Journal: Redox Biology

Article Title: Folic acid modulates VPO1 DNA methylation levels and alleviates oxidative stress-induced apoptosis in vivo and in vitro

doi: 10.1016/j.redox.2018.08.005

Figure Lengend Snippet: The effect of folic acid on reactive oxygen species (ROS) and 8-Hydroxydeoxyguanosine (8-OHdG) level in HUVECs. HUVCE were treated with 0–1000 nmol/L folic acid for 48 h and exposed to 120 μg/mL ox-LDL for the first 24 h. (A) The intracellular ROS was measured by flow cytometry using DCFH-DA staining. The peaks in each panel represent the mean fluorescence intensities. (B) Representative images of unirradiated and irradiated cells obtained by the ImageStream. (C) ROS levels are reported as DCF Florescence, which means percentage of control group cells. (D) Intracellular 8-OHdG level was assayed by ELISA. The plotted values are mean ± SE values for 3 independent experiments. a, p < 0.05 compared with the ox-LDL + FA20. b, p < 0.05 compared with the ox-LDL + FA0.

Article Snippet: The plasma ox-LDL concentration was detected using the mouse ox-LDL ELISA Kit (CUSABIO TECHNOLOGY, Wuhan, China) according to the manufacturer's instructions.

Techniques: Flow Cytometry, Staining, Fluorescence, Irradiation, Control, Enzyme-linked Immunosorbent Assay

Journal: Signal transduction and targeted therapy

Article Title: NPRC deletion mitigated atherosclerosis by inhibiting oxidative stress, inflammation and apoptosis in ApoE knockout mice.

doi: 10.1038/s41392-023-01560-y

Figure Lengend Snippet: Fig. 8 Overexpression of NPRC increased inflammation and apoptosis and aggravated migration and phagocytosis of macrophages. a Representative Western blot images of ICAM1, VCAM1, TNFα, MCP1, and IL-6 expression in oxLDL-stimulated oe-NC and oe-NPRC HAECs. b Quantification of ICAM1, VCAM1, TNFα, MCP1, and IL-6 expression in oxLDL-stimulated oe-NC and oe-NPRC HAECs (n = 5 per group). c Representative images of Raw 264.7 migration in oxLDL-stimulated oe-NC and oe-NPRC HAECs (scale bar = 100 μm). d Quantification of crystal violet positive cells in Raw 264.7 stimulated by medium from si-NC and si-NPRC HAECs (n = 5 per group). e Representative images of phagocytosis of Raw 264.7 from si-NC and oe-NPRC HAECs after oxLDL stimulation (scale bar = 100 μm). f Quantification of relative positive staining area of Oil Red O in (e) (n = 5 per group). g Representative images of TUNEL assay in oxLDL-stimulated oe-NC and oe-NPRC HAECs (scale bar = 100 μm). h Quantification of TUNEL-positive cells in oxLDL-stimulated oe-NC and si-NPRC HAECs (n = 5 per group). i Representative Western blot images of p-AKT1, AKT1, Cleaved-caspase 3, Caspase 3, Cleaved-caspase 7, and Caspase 7 expression in oxLDL-stimulated oe-NC and oe-NPRC HAECs. j Quantification of p-AKT1/AKT1, Cleaved-caspase 3/Caspase 3 and Cleaved-caspase 7/Caspase 7 expression in oxLDL-stimulated oe-NC and oe-NPRC HAECs (n = 5 per group). Normal distributions were tested by Shapiro–Wilk method. Unpaired two-tailed Student’s t tests were applied in (d), (f), (h). Two-way ANOVA was used in (b) and (j)

Article Snippet: Serum lipids levels including total cholesterol (TC) (S03042, Leidu, China), low-density lipoprotein cholesterol (LDL-C) (S03029, Leidu, China), high-density lipoprotein cholesterol (HDL-C) (S03025, Leidu, China), very low-density lipoprotein cholesterol (VLDL-C) (CSBE17089m, Cusabio, China), triglycerides (TG) (S03027, Leidu, China) and oxidized LDL (oxLDL) (CSB-E07933m, Cusabio, China) were measured by enzymatic assay in all mice.

