Journal: Autophagy

Article Title: Kit-mediated autophagy suppression driven by a viral oncoprotein emerges as a crucial survival mechanism in Merkel cell carcinoma

doi: 10.1080/15548627.2025.2477385

Figure Lengend Snippet: MCPyV truncated LT induces paranuclear retention and stabilization of KIT. (A) Immunofluorescence detection of KIT (green) and MCPyV LT (CM2B4, red) or sT (CM8E6, red) in KIT-HEK293 cells transfected with MCPyV T-antigens or vector control ( n = 6). (B) Top : illustration of the expression constructs of LT339 and the VPS39-interaction defective mutant LT339 W209A . Bottom : Representative images showing the effect of LT339 and LT339 W209A mutant on localization of KIT (green). LT was detected by CM2B4 (red). ( n = 3) (C) Immunoblots showing the effect of the LT339 and LT339 W209A on KIT expression. The quantification of KIT level is shown below the immunoblots ( n = 9). (D) Top : immunoblot analysis of the effect LT339 and LT339 W209A on KIT protein stability in the presence of cycloheximide (CHX) up to 4 h. Bottom : quantification of KIT protein stability after normalization to 0 h time point ( n = 4). (E) Immunoblots showing KIT protein stability in KIT-HEK293 cells transfected with LT339, LT339 W209A or plasmid control (CTR) treated with KIT ligand (KITLG, 100 ng/mL) or solvent control (PBS) in the presence of CHX (100 μg/mL). (F) Quantification of KIT protein stability after normalization to 0 h time point ( n = 5). Solid lines represent PBS control (KITLG − ) and dotted lines represent KITLG treatment (KITLG + ). (A and B) Nuclei were stained by DAPI (blue). Scale bar: 10 μm. Numbers below the images refer to the proportion of cells with KIT paranuclear dot-like staining to the total number of cells analyzed. (C, D and F) Error bars represent mean ± SEM. * p < 0.05, *** p < 0.001, ns = not significant were calculated by one-way ANOVA with post-hoc Tukey’s test (C), two-way ANOVA (D) or two-way ANOVA with post-hoc Bonferroni’s test (F).

Article Snippet: For KITLG induced KIT degradation experiments, KIT-293 cells were transfected with LT339, LT339 W209A or CTR for 48 h, followed by addition of CHX (100 μg/mL) and KITLG (100 ng/mL; Proteintech Group, HZ-1024) or PBS.

Techniques: Immunofluorescence, Transfection, Plasmid Preparation, Control, Expressing, Construct, Mutagenesis, Western Blot, Solvent, Staining

Journal: Cancer Immunology, Immunotherapy : CII

Article Title: Cancer-associated fibroblasts promote pro-tumor functions of neutrophils in pancreatic cancer via IL-8: potential suppression by pirfenidone

doi: 10.1007/s00262-025-03946-z

Figure Lengend Snippet: IL-8 secreted by activated neutrophils is suppressed by PFD. A IL-8 secretion by neutrophils measured by ELISA. IL-8 secretion was significantly increased in neutrophils stimulated with CAF-CM compared to non-stimulated neutrophils. PFD administration during neutrophil activation with CAF-CM significantly suppressed IL-8 secretion. B Effect of secretions from activated neutrophils on cancer cell proliferation. The presence of neutrophils significantly increased MIA PaCa-2 proliferation. Mean ± SD is shown. Each dot represents the number of cells per well. Student’s t-test was used. ***, P < 0.001. C Pancreatic cancer cells induce fibroblasts to become iCAFs, and iCAF-CM activates neutrophils, increasing their IL-8 secretion and enhancing the malignant traits of pancreatic cancer cells. Conversely, PFD acts on activated neutrophils, suppressing the malignant traits of pancreatic cancer cells. This figure was created with BioRender.com

Article Snippet: To evaluate IL-8 levels in neutrophil secretions, culture supernatants were assayed using a human IL-8 enzyme-linked immunosorbent assay (ELISA) kit (Proteintech, KE00006) according to the manufacturer’s instructions.

Techniques: Enzyme-linked Immunosorbent Assay, Activation Assay