Journal: Bioactive Materials

Article Title: Glucose/ROS-responsive and redox-gated adaptive hydrogel dressing for accelerating diabetic wound repair via synergistic cGAS/STING pathway inhibition and oxidative stress alleviation

doi: 10.1016/j.bioactmat.2026.03.025

Figure Lengend Snippet: The cellular uptake and anti-inflammatory effect of HPSL in vitro . (A) Flow cytometry analysis and (B) semi-quantitative analysis of cellular uptake of PSL and blank NPs by M1 macrophages. n = 3. (C) Representative Giemsa staining images of LPS and high glucose-stimulated RAW 264.7 cells with different formulations, scale bar = 50 μm. (D) Immunofluorescence staining and semi-quantitative analysis of CD68 (red) and iNOS (green) in RAW 264.7 cells from different treatment groups, scale bar = 50 μm. n = 6. (E) Immunofluorescence staining and semi-quantitative analysis of CD68 (green) and Arg-1 (red) in RAW 264.7 cells from different treatment groups, scale bar = 50 μm. n = 6. Western blotting analysis and corresponding semi-quantitative analysis of (F) STING/ p -STING, (G) TBK1/ p -TBK1, (H) IRF3/ p -IRF3, (I) NF-κB, (J) TNF-α, and (K) IL-6, Lane 1: Normal group, Lane 2: Model group, Lane 3: PSL group, Lane 4: Free H151 group, Lane 5: HPSL group. n = 3. All data are shown as mean ± SEM.

Article Snippet: VEGF-A and TNF-α-specific antibodies were purchased from Wanleibio (Shenyang, China).

Techniques: In Vitro, Flow Cytometry, Staining, Immunofluorescence, Western Blot

Journal: Bioactive Materials

Article Title: Inhalable PD-L1-engineered hybrid cellular vesicles suppress excessive neutrophil activation and restore mitochondrial homeostasis to alleviate ischemia–reperfusion lung injury and pneumonia

doi: 10.1016/j.bioactmat.2026.03.024

Figure Lengend Snippet: Characterization of Res-PD-L1@nmEVs . (A) Schematic illustration of the Res-PD-L1@nmEVs synthesis procedure. (B-D) Representative transmission electron microscopy (TEM) images, dynamic light scattering (DLS) size distributions, and zeta potential measurements of nEVs, PD-L1@mEVs, PD-L1@nmEVs, and Res-PD-L1@nmEVs. (E) PD-L1 expression in PD-L1-overexpressing MSCs (OE-PD-L1) and negative control (NC) MSCs, and CD11b expression in HL60 cells before and after DMSO stimulation, as determined by Western blot. (F) Expression levels of neutrophil membrane markers (CD11b, CXCR2, RAGE, TLR2) and the exosomal marker CD63 in the four EV types. (G) Fluorescence co-localization images of DiO-labeled nEVs (green) and DiL-labeled PD-L1@mEVs (red) after fusion, demonstrating hybrid vesicle formation. (H) Size stability of Res-PD-L1@nmEVs stored at 4 °C and 37 °C for 7 days. (I-K) Binding and neutralization capacity of Res-PD-L1@nmEVs against inflammatory cytokines (TNF-α, IL-6, IL-1β) in vitro. ∗ vs. 0ug/ml; # vs. 100 μg/ml, p < 0.05, n = 5.

Article Snippet: These cells were then stimulated with 40 ng/mL recombinant TNF-α (C008, Novoprotein, China) for 6 h to induce an N1-type neutrophil phenotype.

Techniques: Transmission Assay, Electron Microscopy, Zeta Potential Analyzer, Expressing, Negative Control, Western Blot, Membrane, Marker, Fluorescence, Labeling, Binding Assay, Neutralization, In Vitro

Journal: Bioactive Materials

Article Title: Inhalable PD-L1-engineered hybrid cellular vesicles suppress excessive neutrophil activation and restore mitochondrial homeostasis to alleviate ischemia–reperfusion lung injury and pneumonia

