Journal: eBioMedicine

Article Title: CNPY3 variants associated with a recessive developmental and epileptic encephalopathy affect inflammasome and TLR function

doi: 10.1016/j.ebiom.2026.106506

Figure Lengend Snippet: Functional characterisation of CNPY3 variants. ( A ) Heterologous CNPY3 variant protein expression in HEK293T cells. The indicated HA-tagged CNPY3 constructs were transiently expressed in HEK293T and visualised by western blotting and immunofluorescent detection with the indicated antibodies. A long exposure is shown to illustrate low level expression of two variants. All tested CNPY3 variants except CNPY3 N183X give rise to protein products. A merged image of CNPY3 and HA-tag immunodetections revealed the presence of endogenous CNPY3 in HEK293T cells and the slightly lower mobility of the heterologous variant proteins caused by the appended HA-tag. The CNPY3 Q210∗ protein was detectable by standard enhanced chemiluminescence detection as shown in the lower panel (marked by arrowhead). See main text for details. ( B ) Live cell imaging of THP-1 cell lines expressing CNPY3 wild type or variants. CNPY3 KO cells stably expressing the indicated CNPY3-mCherry variant fusion proteins were stained with a live-cell ER marker and imaged alive by confocal fluorescence microscopy. CNPY3 Q210∗ could not be visualised as the expression construct retained the premature stop codon to exclude any effects of the C-terminally fused mCherry on the variant function. Cells shown are representative of ≥75% of all cells in the conditions imaged. The perimeter of the cell is traced in the mCherry panels for clarity. Magnification bars: 5 μm. ( C ) Immunocytochemical images of THP-1, CNPY3 KO , CNPY3 S156Y and CNPY3 Q210∗ cells. Slides were stained against CNPY3 and the endoplasmic reticulum marker Reticulon 4 (ER). Nuclei are silhouetted by a dotted line and marked with N. Magnification bars: 5 μm. ( D ) Wild type THP-1, CNPY3 KO and stable add-back cell lines expressing the indicated CNPY3 variants were stained for TLR2 or with an isotype control antibody and processed by flow cytometry. The dotted line marks the threshold gate applied for quantification shown in ( E ). ( E ) Quantification of TLR2 surface levels shown in panel C. One-way ANOVA–Šidak's multiple comparisons (∗∗∗∗ P ≤ 0.0001; ns, P > 0.05; n = 3–6). ( F ) Biotinylation assay for analysis of TLR2 cell surface expression. The indicated THP-1 cell lines were treated with a membrane-impermeable biotinylation reagent. Cells were lysed, biotinylated proteins collected on streptavidin beads and samples were processed by western blotting for TLR2. ( G ) Electrophoretic mobility shifts document distinct TLR2 glycosylation patterns in CNPY3 variant cells. Cell extracts from the same THP-1 cell lines as before were separated on a gradient SDS-PAGE with high resolving power to highlight mobility shifts of glycosylated TLR2 species. FLT3 was visualised as negative control. For unclear reasons, the CNPY3 Q210∗ variant was not detectable under these conditions, see text for details. CG, complex glycosylated species; DG, deglycosylated form; HM, high-mannose form. ( H ) Quantification of TLR2 glycosylation in CNPY3 variant lines, expressed as the (CG + HM)/DG ratio. One-way ANOVA–Šidak's multiple comparisons (∗∗∗∗ P ≤ 0.0001, ∗ P < 0.05; n = 2–4). ( I , J ) CNPY3 regulates TLR2 signalling and inflammasome activation. Engineered THP-1 cell lines were challenged with the TLR2 agonist Pam3CSK4 and the inflammasome activator nigericin individually or sequentially as indicated followed by determination of TNF ( I ) or IL-1β ( J ) secretion by ELISA. The same variant/column colour code was used throughout all figure panels. One-way ANOVA–Šidak's multiple comparisons (∗∗∗ P ≤ 0.001, ∗∗∗∗ P ≤ 0.0001; ns, P > 0.05; n = 6).

