Journal: Frontiers in Immunology

Article Title: B-type natriuretic peptide attenuates TLR-induced cytokine and chemokine secretion in monocyte-derived Langerhans cells

doi: 10.3389/fimmu.2026.1791284

Figure Lengend Snippet: The effects of BNP on activated moLCs differentiation and activation markers. (A) Representative density plot of the main markers of moLCs, CD1a and CD207 expression measured by flow cytometry. Monocytes were cultured in the presence of GM-CSF, TNF-α, and TGF-β for 5 days supplemented with IL-4 for the first 48 hrs to differentiate moLCs, and treated with 10 nM BNP throughout the entire differentiation process. On day 4 moLCs were activated with CL075 and poly(I:C) and a combination of both for 24 hrs. (B) A percentage of positive CD207 and CD1a moLCs. N = 6 (C) Histograms of moLCs activation markers, CD83, CD86, and HLA-DQ. (D) A percentage of positive CD83, CD86 and HLA-DQ moLCs. Symbols with different colors represent individual donors. Data are presented as individual values with mean ± SEM. Statistical significance was determined by One-way ANOVA followed by Tukey’s post-hoc test. Homogeneity of variances was confirmed by the Brown-Forsythe test. n=6 biological replicates per group. *P<0.05, **P<0.01, *** P<0.001, **** P< 0.0001. BNP, B-type natriuretic peptide; CL075, TLR7/8 agonist, thiazoquinoline compound; CTRL, Control; Poly(I:C), TLR3 agonist, polyinosinic:polycytidylic acid.

Article Snippet: MoLCs were activated with 0.5 μg/ml CL075 (TLR7/8 agonist) and 20 μg/ml poly(I:C) (TLR3 agonist) and with their combination at day 4 for 24 hours (both were obtained from Invivogen, San Diego, CA, USA).

Techniques: Activation Assay, Expressing, Flow Cytometry, Cell Culture, Control

Journal: Frontiers in Immunology

Article Title: B-type natriuretic peptide attenuates TLR-induced cytokine and chemokine secretion in monocyte-derived Langerhans cells

doi: 10.3389/fimmu.2026.1791284

Figure Lengend Snippet: BNP inhibited the TLR activated moLCs IL-6, TNF-α, IL-12, IL-10 and IFN-β cytokine production. Monocytes were cultured in the presence of GM-CSF, TNF-α, and TGF-β for 5 days supplemented with IL-4 for the first 48 hrs to differentiate moLCs, and treated with 10 nM BNP throughout the entire differentiation process. On day 4 moLCs were activated with CL075 and poly(I:C) and a combination of both for 24 hrs. Supernatants were collected on day 5 from the cells. (A–F) Levels of different pro-inflammatory cytokines IL-6 and IL-8, TNF-α, cytokines during viral response IL-12 and IFN-β, and an anti-inflammatory IL-10, were measured by ELISA. Symbols with different colors represent individual donors, lines marks mean expression levels. Data is presented as Mean ± SEM. Two-way ANOVA followed by Tukey’s multiple comparison test. Normality of residuals was confirmed by QQ plot analysis. n=6–10 biological replicates per group *P<0.05, **P<0.01, *** P<0.001, **** P< 0.0001. BNP, B-type natriuretic peptide; CL075; TLR7/8 agonist, thiazoquinoline compound; CTRL, Control; moLC, monocyte-derived Langerhans cells; Poly(I:C), TLR3 agonist, polyinosinic:polycytidylic acid.

Article Snippet: MoLCs were activated with 0.5 μg/ml CL075 (TLR7/8 agonist) and 20 μg/ml poly(I:C) (TLR3 agonist) and with their combination at day 4 for 24 hours (both were obtained from Invivogen, San Diego, CA, USA).

Techniques: Cell Culture, Enzyme-linked Immunosorbent Assay, Expressing, Comparison, Control, Derivative Assay

Journal: Frontiers in Immunology

Article Title: B-type natriuretic peptide attenuates TLR-induced cytokine and chemokine secretion in monocyte-derived Langerhans cells

