jnk1 sirna  (Santa Cruz Biotechnology)

Journal: Cancer Research

Article Title: Inhibiting Fatty Acid Oxidation Reverses Autophagy-Mediated Acquired Chemotherapy Resistance in Pancreatic Ductal Adenocarcinoma

doi: 10.1158/0008-5472.CAN-25-4517

Figure Lengend Snippet: Autophagy depends on JNK1-induced Beclin-1 activation. A, Pancreatic cancer cell lines MIA PaCa-2 and SU.86.86 were treated with IR (1 μmol/L) for immunoblotting. B, Immunoblot analysis of late-stage autophagy in MIA PaCa-2. ULK1 inhibitor (ULK1i, XST-14, 5 μmol/L; ref. ) and JNK1 inhibitor (JNK1i, SP600125, 20 μmol/L; refs. , ) were administered individually or in combination 48 hours after IR treatment and were maintained for an additional 24 hours. Cell lysates were collected at the 72-hour time point for immunoblotting and ATP assay. Data are shown as the mean ± SD from n = 3 independent experiments. C, co-IP analysis of the Bcl-2 complex in MIA PaCa-2 and SU.86.86 cells treated with IR (1 μmol/L) for 0, 24, 48, and 72 hours. Cell lysates were immunoprecipitated with an anti–Bcl-2 antibody (IP: Bcl-2), and Bcl-2-bound Beclin-1 and Bax were detected by immunoblotting (IB: Beclin-1, Bax). Quantification represents Beclin-1 coimmunoprecipitated with Bcl-2, normalized to immunoprecipitated Bcl-2 (Beclin-1/Bcl-2), and expressed relative to 0 hours (set to 1). Whole-cell lysates (input) from the same samples were analyzed by immunoblotting to determine p-Bcl-2 levels and total Bcl-2, as indicated. Quantification represents p-Bcl-2/total Bcl-2, expressed relative to 0 hours (set to 1). Data are shown as the mean ± SD from n = 3 independent experiments. Statistical comparisons vs. 0 hours were performed using one-way ANOVA. co-IP, coimmunoprecipitation; Ct, control; IR, irinotecan. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001; ns, not significant.

Article Snippet: MIA PaCa-2 cells were treated with irinotecan (1 μmol/L) for 48 hours, followed by treatment with the ULK1 inhibitor XST-14 (5 μmol/L; MedChemExpress, Monmouth Junction; ref. ) or the JNK1 inhibitor SP600125 (20 μmol/L; MedChemExpress, Monmouth Junction; refs. , ) for an additional 24 hours.

Techniques: Activation Assay, Western Blot, ATP Assay, Co-Immunoprecipitation Assay, Immunoprecipitation, Control

Journal: Cancer Research

Article Title: Inhibiting Fatty Acid Oxidation Reverses Autophagy-Mediated Acquired Chemotherapy Resistance in Pancreatic Ductal Adenocarcinoma

doi: 10.1158/0008-5472.CAN-25-4517

Figure Lengend Snippet: The development model of acquired drug resistance by drug treatment. A, When cancer cells are treated with cytotoxic anticancer drugs, they experience cellular stress such as DNA damage, which activates AMPK-dependent early autophagy. At this stage, cells with severe damage undergo autophagic cell death, whereas surviving cells supply FAs via autophagy. B, When cancer cells face sustained stress from anticancer drugs, FAO is activated, leading to increased ATP production. This increase in ATP inactivates AMPK-dependent autophagy and activates mTOR. However, cancer cells also activate JNK1-dependent late autophagy in response to cellular damage, maintaining mTOR activity through FAO. This process results in tumor regrowth during treatment with anticancer drugs. A key feature of this stage is that autophagy and mTOR, which normally inhibit each other via feedback mechanisms, are concurrently activated through independent pathways due to damage caused by anticancer drugs. C, Both AMPK-dependent and JNK1-dependent autophagy promote mTOR activation through FAO induction. Therefore, anticancer drugs combined with FAO-inhibiting drugs induce cell death by inactivating mTOR survival signals and promoting autophagy flux but impairing autolysis due to FA deposits. In this study, we demonstrated this concept by double knockdown of CAC and ACAA1 , the main FAO pathways, or using KN510713 , FAO inhibitors. FA, fatty acid.

