Journal: Cell Communication and Signaling : CCS

Article Title: Signaling pathways regulating VDAC1 overexpression associated with apoptosis, pyroptosis, and ferroptosis

doi: 10.1186/s12964-025-02647-5

Figure Lengend Snippet: Involvement of the MAPK-p38, JNK, and CaMK-II pathways in cisplatin- and erastin-induced VDAC1 overexpression. A HeLa cells were serum-starved for 5 h, pre-incubated (2 h) with the indicated concentrations of SP203580, SP600125 or KN-62, then incubated with or without cisplatin (15 µM). After 16 h, RNA was isolated and subjected to q-RT-PCR using VDAC1 mRNA specific primers (Table S2). B , C HeLa cells were serum-starved for 5 h, pre-incubated with the indicated inhibitor (10 µM, 2 h) then incubated with or without cisplatin (10 or 15 µM, 48 h) and subjected to VDAC1 oligomerization assayed as described in the Methods section. The immunoblot with the positions of VDAC1 monomers, dimers, trimers and multimers are indicated ( B ) and the levels of VDAC1 dimers were quantified ( C ). D , E HeLa cells were serum-starved for 5 h, and pre-incubated with the JNK inhibitor, SP600125 or with the CaMK-II inhibitor, KN-62 (5 or10 µM, 2 h), then incubated with or without cisplatin (15µM, 48 h) and subjected to immunoblotting using specific antibodies against P-c-Jun, P-ATF-1 or b-actin ( D ) and their levels were quantified ( E ). F , G C6 cells were serum starved for 2 h, pre-incubated with p38-MAPK inhibitor, SB203580 (5 and 10 µM, 2 h), then incubated with or without erastin (10 µM, 24 h). Cells were subjected to immunoblotting using specific antibodies against VDAC1, P-c-Jun, P-c-Fos or P-p38. Ponceau S staining is shown as a loading control ( F ). The protein relative levels were then quantified ( G ). H-K HeLa cells were transfected with non-targeting siRNA (si-NT) or si-c-Jun (100 nM) using JetPrime ( H , I ), or with si-p38 (100 nM) using SilentFect transfection reagent ( J , K ), as described in the Methods section. At 24 h post-transfection, cells were treated with cisplatin (15 µM, 48 h) and subjected to immunoblotting for P-c-Jun, P-p38, P-c-Fos or b-actin expression using specific antibodies (H, J) . Protein expression levels were quantified ( I , K ). Cell death was analyzed by PI standing and FACS analysis and is presented in the bottom of the blots ( H , J ). Results are the means ± SEM (n = 3). ∗p ≤ 0.05, ∗∗p ≤ 0.01, ∗∗∗p ≤ 0.001; NS = non-significant

Article Snippet: The nucleotides in italic were 2′-O-methyl modified. si-c-JUN (CST-6203 S) and si-p38-MAPK (CST-6564 S) were purchased from Cell Signaling Technology (Danvers, MS).

Techniques: Over Expression, Incubation, Isolation, Reverse Transcription Polymerase Chain Reaction, Western Blot, Staining, Control, Transfection, Expressing

Journal: Cell Death & Disease

Article Title: Targeting c-MET by Tivantinib through synergistic activation of JNK/c-jun pathway in cholangiocarcinoma

doi: 10.1038/s41419-019-1460-1

Figure Lengend Snippet: a Effect on the expression of JNK and c-jun by co-transfection of JNK-specific siRNA oligonucleotide sequences (each at the concentration of 25 and 50 nM) in HuCC-T1 and EGI-1 cell were assessed by western blotting; medium or non-coding siRNA (Ctrl-siRNA) were used as control. b Cell viability after transfection of siRNA targeting JNK in HuCC-T1 and EGI-1 cell. * p < 0.05 in comparison with cells treated by Tivantinib 0.5 µM for 24 h. FACS analysis of apoptosis **** p , *** p , * * p < 0.01, and * p < 0.05 in comparison with cells with medium or with non-coding siRNA transfection. For this experiment, Tivantinib 0.5 µM was added into cell culture for 3 h after 24 h JNK siRNA transfection. c Cell viability analysis of HGF (25 and 50 ng/mL) stimulated cells treated by Tivantinib. p-MET, MET, p-JNK, and JNK1 were evaluated by western blotting in the same condition. HFG stimulated HuCC-T1 and EGI-1 cell were incubated with HGF for 3 h before being added to Tivantinib 0.5 and 5 µM for 24, 48, and 72 h. d Effect of c-jun silencing by specific siRNA in CC cells. e FACS analysis of apoptosis after transfection of siRNA targeting c-jun.**** p , *** p , ** p < 0.01, and * p < 0.05 in comparison with cells with medium or with non-coding siRNA transfection. f Western blotting analysis of p-MET, MET, p-JNK, JNK1, p-c jun FADD, and cleaved caspase-3 express in HuCC-T1 and EGI-1 cell treated by c-MET neutralizing antibody for 24, 48, and 72 h

Article Snippet: For c-jun N-terminal protein kinase (JNK) and c-jun knockdown in human CC cell lines, the following pre-designed and pre-validated small interfering RNAs (siRNAs) were purchased from Cell Signaling Technology: SignalSilence ® SAPK/JNK siRNA II (catalog number 6233) and SignalSilence ® c-jun siRNA II (catalog number 6204).

