Journal: Cell Reports Medicine

Article Title: Interferon-γ-stimulated antigen-presenting cancer-associated fibroblasts hinder neoadjuvant chemoimmunotherapy efficacy in lung cancer

doi: 10.1016/j.xcrm.2025.102017

Figure Lengend Snippet:

Article Snippet: Phospho-Jak2 (Tyr1007/1008) Rabbit Polyclonal Antibody , Beyotime Biotechnology , Cat# AF5854; RRID: AB_3665514.

Techniques: Recombinant, Staining, Purification, In Vivo, Control, Protease Inhibitor, Activation Assay, Membrane, Modification, Bicinchoninic Acid Protein Assay, Immunohistochemical staining, Enzyme-linked Immunosorbent Assay, Cell Isolation, Sequencing, Negative Control, shRNA, Software

Journal: Neural Regeneration Research

Article Title: Quantitative proteomic and phosphoproteomic analyses of the hippocampus reveal the involvement of NMDAR1 signaling in repetitive mild traumatic brain injury

doi: 10.4103/1673-5374.374654

Figure Lengend Snippet: NMDAR1 helps regulate hippocampus-mediated cognitive dysfunction after rmTBI. (A) Western blot of hippocampal NMDAR1 and pNMDAR1. No change in NMDAR1 protein expression was detected (unpaired t -test, t = 1.406, P = 0.2324), and a relative increase in pNMDAR1 protein expression (unpaired t -test, t = 2.827, P = 0.0475) was detected ( n = 3). NMDAR1: 105 kDa, pNMDAR1: 105 kDa, Tubulin: 55 kDa. (B) Schematic showing how saline and CGP78608 were injected into the hippocampus. (C) The decreased NOR index induced by rmTBI was almost abolished by treatment with CGP78608, but not saline ( n = 6) (unpaired t -test, t = 3.168, P = 0.0100). (D) Interaction network among Grin1 and its upstream kinases. (E) Grin1 PPI network. All data are expressed as the mean ± SEM. * P < 0.05. NMDAR1: N-methyl-D-aspartate receptor 1; NOR: novel object recognition; pNMDAR1: phosphorylation N-methyl-D-aspartate receptor 1; rmTBI: repetitive mild traumatic brain injury.

Article Snippet: Subsequently, the proteins were transferred to PVDF membranes (Millipore, Darmstadt, Germany) that were blocked with 5% nonfat milk for 1 hour and incubated with the following primary antibodies overnight at 4°C: anti-NMDAR1 (rabbit, 1:2000, EPR2481(2), Cat# ab109182, Abcam, Cambridge, UK); anti-pNMDAR1 (rabbit, 1:700, Cat# ab195002, Abcam); β-tubulin (rabbit, 1:2000, ARC0203, Cat# A12289, ABclonal, Wuhan, China).

Techniques: Western Blot, Expressing, Injection

Journal: International Journal of Molecular Sciences

Article Title: 3 β -Hydroxy-12-oleanen-27-oic Acid Exerts an Antiproliferative Effect on Human Colon Carcinoma HCT116 Cells via Targeting FDFT1

doi: 10.3390/ijms241915020

Figure Lengend Snippet: Identification of the antitumor targets of ATA. ( A ) Overlap of the predicted targets of ATA (blue) and disease targets from the transcriptomic data (pink). ( B ) Function analysis of fifteen common targets via GeneMANIA. (C) Binding pattern diagram of ATA and the target proteins FDFT1, PPARA, and PPARG. The yellow dashed line represents the hydrogen bond interaction. ( D – F ) After pre-incubation with or without YM-53601 (2.5 μM and 5 µM, ( D )), GW6471 (2.5 μM and 5 µM, ( E )), or GW9662 (5 μM and 10 µM, ( F )) for 30 min, HCT116 cells were treated with ATA (0 μM, 10 μM, and 15 µM) for 48 h. The cell viabilities were detected via the MTT assay. The data are expressed as means ± SD ( n = 3). ** p < 0.01 and *** p < 0.001 vs. control (Ctrl). ( G ) Venn diagram of predicted targets for ATA, OA, and UA. Five genes in red were identified as ATA-specific targets. ( H ) The cell viabilities of HCT116 cells treated with ATA, UA, and OA for 48 h were detected via the MTT assay and the IC 50 values were calculated using Graphpad Prism 9.0 software.

