Journal: Scientific Reports

Article Title: IL-15-Activated CD38 + HLA-DR + CD8 + T cells induce liver injury in cirrhosis via JAK/STAT5 and PI3K/mTOR pathways

doi: 10.1038/s41598-025-02693-6

Figure Lengend Snippet: The signal pathways involved in IL-15-induced innate cytotoxicity of CD38 + HLA-DR + CD8 + T cells. (A) 1 × 10 6 CD8 + T cells from healthy donors were stimulated with IL-15 (20 ng/mL) for 48 h and 72 h, after which phosphorylation of signaling proteins was assessed by flow cytometry for STAT5 ( n = 10), ERK1/2 ( n = 7) and mTOR ( n = 8). Representative dot plots and the summary data show the expression of signaling proteins in CD38 + HLA-DR + CD8 + T cells. (B) The percentage of CD38 + HLA-DR + CD8 + T cells was analyzed after inhibitors treatment ( n = 4). Representative dot plots from a single donor (left) and summary data (right) are presented. (C-G) The percentage NKG2D, FasL, perforin, and Granzyme B in CD38 + HLA-DR + CD8 + T cells were analyzed after inhibitors treatment ( n = 4). (H) CD8 + T cells from healthy donors were pre-treated with STAT5 inhibitor pimozide, MEK inhibitor PD98059, PI3K inhibitor LY294002 or mTOR inhibitor PP242, followed by IL-15 (20 ng/mL) stimulation for the next 72 h. Then, CD8 + T cells were co-cultured with CFSE-labeled K562 cells at a 10:1 E: T ratio and cytotoxicity against K562 cells evaluated ( n = 4). Representative dot plots and the summary data show expression of PI in the gate of CFSE + . (I) Schematic representation of cytokine-mediated crosstalk between T cells and liver cells in liver cirrhosis. The Wilcoxon matched-pairs signed rank test (A) was used for comparisons among groups. The one-way ANOVA (B , C , D-H) was used for comparisons between groups. Control group was indicated treatment only IL-15. ns, not significant, * p < 0.05, ** p < 0.01.

Article Snippet: CD8 + T cells (1 × 10 6 ) from healthy donors were pre-treated with inhibitors including STAT5 inhibitor pimozide, MEK inhibitor PD98059, PI3K inhibitor LY294002 or mTOR inhibitor PP242 (MedChemExpress, NJ, USA) and stimulated with IL-15 (20 ng/mL) (Thermo, MA, USA) for 72 h. CD8 + T cells were subsequently co-cultured with CFSE-labeled K562 cells at a 10:1 effector-to-target (E: T) ratio for 6 h, and cytotoxicity against K562 cells was assessed.

Techniques: Phospho-proteomics, Flow Cytometry, Expressing, Cell Culture, Labeling, Control

Journal: The Journal of Biological Chemistry

Article Title: CK-666 protects against ferroptosis and renal ischemia-reperfusion injury through a microfilament-independent mechanism

doi: 10.1016/j.jbc.2024.107942

Figure Lengend Snippet: CK-666 attenuates ferroptosis independent of the Arp2/3 complex. A , the molecular structures of CK-666, CK-636, CK-689, CK-869, Benproperine, and Pimozide. B , cell viability of HT1080 cells treated with or without CK-666 (100 μM), CK-636 (100 μM), CK-689 (100 μM), or CK-869 (100 μM) in the presence of different concentrations of RSL3 for 24 h. C , cell viability of HT1080 cells treated with or without Benproperine (10 μM) and Pimozide (10 μM) in the presence of different concentrations of RSL3 for 24 h. D , cell viability of HT1080 cells treated with or without CK-666 (100 μM), CK-636 (100 μM), or CK-689 (100 μM) in the presence of different concentrations of erastin for 24 h. E , cell viability of 293T cells treated with or without CK-666 (100 μM), CK-636 (100 μM), or CK-689 (100 μM) in the presence of different concentrations of RSL3 for 24 h. F , cell viability of Pfa1 cells treated with or without Fer-1 (2 μM), CK-666 (100 μM), CK-636 (100 μM), or CK-689 (100 μM) in the presence of 4-hydroxytamoxifen (4-OHT, 100 μM) for 72 h. G , cell viability of HT1080 cells transfected with siNC, siARP2, or siARPC2 in the presence of different concentrations of RSL3 for 24 h. H , cell viability of HT1080 cells transfected with or without siNC, siARP2, or siARPC2 in the presence of different concentrations of erastin for 24 h. I , cell viability of HT1080 cells treated with or without CK-666 (100 μM) and transfected with siNC or siARP2 in the presence of different concentrations of RSL3 for 24 h. The values were presented as mean ± SD. n = 3 biologically independent replicates.

Article Snippet: The compounds used to treat cells are as follow: RSL3, erastin, CK636, Ferrostatin-1, Liproxstatin-1, Necrostatin-1, and Z-VAD-FMK were purchased from Selleck; Wiskostatin, ML385, iFSP1, Benproperine phosphate, and Pimozide were purchased from Targetmol; and Deferoxamine and viFSP1 (MCE), CK-666 (Abcam), CK-869 (Macklin), CK-689 (Merck), Jasplakinolide (Invitrogen), Cytochalasin D (Aladdin), Latrunculin A (APE x BIO),Latrunculin B (Cayman), SMIFH2 (Sigma), and 4-Hydroxytamoxifen (Sigma).

Techniques: Transfection