Techniques: Over Expression, Migration, Western Blot, Expressing, Staining, TUNEL Assay, Two Tailed Test

Journal: Signal transduction and targeted therapy

Article Title: NPRC deletion mitigated atherosclerosis by inhibiting oxidative stress, inflammation and apoptosis in ApoE knockout mice.

doi: 10.1038/s41392-023-01560-y

Figure Lengend Snippet: Fig. 9 Activation of PKA pathway upregulated eNOS and p-AKT1, and inhibited p-p65 expression. a Representative Western blot images of p-Iκκα/β, Iκκα, Iκκβ, p-P65, and P65 expression in oxLDL-stimulated oe-NC and oe-NPRC HAECs. b Quantification of p-Iκκα/β, Iκκα, Iκκβ, p-P65, and P65 expression in oxLDL-stimulated oe-NC and oe-NPRC HAECs (n = 5 per group). c Representative immunofluorescence images of p-P65 in oe-NC and oe-NPRC HAECs stimulated by oxLDL (scale bar = 50 μm). d Quantification of p-P65 immunofluorescent intensity in nuclei of oe- NC and oe-NPRC HAECs stimulated by oxLDL (n = 5 per group). e Representative Western blot images of p-PKA-substrate expression in HAECs treated by forskolin and H89. f Quantification of p-PKA-substrate expression in HAECs treated by forskolin and H89 (n = 5 per group). g Representative Western blot images of eNOS expression in HAECs treated by forskolin and H89. h Quantification of eNOS expression in HAECs treated by forskolin and H89 (n = 5 per group). i Representative Western blot images of p-P65, P65, p-AKT1, and AKT1 expression in HAECs treated by forskolin and H89. j Quantification of p-P65/P65 expression in HAECs treated by forskolin and H89 (n = 5 per group). k Representative images of TUNEL assay in HAECs pretreated by forskolin and H89, and then stimulated by oxLDL (scale bar = 100 μm). l Quantification of TUNEL-positive cells in HAECs pretreated by forskolin and H89, and then stimulated by oxLDL (n = 5 per group). Normal distributions were tested by Shapiro–Wilk method. Two-way ANOVA was applied in (b). Unpaired two-tailed Student’s t tests were applied in (d). One-way ANOVAs were used in (f), (h), (j), and (l)

Article Snippet: Serum lipids levels including total cholesterol (TC) (S03042, Leidu, China), low-density lipoprotein cholesterol (LDL-C) (S03029, Leidu, China), high-density lipoprotein cholesterol (HDL-C) (S03025, Leidu, China), very low-density lipoprotein cholesterol (VLDL-C) (CSBE17089m, Cusabio, China), triglycerides (TG) (S03027, Leidu, China) and oxidized LDL (oxLDL) (CSB-E07933m, Cusabio, China) were measured by enzymatic assay in all mice.

Techniques: Activation Assay, Expressing, Western Blot, TUNEL Assay, Two Tailed Test

Journal: Signal transduction and targeted therapy

Article Title: NPRC deletion mitigated atherosclerosis by inhibiting oxidative stress, inflammation and apoptosis in ApoE knockout mice.

doi: 10.1038/s41392-023-01560-y

Figure Lengend Snippet: Fig. 10 Schematic diagram showing the mechanism of NPRC-promoted atherosclerosis. Under oxLDL stimulation, NPRC inhibits cAMP/PKA signaling pathway in HAECs, which reduces eNOS expression and promotes oxidative stress, inflammatory cytokine release, and cell apoptosis, leading to foam cell formation and atherosclerosis. Diagram is generated from BioRender

Article Snippet: Serum lipids levels including total cholesterol (TC) (S03042, Leidu, China), low-density lipoprotein cholesterol (LDL-C) (S03029, Leidu, China), high-density lipoprotein cholesterol (HDL-C) (S03025, Leidu, China), very low-density lipoprotein cholesterol (VLDL-C) (CSBE17089m, Cusabio, China), triglycerides (TG) (S03027, Leidu, China) and oxidized LDL (oxLDL) (CSB-E07933m, Cusabio, China) were measured by enzymatic assay in all mice.