Article Snippet: For this study, 3-(4,5-dimethyl thiazolyl-2)-2,5-diphenyltetrazolium bromide (MTT), acridine orange (AO), and dimethyl sulfoxide (DMSO) were purchased from Sigma-Aldrich, St. Louis, MO, USA; cell culture medium, trypsin, penicillin, streptomycin, and fetal bovine serum (FBS) were obtained from Gibco, Burlington, MA, USA; TRIzol was purchased from Invitrogen, Carlsbad, CA, USA; reverse transcriptase, oligo(dT) 18 , and ribonuclease inhibitor were obtained from Shanghai Sangon Biotech Co., Ltd., Shanghai, China; RIPA lysis buffer, BCA protein assay kit, anti-mouse actin mAbs, anti-caspase3 antibody (AF0081), horseradish peroxidase (HRP)-conjugated goat anti-rabbit and anti-mouse IgG (H + L), enhanced chemiluminescence (ECL) kit, and reactive oxygen species (ROS) assay kit were purchased from Beyotime, Shanghai, China; phosphatase inhibitor and protease inhibitor cocktails were obtained from Biotool, Houston, TX, USA; anti-FDFT1 antibody (ab195046) was purchased from Abcam, Cambridge, UK; anti-cyclinD1 antibody (ET1601-31) was obtained from HUABIO, Shanghai, China; anti-GPX4 (67763-1-Ig) antibody was obtained from Proteintech, Rosemont, IL, USA; the antibodies against LC3A/B (4108S) and p62 (39749S) were purchased from CST, Danvers, MA, USA; a monodansylcadaverine (MDC) sensor kit was obtained from KeyGEN BioTECH Corp., Ltd., Nanjing, China; an annexin V-FITC apoptosis detection kit and propidium iodide (PI)/RNase staining buffer were obtained from BD Pharmingen, San Diego, CA, USA; mRFP-GFP-LC3 lentivirus was obtained from Genechem, Shanghai, China.

Techniques: Binding Assay, Incubation, MTT Assay, Software

Journal: International Journal of Molecular Sciences

Article Title: 3 β -Hydroxy-12-oleanen-27-oic Acid Exerts an Antiproliferative Effect on Human Colon Carcinoma HCT116 Cells via Targeting FDFT1

doi: 10.3390/ijms241915020

Figure Lengend Snippet: FDFT1 mediated the cytotoxicity of ATA towards HCT116 cells. ( A ) The gene expression levels of FDFT1 in the HCT116 cells treated with ATA for different times via the RT-qPCR assay. ( B , C ) The protein expression levels of FDFT1 in the HCT116 cells treated with ATA (7.5, 15 and 30 μM) for 24 h via Western blotting. The figure ( B ) shown is representative of three independent experiments. The data ( C ) are expressed as means ± SD ( n = 3). * p < 0.05 and ** p < 0.01 vs. Ctrl (0 μM). ( D – F ) After pre-incubation with or without the FDFT1 inhibitor YM-53601 (2 and 4 µM) for 30 min, HCT116 cells were treated with ATA (0 µM, 5 µM, and 7.5 µM, D), OA (100 μM, E), or UA (40 μM, F) for 24 h. The cell viabilities were detected via the MTT assay. The data are expressed as means ± SD ( n = 3). * p < 0.05 vs. the control (Ctrl). ( G – H ) After pre-incubation with or without YM-53601 (4 µM, 30 min), the HCT116 cells were treated with ATA (20 μM) for 24 h. The autophagic flux was observed through the mRFP-GFP-LC3 assay. The figure ( G ) shown is representative of three independent experiments. The cell apoptosis was determined using FCM. The figure ( H ) is representative of three independent experiments. The apoptotic percentages ( I ) are expressed as means ± SD ( n = 3). *** p < 0.001 vs. Ctrl. ( J – L ) The mRNA ( J ) and protein ( K , L ) expression levels of FDFT1 in HCT116 cells were detected using RT-qPCR and Western blotting after transfection with FDFT1 siRNA for 24 h and 48 h, respectively. siNC—negative control siRNA. The figure ( K ) shown is representative of three independent experiments. The data ( J , L ) are expressed as means ± SD ( n = 3). * p < 0.05 and *** p < 0.001 vs. siNC. ( M ) The HCT116 cells were transfected with FDFT1 siRNAs for 48 h, followed by exposure to ATA at indicated concentrations for another 48 h. The cell viabilities were detected through the MTT assay. The data are expressed as means ± SD ( n = 3). * p < 0.05 and *** p < 0.001 vs. siNC.

