Journal: Journal of Diabetes Research

Article Title: CD36 Regulates PANoptosis in Diabetic Retinopathy via the NOTCH/MAML Pathway

doi: 10.1155/jdr/9324498

Figure Lengend Snippet: NOTCH pathway modulation affects HG‐induced PANoptosis. hRMECs were treated with a NOTCH agonist (Jagged1 peptide) or inhibitor (DAPT) under HG conditions. (A) Cell viability measured by CCK‐8 assay. (B) Caspase‐3/7 activity measured by colorimetric assay. (C) IL‐1 β and IL‐18 levels were quantified by ELISA. (D) Apoptosis rate measured by flow cytometry with quantification. (E) Quantification of Western blot analysis for PANoptosis‐related proteins, including cleaved Caspase‐3, GSDMD‐N, RIPK1, Bax, RIPK3, p‐MLKL, and Bcl‐2. (F) Co‐IP analysis of PANoptosome components (ASC, Caspase‐1, NLRP3, and RIPK3). (G) qPCR analysis of Bax and Bcl-2 mRNA levels. (H, I) IF staining of p‐MLKL (green) with DAPI (blue) and quantification. Scale bar = 150 μ m. Data are presented as mean ± SD ( n = 3). Statistical analysis was performed using one‐way ANOVA followed by Tukey′s post hoc test. ∗ p < 0.01, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ns = not significant.

Article Snippet: To modulate NOTCH signaling under HG conditions, cells were cultured in 25.0 mmol/L D‐glucose for 72 h. During the final 24 h, the HG group was supplemented with either 5 μ g/mL Jagged1 peptide (MedChemExpress, United States) to activate the pathway or 10 μ M DAPT (MedChemExpress, United States) to inhibit it.

Techniques: CCK-8 Assay, Activity Assay, Colorimetric Assay, Enzyme-linked Immunosorbent Assay, Flow Cytometry, Western Blot, Co-Immunoprecipitation Assay, Staining

Journal: International Journal of Molecular Medicine

Article Title: miR-223-3p promotes microglial lactylation and M1 polarization via the FBXW7/Notch1/Hes1/SIRT1 axis

doi: 10.3892/ijmm.2026.5849

Figure Lengend Snippet: miR-223-3p suppresses SIRT1 transcription via the Notch1/Hes1 signaling pathway, thereby promoting microglial lactylation and M1 polarization. (A) RT-qPCR analysis of Notch1 and Hes1 mRNA levels. (B) Western blot analysis of NICD1 and Hes1 protein expression. (C) Western blot analysis of SIRT1 expression following silencing Hes1. (D) Western blot detection of global lactylation levels. (E) RT-qPCR analysis of microglial polarization markers. (F and G) RT-qPCR and western blot detection of SIRT1 following treatment with Notch1 inhibitor DAPT (20 nM; 24 h). (H) Western blot detection of global lactylation levels. (I) RT-qPCR analysis of microglial polarization markers. (J) Western blot analysis of iNOS and Arg-1 protein expression. (K) Flow cytometric analysis of ROS levels. * P<0.05, ** P<0.01 and *** P<0.001. miR, microRNA; SIRT1, sirtuin 1; RT-qPCR, reverse transcription-quantitative PCR; iNOS, inducible nitric oxide synthase; ROS, reactive oxygen species; NC, negative control; ns, not significant.

Article Snippet: The SIRT1 inhibitor EX527 (10 μ M; 49843-98-3; TargetMol) and the SIRT1 activator SRT1720 (5 μ M; 1001645-58-4; TargetMol) were added separately and incubated for 24 h. The γ-secretase inhibitor DAPT (20 nM; cat. no. HY-13027; MedChemExpress) was administered in parallel experiments, while DMSO served as the vehicle control.

Techniques: Quantitative RT-PCR, Western Blot, Expressing, Reverse Transcription, Real-time Polymerase Chain Reaction, Negative Control