Journal: Advanced Science

Article Title: Phosphorylated STYK1 Promotes Gemcitabine Resistance via Autophagy‐Mediated Cancer Stemness in Pancreatic Cancer

doi: 10.1002/advs.202521488

Figure Lengend Snippet: STYK1 promotes gemcitabine resistance in pancreatic cancer cells. (A, B) PANC‐1 cells were treated with different concentrations of gemcitabine for the indicated time periods, and STYK1 mRNA expression was assessed by RT‐qPCR (n = 6). **** p < 0.0001 according to Student's t ‐test. (C) Western blotting analysis of STYK1 expression levels in human pancreatic cancer cell lines PANC‐1 and AsPC‐1, as well as in the gemcitabine‐resistant cell lines PANC‐1/GEM and AsPC‐1/GEM. D) RT‐PCR analysis of STYK1 mRNA levels in PANC‐1, AsPC‐1, PANC‐1/GEM and AsPC‐1/GEM cells. **** p < 0.0001 according to Student's t ‐test. (E) Representative images of IHC staining for STYK1 in gemcitabine‐sensitive and resistant pancreatic cancer PDX tumors tissues. (F–I) Cell viability of PANC‐1/GEM and AsPC‐1/GEM cells with stable STYK1 knockdown, as well as PANC‐1 and AsPC‐1 cells stably overexpressing STYK1, following treatment with different concentrations of gemcitabine for 72 h, as determined by CCK‐8 assay. (J) PANC‐1/GEM cells with stable STYK1 knockdown were treated with gemcitabine (5 µM) for different durations as indicated, and OD values were assessed using the CCK‐8 assay. **** p < 0.0001 according to Student's t ‐test. (K–M) Apoptosis of PANC‐1/GEM and AsPC‐1/GEM cells with stable STYK1 knockdown following gemcitabine treatment (5 µM for PANC‐1/GEM and 40 µM for AsPC‐1/GEM, 24 h), assessed using an Annexin V‐AbFluor 488/PI double staining Apoptosis Detection Kit. Annexin V‐positive cells indicate early apoptosis, whereas PI‐positive cells indicate late apoptosis/necrosis (n = 3). **** p < 0.0001 according to Student's t ‐test. (N) Representative images of resected tumors from nude mice in different groups (n = 5). O) Tumor weights of resected tumors in different groups of mice (n = 5). **** p < 0.0001 according to Student's t ‐test. ** p < 0.01; **** p < 0.0001 according to Student's t ‐test. (P) Tumor volume change curves in different groups of mice (n = 5); Tumor volume was measured every 5 days after cell injection and during gemcitabine treatment (25 mg/kg). **** p < 0.0001 according to Student's t ‐test. (Q) Representative H&E staining and Ki67 immunohistochemical staining images of resected tumor tissues (n = 5). Scale bar: 50 µm. Data were expressed as mean ± SD, * p < 0.05; ** p < 0.01; *** p < 0.001; **** p < 0.0001 according to Student's t ‐test.

Article Snippet: Apoptosis was assessed using an Annexin V‐FITC Apoptosis Detection Kit (Elabscience, E‐CK‐A252).

Techniques: Expressing, Quantitative RT-PCR, Western Blot, Analysis, Reverse Transcription Polymerase Chain Reaction, Immunohistochemistry, Knockdown, Stable Transfection, CCK-8 Assay, Double Staining, Injection, Staining, Immunohistochemical staining

Journal: Redox Report : Communications in Free Radical Research

Article Title: Coordinated mitochondrial quality control underlies oxidative stress resilience in MUSE stem cells

doi: 10.1080/13510002.2026.2698189

Figure Lengend Snippet: MUSE cells activate PINK1-dependent mitophagy to clear damaged mitochondria. A) Representative immunofluorescence images showing the co-localization between lysosomes (LAMP1, green) and mitochondria (MitoTracker, red), with nuclei counterstained with DAPI (blue), in basal conditions and 48 h after 300 μM H₂O₂ exposure (20X magnification). Scale bar 50 µm. The violin plot represents quantification of colocalization using Manders’ coefficient (fraction of MitoTracker signal overlapping with LAMP1), analyzed with the Coloc2 plugin in ImageJ. Positive control was obtained with CCCP, and negative control with bafilomycin A1. Quantification was performed on at least 30 randomly selected fields per condition (B) Representative flow-cytometry dot plots from the Mitophagy Detection Kit and corresponding quantification histogram showing the percentage of cells positive for dual mitochondrial and lysosomal signals. As controls, CCCP was used as a positive inducer of mitophagy, whereas the negative control corresponds to unstained cells. For each condition, at least 10,000 live-cell events were acquired. (C) Western blot analysis of phosphorylated ubiquitin at Ser65 (Ub-S65) and PINK1 in the three populations ± H₂O₂. Densitometric quantifications (adjacent histograms) were normalized to ACTB, which served as the loading control. All data are presented as mean ± SD (n = 3 from independent experiments). ** p < 0.01, *** p < 0.001 vs. respective untreated controls; ## p < 0.01, ### p < 0.001 vs. MSCs + H₂O₂.

Article Snippet: Mitophagy was assessed using the Mitophagy Detection Kit (Dojindo, MD01-10), which measures the colocalization of mitochondria and lysosomes via pH-dependent fluorescence shift.

Techniques: Immunofluorescence, Positive Control, Negative Control, Flow Cytometry, Western Blot, Analysis, Ubiquitin Proteomics, Control