doi: 10.1016/j.bioactmat.2026.03.024

Figure Lengend Snippet: Res-PD-L1@nmEVs Attenuate Inflammation and Oxidative Damage in Lung Epithelial Cells In Vitro . (A-B) Flow cytometric analysis and quantification (B) of DiO-labeled Res-PD-L1@nmEVs uptake by BEAS-2B cells under H/R conditions after pretreatment with different endocytic inhibitors (chlorpromazine, chloroquine, and filipin) or incubation at 4 °C. (C) mRNA expression levels of IL-6, TNF-α, and IL-1β in BEAS-2B cells with or without H/R injury following pretreatment with Res, nEVs, PD-L1@mEVs, PD-L1@nmEVs, or Res-PD-L1@nmEVs. (D-E) Representative fluorescence images (D) and quantitative analysis (E) of cell proliferation assessed by BrdU incorporation (red; nuclei stained with DAPI, blue). Scale bar: 50 μm. (F-G) Apoptosis rates detected by flow cytometry (F) and flow cytometric analysis of Annexin V-positive BEAS-2B cells under the indicated conditions (G). (H–K) Fluorescence microscopy images and quantitative analysis of intracellular nitric oxide (NO, green) (H-I) and reactive oxygen species (ROS, red) (J-K). Scale bar: 100 μm. (L) Flow cytometry analysis of intracellular ROS levels. (M − O) Levels of malondialdehyde (MDA) (M), superoxide dismutase 2 (SOD2) activity (N), and glutathione (GSH) content (O) in cells. (P-Q) Cell migration ability evaluated by wound healing assay under different treatments. ∗ vs. Control; # vs. H/R; & vs. H/R + PD-L1@nmEVs, p < 0.05.

Article Snippet: These cells were then stimulated with 40 ng/mL recombinant TNF-α (C008, Novoprotein, China) for 6 h to induce an N1-type neutrophil phenotype.

Techniques: In Vitro, Labeling, Incubation, Expressing, Fluorescence, BrdU Incorporation Assay, Staining, Flow Cytometry, Microscopy, Activity Assay, Migration, Wound Healing Assay, Control

Journal: Bioactive Materials

Article Title: Inhalable PD-L1-engineered hybrid cellular vesicles suppress excessive neutrophil activation and restore mitochondrial homeostasis to alleviate ischemia–reperfusion lung injury and pneumonia

doi: 10.1016/j.bioactmat.2026.03.024

Figure Lengend Snippet: Res-PD-L1@nmEVs Suppresses Neutrophil Activation HL60 cells were differentiated into neutrophil-like cells using DMSO and subsequently stimulated with TNF-α to induce activation under conditions simulating IRI. The effects of Res, nEVs, PD-L1@mEVs, PD-L1@nmEVs, and Res-PD-L1@nmEVs on neutrophil activation were evaluated. (A) Cell surface PD-1 expression analyzed by flow cytometry. (B) Representative immunofluorescence images of CD206 expression (red). Nuclei were stained with DAPI (blue). Scale bar: 50 μm. (C) Flow cytometric analysis of cell surface CD206 expression. (D) Flow cytometric analysis of cell surface CD95 expression. (E-G) Levels of myeloperoxidase (MPO) (E), neutrophil elastase (NE) (F), and MMP-9 (G) in neutrophil culture supernatants, measured by ELISA. (H-J) BEAS-2B cells were co-cultured with neutrophils in the presence or absence of TNF-α stimulation. Apoptosis levels (I) and migration capacity (J) of BEAS-2B cells were assessed under different treatment conditions. ∗ vs. Control; # vs. TNF-a; & vs. TNF-a+PD-L1@nmEVs, p < 0.05.

Article Snippet: These cells were then stimulated with 40 ng/mL recombinant TNF-α (C008, Novoprotein, China) for 6 h to induce an N1-type neutrophil phenotype.

Techniques: Activation Assay, Expressing, Flow Cytometry, Immunofluorescence, Staining, Enzyme-linked Immunosorbent Assay, Cell Culture, Migration, Control

Journal: Bioactive Materials

Article Title: Inhalable PD-L1-engineered hybrid cellular vesicles suppress excessive neutrophil activation and restore mitochondrial homeostasis to alleviate ischemia–reperfusion lung injury and pneumonia

doi: 10.1016/j.bioactmat.2026.03.024

Figure Lengend Snippet: Res-PD-L1@nmEVs Effectively Attenuates MRSA-Induced Pneumonia (A-B) Rats with MRSA-induced pneumonia received three bronchial nebulization treatments over one week with different formulations (Res, nEVs, PD-L1@mEVs, PD-L1@nmEVs, or Res-PD-L1@nmEVs). (A) Representative H&E-stained lung sections and (B) corresponding lung injury scores are shown (n = 5). (C) TUNEL staining of lung tissues to assess apoptosis. (D) Representative micro-CT images of anesthetized rats. (E-G) Flow cytometric analysis of immune cell proportions in lung single-cell suspensions: CD8 + T cells (E), neutrophils (F), and classical monocytes (G). (H-J) Plasma levels of inflammatory cytokines IL-6 (H), IL-1β (I), and TNF-α (J) (n = 5). (K) Immunofluorescence staining of tight junction proteins Occludin (green) and ZO-1 (red) in lung tissues (nuclei stained with DAPI). Scale bar: 50 μm. (L-N) Pulmonary function parameters: lung compliance (L), airway resistance (M), and oxygenation index (N) (n = 4). ∗ vs. Sham; # vs. MRSA; & vs. MRSA + PD-L1@nmEVs, p < 0.05.

