Journal: iScience

Article Title: 3-Hydroxypropionic acid converts inflammatory macrophage glycolysis into mitochondrial oxidation through GAPDH carboxyethylation

doi: 10.1016/j.isci.2026.116258

Figure Lengend Snippet: 3-HPA inhibits the secretion of inflammatory factors and glycolysis in macrophages (A and B) Schematic diagram of THP-1 cell (A) and BMDMs (B) activation into pro-inflammatory macrophages. Created with BioRender.com. (C) The concentrations of IL-6, TNF-α, and IL-1β in THP-1 cell supernatants were quantified by ELISA after 24 h treatment with LPS (100 ng/mL), or LPS+3-HPA (5 mM). Data are the means ± SD and n = 3 per group. Statistical significance was determined using one-way ANOVA followed by Tukey’s multiple comparisons test with ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, and ∗∗∗∗p < 0.0001. (D) The concentrations of IL-6, TNF-α, and IL-1β in supernatants of THP-1 cells treated with different concentrations of 3-HPA (0, 0.625, 1.25, 5 mM) followed by LPS stimulation. Data are the means ± SD and n = 3 per group. Statistical significance was determined using one-way ANOVA followed by Dunnett’s multiple comparisons test ∗p < 0.05, ∗∗p < 0.01, and ∗∗∗p < 0.001. (E) The concentrations of IL-6, TNF-α, and IL-1β in BMDM cell supernatants were quantified by ELISA after 24 h treatment with LPS (100 ng/mL) or LPS+3-HPA (5 mM). Data are the means ± SD and n = 3 per group. Statistical significance was determined using one-way ANOVA followed by Tukey’s multiple comparisons test with ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, and ∗∗∗∗p < 0.0001. (F) Bubble plot of KEGG pathway enrichment analysis for differentially expressed genes between LPS (100 ng/mL) and LPS+3-HPA (5 mM) treated in THP-1 cells. The size of each bubble represents the number of differentially expressed genes, and the color indicates the enrichment factor. (G and H) Pyruvate and lactate levels in THP-1 cells (G) and BMDMs (H) treated with LPS (100 ng/mL), or LPS+3-HPA (5 mM). (I) Immunoblots of protein expression levels of HK, GAPDH, PKM, and LDHA in THP-1 cells treated with LPS (100 ng/mL), or LPS+3-HPA (5 mM), and quantitative results of GAPDH. (J) The mRNA levels of GAPDH in THP-1 cells treated with LPS (100 ng/mL) or LPS+3-HPA (5 mM). (K) GAPDH activity assay in THP-1 cells and BMDMs treated with PBS, LPS (100 ng/mL), or LPS+3-HPA (5 mM). Data in (G–K) are the means ± SD and n = 3 per group. Statistical significance was determined using one-way ANOVA followed by Tukey’s multiple comparisons test with ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, and ∗∗∗∗p < 0.0001; ns, not significant.

Article Snippet: Mouse IL-1β Precoated ELISA Kit , Dakewe , 1210122.

Techniques: Activation Assay, Enzyme-linked Immunosorbent Assay, Western Blot, Expressing, Activity Assay

Journal: iScience

Article Title: 3-Hydroxypropionic acid converts inflammatory macrophage glycolysis into mitochondrial oxidation through GAPDH carboxyethylation

