Journal: Cell Death Discovery

Article Title: Inhibiting HSP27 activates the XBP1s/CerS1 interplay, which triggers DRP1-driven mitophagy, thereby protecting against cell death and promoting the KSHV lytic cycle in primary effusion lymphoma cells

doi: 10.1038/s41420-026-02979-2

Figure Lengend Snippet: 4μ8 C ( F ) or P053 ( G ) or Mdivi-1 (DRP1 inhibitor) ( H ). Mitophagy was evaluated following treatment with J2 with or without Mdivi-1 (DRP1inhibitor) ( A ). Cells treated with CCCP (Mitophagy inducer) were used as positive control. BC3 cells were stained with MitoTracker (a mitochondrial marker, green) and Lysotracker (a lysosomal marker, red). Yellow puncta resulting from the co-localization of MitoTracker (a mitochondrial marker green) and LysoTracker (a lysosomal marker red) were evaluated in control and J2-treated cells. The histograms indicate yellow puncta/cell. The data are represented as the mean plus S.D. of three different experiments. p value * <0.05, as calculated by ANOVA test. Scale bar = 10 µm. B Western blot analysis of p62 and LC3I/II in BC3 cells cultured with Mdivi-1 (DRP1 inhibitor) and subsequently treated with J2 or with J2 alone. The histograms represent the densitometric analysis of the ratio of p62/GAPDH and LC3II/GAPDH. The data are represented as the mean plus S.D. of three different experiments. p value * <0.05, as calculated by ANOVA test. C Expression of HADHA as evaluated by western blot analysis in PEL cell lines pretreated with Mdivi-1 (DRP1 inhibitor) before J2 treatment (20 μM). Histone H3 and GAPDH were used as loading controls. The histograms represent the densitometric analysis of the ratio of HADHA /H3 and HADHA /GAPDH. The data are represented as the mean plus S.D. of three different experiments. p value * <0.05, as calculated by ANOVA test. D BC3 and BCBL1 cell lines were knocked down for DRP1 (siDRP1) or scramble-treated for 24 h, exposed or not to J2 and finally evaluated for the expression of DRP1 and HADHA by western blot analysis. The histograms represent the densitometric analysis of the ratio of DRP1/GAPDH and HADHA/GAPDH. The data are represented as the mean plus S.D. of three different experiments. p value * <0.05, as calculated by ANOVA test. E Western blot analysis of HADHA in BC3 cells cultured with 4μ8 C (Ire1α endoribonuclease activity inhibitor) and subsequently treated with J2 or with J2 alone for 4 h and ( F ) in BC3 cells cultured with P053 (CerS1 inhibitor) and then treated with J2 or with J2 alone for 4 h. The histograms represent the densitometric analysis of the ratio HADHA/GAPDH. The data are represented as the mean plus S.D. of three different experiments. p value * <0.05, as calculated by ANOVA test.

Article Snippet: The following primary antibodies were used in western blots: rabbit polyclonal anti-PARP (1:1000) (Cell Signaling, Danvers, MA, USA, cat n. 9542), mouse monoclonal anti-CerS1 (LASS1) (1:500) (Santa Cruz Biotechnology Inc., Dallas, TX, USA, cat. n. 293497), rabbit polyclonal anti-BiP/GRIP78 (1:5000) (; Proteintech, Rosemont, IL, USA, cat n. 11587-1-AP), rabbit polyclonal anti-CHOP (GADD153) (1:1000) (Proteintech, Rosemont, IL, USA, cat n. 15204-1-AP), rabbit polyclonal anti-XBP1 (1:500) (Novus Biologicals, Centennial, CO, USA; cat n. NBP1-77681SS), mouse monoclonal anti-DRP1 (1:500) (Santa Cruz Biotechnology Inc., Dallas, TX, USA, cat. n. sc-271583), mouse monoclonal anti-HADHA (1:500) (Santa Cruz Biotechnology Inc., Dallas, TX, USA, cat. n. sc-374497), mouse monoclonal anti-Parkin (1:500) (Santa Cruz Biotechnology Inc., Dallas, TX, USA, cat. n. sc-32283), rabbit polyclonal anti-LC3 I/II (1:1000) (Novus Biologicals, Centennial, CO, USA cat. n. NB100–2220), rabbit polyclonal anti-SQSTM1/p62 (1:500) (Novus Biologicals, Centennial, CO, USA cat. n. NBP-48-320), mouse monoclonal anti-HSP27 (1:1000) (Proteintech, Rosemont, IL, USA, cat n. 182841), rabbit polyclonal anti-HSP110 (1:200) (Abcam, Cambridge UK, cat. n. ab24503).