Article Snippet: These cells were then stimulated with 40 ng/mL recombinant TNF-α (C008, Novoprotein, China) for 6 h to induce an N1-type neutrophil phenotype.

Techniques: Staining, TUNEL Assay, Micro-CT, Single Cell, Clinical Proteomics, Immunofluorescence

Journal: International Journal for Parasitology: Drugs and Drug Resistance

Article Title: Terpenic compounds possess anthelmintic and immunomodulatory properties with potential for controlling equine cyathostomin infections

doi: 10.1016/j.ijpddr.2026.100642

Figure Lengend Snippet: Scatter plots of the association between TNF-α production and the IC 50 values (LDA and LMA) Scatter plots showing the association between TNF-α production percentage (expressed relative to the control) and the IC 50 values obtained in (left) the Larval Development Assay (LDA) and (right) the Larval Migration Assay (LMA) for six terpene compounds (anethole, cinnamaldehyde, menthol, carvacrol, eugenol and thymol). Each point represents the mean IC 50 and TNFα production for a given compound, and horizontal/vertical bars indicate the corresponding confidence intervals. Lower IC 50 values reflect higher antiparasitic potency.

Article Snippet: The cells were then incubated at +37 °C (5% CO 2 ) for 24 h. After incubation, the concentration of TNF-α in the medium for each condition was quantified by ELISA using mouse TNF-α paired antibodies (R and D Systems DY410).

Techniques: Control, Migration

Journal: International Journal for Parasitology: Drugs and Drug Resistance

Article Title: Terpenic compounds possess anthelmintic and immunomodulatory properties with potential for controlling equine cyathostomin infections

doi: 10.1016/j.ijpddr.2026.100642

Figure Lengend Snippet: Anti-inflammatory activity of carvacrol and cinnamaldehyde on equine PBMC Boxplots showing TNF-α concentrations (ng/mL) measured in equine peripheral blood mononuclear cells (PBMCs) exposed to DMSO (0.05%), LPS (125 ng/mL), the combination of DMSO and LPS, carvacrol (5 μg/mL), cinnamaldehyde (5 μg/mL), the combination of either compound with LPS, and the untreated condition (control). Points represent individual replicates from four independent assays. Asterisks indicate significant differences relative to the corresponding control condition (∗ P = 0.01, ∗∗ P < 0.001).

Article Snippet: The cells were then incubated at +37 °C (5% CO 2 ) for 24 h. After incubation, the concentration of TNF-α in the medium for each condition was quantified by ELISA using mouse TNF-α paired antibodies (R and D Systems DY410).

Techniques: Activity Assay, Control

Journal: Bioactive Materials

Article Title: A safe and anti-inflammatory plant-derived nanovesicle platform for targeted delivery in acute lung injury

doi: 10.1016/j.bioactmat.2026.03.033

Figure Lengend Snippet: The anti-inflammatory effect of the nanoparticles in vitro (a) The proportion of CD86 M1 and CD206 M2 in RAW264.7 cells by flow cytometry. (b-c) Level of inflammatory cytokines IL-6, TNF-α and IL-10 in LPS-stimulated RAW 264.7 cells (b) and MH-S cells (c) after different treatments (n = 5). (d-e) NF-κB p65 nuclear translocation observed by CLSM (n = 5). ①control group; ② PBS treated group; ③ LEVs treated group; ④ GRb1 treated group; ⑤ GRb1@LEVs treated group; ⑥ GRb1@LEVs-cRGD treated group. (f) The protein expressions of key members in the NF-κB pathway by Western blot, including the phosphorylated (p-p65) and basal NF-κB p65, p-IκBα, and IκBα (n = 3). (g) Heat map showing the hierarchical clustering results of the DEGs detected between LPS group and LPS treated with GRb1@LEVs-cRGD group. DEGs were identified based on a fold change greater than 1.5 and an adjusted P-value less than 0.05. (g) Gene Ontology (GO) term enrichment analysis was performed, and the top 30 significantly enriched GO terms were selected based on an FDR <0.05. (h) Top 20 enriched pathways identified using Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis of DEGs in cells triggered with LPS. (i-j) Gene Set Enrichment Analysis (GSEA) images. (l) Schematic illustration of the mechanism in the reprogramming of RAW 264.7 cells by GRb1@LEVs-cRGD.