Article Snippet: For this study, 3-(4,5-dimethyl thiazolyl-2)-2,5-diphenyltetrazolium bromide (MTT), acridine orange (AO), and dimethyl sulfoxide (DMSO) were purchased from Sigma-Aldrich, St. Louis, MO, USA; cell culture medium, trypsin, penicillin, streptomycin, and fetal bovine serum (FBS) were obtained from Gibco, Burlington, MA, USA; TRIzol was purchased from Invitrogen, Carlsbad, CA, USA; reverse transcriptase, oligo(dT) 18 , and ribonuclease inhibitor were obtained from Shanghai Sangon Biotech Co., Ltd., Shanghai, China; RIPA lysis buffer, BCA protein assay kit, anti-mouse actin mAbs, anti-caspase3 antibody (AF0081), horseradish peroxidase (HRP)-conjugated goat anti-rabbit and anti-mouse IgG (H + L), enhanced chemiluminescence (ECL) kit, and reactive oxygen species (ROS) assay kit were purchased from Beyotime, Shanghai, China; phosphatase inhibitor and protease inhibitor cocktails were obtained from Biotool, Houston, TX, USA; anti-FDFT1 antibody (ab195046) was purchased from Abcam, Cambridge, UK; anti-cyclinD1 antibody (ET1601-31) was obtained from HUABIO, Shanghai, China; anti-GPX4 (67763-1-Ig) antibody was obtained from Proteintech, Rosemont, IL, USA; the antibodies against LC3A/B (4108S) and p62 (39749S) were purchased from CST, Danvers, MA, USA; a monodansylcadaverine (MDC) sensor kit was obtained from KeyGEN BioTECH Corp., Ltd., Nanjing, China; an annexin V-FITC apoptosis detection kit and propidium iodide (PI)/RNase staining buffer were obtained from BD Pharmingen, San Diego, CA, USA; mRFP-GFP-LC3 lentivirus was obtained from Genechem, Shanghai, China.

Techniques: Expressing, Quantitative RT-PCR, Western Blot, Incubation, MTT Assay, Transfection, Negative Control

Journal: International Journal of Molecular Sciences

Article Title: 3 β -Hydroxy-12-oleanen-27-oic Acid Exerts an Antiproliferative Effect on Human Colon Carcinoma HCT116 Cells via Targeting FDFT1

doi: 10.3390/ijms241915020

Figure Lengend Snippet: ATA inhibited the tumor growth of HCT116 xenografts in nude mice. ( A ) Tumor anatomy of HCT116 subcutaneous tumor-bearing mice. Scale bar = 10 mm. ( B ) Growth curve of tumor volume. ( C ) Average final tumor weights of each group. ( D ) Body weights of tumor-bearing mice. ( E , F ) Expression levels of FDFT1, LC3, p62, cyclinD1, pro-caspase3, and GPX4 in tumor tissues from Ctrl and ATA (60 mg/kg) groups via Western blotting. The data were expressed as means ± SD ( n = 4). * p < 0.05, ** p < 0.01, and *** p < 0.001 vs. Ctrl.