doi: 10.3389/fimmu.2026.1791284

Figure Lengend Snippet: BNP reduced the T cell proliferation induction ability in Poly(I:C) and CL075 activated moLCs. (A) Proliferating T cells were measured with flow cytometry by the CFSE intensity at day 5 of coculture. Monocytes were cultured in the presence of GM-CSF, TNF-α, and TGF-β for 5 days, supplemented with IL-4 for the first 48 hrs to differentiate moLCs, and treated with 10 nM BNP throughout the differentiation process. The moLCs were activated for 24 hrs with CL075, and poly(I:C), and a combination of both. Naїve CD4 + T cells were isolated with CD4 + magnetic beads and made a coculture with fully differentiated moLCs for 5 days. n=6 (B) Representative histograms of CFSE intensity from a coculture of T cells and moLCs. (C) Representative Western blot of IDO1 expression of moLCs. Cells were treated with 10 nM BNP and activated for 24 hrs in the presence of CL075 and poly(I:C) at day 4. Symbols with different colors represent individual donors. Data are presented as individual values with mean ± SEM. Statistical significance was determined by One-way ANOVA followed by Tukey’s post-hoc test. Homogeneity of variances was confirmed by the Brown-Forsythe test. n=2 biological replicates per group *P<0.05, **P<0.01, *** P<0.001, **** P< 0.0001. BNP, B-type natriuretic peptide; CFSE, Carboxyfluorescein succinimidyl ester; CL075; TLR7/8 agonist, thiazoquinoline compound; CTRL, Control; IDO1, Indoleamine 2,3-dioxygenase; Poly(I:C), TLR3 agonist, polyinosinic:polycytidylic acid.

Article Snippet: MoLCs were activated with 0.5 μg/ml CL075 (TLR7/8 agonist) and 20 μg/ml poly(I:C) (TLR3 agonist) and with their combination at day 4 for 24 hours (both were obtained from Invivogen, San Diego, CA, USA).

Techniques: Flow Cytometry, Cell Culture, Isolation, Magnetic Beads, Western Blot, Expressing, Control

Journal: Frontiers in Immunology

Article Title: B-type natriuretic peptide attenuates TLR-induced cytokine and chemokine secretion in monocyte-derived Langerhans cells

doi: 10.3389/fimmu.2026.1791284

Figure Lengend Snippet: RNA-Seq analysis showed an upregulation of migratory genes and the downregulation of inflammatory cytokines and chemokines in activated moLCs. Monocytes were cultured in the presence of GM-CSF, TNF-α, and TGF-β for 5 days supplemented with IL-4 for the first 48 hrs to differentiate moLCs, and treated with 10 nM BNP throughout the differentiation process. The moLCs were activated for 24 hrs with CL075, and poly(I:C), and a combination of both. [( A–C) , left] Volcano plot analysis of poly(I:C) and CL075 and BNP treated moLCs compared to vehicle-treated controls or each other. The top 15 most significantly changed gene symbols are highlighted. [ (A–C , middle, right] GO annotation of the biological process of up- and down-regulated genes between the groups marked. The size of the circle corresponds to the number of induced genes, while the color of the line corresponds to the -log10 of the False Discovery Rate (FDR). (A) Poly(I:C) and CL075 vs. CTRL (B) BNP, Poly(I:C) and CL075 vs. CTRL (C) BNP, Poly(I:C) and CL075 vs. Poly(I:C) and CL075. BNP, B-type natriuretic peptide; CL075; TLR7/8 agonist, thiazoquinoline compound; CTRL, Control; GO, gene ontology; Poly(I:C), TLR3 agonist, polyinosinic:polycytidylic acid.

Article Snippet: MoLCs were activated with 0.5 μg/ml CL075 (TLR7/8 agonist) and 20 μg/ml poly(I:C) (TLR3 agonist) and with their combination at day 4 for 24 hours (both were obtained from Invivogen, San Diego, CA, USA).

Techniques: RNA Sequencing, Cell Culture, Control

Journal: Frontiers in Immunology

Article Title: B-type natriuretic peptide attenuates TLR-induced cytokine and chemokine secretion in monocyte-derived Langerhans cells

doi: 10.3389/fimmu.2026.1791284

Figure Lengend Snippet: BNP treatment increased moLCs migration toward CCL21 and reduced chemokine production in activated moLCs. Monocytes were cultured in the presence of GMCSF, TNF-α, and TGF-β for 5 days supplemented with IL-4 for the first 48 hrs to differentiate moLCs, and treated with 10 nM BNP throughout the entire differentiation process. On day 4, moLCs were activated with CL075 and poly(I:C) and a combination of both for 24 hrs. (A) In Transwell migration assay, 1*10 6 cells were applied in the upper well of the Transwell plate, and in the bottom of the plate, it contains RPMI supplemented with CCL19 and CCL21. Migration of the cells was measured with flow cytometry. Data are presented as individual values with mean ± SEM. Statistical significance was determined by Two-way ANOVA followed by Tukey’s multiple comparison test. Normality of residuals was confirmed by QQ plot analysis. n=8 biological replicates per group (B) A chemokine array was performed using supernatants from BNP treated and TLR activated cells. Symbols with different colors represent individual donors. Data is presented as Mean ± SEM. One-way ANOVA with Tukey’s multiple comparison test was used for statistical analysis. n=2 biological replicates per group *P<0.05, **P<0.01, *** P<0.001, **** P< 0.0001. BNP, B-type natriuretic peptide; CL075; TLR7/8 agonist, thiazoquinoline compound; CTRL, Control; moLC, monocyte-derived Langerhans cells; Poly(I:C), TLR3 agonist, polyinosinic:polycytidylic acid.