Techniques: Expressing, Generated

Journal: Cell Reports Medicine

Article Title: TRPM2 overactivation drives hyperlipidemia-induced dysfunction of myeloid cells and neurovascular units

doi: 10.1016/j.xcrm.2025.101998

Figure Lengend Snippet: Global TRPM2 knockout abolishes the exacerbation of ischemic brain injury by hyperlipidemia (A) The presence of atherosclerotic plaque (indicated by black arrows) at the bifurcation of common carotid artery (CCA) and in the internal carotid artery (ICA) (scale bar size: 2 mm). (B–E) Triphenyl tetrazolium chloride (TTC) staining 24 h after MCAO (60-min) and neurological deficit score in WT, Apoe −/− mice, and Apoe −/− gM2KO mice (B and C) and in Trpm2 fl/fl mice, Apoe −/− Trpm2 fl/fl cre − mice, and Apoe −/− Trpm2 fl/fl cre + mice (D and E) fed with or without HFD ( n = 12–20 mice per group) (scale bar size: 5 mm). (F and G) TRPM2 expression in brains from WT mice fed with or without HFD ( n = 5 and 5 mice). (H and I) TRPM2 and CD36 expression in primary neurons, CECs, BMDMs, and peripheral leukocytes isolated from WT mice ( n = 5, 5, 5, and 5 dishes of cells isolated from at least 5 mice). (J and K) TRPM2 expression in peripheral leukocytes isolated from WT mice fed with HFD for 0, 1, 2, 3, and 4 months ( n = 5, 5, 5, 5, and 5 dishes of cells isolated from at least 5 mice). (L) ELISA measurement of TRPM2 levels in lysates from B cells, T cells, monocytes, and neutrophils after cell sorting of peripheral leukocytes from WT mice with or without HFD treatment. (M and N) ELISA measurement of plasma oxLDL and IL-1β levels in WT mice fed with HFD for 0, 1, 2, 3, and 4 months ( n = 5, 5, 5, 5, and 5 plasma samples from different mice). (O and P) Correlation of plasma IL-1β level with plasma oxLDL level (O) and TRPM2 expression (P) in peripheral leukocytes ( n = 5 mice). (Q) Correlation of plasma oxLDL level with TRPM2 expression in peripheral leukocytes ( n = 5 mice). Error bars: mean ± SEM; ns, no statistical significance, ∗, p < 0.05, ∗∗, p < 0.01, ∗∗∗, p < 0.001. See also Figure S2 and Table S1 .

Article Snippet: Experiments were performed following the manufacturer’s instructions for the oxLDL (Mouse Oxidized Low-Density Lipoprotein ELISA Kit, Abclonal, RK03096) and IL-1β (LEGEND MAX Human IL-1β ELISA Kit, BioLegend, 437007) kits.

Techniques: Knock-Out, Staining, Expressing, Isolation, Enzyme-linked Immunosorbent Assay, FACS, Clinical Proteomics

Journal: Cell Reports Medicine

Article Title: TRPM2 overactivation drives hyperlipidemia-induced dysfunction of myeloid cells and neurovascular units