Techniques: Positive Control, Staining, Marker, Control, Western Blot, Cell Culture, Expressing, Activity Assay

Journal: Journal of Neuroinflammation

Article Title: Selective vulnerability of the aging cholinergic system to amyloid pathology revealed by induced APP overexpression

doi: 10.1186/s12974-025-03682-2

Figure Lengend Snippet: Autophagic activity in the mouse brain was modulated by both aging and mutant APP transgene expression. a Representative western blot images are shown. Autophagy-related targets assessed include ( b ) LC3A-II/LC3A-I ratio, ( c ) total LC3A, ( d ) LC3B-II/LC3B-I ratio, ( e ) total LC3B, ( f ) ATG3, ( g ) ATG5, ( h ) ATG7, ( i ) ATG12, ( j ) Beclin-1, and ( k ) SQSTM1/p62. Sample size was 5 male and 5 female 18mo controls, 4–5 male and 6 female APP 6 → 18mo, 4–5 male and 5 female 24mo controls, and 5 male and 5–6 female APP 12 → 24mo. Individual data points and group means ± S.E.M. are presented. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001

Article Snippet: The following primary antibodies were used: mouse monoclonal anti-human Aβ (#SIG-39300, clone 6E10, 1:2,000; Covance, Princeton, NJ, USA), rabbit monoclonal anti-BACE1 (#5606, clone D10E5, 1:2,000; Cell Signaling Technology, Danvers, MA, USA), rabbit monoclonal anti-presenilin 1 (#5643, clone D39D1, 1:3,000; Cell Signaling Technology), rabbit monoclonal anti-LC3A (#4599, clone D50G8, 1:2,000; Cell Signaling Technology), rabbit polyclonal anti-LC3B (#2775, 1:2,000; Cell Signaling Technology), rabbit polyclonal anti-ATG3 (#3415, 1:2,000; Cell Signaling Technology), rabbit monoclonal anti-ATG5 (#8540, clone D1G9, 1:2,000; Cell Signaling Technology), rabbit monoclonal anti-ATG7 (#8558, clone D12B11, 1:2,000; Cell Signaling Technology), rabbit monoclonal anti-ATG12 (#4180, clone D88H11, 1:2,000; Cell Signaling Technology), rabbit monoclonal anti-beclin 1 (#3495, clone D40C5, 1:2,000; Cell Signaling Technology), rabbit polyclonal anti-p62 (#5114, 1:1,500; Cell Signaling Technology), rabbit monoclonal anti-ChAT (#ab181023, clone EPR13024 (B), 1:2,000; Abcam, Cambridge, UK), rabbit polyclonal anti-SLC5A7 (#ab135043, 1:3,000; Abcam), rabbit polyclonal anti-PSD95 (#2507, 1:2,000; Cell Signaling Technology), mouse monoclonal anti-synaptophysin (#ab8049, clone SY38, 1:2,000; Abcam), rabbit anti GFAP (#12389, clone D1F4Q, 1:2,000; Cell Signaling Technology), rabbit anti CD11b (#17800, clone E6E1M, 1:2,000; Cell Signaling Technology), rabbit anti CD68 (#97778, clone E3O7V, 1:2,000; Cell Signaling Technology), rabbit anti Iba1/AIF-1 (#17198, clone E4O4W, 1:2,000; Cell Signaling Technology), rabbit anti TREM2 (#59621, clone E9O9F, 1:2,000; Cell Signaling Technology), mouse monoclonal anti-actin (#869100, clone C4, 1:20,000; MP Biomedicals, Santa Ana, CA, USA).

Techniques: Activity Assay, Mutagenesis, Expressing, Western Blot