Article Snippet: Mouse precoated ELISA kits for TNF-α, IL-6, IL-1β were purchased from Beijing Dakewe Biotechnology Co., Ltd (China).NF-kB p65/RelA Rabbit mAb (A22331) and ZO-1 Rabbit mAb (A25306) were purchased from ABclonal Technology Co., Ltd (China).

Techniques: In Vitro, Flow Cytometry, Translocation Assay, Western Blot

Journal: Bioactive Materials

Article Title: A safe and anti-inflammatory plant-derived nanovesicle platform for targeted delivery in acute lung injury

doi: 10.1016/j.bioactmat.2026.03.033

Figure Lengend Snippet: GRb1@LEVs-cRGD alleviated LPS-induced lung injury and inflammatory responses in the lung. (a) Schematic illustration of animal experimental design. (b-d) lung wet/dry ratio (b), levels of protein (c), total cell number (d) in BALF in LPS induced ALI mice treated with PBS, LEVs, GRb1, GRb1@LEVs, GRb1@LEVs-cRGD (n = 5). (e-h) Level of IL-6 and TNF-α in BALF (e) and serum (h) of mice after different treatments (n = 5). (i-j) Representative H&E images of lungs after different treatments (i) and the corresponding analysis of lung injury score (j) (n = 5). (k) Lung tissue analysis TUNEL staining in each group (n = 5). (l-n) The distribution of tight junction proteins, ZO-1 and VE-Cadherin, by immunofluorescent staining in lung tissues and corresponding quantification.

Article Snippet: Mouse precoated ELISA kits for TNF-α, IL-6, IL-1β were purchased from Beijing Dakewe Biotechnology Co., Ltd (China).NF-kB p65/RelA Rabbit mAb (A22331) and ZO-1 Rabbit mAb (A25306) were purchased from ABclonal Technology Co., Ltd (China).

Techniques: TUNEL Assay, Staining

Journal: Bioactive Materials

Article Title: A safe and anti-inflammatory plant-derived nanovesicle platform for targeted delivery in acute lung injury

doi: 10.1016/j.bioactmat.2026.03.033

Figure Lengend Snippet: TIG/GRb1@LEVs-cRGD alleviated Kp NDM-induced lung injury and inflammatory responses in the lung. (a) Experimental design of in vivo assessment using a Kp NDM-induced model. (b) Growth curves of KP NDM co-incubated with various preparations (n = 3). (c) Corresponding quantification of bacterial load in lung tissue homogenate (n = 5). (d-e) Lung wet/dry ratio (d) and levels of protein (e) in Kp NDM-induced ALI mice treated with PBS, TIG, TIG/GRb1@LEVs-cRGD and negative control (n = 5). (f-h) IL-6 (f) and TNF-α (g) IL-1β (h) of BALF in above groups (n = 5). (i-j) Representative H&E images of the lung after different treatments (i) and corresponding lung injury score analysis (j) (n = 5). (k-p) Immunofluorescence staining images of IL-6 (k) and corresponding quantitative analysis (n), TNF-α (l) and corresponding quantitative analysis (o), IL-1β(m) and corresponding quantitative analysis (p) (n = 5).

Article Snippet: Mouse precoated ELISA kits for TNF-α, IL-6, IL-1β were purchased from Beijing Dakewe Biotechnology Co., Ltd (China).NF-kB p65/RelA Rabbit mAb (A22331) and ZO-1 Rabbit mAb (A25306) were purchased from ABclonal Technology Co., Ltd (China).