doi: 10.1016/j.xcrm.2025.101998

Figure Lengend Snippet: Increase of TRPM2 expression by oxLDL compromises resistance of neurons to ischemia (A) ELISA measurement of oxLDL amount in the brain tissue of the contralateral (left) and ipsilateral (right) hemisphere 24 h after MCAO. (B–E) TRPM2, pp65, and caspase-3 expression in primary cortical neurons isolated from WT mouse treated with oxLDL (B and C) and LDL (D and E) for 0, 6, 12, 18, 24, and 48 h ( n = 5 dishes of cells isolated from at least 5 mice). (F and G) TRPM2 expression in WT mouse primary cortical neurons treated with oxLDL for 48 h with the co-treatment of DMSO, SSO, and scavengers Mn (III) TBAP and L-NMA ( n = 5 dishes of cells isolated from at least 5 mice). (H and I) TRPM2, pp65, and caspase-3 expression in WT and M2KO mouse primary cortical neurons treated with oxLDL for 48 h with the co-treatment of DMSO, MK801/AP5, SN50, ACA, and TAT-M2 ( n = 5 dishes of cells isolated from at least 5 mice). (J and K) Graphic illustration of the administration strategy, as well as quantification and representative images of TTC staining 7 days after MCAO, neurological deficit score, and rotarod performance in Apoe −/− mice fed with or without HFD. TAT-SC and TAT-M2 were injected intraperitoneally (10 nmol/kg) right after reopening of MCA ( n = 7, 7, and 7 mice) (scale bar size: 5 mm). (L) Quantification of body weight of the mice used. Error bars: mean ± SEM; ns, no statistical significance, ∗, p < 0.05, ∗∗, p < 0.01, ∗∗∗, p < 0.001.

Article Snippet: Experiments were performed following the manufacturer’s instructions for the oxLDL (Mouse Oxidized Low-Density Lipoprotein ELISA Kit, Abclonal, RK03096) and IL-1β (LEGEND MAX Human IL-1β ELISA Kit, BioLegend, 437007) kits.

Techniques: Expressing, Enzyme-linked Immunosorbent Assay, Isolation, Staining, Injection

Journal: Cell Reports Medicine

Article Title: TRPM2 overactivation drives hyperlipidemia-induced dysfunction of myeloid cells and neurovascular units

doi: 10.1016/j.xcrm.2025.101998

Figure Lengend Snippet:

Article Snippet: Experiments were performed following the manufacturer’s instructions for the oxLDL (Mouse Oxidized Low-Density Lipoprotein ELISA Kit, Abclonal, RK03096) and IL-1β (LEGEND MAX Human IL-1β ELISA Kit, BioLegend, 437007) kits.

Techniques: Recombinant, Clinical Proteomics, Sequencing, Enzyme-linked Immunosorbent Assay, Bicinchoninic Acid Protein Assay, Knock-Out, Generated, Software, Microscopy, Imaging

Journal: Molecular medicine reports

Article Title: Erythropoietin ameliorates renal interstitial fibrosis via the inhibition of fibrocyte accumulation.

doi: 10.3892/mmr.2015.3157

Figure Lengend Snippet: Figure 5. CXCL12, CCL21 and CXCL16 expression analysis. (A) Representative western blots for CXCL12, CCL21 and CXCL16. Semi‑quantitative analysis of protein expression levels of (B) CXCL12, (C) CCL21 and (D) CXCL16. Actin was used as an internal control. *P<0.05 vs. Sh; †P<0.05 vs. U+V and ‡P>0.05 vs. U+V. UUO, unilateral ureteral obstruction; Sh, control; U+V, UUO+vehicle; U+E1, UUO treated with 300 U/kg rhEPO; rhEPO, recombinant human erythropoietin; U+E2, UUO treated with 1,000 U/kg rhEPO; CXCL16, CXC chemokine ligand 16; CCL21, CC chemokine ligand 21.

Article Snippet: Proteins were electrophoretically transferred to nitrocellulose membranes (Millipore, Billerica, MA, USA) which were subsequently incubated with antibodies specific for α‐SMA (1:500), collagen I (1:1,000), fibronectin (1:400), CXCL12 (1:1,000) , CCL21 (1:600), CXCL16 (1:500) and β-actin (1:1,000), followed by incubation with secondary antibody conjugated with IRDye® infrared dye (Rockland Immunochemicals, Inc.).

Techniques: Expressing, Western Blot, Control, Recombinant