Techniques: Expressing, Cotransfection, Concentration Assay, Western Blot, Control, Transfection, Comparison, Cell Culture, Incubation

Journal: Journal of Cellular and Molecular Medicine

Article Title: Inhibition of miR‐148a‐3p resists hepatocellular carcinoma progress of hepatitis C virus infection through suppressing c‐Jun and MAPK pathway

doi: 10.1111/jcmm.14045

Figure Lengend Snippet: miRNA mimics and inhibitor, c‐Jun siRNA sequence

Article Snippet: SiRNA 1 and siRNA 2 for silencing c‐Jun (si‐ c‐Jun ‐1 and si‐ c‐Jun ‐2) were purchased from Ribobio.

Techniques: Sequencing

Journal: BMC Cancer

Article Title: TNF-alpha promotes lymphangiogenesis and lymphatic metastasis of gallbladder cancer through the ERK1/2/AP-1/VEGF-D pathway

doi: 10.1186/s12885-016-2259-4

Figure Lengend Snippet: TNF-α upregulated VEGF-D expression and VEGF-D promoter activity downstream of the ERK1/2/AP-1 pathway. a , c The effect of the TNF-α⁄AP-1 signaling pathway on the promoter activity and protein expression of the VEGF-D gene. Transfection with AP-1 siRNA effectively knocked down the expression of AP-1 and p-AP-1 in both NOZ and GBC-SD cells. The protein level and promoter activity of VEGF-D were accordingly reduced irrespective of treatment with TNF-α. b , d The effect of inhibition of MAPK pathway members on the protein expression and promoter activity of VEGF-D. When treated with SP600125 (10 μM), SB203580 (20 μM) or PD98059 (50 μM), the expression of AP-1 and p-AP-1 in both NOZ and GBC-SD cells were reduced. However, the protein expression and promoter activity of VEGF-D were significantly reduced only in the PD98059-treated group. * P < 0.05

Article Snippet: The AP-1 (c-Jun) siRNA interference sequence has been described previously [ ] (named siAP-1, sense: 5′-GAUGGAAACGACCUUCUAUdTdT-3′, anti-sense: 5′-AUAGAAGGUCGUUUCCAUCdTdT-3′), and the non-targeting control (named siNC) were synthesized chemically by GenePharma Co., Ltd. (Suzhou, China).

Techniques: Expressing, Activity Assay, Transfection, Inhibition

Journal: BMC Cancer

Article Title: TNF-alpha promotes lymphangiogenesis and lymphatic metastasis of gallbladder cancer through the ERK1/2/AP-1/VEGF-D pathway

doi: 10.1186/s12885-016-2259-4

Figure Lengend Snippet: The TNF-α - VEGF-D axis promoted the tube formation of human dermal lymphatic endothelial cells (HDLECs) in vitro . a , b Construction of a NOZ cell line and a GBC-SD cell line stably expressing lentiviral VEGF-D shRNA and a green fluorescent protein sequence. The cells were observed under a fluorescence microscope with bright or blue light. c , d VEGF-D mRNA and protein expression of NOZ or GBC-SD cells stably transfected with LV-siVEGF-D were analyzed by real-time reverse transcription-polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA), respectively. GAPDH served as an internal control. e , f , g , h DiI-labeled HDLECs (emit red fluorescence) were cocultured with the three NOZ (or GBC-SD) cell lines and were treated with TNF-α (50 ng⁄ mL) for 5 h. HDLEC tube formation was observed under fluorescence microscopy, and the tube number was counted. (* P < 0.05; ** P < 0.01; *** P < 0.001)

Article Snippet: The AP-1 (c-Jun) siRNA interference sequence has been described previously [ ] (named siAP-1, sense: 5′-GAUGGAAACGACCUUCUAUdTdT-3′, anti-sense: 5′-AUAGAAGGUCGUUUCCAUCdTdT-3′), and the non-targeting control (named siNC) were synthesized chemically by GenePharma Co., Ltd. (Suzhou, China).

Techniques: In Vitro, Stable Transfection, Expressing, shRNA, Sequencing, Fluorescence, Microscopy, Transfection, Reverse Transcription, Polymerase Chain Reaction, Reverse Transcription Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay, Control, Labeling