Article Snippet: MIA PaCa-2 cells were treated with irinotecan (1 μmol/L) for 48 hours, followed by treatment with the ULK1 inhibitor XST-14 (5 μmol/L; MedChemExpress, Monmouth Junction; ref. ) or the JNK1 inhibitor SP600125 (20 μmol/L; MedChemExpress, Monmouth Junction; refs. , ) for an additional 24 hours.

Techniques: Activity Assay, Activation Assay, Knockdown

Journal: iScience

Article Title: Induction of adipocyte thermogenic program by Rho-associated coiled-coil containing kinase 2

doi: 10.1016/j.isci.2026.115225

Figure Lengend Snippet: ROCK2-mediated thermogenic gene expression may occur through the JNK/FOXO1/UCP1 axis (A) Western blotting and quantification of FOXO1 in sWAT from male WT mice housed at room or cold temperature for 3 h ( n = 4 mice per group). (B) Ucp1 expression in sWAT from male WT mice housed at 4 °C for 0, 3, 6, or 12 h ( n = 4 mice). (C) Ucp1 expression in sWAT harvested from control mice and then treated with 40 μM isoproterenol (Iso) and/or 4 μM FOXO1 inhibitor (AS1842856) for 6 h ( n = 4 wells per group). (D) Foxo1 expression in sWAT from male control and AdR2KO mice housed at RT or cold for 3 h ( n = 4 mice). (E) Western blotting and quantification of nuclear FOXO1 (nFOXO1), cytoplasmic FOXO1 (cFOXO1), and total FOXO1 (tFOXO1) in sWAT from male control and AdR2KO mice housed at RT or cold for 3 h ( n = 3 mice). (F) Western blotting and quantification of phosphorylated JNK (pJNK) and total JNK (tJNK) in sWAT from male WT mice housed at RT or cold for 3 h ( n = 3 mice). (G) Foxo1 expression in sWAT harvested from control mice and then treated with 40 μM isoproterenol (Iso) and/or 100 μM JNK inhibitor (SP600125) for 6 h ( n = 6 wells per group). (H) Western blotting and quantification of pJNK and tJNK in sWAT from control and AdR2KO mice housed at RT or cold for 3 h ( n = 3 mice). (I) Foxo1 and (J) Ucp1 expression in differentiated primary adipocytes from sWAT of control and AdR2KO mice. After differentiation for 8 days, mature adipocytes were treated with or without 10 μM isoproterenol (Iso), 1 μM AS1842856, or 50 μM SP600125 for 6 h n = 5–6 wells in (I); n = 3–8 wells in (J). (K) The mRNA levels of Mapk8 (encodes JNK1), Foxo1 , Ppargc1a , and Ucp1 in mature mouse adipocyte cells, 3T3-L1. Adipocytes were transfected with siRNAs for 72 h and then treated with 10 μM isoproterenol (Iso) for 6 h ( n = 6 wells per group). (L) The mRNA levels of Foxo1 , Ppargc1a , and Ucp1 in differentiated mouse primary adipocytes from sWAT of control and AdR2KO mice. Adipocytes were transduced with low ( Foxo1L ) and high ( Foxo1H ) MOI of AAV-8 containing mouse Foxo1 plasmid for 7 days and then treated with 20 μM isoproterenol (Iso) for 16 h ( n = 8 wells per group). The immunoblot images are representative of at least three independent experiments. p values were determined by the two-tailed Student’s t test (A and F) or one-way ANOVA with Tukey’s multiple comparisons test (B-E and G-L). All data are presented as the mean ± SEM of at least three independent biological replicates. See also and . AS: AS1842856; SP: SP600125.

Article Snippet: JNK1 siRNA (m) , Santa Cruz Biotechnology , Cat# sc-29381.

Techniques: Gene Expression, Western Blot, Expressing, Control, Transfection, Transduction, Plasmid Preparation, Two Tailed Test