Techniques: In Vivo, Incubation, Negative Control, Immunofluorescence, Staining

Journal: Bioactive Materials

Article Title: A safe and anti-inflammatory plant-derived nanovesicle platform for targeted delivery in acute lung injury

doi: 10.1016/j.bioactmat.2026.03.033

Figure Lengend Snippet: Vanc/GRb1@LEVs-cRGD alleviated MRSA-induced lung injury and inflammatory responses in the lung. (a) Experimental design of in vivo assessment using a MRSA-induced model. (b) Growth curves of MRSA co-incubated with various preparations (n = 3). (c) Corresponding quantification of bacterial load in lung tissue homogenate (n = 5). (d-e) Lung wet/dry ratio (d) and levels of protein (e) in MRSA-induced ALI mice treated with PBS, Vanc, Vanc/GRb1@LEVs-cRGD and negative control (n = 5). (f-h) IL-6 (f) and TNF-α (g) IL-1β (h) of BALF in above groups (n = 5). (i-j) Representative H&E images of the lung after different treatments (i) and corresponding lung injury score analysis (j) (n = 5). (k-p) Immunofluorescence staining images of IL-6 (k) and corresponding quantitative analysis (n), TNF-α (l) and corresponding quantitative analysis (o), IL-1β (m) and corresponding quantitative analysis (p) (n = 5).

Article Snippet: Mouse precoated ELISA kits for TNF-α, IL-6, IL-1β were purchased from Beijing Dakewe Biotechnology Co., Ltd (China).NF-kB p65/RelA Rabbit mAb (A22331) and ZO-1 Rabbit mAb (A25306) were purchased from ABclonal Technology Co., Ltd (China).

Techniques: In Vivo, Incubation, Negative Control, Immunofluorescence, Staining

Journal: Translational Oncology

Article Title: Combination of APS, HMGN1, and anti-TNFR2 antibody remodels the tumor immune microenvironment to enhance antitumor immunity in colorectal cancer

doi: 10.1016/j.tranon.2026.102814

Figure Lengend Snippet: Triple combination therapy alleviates colorectal cancer immunosuppression by targeting immune cells and cytokine networks. (A-E) The ratio of CD4 + CD25 + Foxp3 + Tregs among CD4 + cells in the tumor, spleen, TDLNs, and NTDLNs (n=5). (F-J) Percentage of Tim3 + cells among CD8 + T cells in the tumor, spleen, TDLNs, and NTDLNs(n=5). (K-O) Serum circulating cytokines were quantified from the serum samples, including IL-2, IL-10, TNF-α, TGF-β, IL-6 with 5 samples each in an experimental group. (All data are presented as mean ± SD and were analyzed using one-way ANOVA. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001, ns = no significant difference, P > 0.05.).

Article Snippet: Serum levels of certain cytokines (IL-2(Cat. No. GEM0038), IL-6(Cat. No. GEM0001), IL-10(Cat. No. GEM0003), TNF-α (Cat. No. GEM0004), TGF-β (Cat. No. GEM0051) were detected by ELISA kits (Servicebio, Wuhan, China) according to the instructions from the manufacturer.

Techniques:

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: Human TNF-α Precoated ELISA Kit , Dakewe , 1117202.

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: GAPDH is modified with carboxyethylation at Cys 247 (A) Mass spectrometry analysis of GAPDH peptide (234–260) with carboxyethylation and 3-HPA (5 mM) incubated with the GAPDH peptide (234–260) at 37 °C for 4 h. (B) SPR analysis of the affinity of the anti-ceC247 antibody for the carboxyethylation modified GAPDH peptide (234–260) and unmodified GAPDH peptide (234–260). (C) ELISA-based binding curve of anti-ceC247 antibody to modified GAPDH peptide (GAPDH ce (234–260)) and unmodified GAPDH peptide (234–260). Data are the means ± SD and n = 3 per group. Statistical significance was determined using two-way ANOVA followed by ∗∗p < 0.01. (D) Chemical structures of cysteine carboxyethylation and cysteine lactylation. (E) Unmodified GAPDH peptide (234–260), carboxyethylated peptide (GAPDH ce (234–260)), and lactylated peptide (GAPDH lac (234–260)) were tested with the anti-ceC247 antibody in dot blot assays. (F) Immunoblots of lysates from 293 T cells overexpressing GAPDH, which were treated with 5 mM 3-HPA and 5 μM MG132. The blots were probed with the anti-ceC247 antibody. (G) 3-HPA incubated with the GAPDH peptide (234–260) at 37 °C for 4 h. An anti-ceC247 antibody and the anti-wtC247 antibody were used in dot blot assays.

Article Snippet: Human TNF-α Precoated ELISA Kit , Dakewe , 1117202.

Techniques: Modification, Mass Spectrometry, Incubation, Enzyme-linked Immunosorbent Assay, Binding Assay, Dot Blot, Western Blot

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: Human TNF-α Precoated ELISA Kit , Dakewe , 1117202.

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: Human TNF-α Precoated ELISA Kit , Dakewe , 1117202.

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