doi: 10.1016/j.isci.2026.116258

Figure Lengend Snippet: 3-HPA enhances the metabolite content of the TCA cycle and mitochondrial oxidation (A) Schematic diagram of THP-1 with 2-DG treatment. Created with BioRender.com. (B) Concentrations of IL-6, TNF-α, and IL-1β in supernatants of THP-1 cells treated with LPS, LPS+3-HPA, LPS+2-DG, or LPS+3-HPA+2-DG. Data are the means ± SD and n = 3 per group. Statistical significance was determined using one-way ANOVA followed by Tukey’s multiple comparisons test with ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, and ∗∗∗∗p < 0.0001; ns, not significant. (C) Schematic diagram of THP-1 with high glucose treatment. Created with BioRender.com. (D) Concentrations of IL-6, TNF-α, and IL-1β in supernatants of THP-1 cells treated with LPS+3-HPA, LPS+3-HPA+glucose. Data are the means ± SD and n = 3 per group. Statistical significance was determined using unpaired Student’s t test with ∗∗∗p < 0.001; ns, not significant. (E) KEGG pathway enrichment analysis of metabolic pathways in BMDM cells treated with LPS or LPS+3-HPA. (F) Relative abundance of metabolite (ornithine, citrulline, L-malate, succinic acid, trans-aconitic acid, cis-aconitic acid) in BMDM cells treated with LPS or LPS+3-HPA. Data are the means ± SD and n = 4 per group. Statistical significance was determined using unpaired Student’s t test with ∗p < 0.05; ∗∗p < 0.01. (G) Correlation network of metabolites and genes in the metabolic pathway. Nodes represent metabolites (blue squares) and genes (colored circles). Gray edges indicate pairwise correlations between metabolites and genes. The colors similarly represent expression levels, with red typically indicating higher expression and blue indicating lower expression compared to the mean. (H) Schematic diagram of arginine metabolism and the TCA cycle. (I) Heatmap of mitochondrial oxidation-related gene expression associated with differentially expressed metabolites. Red indicates relatively high gene expression, while blue indicates relatively low gene expression within each row. (J) Schematic diagram of the catalytic function of the GAPDH enzyme. (K) Concentrations of the NAD + /NADH ratio in THP-1 cells and BMDM cells treated with PBS, LPS, or LPS+3-HPA. Data are the means ± SD and n = 4 per group. Statistical significance was determined using one-way ANOVA followed by Tukey’s multiple comparisons test with ∗ p < 0.05, ∗∗ p < 0.01, and ∗∗∗∗ p < 0.0001; (L) ATP concentrations in THP-1 cells and BMDMs treated with PBS, LPS, or LPS+3-HPA. Data are the means ± SD and n = 3 per group. Statistical significance was determined using one-way ANOVA followed by Tukey’s multiple comparisons test with ∗ p < 0.05; ∗∗∗∗ p < 0.0001; ns, not significant. (M) Representative images and mitochondrial analysis of BMDM cells treated with PBS, LPS, or LPS+3-HPA. Scale bars, 1 μm (upper) and 0.25 μm (lower). For mitochondrial number, n = 8–12 ( n = 12 for PBS, n = 8 for LPS, n = 9 for LPS+3-HPA group).

Article Snippet: Mouse IL-1β Precoated ELISA Kit , Dakewe , 1210122.

Techniques: Expressing, Gene Expression

Journal: iScience

Article Title: 3-Hydroxypropionic acid converts inflammatory macrophage glycolysis into mitochondrial oxidation through GAPDH carboxyethylation

