins were used: GAPDH (1:10,000; 36 KDa; cat. no. 10494-1-AP; Wuhan Sanying Biotechnology); Bax (1:5,000; 21 kDa; cat. no. GR3275117-D; Abcam) and Bcl2 (1:2,000; 26 kDa; cat. no. GR3325129-9; Abcam); glucose regulated protein 78 (GRP78; 1:1,000; 78kDa; cat. no. 11587-1-AP; Wuhan Sanying Biotechnology), CHOP (27 kDa; cat. no. 44266; 1:1,000; GeneTex, Inc.), activating transcription factor 4 (ATF4; 42 kDa; cat. no. 39568; 1:1,000; GeneTex, Inc.) and X-box binding protein 1 splicing form (XBP1-s; 55 kDa; cat. no. 44286; 1:1,000; GeneTex, Inc.); IRE1 (1:1,000; 130 kDa; cat. no. 3294S; Cell Signaling Technology, Inc.); phosphorylated (p)-protein kinase R-like ER kinase (PERK; 1:1,000; 125 kDa; cat. no. 6N04; SAB Biotherapeutics, Inc.); PERK (1;1,000; 115 kDa; cat. no. 5683S; Cell Signaling Technology, Inc.); p-eukaryotic translation initiation factor 2α (eIF2α; 1:1,000; 32 kDa; cat. no. AP0692; ABclonal Biotech Co., Ltd.) and eIF2α (1:1,000; 38 kDa; cat. no. 42774; GeneTex, Inc.).

Techniques: Produced, Concentration Assay, Expressing, Western Blot, Binding Assay