Article Snippet: MoLCs were activated with 0.5 μg/ml CL075 (TLR7/8 agonist) and 20 μg/ml poly(I:C) (TLR3 agonist) and with their combination at day 4 for 24 hours (both were obtained from Invivogen, San Diego, CA, USA).

Techniques: Migration, Cell Culture, Transwell Migration Assay, Flow Cytometry, Comparison, Control, Derivative Assay

Journal: Frontiers in Immunology

Article Title: B-type natriuretic peptide attenuates TLR-induced cytokine and chemokine secretion in monocyte-derived Langerhans cells

doi: 10.3389/fimmu.2026.1791284

Figure Lengend Snippet: BNP treated moLCs supernatant inhibited NK cell migration. Monocytes were cultured in the presence of GM-CSF, TNF-α, and TGF-β for 5 days supplemented with IL-4 for the first 48 hrs to differentiate moLCs, and treated with 10 nM BNP throughout the entire differentiation process. On day 4 moLCs were activated with CL075 and poly(I:C) and a combination of both for 24 hrs. Transwell plates were used to perform the migration experiments, and cell numbers were determined by flow cytometry. (A) A flow cytometry gating strategy was used to distinguish between the PBL and monocyte populations based on the FSC-H and SSC-H. In the histograms, CD3 + cells represent the T cell population. CD3 - cells selected to determine the CD56 + population, which represented the NK cells. (B–F) Flow cytometry was used to measure the number of migrating cells in the lower wells of the Transwell plate. (B) PBL, (C) CD3 + , (D) CD3 - , CD56 + , (E) CD3 - , CD56 - , (F) monocytes. Symbols represent individual donors. Data are presented as individual values with mean ± SEM. Statistical significance was determined by Two-way ANOVA followed by Tukey’s multiple comparison test. Normality of residuals was confirmed by QQ plot analysis. n=10–12 biological replicates per group *P<0.05, **P<0.01, **** P< 0.0001. BNP, B-type natriuretic peptide; CL075; TLR7/8 agonist, thiazoquinoline compound; CTRL, Control; moLC, monocyte-derived Langerhans cells; NK cells, Natural Killer cells; Poly(I:C), TLR3 agonist, polyinosinic:polycytidylic acid.

Article Snippet: MoLCs were activated with 0.5 μg/ml CL075 (TLR7/8 agonist) and 20 μg/ml poly(I:C) (TLR3 agonist) and with their combination at day 4 for 24 hours (both were obtained from Invivogen, San Diego, CA, USA).

Techniques: Migration, Cell Culture, Flow Cytometry, Comparison, Control, Derivative Assay

Journal: The Journal of Allergy and Clinical Immunology

Article Title: Toll-like receptor 8 agonist nanoparticles mimic immunomodulating effects of the live BCG vaccine and enhance neonatal innate and adaptive immune responses

doi: 10.1016/j.jaci.2016.12.985

Figure Lengend Snippet: CL075:Ag85Bp25-PS and BCG induce comparable Ag85B-specific CD4 + effector T cells in the spleens of neonatal huTLR8Tg mice. A, Experimental design: 7-day-old huTLR8Tg and matched littermate WT mice were immunized subcutaneously and killed 14 days later before harvesting spleens and staining splenocytes with a control versus Ag85B 280–294 MHCII tetramer and for surface marker expression. B, Representative flow cytometric plots showing Tet + T cells as CD44 + Neg Tet + or CD44 + Ag85B Tet + cells gated on live CD4 + splenic lymphocytes of BCG- or CL075-Ag85Bp25-PS–vaccinated mice 2 weeks after immunization. C and D, Similar to BCG, CL075-Ag85Bp25-PS induced a significant increase in CD44 + Ag85B Tet + cells compared with CD44 + Neg Tet + cells. Results represent means ± SEMs (n = 14-15), with significance relative to unspecific control tetramer. E, Neonatal humanized TLR8 mice have greater numbers of splenic Ag85B-specific CD4 + T cells versus their WT littermates. The unpaired Mann-Whitney test was applied, and statistical significance was denoted as follows: ** P < .01 and *** P < .001. NS , Not significant.

Article Snippet: Mice were immunized subcutaneously at the scruff of the neck with either 0.2 × 10 6 colony-forming units of BCG (Organon Teknika/Merck, Durham, NC) or CL075:Ag85Bp25-PS containing approximately 8 to 10 μg of Ag85B/p25 (Biomatik, Cambridge, Ontario, Canada) and approximately 0.1 mg/kg CL075 in a total volume of 25 μL.

Techniques: Staining, Marker, Expressing, MANN-WHITNEY