doi: 10.1016/j.isci.2026.116258

Figure Lengend Snippet: GAPDH carboxyethylation inhibited macrophage glycolysis and the release of inflammatory factors (A) Schematic workflow illustrating the strategy of silencing endogenous GAPDH via 3′UTR-targeting siRNA and overexpressing exogenous GAPDH. (B) Immunoblot and quantitative analysis of GAPDH protein in 293 T cells transfected with GAPDH 3′UTR-targeting siRNAs (siGAPDH 1, siGAPDH 2) or siRNA NC. Data are the means ± SD and n = 3 per group. Statistical significance was determined using one-way ANOVA followed by Dunnett’s multiple comparisons test ∗∗p < 0.01. (C) Relative mRNA expression of GAPDH in 293 T cells transfected with GAPDH 3′UTR-targeting siRNAs (siGAPDH 1, siGAPDH 2) or siRNA NC. Data are the means ± SD and n = 3 per group. Statistical significance was determined using one-way ANOVA followed by Dunnett’s multiple comparisons test ∗∗∗∗p < 0.0001. (D) Immunoblot analysis of FLAG-tagged exogenous GAPDH(E) and GAPDH in 293 T cells. Knockdown of endogenous GAPDH with siRNA followed by the overexpression of GAPDH (E). Data are the means ± SD and n = 3 per group. Statistical significance was determined using one-way ANOVA followed by Dunnett’s multiple comparisons test with ∗p < 0.05; ns, not significant. (E) Relative mRNA expression of GAPDH in 293 T cells knockdowned endogenous GAPDH (siGAPDH) and overexpressed GAPDH (C) and GAPDH (E), respectively. Data are the means ± SD and n = 3 per group. Statistical significance was determined using one-way ANOVA followed by Tukey’s multiple comparisons test with ∗∗p < 0.01; ∗∗∗p < 0.001; ns, not significant. (F) GAPDH activity assay in 293 T cells transfected with GAPDH 3′UTR siRNA, and overexpressing GAPDH(C), GAPDH(E). Data are the means ± SD and n = 4 per group. Statistical significance was determined using one-way ANOVA followed by Tukey’s multiple comparisons test with ∗p < 0.05; ∗∗∗∗p < 0.0001; ns, not significant. (G) Concentrations of lactate and pyruvate in 293 T cells transfected with GAPDH 3′UTR siRNA, and overexpressing GAPDH(C), GAPDH(E). Data are the means ± SD and n = 4 per group. Statistical significance was determined using one-way ANOVA followed by Tukey’s multiple comparisons test with ∗p < 0.05, ∗∗p < 0.01, and ∗∗∗∗p < 0.0001. (H) Relative mRNA expression of IL-6 , TNF-α , and IL-1β in THP-1 cells which transfected with GAPDH 3′UTR siRNA, and overexpressing GAPDH(C) or GAPDH(E). Data are the means ± SD and n = 3 per group. Statistical significance was determined using one-way ANOVA followed by Tukey’s multiple comparisons test with ∗p < 0.05; ∗∗p < 0.01; ∗∗∗p < 0.001; ns, not significant. (I) The concentration of TNF-α in THP-1 cells that overexpressed GAPDH(C) or GAPDH(E). Data are the means ± SD and n = 3 per group. Statistical significance was determined using one-way ANOVA followed by Tukey’s multiple comparisons test with ∗∗∗p < 0.001; ∗∗∗∗p < 0.0001; ns, not significant.

Article Snippet: Mouse IL-1β Precoated ELISA Kit , Dakewe , 1210122.

Techniques: Western Blot, Transfection, Expressing, Knockdown, Over Expression, Activity Assay, Concentration Assay

Journal: Bioactive Materials

Article Title: M2 macrophage-derived exosomes delivering haptoglobin and interleukin-10 plasmids for synergistic therapy of intracerebral hemorrhage

doi: 10.1016/j.bioactmat.2026.01.047

Figure Lengend Snippet: M2-exo@HI promotes in vitro microglia polarization, BBB repair and neuroprotection. (a) Flow cytometry analysis of M1-type (CD86 + ) and M2-type microglia (CD163 + ) following treatment with different formulations. (b,c) Percentages of CD86 + and CD163 + microglia populations (n = 3). (d–g) The cytokine levels of IL-10, TGF-β, TNF-α, and IL-1β in treated microglia (n = 3). (h) Fluorescence microscopy images showing erythrophagocytosis by microglia across treatment groups. (i) Schematic of the in vitro BBB model assessing FITC-dextran permeability using a transwell assay. (j) Quantitative analysis of FITC-dextran penetration (n = 7). (k) Flow cytometry analysis of neuronal apoptosis across treatments (n = 3). (l) Quantitative analysis of neuronal apoptosis (n = 3). Data are presented as mean ± SD. Statistical significance was tested by one-way ANOVA with Tukey's multiple comparisons test.

Article Snippet: ELISA kits included mouse TNF- α ELISA kit (Solarbio), mouse IL-10 ELISA kit (Elabscience), mouse Hp ELISA kit (Elabscience), mouse IL-1β ELISA kit (Solarbio), and TGF-β ELISA kit (Solarbio).

Techniques: In Vitro, Flow Cytometry, Fluorescence, Microscopy, Permeability, Transwell Assay