Journal: Bioactive Materials

Article Title: Dynamic feedback BacGuard anchors microbial metabolism to host symbiosis in real-time ulcerative colitis therapy

doi: 10.1016/j.bioactmat.2026.05.060

Figure Lengend Snippet: BacGuard promotes colon tissue repairment. a) Scheme of microbiota-dependent epithelial repair mechanism orchestrated by BacGuard. b) Short-chain fatty acid (SCFA) profile alterations. n = 6. c) BacGuard-induced probiotic proliferation and d) quantitative results. n = 3. e) Immunofluorescence analysis of ILC3 (ROR γt + CD3 − cells) in colon tissue. f) Flow cytometric analysis of lamina propria lymphocytes (ROR γt + ). n = 3. g) Concentration of IL-22 in MNK-3 cells. n = 3. h) Representative PAS-staining (upper panel) and MUC-2 immunohistochemistry (lower panel) images of colon tissues. n = 5. ns, not significant; ∗p < 0.05, ∗∗p < 0.01 and ∗∗∗p < 0.001.

Article Snippet: For the detection of IL-22 and LPS concentrations, the Mouse IL-22 Precoated ELISA Kit (DAKEWE, China) and Mouse LPS ELISA Kit (JONLNBIO, China) were employed correspondingly.

Techniques: Immunofluorescence, Concentration Assay, Staining, Immunohistochemistry

Journal: iScience

Article Title: IL-4 and arginine synergy accelerates early bone regeneration via enhanced efferocytosis and ameliorated microenvironment

doi: 10.1016/j.isci.2026.117739

Figure Lengend Snippet: Simultaneous application of IL-4 and Arg promotes efferocytosis and improves the pro-regenerative microenvironment (A) Schematic illustration of the in vitro efferocytosis assessment. (B and C) Macrophage efferocytosis assessment via flow cytometry, showing representative scatterplots of macrophages engulfing apoptotic cells (B) and quantification of the efferocytosis rate after 24 h (C). (D) Morphological evaluation of macrophage efferocytosis, showing representative cell tracking images of macrophages (labeled with CMFDA) engulfing apoptotic cells (labeled with CM-Dil) after 24 h of co-culture. (E) Quantification of the efferocytosis rate after 6, 24, and 48 h determined by flow cytometry under Control and IL-4 + Arg conditions. (F) Experimental design for detecting microenvironmental changes following efferocytosis. (G) ROS quantification following efferocytosis. (H) Enzyme-linked immunosorbent assay (ELISA) quantification of TGF-β1 and IL-10 in the culture supernatants. Error bars represent SD. Data were analyzed by one-way ANOVA followed by Tukey’s post hoc test (C, G, and H) or two-way ANOVA followed by Sidak’s multiple comparisons test (E). Sample sizes: n = 3 biological replicates (B–E); n = 5 biological replicates, each performed with 3 technical replicates (G) and n = 3 biological replicates, each performed with 3 technical replicates (H). Scale bars, 100 μm (D) and 20 μm (inset).

Article Snippet: Mouse TGF-β1 ELISA Kit , ABclonal , Cat# RK00057.

Techniques: In Vitro, Flow Cytometry, Cell Tracking Assay, Labeling, Co-culture Assay, Control, Enzyme-linked Immunosorbent Assay

Journal: iScience

Article Title: IL-4 and arginine synergy accelerates early bone regeneration via enhanced efferocytosis and ameliorated microenvironment

doi: 10.1016/j.isci.2026.117739

Figure Lengend Snippet: Transcriptomic changes underlying synergistic regulation of macrophages by IL-4 and Arg (A) RT-qPCR analysis of M2-related marker gene expression ( Tgfb1 , Il10 , Arg1 , and Cd206 ) in macrophages. (B) Schematic illustration of the bone marrow-derived macrophages (BMDMs) isolation, total RNA extraction, and transcriptomic sequencing workflow. (C) Functional annotation of upregulated genes using Gene Ontology (GO) enrichment analysis. (D) Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis emphasizing significant signaling pathways. (E) RT-qPCR analysis of Tgfb1 and Il10 mRNA expression levels of macrophage after treatment with the IL-4 + Arg combination in the presence or absence of nor-NOHA. (F) ELISA quantification of secreted TGF-β1 and IL-10 protein concentrations in the culture supernatants under nor-NOHA inhibition. Error bars represent SD. Data were analyzed by one-way ANOVA followed by Tukey’s post hoc test (A, E, and F). Sample sizes: n = 3 biological replicates (C and D); n = 3 (A and F) and 4 (E) biological replicates, each performed with 3 technical replicates.

Article Snippet: Mouse TGF-β1 ELISA Kit , ABclonal , Cat# RK00057.

Techniques: Quantitative RT-PCR, Analysis, Marker, Gene Expression, Derivative Assay, Isolation, RNA Extraction, Sequencing, Functional Assay, Protein-Protein interactions, Expressing, Enzyme-linked Immunosorbent Assay, Inhibition

Journal: iScience

Article Title: IL-4 and arginine synergy accelerates early bone regeneration via enhanced efferocytosis and ameliorated microenvironment

doi: 10.1016/j.isci.2026.117739

Figure Lengend Snippet:

Article Snippet: Mouse TGF-β1 ELISA Kit , ABclonal , Cat# RK00057.

Techniques: Cell-Signaling, Recombinant, Enzyme-linked Immunosorbent Assay, TUNEL Assay, Software

Journal: iScience

Article Title: IL-4 and arginine synergy accelerates early bone regeneration via enhanced efferocytosis and ameliorated microenvironment

doi: 10.1016/j.isci.2026.117739

Figure Lengend Snippet: IL-4 and Arg remodel macrophages and promote their migration (A) Schematic illustration of the regulatory role of IL-4 on macrophage phenotype. (B) Volcano plot displaying differentially expressed genes (DEGs) between the control and IL-4-treated macrophages. (C) Heatmap demonstrating the expression of key macrophage polarization markers. (D) Multi-pathway Gene Set Enrichment Analysis (GSEA) demonstrating the integrated enrichment of indicated pathways regulating macrophage functions. (E) Schematic illustration of macrophage priming for efferocytosis mediated by IL-4 and Arg. (F) Relative cell viability of macrophages treated with different concentrations of Arg at 24, 48, and 72 h. (G and H) Macrophage proliferation evaluated by 5-ethynyl-2′-deoxyuridine (EdU) assay, showing representative fluorescence images of EdU and 4',6-diamidino-2-phenylindole (DAPI) staining at 24, 48, and 72 h (G) and quantification of EdU-positive cells treated with 2 mM Arg (H). (I) Schematic illustration of the Transwell migration assay. (J and K) Macrophage recruitment evaluated by Transwell assay, showing representative crystal violet staining images of migrated macrophages after 48 h (J) and quantification of the number of migrated macrophages (K). Error bars represent SD. Data were analyzed by two-way ANOVA followed by Sidak’s multiple comparisons test (F and H) or one-way ANOVA followed by Tukey’s post hoc test (K). Sample sizes: n = 3 biological replicates (B–D and F); n = 5 biological replicates, each performed with 3 technical replicates (G, H, J, and K). Scale bars, 25 μm (G), 500 μm (J, top), and 100 μm (J, bottom).

Article Snippet: The cumulative release of Arg was quantified using an Arg Content Assay Kit (Solarbio, Beijing, China), and the release of IL-4 was determined using a Mouse IL-4 ELISA Kit (Elabscience, Wuhan, China).

Techniques: Migration, Control, Expressing, Analysis, EdU Assay, Fluorescence, Staining, Transwell Migration Assay, Transwell Assay

Journal: iScience

Article Title: IL-4 and arginine synergy accelerates early bone regeneration via enhanced efferocytosis and ameliorated microenvironment

doi: 10.1016/j.isci.2026.117739

Figure Lengend Snippet: Simultaneous application of IL-4 and Arg promotes efferocytosis and improves the pro-regenerative microenvironment (A) Schematic illustration of the in vitro efferocytosis assessment. (B and C) Macrophage efferocytosis assessment via flow cytometry, showing representative scatterplots of macrophages engulfing apoptotic cells (B) and quantification of the efferocytosis rate after 24 h (C). (D) Morphological evaluation of macrophage efferocytosis, showing representative cell tracking images of macrophages (labeled with CMFDA) engulfing apoptotic cells (labeled with CM-Dil) after 24 h of co-culture. (E) Quantification of the efferocytosis rate after 6, 24, and 48 h determined by flow cytometry under Control and IL-4 + Arg conditions. (F) Experimental design for detecting microenvironmental changes following efferocytosis. (G) ROS quantification following efferocytosis. (H) Enzyme-linked immunosorbent assay (ELISA) quantification of TGF-β1 and IL-10 in the culture supernatants. Error bars represent SD. Data were analyzed by one-way ANOVA followed by Tukey’s post hoc test (C, G, and H) or two-way ANOVA followed by Sidak’s multiple comparisons test (E). Sample sizes: n = 3 biological replicates (B–E); n = 5 biological replicates, each performed with 3 technical replicates (G) and n = 3 biological replicates, each performed with 3 technical replicates (H). Scale bars, 100 μm (D) and 20 μm (inset).

Article Snippet: The cumulative release of Arg was quantified using an Arg Content Assay Kit (Solarbio, Beijing, China), and the release of IL-4 was determined using a Mouse IL-4 ELISA Kit (Elabscience, Wuhan, China).

Techniques: In Vitro, Flow Cytometry, Cell Tracking Assay, Labeling, Co-culture Assay, Control, Enzyme-linked Immunosorbent Assay

Journal: iScience

Article Title: IL-4 and arginine synergy accelerates early bone regeneration via enhanced efferocytosis and ameliorated microenvironment

doi: 10.1016/j.isci.2026.117739

Figure Lengend Snippet: Transcriptomic changes underlying synergistic regulation of macrophages by IL-4 and Arg (A) RT-qPCR analysis of M2-related marker gene expression ( Tgfb1 , Il10 , Arg1 , and Cd206 ) in macrophages. (B) Schematic illustration of the bone marrow-derived macrophages (BMDMs) isolation, total RNA extraction, and transcriptomic sequencing workflow. (C) Functional annotation of upregulated genes using Gene Ontology (GO) enrichment analysis. (D) Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis emphasizing significant signaling pathways. (E) RT-qPCR analysis of Tgfb1 and Il10 mRNA expression levels of macrophage after treatment with the IL-4 + Arg combination in the presence or absence of nor-NOHA. (F) ELISA quantification of secreted TGF-β1 and IL-10 protein concentrations in the culture supernatants under nor-NOHA inhibition. Error bars represent SD. Data were analyzed by one-way ANOVA followed by Tukey’s post hoc test (A, E, and F). Sample sizes: n = 3 biological replicates (C and D); n = 3 (A and F) and 4 (E) biological replicates, each performed with 3 technical replicates.

Article Snippet: The cumulative release of Arg was quantified using an Arg Content Assay Kit (Solarbio, Beijing, China), and the release of IL-4 was determined using a Mouse IL-4 ELISA Kit (Elabscience, Wuhan, China).

Techniques: Quantitative RT-PCR, Analysis, Marker, Gene Expression, Derivative Assay, Isolation, RNA Extraction, Sequencing, Functional Assay, Protein-Protein interactions, Expressing, Enzyme-linked Immunosorbent Assay, Inhibition

Journal: iScience

Article Title: IL-4 and arginine synergy accelerates early bone regeneration via enhanced efferocytosis and ameliorated microenvironment

doi: 10.1016/j.isci.2026.117739

Figure Lengend Snippet: Simultaneous application of IL-4 and Arg promotes bone regeneration in vivo (A) Schematic illustration of the surgical procedure and timeline. (B and C) Evaluation of early macrophage polarization in vivo , showing representative immunofluorescence images of macrophages (F4/80, red) and M2 marker (Arg1, green) at days 1 and 3 post-surgery (B) and quantification of the percentage of M2 macrophages (Arg1 + /F4/80 + ) (C). (D–F) Bone regeneration assessment via micro-CT, showing representative 3D reconstruction images of the bone defect area at day 7 and day 14 post-surgery (D) and quantification of BV (E) and BV/TV (F). (G and H) Histological evaluation of bone repair, showing representative images of H&E and Masson’s trichrome staining at day 7 and day 14 (G) and quantification of new bone area (%) (H). Error bars represent SD. Data were analyzed by two-way ANOVA followed by Sidak’s multiple comparisons test (C, E, F, and H). Sample sizes: n = 4 animals per group. Scale bars, 50 μm (B) and 10 μm (inset), 0.5 mm (D) and 200 μm (inset), 250 μm (G).

Article Snippet: The cumulative release of Arg was quantified using an Arg Content Assay Kit (Solarbio, Beijing, China), and the release of IL-4 was determined using a Mouse IL-4 ELISA Kit (Elabscience, Wuhan, China).

Techniques: In Vivo, Immunofluorescence, Marker, Micro-CT, Staining

Journal: iScience

Article Title: IL-4 and arginine synergy accelerates early bone regeneration via enhanced efferocytosis and ameliorated microenvironment

doi: 10.1016/j.isci.2026.117739

Figure Lengend Snippet:

Article Snippet: The cumulative release of Arg was quantified using an Arg Content Assay Kit (Solarbio, Beijing, China), and the release of IL-4 was determined using a Mouse IL-4 ELISA Kit (Elabscience, Wuhan, China).

Techniques: Cell-Signaling, Recombinant, Enzyme-linked Immunosorbent Assay, TUNEL Assay, Software

Journal: iScience

Article Title: Topical application of urolithin A activates mitophagy and enhances mitochondrial function to promote skin longevity

doi: 10.1016/j.isci.2026.117646

Figure Lengend Snippet: UA beneficial impact on human keratinocytes and reconstituted human epidermis (A) Mitochondrial respiration (basal and maximal) assessed as oxygen consumption rate (OCR) in human dermal keratinocytes after 72 h of differentiation from young donor treated with DMSO for 24 h or the indicated dose of UA. ∗ p < 0.05 and ∗∗∗ p < 0.001 after one-way ANOVA. Error bars represent mean ± SEM. (B) TOMM20 immunostaining of skin epidermis in a human full-thickness skin model from an old donor treated with or without UA 1 μM for 25 days. Data expressed as antibody staining signal over imaging field. ∗ p < 0.05 and ∗∗∗∗ p < 0.0001 after unpaired t test one-way ANOVA. Error bars represent mean ± SEM. Scale bars, 50 μm. (C) Relative gene expression of PINK1 and PARK2 in reconstructed human epidermis (RHE). RHE tissues were moisturized for 24 h with or without 1% Mitopure, and relative expression levels of PINK1 and PARK2 were quantified. Statistical analysis was performed using one-way ANOVA (∗ p < 0.05; ∗∗ p < 0.01). Error bars represent mean ± SEM. (D) Interleukin 6 levels normalized to cell viability in keratinocytes. Keratinocytes were stimulated with a pro-inflammatory cytokine mix for 24 h then treated for an additional 24 h with either DMSO or 1 μM UA. IL-6 secretion was quantified and normalized to cell viability. Statistical analysis was performed using one-way ANOVA (∗∗∗∗ p < 0.0001). Error bars represent mean ± SEM. (E) p16 immunostaining in a human full-thickness skin model from an old donor treated with or without UA 1 μM for 25 days. Data expressed as antibody staining signal over imaging field. ∗∗∗∗ p < 0.0001 after unpaired t test. Error bars represent mean ± SEM. Scale bars, 50 μm. (F) Ki67 immunostaining in a human full-thickness skin model from an old donor treated with or without UA 1 μM for 25 days. Data expressed as number of Ki67 positive cells normalized over μm of dermal epidermal junction (count/μm) ∗∗∗∗ p < 0.0001 after unpaired t test. Error bars represent mean ± SEM. Scale bars, 50 μm. (G) Keratin 10 immunostaining in a human full-thickness skin model from an old donor treated with or without UA 1 μM for 25 days. Data expressed as fluorescence intensity over epidermis area. ∗∗∗∗ p < 0.0001 after unpaired t test. Error bars represent mean ± SEM. Scale bars, 50 μm.

Article Snippet: IL-6 ELISA Kit , Proteintech , Cat#KE10007.

Techniques: Immunostaining, Staining, Imaging, Gene Expression, Expressing, Analysis, Fluorescence

Journal: iScience

Article Title: Topical application of urolithin A activates mitophagy and enhances mitochondrial function to promote skin longevity

doi: 10.1016/j.isci.2026.117646

Figure Lengend Snippet:

Article Snippet: IL-6 ELISA Kit , Proteintech , Cat#KE10007.

Techniques: Binding Assay, Ex Vivo, Recombinant, Metals, Enzyme-linked Immunosorbent Assay, Reverse Transcription, Cell Viability Assay, Staining, DC Protein Assay, Gene Expression, Glass, Software, Imaging, Microscopy, Real-time Polymerase Chain Reaction

Journal: Biomarker Research

Article Title: ZDHHC5-mediated S-palmitoylation of NOD2 drives neutrophil extracellular trap formation via mitophagy to exacerbate inflammation in COPD

doi: 10.1186/s40364-026-01001-2

Figure Lengend Snippet: NOD2 deficiency ameliorates cigarette smoke-induced lung inflammation and injury in vivo The functional role of NOD2 was evaluated in a cigarette smoke (CS)-induced mouse model of COPD using wild-type (wt) and NOD2-knockout (NOD2-/-) mice, with lentiviral-mediated NOD2 re-expression (NOD2-OE) in NOD2-/- COPD mice. ( A ) Schematic of the in vivo experimental design. ( B ) qRT-PCR analysis of NOD2 mRNA expression in lung tissue. ( C ) Representative images of hematoxylin and eosin (H&E) staining of lung tissue sections. Scale bars: 200 μm in the upper panels and 100 μm in the enlarged lower panels. ( D ) Representative images of Masson’s trichrome staining for collagen deposition in lung tissue sections. Scale bar: 100 μm. ( E ) Quantification of total cells, neutrophils, and macrophages in bronchoalveolar lavage fluid (BALF), and ELISA measurements of TNF-α, IL-6, and CXCL1 levels in BALF supernatant. ( F ) Representative images of IHC staining for E-cadherin and N-cadherin in lung tissue. Scale bar: 50 μm. ( G ) Quantification of the average optical density (AOD) for E-cadherin and N-cadherin IHC staining. ( H ) qRT-PCR analysis of senescence-associated secretory phenotype (SASP) factors (CSF3, CCL3, and IL1α) in lung tissue. Data are presented as mean ± SD. Data were analyzed using one-way ANOVA followed by Tukey’s test. * p < 0.05 and ** p < 0.01 Control vs. wt-COPD groups; # p < 0.05 and ## p < 0.01 wt-COPD vs. NOD2 −/− -COPD groups; & p < 0.05 and && p < 0.01 NOD2 −/− -COPD + vector vs. NOD2 −/− -COPD + NOD2-OE groups

Article Snippet: Additionally, the commercial ELISA kits (Human and mouse MPO ELISA Kits, Cat. # CSB-E08721h, CSB-E08723m, CUSABIO, Wuhan, China; Human and mouse neutrophil elastase ELISA Kit, Cat. # CSB-EL007587HU, CSB-E04804m, CUSABIO) were applied to quantify the levels of MPO and neutrophil elastase released into cell culture supernatants as per the instructions of the manufacturer.

Techniques: In Vivo, Functional Assay, Knock-Out, Expressing, Quantitative RT-PCR, Analysis, Tissue, Staining, Enzyme-linked Immunosorbent Assay, Immunohistochemistry, Control, Plasmid Preparation

Journal: Biomarker Research

Article Title: ZDHHC5-mediated S-palmitoylation of NOD2 drives neutrophil extracellular trap formation via mitophagy to exacerbate inflammation in COPD

doi: 10.1186/s40364-026-01001-2

Figure Lengend Snippet: NOD2 promotes Neutrophil Extracellular Trap (NET) formation in the CS-induced COPD mouse model The association between NOD2, autophagy, and Neutrophil Extracellular Trap (NET) formation was investigated in the lung tissue of mice from the CS-induced COPD model. ( A) Representative immunofluorescence images showing co-localization of the neutrophil marker Ly6G (green) and the autophagy marker LC3B (red), with nuclei counterstained with DAPI (blue). Scale bars: 20 μm in the upper panels and 10 μm in the enlarged lower panels. ( B ) Representative immunofluorescence images showing staining for the NET markers citrullinated histone H3 (CitH3, green) and myeloperoxidase (MPO, red), with nuclei counterstained with DAPI (blue). Scale bars: 20 μm in the upper panels and 10 μm in the enlarged lower panels. ( C ) Quantification of the NET biomarkers CitH3-DNA and MPO-DNA complexes in serum and lung tissue homogenates by ELISA. ( D ) Immunoblotting analysis of the neutrophil marker Ly6G and the NET marker CitH3 in whole-lung tissue lysates. ( E ) Representative immunofluorescence images showing colocalization of the neutrophil marker Ly6G (red), the NET marker CitH3 (green), and NOD2 (pink) in lung tissue, with nuclei counterstained with DAPI (blue). Scale bars: 20 μm in the upper panels and 10 μm in the enlarged lower panels. Data are presented as mean ± SD. C and D were analyzed using one-way ANOVA followed by Tukey’s test. ** p < 0.01 Control vs. wt -COPD groups; and ## p < 0.01 wt -COPD vs. NOD2 −/− -COPD groups; && p < 0.01 NOD2 −/− -COPD + vector vs. NOD2 −/− -COPD + NOD2-OE groups

Article Snippet: Additionally, the commercial ELISA kits (Human and mouse MPO ELISA Kits, Cat. # CSB-E08721h, CSB-E08723m, CUSABIO, Wuhan, China; Human and mouse neutrophil elastase ELISA Kit, Cat. # CSB-EL007587HU, CSB-E04804m, CUSABIO) were applied to quantify the levels of MPO and neutrophil elastase released into cell culture supernatants as per the instructions of the manufacturer.

Techniques: Tissue, Immunofluorescence, Marker, Staining, Enzyme-linked Immunosorbent Assay, Western Blot, Analysis, Control, Plasmid Preparation

Journal: Biomarker Research

Article Title: ZDHHC5-mediated S-palmitoylation of NOD2 drives neutrophil extracellular trap formation via mitophagy to exacerbate inflammation in COPD

doi: 10.1186/s40364-026-01001-2

Figure Lengend Snippet: Neutrophil depletion minimizes genotype-dependent differences in CS-induced lung injury, inflammation, fibrosis, and NET formation The dependence of NOD2-associated COPD pathology on neutrophils was evaluated by depleting neutrophils in CS-exposed wild-type (wt) and NOD2 −/− mice. Mice were randomly assigned to the wt -COPD + IgG, wt -COPD + Anti-Ly6G, NOD2 −/− -COPD + IgG, and NOD2 −/− -COPD + Anti-Ly6G groups ( n = 6 per group). ( A ) Schematic of the experimental design. Mice were exposed to CS for 12 weeks and received 200 µg of anti-Ly6G antibody or an equivalent dose of isotype IgG intraperitoneally three times per week from week 2 until the end of the experiment. ( B ) Representative flow-cytometry plots and quantification of CD11b + Ly6G + neutrophils in lung tissue and peripheral blood. ( C ) Representative H&E-stained images of lung tissue sections. Scale bars: 200 μm in the upper panels and 100 μm in the enlarged lower panels. ( D ) Representative images of Masson’s trichrome staining for pulmonary collagen deposition. Scale bars: 100 μm. ( E ) Quantification of total cells, neutrophils, and macrophages in bronchoalveolar lavage fluid (BALF), together with ELISA measurements of TNF-α, IL-6, and CXCL1 levels in BALF supernatants. ( F ) Quantification of CitH3-DNA and MPO-DNA complexes in serum and lung tissue homogenates by ELISA. Data are presented as mean ± SD. Data were analyzed using two-way ANOVA followed by Tukey’s test. ** p < 0.01 vs. the wt -COPD + IgG group; ## p < 0.01 vs. the wt-COPD + Anti-Ly6G group; & p < 0.05, & p < 0.01 vs. the NOD2 −/− -COPD + IgG group

Article Snippet: Additionally, the commercial ELISA kits (Human and mouse MPO ELISA Kits, Cat. # CSB-E08721h, CSB-E08723m, CUSABIO, Wuhan, China; Human and mouse neutrophil elastase ELISA Kit, Cat. # CSB-EL007587HU, CSB-E04804m, CUSABIO) were applied to quantify the levels of MPO and neutrophil elastase released into cell culture supernatants as per the instructions of the manufacturer.

Techniques: Pathology, Flow Cytometry, Tissue, Staining, Enzyme-linked Immunosorbent Assay

Journal: Biomarker Research

Article Title: ZDHHC5-mediated S-palmitoylation of NOD2 drives neutrophil extracellular trap formation via mitophagy to exacerbate inflammation in COPD

doi: 10.1186/s40364-026-01001-2

Figure Lengend Snippet: Knockdown of NOD2 inhibits CSE-induced NET formation, ROS production, and mitophagy in human neutrophils The role of NOD2 in cigarette smoke extract (CSE)-induced Neutrophil Extracellular Trap (NET) formation, reactive oxygen species (ROS) accumulation, and lysosome–mitochondrion spatial relationships were evaluated in differentiated HL-60-derived neutrophil-like cells. ( A ) Schematic of the in vitro experimental design and treatment groups. ( B ) Representative DAPI-stained images of extracellular DNA structures and quantification of NET formation based on DAPI-stained nuclear morphology. Scale bars: 20 μm. ( C ) Extracellular DNA levels in cell culture supernatants determined using a PicoGreen dsDNA assay. ( D ) Myeloperoxidase (MPO) levels in cell culture supernatants determined by ELISA. ( E ) Neutrophil elastase (NE) levels in cell culture supernatants determined by ELISA. ( F ) Representative flow-cytometry histograms and quantitative analysis of intracellular ROS levels measured using DCFH-DA staining. ( G ) Representative immunofluorescence images showing the spatial relationship between LAMP1-positive lysosomes (green) and TOM20-positive mitochondria (red), with nuclei counterstained with DAPI (blue). The colocation efficients for LAMP1 and TOM20 was determined by ImageJ and shown on the right. Scale bars: 20 μm. Data are presented as mean ± SD. Quantitative data were analyzed using one-way ANOVA followed by Tukey’s test. ** p < 0.01 vs. Control group; ## p < 0.01 vs. CSE + sh-NC group

Article Snippet: Additionally, the commercial ELISA kits (Human and mouse MPO ELISA Kits, Cat. # CSB-E08721h, CSB-E08723m, CUSABIO, Wuhan, China; Human and mouse neutrophil elastase ELISA Kit, Cat. # CSB-EL007587HU, CSB-E04804m, CUSABIO) were applied to quantify the levels of MPO and neutrophil elastase released into cell culture supernatants as per the instructions of the manufacturer.

Techniques: Knockdown, Derivative Assay, In Vitro, Staining, Morphology, Cell Culture, Picogreen Assay, Enzyme-linked Immunosorbent Assay, Flow Cytometry, Analysis, Immunofluorescence, Control

Journal: Biomarker Research

Article Title: ZDHHC5-mediated S-palmitoylation of NOD2 drives neutrophil extracellular trap formation via mitophagy to exacerbate inflammation in COPD

doi: 10.1186/s40364-026-01001-2

Figure Lengend Snippet: ZDHHC5 overexpression partially counteracts the effects of NOD2 knockdown in human neutrophil-like cells The ability of ZDHHC5 overexpression to counteract the effects of NOD2 knockdown was evaluated in CSE-stimulated, differentiated HL-60-derived neutrophil-like cells. ( A ) Schematic of the in vitro experimental design and treatment groups. ( B ) Representative DAPI-stained images of extracellular DNA structures and quantification of NET formation based on DAPI-stained nuclear morphology. Scale bars: 20 μm. ( C ) Extracellular DNA levels in cell culture supernatants determined using a PicoGreen dsDNA assay. ( D ) MPO levels in cell culture supernatants determined by ELISA. ( E ) NE levels in cell culture supernatants determined by ELISA. ( F ) Representative flow cytometry histograms and quantitative analysis of intracellular ROS levels measured using DCFH-DA staining. ( G ) Representative immunofluorescence images showing LAMP1-positive lysosomes (green) and TOM20-positive mitochondria (red), with nuclei counterstained with DAPI (blue). The colocalization coefficients for LAMP1 and TOM20 were determined by ImageJ and shown on the right. Scale bars: 20 μm. Data are presented as mean ± SD. Data were analyzed using two-way ANOVA followed Tukey’s test. ** p < 0.01 versus the CSE + sh-NC + Vector group; ## p < 0.01 versus the CSE + sh-NOD2 + ZDHHC5 group

Article Snippet: Additionally, the commercial ELISA kits (Human and mouse MPO ELISA Kits, Cat. # CSB-E08721h, CSB-E08723m, CUSABIO, Wuhan, China; Human and mouse neutrophil elastase ELISA Kit, Cat. # CSB-EL007587HU, CSB-E04804m, CUSABIO) were applied to quantify the levels of MPO and neutrophil elastase released into cell culture supernatants as per the instructions of the manufacturer.

Techniques: Over Expression, Knockdown, Derivative Assay, In Vitro, Staining, Morphology, Cell Culture, Picogreen Assay, Enzyme-linked Immunosorbent Assay, Flow Cytometry, Analysis, Immunofluorescence, Plasmid Preparation

Journal: Biomarker Research

Article Title: ZDHHC5-mediated S-palmitoylation of NOD2 drives neutrophil extracellular trap formation via mitophagy to exacerbate inflammation in COPD

doi: 10.1186/s40364-026-01001-2

Figure Lengend Snippet: ZDHHC5 overexpression partially counteracts the protective effects of NOD2 knockdown in a mouse model of COPD The ability of ZDHHC5 overexpression to counteract the effects of NOD2 knockdown was evaluated in a cigarette smoke (CS)-induced mouse model of COPD. ( A ) Schematic of the in vivo experimental design. ( B ) qRT-PCR analysis of NOD2 and ZDHHC5 mRNA expression in lung tissue. ( C ) Representative images of H&E staining of lung sections. Scale bar: 100 μm. ( D ) Representative images of Masson’s trichrome staining for collagen deposition. Scale bar: 100 μm. ( E ) Quantification of inflammatory cells and TNF-α, IL-6, and CXCL1 levels in BALF. ( F ) Representative images of IHC staining for E-cadherin and N-cadherin. Scale bar: 50 μm. ( G ) Quantitative analysis of IHC staining. ( H ) Immunofluorescence images showing spatial overlap between Ly6G (green) and LC3B (red). Scale bars: 20 μm in the upper panels and 10 μm in the enlarged lower panels. ( I ) Immunofluorescence images showing NETs stained for CitH3 (green) and MPO (red). Scale bars: 20 μm in the upper panels and 10 μm in the enlarged lower panels. ( J ) Quantification of CitH3-DNA and MPO-DNA complexes in serum and lung tissue homogenates by ELISA. Data are presented as mean ± SD. Quantitative data were analyzed using two-way ANOVA followed by Tukey’s test. * p < 0.05, ** p < 0.01 vs. COPD + sh-NC + Vector group; # p < 0.05, ## p < 0.01 vs. COPD + sh-NOD2 + ZDHHC5 group

Article Snippet: Additionally, the commercial ELISA kits (Human and mouse MPO ELISA Kits, Cat. # CSB-E08721h, CSB-E08723m, CUSABIO, Wuhan, China; Human and mouse neutrophil elastase ELISA Kit, Cat. # CSB-EL007587HU, CSB-E04804m, CUSABIO) were applied to quantify the levels of MPO and neutrophil elastase released into cell culture supernatants as per the instructions of the manufacturer.

Techniques: Over Expression, Knockdown, In Vivo, Quantitative RT-PCR, Analysis, Expressing, Tissue, Staining, Immunohistochemistry, Immunofluorescence, Enzyme-linked Immunosorbent Assay, Plasmid Preparation

Journal: PCN Reports: Psychiatry and Clinical Neurosciences

Article Title: Acute trazodone treatment ameliorates behavioral abnormalities and improves subsequent short‐term memory in a lipopolysaccharide‐induced mouse model of acute brain dysfunction

doi: 10.1002/pcn5.70426

Figure Lengend Snippet: Plasma interleukin‐6 (IL‐6) concentrations at Days 1, 3, and 7. (a–c) Plasma IL‐6 concentrations at Days 1, 3, and 7, respectively. Plasma IL‐6 concentrations were measured by enzyme‐linked immunosorbent assay at Days 1, 3, and 7. Data are shown as box‐and‐whisker plots with individual data points ( n = 8 per group). Statistical analyses were performed separately at each time point using the Kruskal–Wallis test followed by the Steel–Dwass post hoc test. * p < 0.05, ** p < 0.01. LPS, lipopolysaccharide; Sal, saline; TZD, trazodone.

Article Snippet: Plasma interleukin‐6 (IL‐6) and tumor necrosis factor‐alpha (TNF‐α) concentrations were measured using commercial ELISA kits according to the manufacturers' instructions (Mouse IL‐6 ELISA Kit, KE10007; Mouse TNF‐α ELISA Kit, KE10002; Proteintech Group, Inc.).

Techniques: Clinical Proteomics, Enzyme-linked Immunosorbent Assay, Whisker Assay, Saline