Journal: Frontiers in Immunology

Article Title: Macrophage IL-1β turns meningeal fibroblasts into inflammatory amplifiers in pneumococcal infection

doi: 10.3389/fimmu.2026.1808185

Figure Lengend Snippet: Pneumococcal challenge in human CSF generated higher release of LDH, IL-6 and IL-8 by Ben-Men-1 cells. Ben-Men-1 cells were challenged with increasing concentrations of SP D39 (0.25/0.5/1/2x10 7 CFU/ml) while being held in human CSF from 8 different donors (n=3-6, control: MCM) for 24h. Negative control was pure bacterial THY growth medium. Supernatants were analyzed for LDH (positive control: triton 1%), IL-6 and IL-8 (A, C) . (B) = Summary of data from (A–C) . During pneumococcal challenge (2x10 7 CFU/ml, MCM, negative controls: THY medium, PBS), cells were treated with the following inhibitors: TLR-cocktail (25µg/ml anti-TLR2 antibody + 5µM TLR4 inhibitor + 20µg/ml chloroquine), 0.1-10 µM BAY 11-7082, 0.01-1µM 666-15, 0.01-1µM JQ1 or DMSO control. (D) Supernatants were analyzed for LDH and IL-6. Data are given as individual values as well as means ± SD.

Article Snippet: In addition, TLR signaling was assessed using an anti-TLR2 mAB T2.5 (25μg/ml, Hycult), TLR4 inhibitor TAK-242 (5μM, Calbiochem, USA) and chloroquine (20μg/ml, Sigma-Aldrich, USA) to inhibit endosomal TLRs ( ).

Techniques: Generated, Control, Negative Control, Positive Control

Journal: Frontiers in Immunology

Article Title: Macrophage IL-1β turns meningeal fibroblasts into inflammatory amplifiers in pneumococcal infection

doi: 10.3389/fimmu.2026.1808185

Figure Lengend Snippet: Primary human meningeal cells display partially PLY-dependent cell death and cytokine release upon pneumococcal challenge. HMC were challenged with P/S-lysed S. pneumoniae serotype 2 (D39), D39Δ ply , serotypes 6B and 7F (0.25/1/4x10 7 CFU/ml, negative control: THY bacterial medium without bacteria) for 24h. Supernatants were analyzed for LDH (C, D) and IL-6 release (B, E) After exposure to THY, D39 or D39Δ ply (4x10 7 CFU/ml), supernatants were pooled and analyzed with antibody array. (A) HMC were treated during pneumococcal challenge (4x10 7 CFU/ml) with the following inhibitors: TLR-cocktail (anti-TLR2 antibody T2.5 (25µg/ml) + TLR4 inhibitor TAK-242 (5µM) + endosomal TLR inhibitor chloroquine (20µg/ml)), NF-κB inhibitors parthenolide (0.5µM) and BAY 11-7082 (1-10 µM), AP-1 inhibitor SR11302 (1µM), BRD4 inhibitors I-BET (10µM) and JQ1 (1µM), CREB inhibitors KG-501 (10µM) and 666-15 (1µM), or DMSO control. (D, E) Data are given as individual values as well as means ± SD. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001. Statistical analysis was calculated with ANOVA with Tukey’s multiple comparison test.

Article Snippet: In addition, TLR signaling was assessed using an anti-TLR2 mAB T2.5 (25μg/ml, Hycult), TLR4 inhibitor TAK-242 (5μM, Calbiochem, USA) and chloroquine (20μg/ml, Sigma-Aldrich, USA) to inhibit endosomal TLRs ( ).

Techniques: Negative Control, Bacteria, Ab Array, Control, 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: 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: iScience

Article Title: cGAS/STING sensing in dendritic cells discriminates between daptomycin sensitive and resistant Staphylococcus aureus clinical isolates

doi: 10.1016/j.isci.2026.115854

Figure Lengend Snippet: TLR2 is required for MutuDC sensing of, but not internalization of MRSA (A) Relative pHrodo labeled MRSA internalization by MutuDC over 4 h following stimulation with DapS A8819 (light blue symbols) or DapR A8817 (dark blue symbols), or media alone (white squares). Prior to stimulation, MutuDC were pre-treated for 1 h with TLR2 blocking antibody (clone T2.5; triangles with dashed lines) or media alone (circles with filled lines). Relative MRSA internalization by each DC subset is expressed as the gMFI of pHrodo. Results show the mean and (SD) of duplicates from one experiment, representative of two independent experiments. (B) Cytokine secretion (pg/mL) by MutuDC stimulated with TLR2 ligand peptidoglycan of S. aureus (PGN-SA) (10 μg/mL) or (C) DapS (A8819; light blue) or DapR (A8817; dark blue) MRSA (MOI of 10) for 18 h. MutuDC were first pre-treated with either TLR2 blocking antibody (dot-filled bars) or media alone (filled bars) as in A, or an isotype control (clone 163D3, empty bars) at 1 μg/mL. Results pooled from four (B) or three (C) independent experiments and expressed as the mean ± SEM, with each symbol (circle, square, and directional triangles) representing paired experimental replicates ( n = 3). Statistical significance determined using paired t test and reported as indicated by an ∗ when p ≤ 0.05. (D) Expression of surface activation markers by MutuDC stimulated with DapS A8819 MRSA. DC were pre-treated with TLR2 blocking antibody (black trace), isotype control (dashed red trace), and media alone (light blue shaded). Unstained control sample is shown for each marker (black dashed trace). Data shown from one experiment, representative of three independent experiments.

Article Snippet: mAB mTLR2- anti-mouse/human TLR2 , InvivoGen , Cat# mab-mtlr2; RRID: AB_763722.

Techniques: Labeling, Blocking Assay, Control, Expressing, Activation Assay, Marker, FACS

Journal: iScience

Article Title: cGAS/STING sensing in dendritic cells discriminates between daptomycin sensitive and resistant Staphylococcus aureus clinical isolates

doi: 10.1016/j.isci.2026.115854

Figure Lengend Snippet:

Article Snippet: mAB mTLR2- anti-mouse/human TLR2 , InvivoGen , Cat# mab-mtlr2; RRID: AB_763722.

Techniques: Purification, Cell-Signaling, Enzyme-linked Immunosorbent Assay, Western Blot, Virus, Recombinant, Transfection, Derivative Assay, Expressing, Clone Assay, Plasmid Preparation, Software