Article Snippet: Antibodies for Western blot: CNPY3 (Proteintech, Cat#15215-1-AP, RRID: AB_11182172 ), TLR2 (Proteintech, Cat#66645, RRID: AB_3084262 ), FLT3 clone C-20 (Santa Cruz Biotech), HA-tag (Biolegend Cat#901513, RRID: AB_2565335 ).

Techniques: Functional Assay, Variant Assay, Expressing, Construct, Western Blot, Live Cell Imaging, Stable Transfection, Staining, Marker, Confocal, Fluorescence, Microscopy, Control, Flow Cytometry, Biotinylation, Analysis, Membrane, Glycoproteomics, SDS Page, Negative Control, Activation Assay, Enzyme-linked Immunosorbent Assay

Journal: Journal of Extracellular Vesicles

Article Title: Helicobacter pylori Outer Membrane Vesicles Engage TLR2 Through NapA to Promote Epithelial Association and Inflammatory Signalling

doi: 10.1002/jev2.70376

Figure Lengend Snippet: Lack of functional TLR2 heterodimers and NF‐κB responsiveness in gastric epithelial cells. (A) Schematic representation of the experimental workflow used to assess NF‐κB–dependent luminescent reporter activity in gastric epithelial cells, including stimulation in 96‐well plates followed by luminescence readout. (B, C) AGS–NF‐κB RE and GES‐1–NF‐κB RE reporter cell pools were treated for 24 h with increasing concentrations (5–100 µg/mL) of outer membrane vesicles (OMVs) isolated from H. pylori strains 26695 or 60190. Tumour necrosis factor (TNF; 20 ng/mL, 24 h) was included as a positive control. NF‐κB activation was quantified by luciferase assay and expressed as relative light units (RLU) relative to unstimulated control cells. (D, E) AGS–NF‐κB RE and GES‐1–NF‐κB RE reporter cells were stimulated with the synthetic TLR2 agonists Pam2CSK4 (PAM2) and Pam3CSK4 (PAM3) (0.01–10 µg/mL), which selectively activate TLR2/6 and TLR2/1 heterodimers, respectively. NF‐κB activation was quantified by luciferase assay and expressed as RLU relative to unstimulated control cells. (F) Western blot analysis of TLR2, TLR1, TLR6 and TLR4 expression in a panel of gastric epithelial cell lines. Protein lysates from HEK293 cells transiently transfected with pDuo‐TLR2/1 or pDuo‐TLR2/6 plasmids were included as positive controls. The lower–molecular–weight TLR1‐reactive species, consistent with a non‐glycosylated form, is indicated by a red arrow. A representative blot is shown. Data are presented as mean ± SD ( n = 3).

Article Snippet: To validate the responsiveness of the reporter clones, TLR2/1–NF‐κB cells were stimulated with Pam3CSK4 (InvivoGen) and CU‐T12‐9 (Tocris), and TLR2/6–NF‐κB cells were stimulated with Pam2CSK4 and FSL‐1 (InvivoGen).

Techniques: Functional Assay, Activity Assay, 96 Well Plates, Membrane, Isolation, Positive Control, Activation Assay, Luciferase, Control, Western Blot, Analysis, Expressing, Transfection, Molecular Weight

Journal: Journal of Extracellular Vesicles

Article Title: Helicobacter pylori Outer Membrane Vesicles Engage TLR2 Through NapA to Promote Epithelial Association and Inflammatory Signalling

doi: 10.1002/jev2.70376

Figure Lengend Snippet: Functional reconstitution of TLR2 heterodimers enables NF‐κB activation by synthetic TLR2 agonists in gastric epithelial cells. (A, D) GES‐1–NF‐κB RE (A) and AGS–NF‐κB RE (D) reporter cells transfected with the empty control plasmid (pDuo‐MCS) were stimulated with synthetic TLR2 agonists (Pam2CSK4, PAM2, 10 µg/mL; Pam3CSK4, PAM3, 10 µg/mL; CU‐T12‐9, 10 µM; or FSL‐1, 10 µg/mL) or with Escherichia coli LPS (1 µg/mL) and analyzed for NF‐κB–dependent reporter activity. Tumour necrosis factor (TNF; 20 ng/mL) was included as a positive control. (B, E) GES‐1–NF‐κB RE (B) and AGS–NF‐κB RE (E) reporter cells constitutively expressing the TLR2/6 heterodimer were stimulated with the indicated TLR2 agonists, E. coli LPS, or TNF. (C, F) GES‐1–NF‐κB RE (C) and AGS–NF‐κB RE (F) reporter cells constitutively expressing the TLR2/1 heterodimer were stimulated with the indicated agonists under the same conditions. NF‐κB activation was quantified by luciferase assay and expressed as relative light units (RLU) relative to unstimulated control cells. Bars represent the mean ± SD ( n = 3).

Article Snippet: To validate the responsiveness of the reporter clones, TLR2/1–NF‐κB cells were stimulated with Pam3CSK4 (InvivoGen) and CU‐T12‐9 (Tocris), and TLR2/6–NF‐κB cells were stimulated with Pam2CSK4 and FSL‐1 (InvivoGen).

Techniques: Functional Assay, Activation Assay, Transfection, Control, Plasmid Preparation, Activity Assay, Positive Control, Expressing, Luciferase

Journal: Journal of Extracellular Vesicles

Article Title: Helicobacter pylori Outer Membrane Vesicles Engage TLR2 Through NapA to Promote Epithelial Association and Inflammatory Signalling

doi: 10.1002/jev2.70376

Figure Lengend Snippet: Functional validation of TLR2/1‐ and TLR2/6‐specific NF‐κB reporter cells using heterodimer‐selective agonists and purified LPS. (A, B, D, E) GES‐1 (A, B) and AGS (D, E) NF‐κB reporter cells stably expressing either the TLR2/6 or TLR2/1 heterodimer were stimulated for 5 h with the TLR2/6‐selective agonists FSL‐1 or Pam2CSK4 (Pam2), or with the TLR2/1‐selective agonists CU‐T12‐9 or Pam3CSK4 (Pam3), over the indicated concentration ranges (1–1,000 ng/mL for peptide agonists and 1–1,000 nM for CU‐T12‐9). (C, F) GES‐1 (C) and AGS (F) reporter cells expressing either TLR2/1 or TLR2/6 were stimulated for 5 h with increasing concentrations of purified E. coli LPS (1–1,000 ng/mL; 0.14–140 nM Kdo equivalents). (G) Representative colorimetric Kdo assay used for LPS quantification. (H) Standard curve generated with commercially available 2‐keto‐3‐deoxyoctonate (Kdo) and used for normalization of purified LPS preparations. (I) AGS NF‐κB reporter cells expressing either the TLR2/1 or TLR2/6 heterodimer were stimulated for 5 h with purified H. pylori strain 26695 LPS normalized according to Kdo content (0.14–1,400 nM Kdo equivalents). NF‐κB‐dependent luminescent reporter activity was measured after stimulation and expressed as relative light units (RLUs) normalized to untreated controls. Bars represent the mean ± SD of three independent experiments. Statistical significance is indicated as * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001.

Article Snippet: To validate the responsiveness of the reporter clones, TLR2/1–NF‐κB cells were stimulated with Pam3CSK4 (InvivoGen) and CU‐T12‐9 (Tocris), and TLR2/6–NF‐κB cells were stimulated with Pam2CSK4 and FSL‐1 (InvivoGen).

Techniques: Functional Assay, Biomarker Discovery, Purification, Stable Transfection, Expressing, Concentration Assay, Standard Curve, Generated, Activity Assay

Journal: iScience

Article Title: Hippuric acid contributes to NK-cell activation with dependence on TLR2 in chronic hepatitis B complicated by MASLD

doi: 10.1016/j.isci.2026.117362

Figure Lengend Snippet: TLR2-expressing NK cells exhibit an activated phenotype associated with metabolic parameters and liver stiffness in CHB-MASLD (A) Heatmap: DEGs identified from exploratory transcriptomic profiling of sorted NK cells from patients with CHB ( n = 4) and CHB-MASLD ( n = 3), including TLR2. (B) Frequency of TLR2 + NK cells in peripheral blood from patients with CHB ( n = 12) and CHB-MASLD ( n = 12). (C) TLR2 expression in NK/T-like cells, T cells, and B cells from patients with CHB ( n = 5) and CHB-MASLD ( n = 5) with complete staining data. (D and E) Phenotypic comparison between TLR2 − and TLR2 + NK-cell subsets: (D) CD226 expression and (E) PD-1 expression. (F and G) Intracellular cytokine production in TLR2 − and TLR2 + NK cells: (F) IFN-γ and (G) TNF-α. (D–G) were generated from patients with CHB-MASLD ( n = 12). (H–K) Correlation analyses between the frequency of TLR2 + NK cells and metabolic or liver-related clinical parameters: (H) TG, (I) HDL-C, (J) GLU, and (K) LSM ( n = 24 participants in total). TLR2-focused analyses were restricted to samples with complete TLR2 panel measurements, and correlation analyses were performed in the participants with complete paired clinical and flow cytometry data. Data are presented as mean ± SD. Statistical comparisons were performed using student’s t test or Mann-Whitney U test. Correlations were analyzed using Pearson correlation analysis. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001.

Article Snippet: Anti-TLR2 neutralizing antibody (MAb-mTLR2, clone T2.5) , InvivoGen , Cat# mab2-mtlr2.

Techniques: Expressing, Staining, Comparison, Generated, Flow Cytometry, MANN-WHITNEY

Journal: iScience

Article Title: Hippuric acid contributes to NK-cell activation with dependence on TLR2 in chronic hepatitis B complicated by MASLD

doi: 10.1016/j.isci.2026.117362

Figure Lengend Snippet: Metabolic-stress modeling increases TLR2 expression and NK-cell activation-related features in vitro (A) Schematic overview of the in vitro metabolic stress model. PBMCs were treated with PA and/or high glucose (HG) to model in vitro metabolic-stress conditions associated with CHB-MASLD. BSA-matched vehicle controls were used for PA stimulation, and mannitol was used as the osmotic control for high-glucose stimulation. Flow cytometry was performed to assess NK-cell activation and function ( n = 10). (B) Flow cytometry analysis of TLR2 expression in NK cells following PA stimulation alone or in combination with HG (PAO). (C–F) Comparison of CD226, PD-1, and IFN-γ expression in NK cells: (C) following PA stimulation; (D) following PAO stimulation; (E) between TLR2 − and TLR2 + NK-cell subsets after PA treatment; and (F) between TLR2 − and TLR2 + NK-cell subsets after PAO treatment. In vitro metabolic-stress experiments were performed using PBMCs from an independent experimental subset of 10 donors ( n = 10), as indicated in the figure labels. Each data point represents one independent donor sample. Technical replicate wells, where applicable, were averaged before statistical analysis. Data are presented as mean ± SD. Paired comparisons were performed using the paired student’s t test. ∗ p < 0.05,∗∗ p < 0.01,∗∗∗ p < 0.001,∗∗∗∗ p < 0.0001.

Article Snippet: Anti-TLR2 neutralizing antibody (MAb-mTLR2, clone T2.5) , InvivoGen , Cat# mab2-mtlr2.

Techniques: Expressing, Activation Assay, In Vitro, Control, Flow Cytometry, Comparison

Journal: iScience

Article Title: Hippuric acid contributes to NK-cell activation with dependence on TLR2 in chronic hepatitis B complicated by MASLD

doi: 10.1016/j.isci.2026.117362

Figure Lengend Snippet: Hippuric acid is associated with partially TLR2-dependent NK-cell activation and increased LX-2 activation-related readouts (A) Pearson correlation between normalized plasma hippuric acid intensity and the proportion of NK cells among PBMCs in participants with complete paired measurements ( n = 24). The line represents the simple linear regression fit. (B and C) Dose-dependent effects of hippuric acid stimulation at 0, 5, and 10 μM on NK-cell activation in PBMC cultures from 5 independent donors ( n = 5): (B) frequency of TLR2 + NK cells and (C) percentage of IFN-γ-producing NK cells. Measurements were paired within donors. (D) Effect of hippuric acid stimulation on the frequencies of TLR2 + NK cells, CD226 + NK cells, and IFN-γ + NK cells in PBMC cultures from 15 independent donors ( n = 15). (E) Paired comparison of CD226 expression and IFN-γ production between TLR2 − and TLR2 + NK-cell subsets from 10 independent donors ( n = 10). (F) PBMCs were pretreated with a TLR2-neutralizing antibody or an isotype control before hippuric acid stimulation. TLR2 blockade reduced the frequencies of TLR2 + NK cells, CD226 + NK cells, and IFN-γ + NK cells in PBMC cultures from 6 independent donors ( n = 6). (G) FACS-sorted NK cells were treated with a TLR2-neutralizing antibody or an isotype control, followed by hippuric acid stimulation. TLR2 blockade reduced TLR2 expression, CD226 expression, and IFN-γ production. Experiments were performed using cells from 5 independent donors ( n = 5), with two technical replicate wells per condition. (H) Baseline and direct-exposure controls in the NK-LX-2 co-culture system. LX-2 cells were co-cultured with untreated CHB-derived NK cells or hippuric acid-treated NK cells, or were directly exposed to hippuric acid alone. α-SMA expression was quantified by flow cytometry. (I) Representative histogram and quantification of α-SMA expression in LX-2 cells after co-culture with control NK cells, hippuric acid-treated NK cells, or hippuric acid-treated NK cells subjected to TLR2 blockade. (J and K) Supernatant analysis of fibrogenic markers in the NK-LX-2 co-culture system: (J) hyaluronic acid (HA) and (K) procollagen III N-terminal peptide (PIIINP). Both markers were increased after hippuric acid stimulation and partially reduced following TLR2 blockade. (H–K) were generated using the same experimental cohort of NK cells from 5 independent donors ( n = 5). PBMC stimulation assays, sorted NK-cell assays, and NK-LX-2 co-culture experiments were performed in independent experimental subsets. Unless otherwise specified, n represents the number of independent participants or donors. Technical replicate measurements were averaged to generate one value per biological sample and were not treated as independent observations. Data are presented as mean ± SD or median with interquartile range, as appropriate. Paired two-condition comparisons were performed using the paired student’s t test or Wilcoxon matched-pairs signed-rank test, as appropriate. Comparisons across three or more matched conditions were performed using one-way repeated-measures ANOVA with appropriate multiple-comparison testing or the Friedman test, as appropriate. ns, not significant; ∗ p < 0.05; ∗∗ p < 0.01; ∗∗∗ p < 0.001; ∗∗∗∗ p < 0.0001.

Article Snippet: Anti-TLR2 neutralizing antibody (MAb-mTLR2, clone T2.5) , InvivoGen , Cat# mab2-mtlr2.

Techniques: Activation Assay, Clinical Proteomics, Comparison, Expressing, Control, Co-Culture Assay, Cell Culture, Derivative Assay, Flow Cytometry, Generated

Journal: iScience

Article Title: Hippuric acid contributes to NK-cell activation with dependence on TLR2 in chronic hepatitis B complicated by MASLD

doi: 10.1016/j.isci.2026.117362

Figure Lengend Snippet:

Article Snippet: Anti-TLR2 neutralizing antibody (MAb-mTLR2, clone T2.5) , InvivoGen , Cat# mab2-mtlr2.

Techniques: Control, Recombinant, Software

Journal: Frontiers in Microbiology

Article Title: Integrated dual transcriptome sequencing and experimental validation reveal potential mechanisms of baicalin against pneumocystis pneumonia in immunosuppressed rats

doi: 10.3389/fmicb.2026.1923465

Figure Lengend Snippet: Validation and identification of core therapeutic targets through transcriptomic integration. (A) Venn diagram of overlapping targets. Eleven candidate targets were identified at the intersection of four gene sets: drug-disease targets from network pharmacology ( n = 93), and three sets of differentially expressed genes (|log₂FC| > 1.2, p < 0.05) from RNA-Seq comparisons (PCP vs. control, Ba-treated vs. control, PCP vs. Ba-treated). (B, C) PPI network of candidate targets. (B) Initial PPI network of the 11 genes constructed using STRING (confidence > 0.4). Two genes (Cftr, Prcp) were disconnected and excluded. (C) Final PPI network consisting of nine interconnected core proteins (Xdh, Nfe2l2, Cyp1a1, Tlr8, Tlr2, Ctss, Lgals3, Npc1, Ctsd), representing key mediators of Ba′s anti-PCP effects. (D) Venn diagram of candidate targets identified under stringent criteria. The diagram illustrates the intersection of four gene sets: differentially expressed genes (|log₂FC| > 2, p < 0.05) from three RNA-Seq comparisons (PCP vs. control, Ba-treated vs. control, PCP vs. Ba-treated) and Ba′s predicted targets from the SwissTargetPrediction database. This intersection highlights high-confidence targets likely involved in Ba′s anti-PCP mechanism. PPI, protein–protein interaction, Ba, baicalin; PCP, Pneumocystis pneumonia.

Article Snippet: Subsequently, the sections were blocked with goat serum for 30 min at 37 °C and incubated with primary antibodies targeting nuclear factor erythroid 2-related factor 2 (Nrf2, Cat No. 16396-1-AP; dilution 1:300; Proteintech, Wuhan, China), indoleamine 2,3-dioxygenase 1 (IDO1, Cat No. DF6502; dilution 1:100; Affinity Biosciences, Wuhan, China), toll-like receptor (TLR4, Cat No. bs-1021R; dilution 1:200; Bioss Biosciences, Beijing, China), and TLR2 (Cat No. A11225; dilution 1:100; ABclonal Biotechnology, Wuhan, China) overnight at 4 °C.

Techniques: Biomarker Discovery, RNA Sequencing, Control, Construct

Journal: Frontiers in Microbiology

Article Title: Integrated dual transcriptome sequencing and experimental validation reveal potential mechanisms of baicalin against pneumocystis pneumonia in immunosuppressed rats

doi: 10.3389/fmicb.2026.1923465

Figure Lengend Snippet: The anti-PCP effect of Ba activates the protein expression of TLR4 and TLR2 in lung tissue. (A,B) IHC assay data showed the expression levels and distribution of TLR4/2 protein in pulmonary tissue derived from various groups (magnification, ×200; scale bar = 50 μm) and (C,D) corresponding statistical results. Ba, baicalin; PCP, Pneumocystis pneumonia; TLR, Toll-like receptor. ** p < 0.01, n = 7/group.

Article Snippet: Subsequently, the sections were blocked with goat serum for 30 min at 37 °C and incubated with primary antibodies targeting nuclear factor erythroid 2-related factor 2 (Nrf2, Cat No. 16396-1-AP; dilution 1:300; Proteintech, Wuhan, China), indoleamine 2,3-dioxygenase 1 (IDO1, Cat No. DF6502; dilution 1:100; Affinity Biosciences, Wuhan, China), toll-like receptor (TLR4, Cat No. bs-1021R; dilution 1:200; Bioss Biosciences, Beijing, China), and TLR2 (Cat No. A11225; dilution 1:100; ABclonal Biotechnology, Wuhan, China) overnight at 4 °C.

Techniques: Expressing, Derivative Assay

Journal: Frontiers in Microbiology

Article Title: Integrated dual transcriptome sequencing and experimental validation reveal potential mechanisms of baicalin against pneumocystis pneumonia in immunosuppressed rats

doi: 10.3389/fmicb.2026.1923465

Figure Lengend Snippet: PPI interaction network analysis of the 14 DEGs detected in our studies including IDO1, Nrf2, chemokines (CCL2, CCL9, CXCR3, CXCL9, CXCL10, CXCL11)—oxidative stress biomarkers (SOD, GSH-Px), key members in the TLR signaling pathway (TLR2 and TLR4) and matrix metalloproteinases (MMPs, MMP2 and MMP9) using the String online database (A) and corresponding final PPI network (B) . IDO1, indoleamine 2,3 dioxygenase 1; Nrf2, nuclear factor erythroid 2-related factor 2; CCL2, chemokine (C-C motif) ligand 2 (also known as monocytic chemotactic protein 1, MCP-1); CCL9, chemokine (C-C motif) ligand 9; CXCR3, C-X-C motif receptor 3; CXCL9, C-X-C motif ligand 9; CXCL10, C-X-C motif ligand 10; CXCL11, C-X-C motif ligand 11; SOD, superoxide dismutase; GSH-Px, glutathione peroxidase; TLR, toll-like receptor; matrix metalloproteinase-2; MMP9, matrix metalloproteinase-9.

Article Snippet: Subsequently, the sections were blocked with goat serum for 30 min at 37 °C and incubated with primary antibodies targeting nuclear factor erythroid 2-related factor 2 (Nrf2, Cat No. 16396-1-AP; dilution 1:300; Proteintech, Wuhan, China), indoleamine 2,3-dioxygenase 1 (IDO1, Cat No. DF6502; dilution 1:100; Affinity Biosciences, Wuhan, China), toll-like receptor (TLR4, Cat No. bs-1021R; dilution 1:200; Bioss Biosciences, Beijing, China), and TLR2 (Cat No. A11225; dilution 1:100; ABclonal Biotechnology, Wuhan, China) overnight at 4 °C.

Techniques:

Journal: Frontiers in Microbiology

Article Title: Integrated dual transcriptome sequencing and experimental validation reveal potential mechanisms of baicalin against pneumocystis pneumonia in immunosuppressed rats

doi: 10.3389/fmicb.2026.1923465

Figure Lengend Snippet: Molecular docking results of Ba into the protein crystal structures of Nrf2, IDO1, TLR2, TLR4, and PcRtt109 . IDO1, Indoleamine 2,3 Dioxygenase 1; Nrf2, Nuclear Factor Erythroid 2-Related Factor 2; PcRtt109, Pneumocystis carinii Rtt 109 ; IDO1, indoleamine 2,3 dioxygenase 1; TLR, toll-like receptor; Ba, baicalin.

Article Snippet: Subsequently, the sections were blocked with goat serum for 30 min at 37 °C and incubated with primary antibodies targeting nuclear factor erythroid 2-related factor 2 (Nrf2, Cat No. 16396-1-AP; dilution 1:300; Proteintech, Wuhan, China), indoleamine 2,3-dioxygenase 1 (IDO1, Cat No. DF6502; dilution 1:100; Affinity Biosciences, Wuhan, China), toll-like receptor (TLR4, Cat No. bs-1021R; dilution 1:200; Bioss Biosciences, Beijing, China), and TLR2 (Cat No. A11225; dilution 1:100; ABclonal Biotechnology, Wuhan, China) overnight at 4 °C.

Techniques: Docking Assay

Journal: Journal of Inflammation Research

Article Title: Nasal Streptococcus pyogenes Infection Exacerbates Psoriasiform Relapse in Mice Through TLR2-Dependent Dendritic Cell Metabolic Reprogramming and γδ T Cell Activation

doi: 10.2147/JIR.S596821

Figure Lengend Snippet: Streptococcal LTA promotes TLR2-dependent metabolic reprogramming and hyper-migration in BMDCs. ( A ) qPCR analysis of pro-inflammatory, metabolic (HK2, LDHA), and motility-related (MYPT1) genes in BMDCs stimulated with LTA (100 ng/mL) ± TLR2 inhibitor (TL2-C29). ( B ) Real-time metabolic profiles showing OCR and ECAR. ( C ) Representative images and quantification of the scratch wound healing assay. Red dashed lines indicate the wound edge at the endpoint (24 h), marking the position to which cells have migrated. Data are mean ± SEM; error bars indicate standard error of the mean. Statistical significance was determined by one-way ANOVA. Data are representative of five independent experiments; * P < 0.05, ** P < 0.01, **** P < 0.0001 vs Ctrl; # P < 0.05, ## P < 0.01, ### P < 0.001, #### P < 0.0001 vs LTA.

Article Snippet: Stimulation: BMDCs were pretreated with 100 ng/mL purified S. pyogenes lipoteichoic acid (LTA; Merk, NO. L3140-5MG) for 6 hours as, with or without an incubation with the TLR2 inhibitor TL2-C29 (25 μM; InvivoGen, Cat.code: inh-c29).

Techniques: Migration, Wound Healing Assay

Journal: Journal of Inflammation Research

Article Title: Nasal Streptococcus pyogenes Infection Exacerbates Psoriasiform Relapse in Mice Through TLR2-Dependent Dendritic Cell Metabolic Reprogramming and γδ T Cell Activation

doi: 10.2147/JIR.S596821

Figure Lengend Snippet: TLR2 ligation activates the canonical NF-κB pathway and actomyosin contractility in DCs. Representative Western blots and densitometric quantification of p-P65, p-IKBα, NOS2, TLR2, p-MYPT1, and p-MLC in BMDCs following LTA treatment ± TL2-C29. Tubulin or β-actin served as the loading control. Data are mean ± SEM from n = 3 independent experiments. Statistical significance was determined by one-way ANOVA; * P < 0.05, ** P < 0.01vs Ctrl; # P < 0.05, ## P < 0.01vs LTA.

Article Snippet: Stimulation: BMDCs were pretreated with 100 ng/mL purified S. pyogenes lipoteichoic acid (LTA; Merk, NO. L3140-5MG) for 6 hours as, with or without an incubation with the TLR2 inhibitor TL2-C29 (25 μM; InvivoGen, Cat.code: inh-c29).

Techniques: Ligation, Western Blot, Control

Journal: Journal of Inflammation Research

Article Title: Nasal Streptococcus pyogenes Infection Exacerbates Psoriasiform Relapse in Mice Through TLR2-Dependent Dendritic Cell Metabolic Reprogramming and γδ T Cell Activation

doi: 10.2147/JIR.S596821

Figure Lengend Snippet: Mechanistic model of infection-triggered psoriasis relapse. Schematic representation of the “nasal-skin” immune axis: Nasal SP infection → TLR2-driven DC glycolytic reprogramming → DC migration to skin → Conjugation with γδ T cells → IL-17A-driven relapse. Blue arrows indicate downregulation, and Orange arrows indicate upregulation following SP intervention.

Article Snippet: Stimulation: BMDCs were pretreated with 100 ng/mL purified S. pyogenes lipoteichoic acid (LTA; Merk, NO. L3140-5MG) for 6 hours as, with or without an incubation with the TLR2 inhibitor TL2-C29 (25 μM; InvivoGen, Cat.code: inh-c29).

Techniques: Infection, Migration, Conjugation Assay