Article Snippet: For this study, 3-(4,5-dimethyl thiazolyl-2)-2,5-diphenyltetrazolium bromide (MTT), acridine orange (AO), and dimethyl sulfoxide (DMSO) were purchased from Sigma-Aldrich, St. Louis, MO, USA; cell culture medium, trypsin, penicillin, streptomycin, and fetal bovine serum (FBS) were obtained from Gibco, Burlington, MA, USA; TRIzol was purchased from Invitrogen, Carlsbad, CA, USA; reverse transcriptase, oligo(dT) 18 , and ribonuclease inhibitor were obtained from Shanghai Sangon Biotech Co., Ltd., Shanghai, China; RIPA lysis buffer, BCA protein assay kit, anti-mouse actin mAbs, anti-caspase3 antibody (AF0081), horseradish peroxidase (HRP)-conjugated goat anti-rabbit and anti-mouse IgG (H + L), enhanced chemiluminescence (ECL) kit, and reactive oxygen species (ROS) assay kit were purchased from Beyotime, Shanghai, China; phosphatase inhibitor and protease inhibitor cocktails were obtained from Biotool, Houston, TX, USA; anti-FDFT1 antibody (ab195046) was purchased from Abcam, Cambridge, UK; anti-cyclinD1 antibody (ET1601-31) was obtained from HUABIO, Shanghai, China; anti-GPX4 (67763-1-Ig) antibody was obtained from Proteintech, Rosemont, IL, USA; the antibodies against LC3A/B (4108S) and p62 (39749S) were purchased from CST, Danvers, MA, USA; a monodansylcadaverine (MDC) sensor kit was obtained from KeyGEN BioTECH Corp., Ltd., Nanjing, China; an annexin V-FITC apoptosis detection kit and propidium iodide (PI)/RNase staining buffer were obtained from BD Pharmingen, San Diego, CA, USA; mRFP-GFP-LC3 lentivirus was obtained from Genechem, Shanghai, China.

Techniques: Expressing, Western Blot

Journal: International Journal of Molecular Sciences

Article Title: 3 β -Hydroxy-12-oleanen-27-oic Acid Exerts an Antiproliferative Effect on Human Colon Carcinoma HCT116 Cells via Targeting FDFT1

doi: 10.3390/ijms241915020

Figure Lengend Snippet: Hypothetical pathway of FDFT1 in mediating the cytotoxic effect of ATA against HCT116 cells.

Article Snippet: For this study, 3-(4,5-dimethyl thiazolyl-2)-2,5-diphenyltetrazolium bromide (MTT), acridine orange (AO), and dimethyl sulfoxide (DMSO) were purchased from Sigma-Aldrich, St. Louis, MO, USA; cell culture medium, trypsin, penicillin, streptomycin, and fetal bovine serum (FBS) were obtained from Gibco, Burlington, MA, USA; TRIzol was purchased from Invitrogen, Carlsbad, CA, USA; reverse transcriptase, oligo(dT) 18 , and ribonuclease inhibitor were obtained from Shanghai Sangon Biotech Co., Ltd., Shanghai, China; RIPA lysis buffer, BCA protein assay kit, anti-mouse actin mAbs, anti-caspase3 antibody (AF0081), horseradish peroxidase (HRP)-conjugated goat anti-rabbit and anti-mouse IgG (H + L), enhanced chemiluminescence (ECL) kit, and reactive oxygen species (ROS) assay kit were purchased from Beyotime, Shanghai, China; phosphatase inhibitor and protease inhibitor cocktails were obtained from Biotool, Houston, TX, USA; anti-FDFT1 antibody (ab195046) was purchased from Abcam, Cambridge, UK; anti-cyclinD1 antibody (ET1601-31) was obtained from HUABIO, Shanghai, China; anti-GPX4 (67763-1-Ig) antibody was obtained from Proteintech, Rosemont, IL, USA; the antibodies against LC3A/B (4108S) and p62 (39749S) were purchased from CST, Danvers, MA, USA; a monodansylcadaverine (MDC) sensor kit was obtained from KeyGEN BioTECH Corp., Ltd., Nanjing, China; an annexin V-FITC apoptosis detection kit and propidium iodide (PI)/RNase staining buffer were obtained from BD Pharmingen, San Diego, CA, USA; mRFP-GFP-LC3 lentivirus was obtained from Genechem, Shanghai, China.

Techniques: