Journal: Journal of ethnopharmacology

Article Title: Effect of Echinacea purpurea (L.) Moench and its extracts on the immunization outcome of avian influenza vaccine in broilers.

doi: 10.1016/j.jep.2023.117306

Figure Lengend Snippet: Fig. 4. EP and EE affects the immune gene expression in jejunum: a) relative TLR4 mRNA expression, b) relative MyD88 mRNA expression, c) relative TPAF6 mRNA expression, d) relative AP-1 mRNA expression. All data were presented as mean ± SD(n = 6). *p < 0.05,**p < 0.01,***p < 0,001 compared to the group C; #p < 0.05,##p < 0.01,###p < 0,001 compared to the group V and intergroup.

Article Snippet: Name Company Cat. no. Dilution TLR4 Rabbit pAb Bioss bs-20379R 1:1000 MyD88 Rabbit pAb Novus Biologicals NB100-56698 1:1000 ERK1/2Rabbit pAb Abmart T55487 1:1000 JNK1/2/3Rabbit pAb Abmart T40073 1:1000 TRAF6Rabbit pAb ABclonal A16991 1:1000 GAPDHRabbit pAb ABclonal AC001 1:9000 Goat anti-Rabbit IgG Zenbio H&L 1:5000 X. Wang et al. Journal of Ethnopharmacology 319 (2024) 117306 On day 7 after vaccination, the FCR of the VE and VEE groups was significantly lower compared to the groups C and V (P < 0.05).

Techniques: Gene Expression, Expressing

Journal: Journal of ethnopharmacology

Article Title: Effect of Echinacea purpurea (L.) Moench and its extracts on the immunization outcome of avian influenza vaccine in broilers.

doi: 10.1016/j.jep.2023.117306

Figure Lengend Snippet: Fig. 5. Effects of EP and EE on the relative expression of jejunal proteins in broilers after immunization: a), b), c) is Western blot detects protein expression bands on days 7,21,35; d), e), f), g), h) is the level of the protein expression: TLR4,MyD88,TPAF6,ERK,JNK. All data were presented as mean ± SD(n = 6). *p < 0.05, **p < 0.01,***p < 0,001 compared to the group C; #p < 0.05,##p < 0.01,###p < 0,001 compared to the group V and intergroup.

Article Snippet: Name Company Cat. no. Dilution TLR4 Rabbit pAb Bioss bs-20379R 1:1000 MyD88 Rabbit pAb Novus Biologicals NB100-56698 1:1000 ERK1/2Rabbit pAb Abmart T55487 1:1000 JNK1/2/3Rabbit pAb Abmart T40073 1:1000 TRAF6Rabbit pAb ABclonal A16991 1:1000 GAPDHRabbit pAb ABclonal AC001 1:9000 Goat anti-Rabbit IgG Zenbio H&L 1:5000 X. Wang et al. Journal of Ethnopharmacology 319 (2024) 117306 On day 7 after vaccination, the FCR of the VE and VEE groups was significantly lower compared to the groups C and V (P < 0.05).

Techniques: Expressing, Western Blot

Journal: Theranostics

Article Title: Membrane TLR9 Positive Neutrophil Mediated MPLA Protects Against Fatal Bacterial Sepsis

doi: 10.7150/thno.37139

Figure Lengend Snippet: The effect of MPLA mediated inflammatory preconditioning (InP) on cav-1 -/- mice. (A) Survival curves, n=8 mice per group. The mice were first given 0.1×10 8 CFUs E. coli , followed by the lethal dose of E. coli (3×10 8 CFUs) 2 h later,* vs. Saline. (B) Experimental procedures. (1) The WT mice were injected i.p. with 0.1×10 8 CFUs E. coli, and 2 h later were injected with 3×10 8 CFUs E. coli per mouse, and all 8 mice survived (InP). (2) C av-1 -/ - mice were injected i.p. with 0.1×10 8 E. coli, and 2 h later were injected with 3×10 8 CFUs E. coli, and all 8 mice died. (C) Intestine and liver tissues of mice stained with H&E. (D) ELISA of TNF-α/IL-6 in the supernatants of peripheral blood neutrophils in mice stimulated with 0.1×10 8 CFUs E. coli for 2 h. ** P <0.01 (Student's t test). (E) Western blot shows that InP did not promote translocation of TLR9 from cytosol to cell membrane in cav -/- mice. (F) Western blot shows significant reduction of endogenous Cav-1 by shRNA against Cav-1 (Cav-1 shRNA). (G) Immunoblot of TLR9, MyD88, TRAF3 and IRF3 in HL60 cells transfected Cav-1 shRNA and then stimulated with 0.1×10 8 CFUs E. coli for indicated times. Similar results were obtained in three independent experiments. (H) ELISA of TNF-α/IL-6 in the supernatants of HL60 cells transfected Cav-1 shRNA, stimulated with 0.1×10 8 CFUs E. coli for 2 h. ** P <0.01 (Student's t test).

Article Snippet: MyD88 homodimerization inhibitor peptide (NBP2-29328) was purchased from Novus Biologicals (Novus, USA).

Techniques: Saline, Injection, Staining, Enzyme-linked Immunosorbent Assay, Western Blot, Translocation Assay, Membrane, shRNA, Transfection

Journal: Theranostics

Article Title: Membrane TLR9 Positive Neutrophil Mediated MPLA Protects Against Fatal Bacterial Sepsis

doi: 10.7150/thno.37139

Figure Lengend Snippet: Expression of TLR9-Cav-1 signaling proteins in the neutrophils of patients with sepsis. (A) Membrane TLR9 expression. (B) Cav-1 expression. (C) ROC curve for mTLR9. P <0.05. (D) ROC curve for Cav-1. P <0.05. (E) Association of Cav-1 with surface TLR9 expression in neutrophils. r2 =0.5791. (F) MyD88 expression. (G) TRAF3 expression. (H) IRF3 expression.

Article Snippet: MyD88 homodimerization inhibitor peptide (NBP2-29328) was purchased from Novus Biologicals (Novus, USA).

Techniques: Expressing, Membrane

Journal: Oncogene

Article Title: Cancer cell-derived long pentraxin 3 (PTX3) promotes melanoma migration through a toll-like receptor 4 (TLR4)/NF-κB signaling pathway.

doi: 10.1038/s41388-019-0848-9

Figure Lengend Snippet: Fig. 8 TLR4/MYD88 expression is required for PTX3- induced melanoma cell migration. a Transwell migration assays were performed on 1205Lu cells transfected with siCTRL or two different siTLR4. Immunoblot analysis shows TLR4 depletion. HSP60, loading control. Bar graphs represent mean of migrated cells ± s.e.m (n = 3 independent experiments). b 1205Lu cells were transfected with siPTX3 or siTLR4. After 3 days, cells were stimulated or not with rhPTX3 for 4 h and Transwell migration assays were performed. Bar graphs show migration mean ± s.e.m (n = 2 independent experiments). c Transwell migration assays on control and MYD88-depleted 1205Lu cells. Bar graphs show migration mean ± s.e.m (n = 2 independent experiments). MYD88 and TWIST1 levels were controlled by immunoblot. d 1205Lu cells were treated with the IKK inhibitor BMS345541 and PTX3 expression was assessed by immunoblot. e Schematic model of PTX3 autocrine action on melanoma cell migration. ***P < 0.001, **P < 0.01 as determined by two-way ANOVA and Bonferroni post-tests

Article Snippet: MITF (sc-35934), MYD88 (sc-35986), and TWIST1 (sc-38604) siRNAs were from Santa Cruz Biotechnology.

Techniques: Expressing, Migration, Transfection, Western Blot, Control

Journal: Molecular Pharmacology

Article Title: TCL1A Single-Nucleotide Polymorphisms and Estrogen-Mediated Toll-Like Receptor-MYD88–Dependent Nuclear Factor- κ B Activation: Single-Nucleotide Polymorphism– and Selective Estrogen Receptor Modulator–Dependent Modification of Inflammation and Immune Response

doi: 10.1124/mol.117.108340

Figure Lengend Snippet: TLR intracellular localization and signaling. TLR3, TLR7, TLR8, and TLR9 are located on the membranes of intracellular compartments, such as the endoplasmic reticulum and endosomes. The myeloid differentiation primary response gene 88 (MYD88) is the one of the functional adapter molecules that have been reported to interact with all TLRs except TLR3. MYD88 recruits IL-IRAK1, IRAK2, IRAK4, and tumor necrosis factor–receptor associated factor 6 (TRAF6), leading ultimately to NF-κB activation and proinflammatory cytokine secretion and an inflammatory response. UNC93B1 is a multitransmembrane-domain-containing protein and plays a critical role in trafficking TLR7 and TLR9 from the endoplasmic reticulum to endosomes, where TLR7 and TLR9 transmit signals via MYD88/TRIF-dependent pathways (Kim et al., 2008).

Article Snippet: The membranes were incubated overnight with primary antibodies: TCL1A, TLR2, TLR7, TLR9, TLR10, MYD88, UNC93B1 (Novus Biologicals, Littleton, CO), and ACTB at a 1:500 dilution at 4°C.

Techniques: Functional Assay, Activation Assay

Journal: Molecular Pharmacology

Article Title: TCL1A Single-Nucleotide Polymorphisms and Estrogen-Mediated Toll-Like Receptor-MYD88–Dependent Nuclear Factor- κ B Activation: Single-Nucleotide Polymorphism– and Selective Estrogen Receptor Modulator–Dependent Modification of Inflammation and Immune Response

doi: 10.1124/mol.117.108340

Figure Lengend Snippet: SNP- and estrogen-dependent mRNA expression of TCL1A (A), TLR2 (B), TLR7 (C), TLR9 (D), TLR10 (E), MYD88 (F), and UNC93B1 (G) in LCLs (H). Western blot analysis was performed for TCL1A, TLR2, TLR7, TLR9 TLR10, MYD88, UNC93B1, and ACTB in LCLs with known TCL1A SNP genotypes. The cells were treated with 0.1 nM E2 or with 0.1 nM E2 plus 10−7µM 4-hydroxytamoxifen (4OH-TAM) for an additional 24 hours. Eight cell lines homozygous for the variant (V) genotypes for all three of the TCL1A SNPs and eight cell lines homozygous for WT genotypes were used in these experiments. ***P < 0.0001.

Article Snippet: The membranes were incubated overnight with primary antibodies: TCL1A, TLR2, TLR7, TLR9, TLR10, MYD88, UNC93B1 (Novus Biologicals, Littleton, CO), and ACTB at a 1:500 dilution at 4°C.

Techniques: Expressing, Western Blot, Variant Assay

Journal: Molecular Pharmacology

Article Title: TCL1A Single-Nucleotide Polymorphisms and Estrogen-Mediated Toll-Like Receptor-MYD88–Dependent Nuclear Factor- κ B Activation: Single-Nucleotide Polymorphism– and Selective Estrogen Receptor Modulator–Dependent Modification of Inflammation and Immune Response

doi: 10.1124/mol.117.108340

Figure Lengend Snippet: Correlations of TCL1A mRNA expression and those of toll-like receptors and MYDBB in the Human Variation Panel of 300 LCLs

Article Snippet: The membranes were incubated overnight with primary antibodies: TCL1A, TLR2, TLR7, TLR9, TLR10, MYD88, UNC93B1 (Novus Biologicals, Littleton, CO), and ACTB at a 1:500 dilution at 4°C.

Techniques: Expressing

Journal: Molecular Pharmacology

Article Title: TCL1A Single-Nucleotide Polymorphisms and Estrogen-Mediated Toll-Like Receptor-MYD88–Dependent Nuclear Factor- κ B Activation: Single-Nucleotide Polymorphism– and Selective Estrogen Receptor Modulator–Dependent Modification of Inflammation and Immune Response

doi: 10.1124/mol.117.108340

Figure Lengend Snippet: Correlations of MYD88 mRNA expression with those of toll-like receptors and UNC93B1 in the Human Variation Panel of 300 LCLs

Article Snippet: The membranes were incubated overnight with primary antibodies: TCL1A, TLR2, TLR7, TLR9, TLR10, MYD88, UNC93B1 (Novus Biologicals, Littleton, CO), and ACTB at a 1:500 dilution at 4°C.

Techniques: Expressing

Journal: Molecular Pharmacology

Article Title: TCL1A Single-Nucleotide Polymorphisms and Estrogen-Mediated Toll-Like Receptor-MYD88–Dependent Nuclear Factor- κ B Activation: Single-Nucleotide Polymorphism– and Selective Estrogen Receptor Modulator–Dependent Modification of Inflammation and Immune Response

doi: 10.1124/mol.117.108340

Figure Lengend Snippet: TCL1A could modulate TLR2, TLR7, TLR9, TLR10, MYD88, and UNC93B1 expression in LCLs. Relative mRNA expression (A and B) of TCL1A, TLR2, TLR7, TLR9, TLR10, MYD88, and UNC93B1 after knockdown of TCL1A in LCLs with known TCL1A SNP genotypes using pooled siRNA. Eight cell lines of each genotype were used in these experiments. *P < 0.05. Student’s t test was performed to compare gene expression in LCLs with differing TCL1A SNP genotypes before and after gene knockdown, *P value ≤ 0.05 was considered statistically significant. All values are mean ±S.E.M for three separate independent assays. Protein expression was determined by Western blot analysis (C). Relative mRNA expression (D and E) of TCL1A, TLR2, TLR7, TLR9, TLR10, MYD88, and UNC93B1 after knockdown of MYD88 in LCLs with known TCL1A SNP genotypes using pooled siRNA. (F) Western blot analysis was performed for TCL1A, TLR2, TLR7, TLR9, TLR10, MYD88, and UNC93B1 after knockdown of MYD88.

Article Snippet: The membranes were incubated overnight with primary antibodies: TCL1A, TLR2, TLR7, TLR9, TLR10, MYD88, UNC93B1 (Novus Biologicals, Littleton, CO), and ACTB at a 1:500 dilution at 4°C.

Techniques: Expressing, Knockdown, Gene Expression, Western Blot

Journal: Molecular Pharmacology

Article Title: TCL1A Single-Nucleotide Polymorphisms and Estrogen-Mediated Toll-Like Receptor-MYD88–Dependent Nuclear Factor- κ B Activation: Single-Nucleotide Polymorphism– and Selective Estrogen Receptor Modulator–Dependent Modification of Inflammation and Immune Response

doi: 10.1124/mol.117.108340

Figure Lengend Snippet: TCL1A SNP- and estrogen-dependent NF-κB activation as determined by NF-κB reporter assays could be altered by the knockdown or inhibition of MYD88. ER blockade by fulvestrant (ICI) or 4-hydroxytamoxifen (4OH) treatment resulted in TCL1A SNP -dependent NF-κB activation (A and B). TCL1A SNP and estrogen-dependent NF-κB activation could be blocked by MYD88 siRNA knockdown (C) or by exposure to MYD88 inhibitory peptide (100 µM) (D). Specifically, LCLs were cotransfected with an NF-κB reporter construct and siRNA (MYD88 or control siRNA); 24 hours after transfection, cells were treated with either vehicle or 0.1 nM E2 for 24 hours, followed by 10−7µM 4OH or ICI for an additional 24 hours. In some experiments, cells were exposed to MYD88 inhibitory peptide for 24 hours before E2 treatment. Luciferase activity was measured 72 hours after transfection. The firefly luciferase activity derived from the NF-κB responsive reporter was normalized by the use of Renilla luciferase activity as a control to correct for possible variation in transfection efficiency. All experiments were repeated three times in triplicate. **P < 0.001. (E) Coimmunoprecipitation was used to determine whether TCL1A protein could interact with MYD88 in LCLs. Whole-cell lysates from 1 × 107 LCLs were immunoprecipitated with anti-TCL1A (1:50) antibodies or anti-IgG antibodies. Whole-cell lysate (input, left panel) and immunoprecipitated samples (middle and right panels) were immunoblotted and probed with antibodies against TCL1A and MYD88.

Article Snippet: The membranes were incubated overnight with primary antibodies: TCL1A, TLR2, TLR7, TLR9, TLR10, MYD88, UNC93B1 (Novus Biologicals, Littleton, CO), and ACTB at a 1:500 dilution at 4°C.

Techniques: Activation Assay, Knockdown, Inhibition, Construct, Control, Transfection, Luciferase, Activity Assay, Derivative Assay, Immunoprecipitation

Journal: International Journal of Molecular Sciences

Article Title: Uncovering the Potential Mechanisms of Ergothioneine in Neuroinflammation Through Network Pharmacology, Molecular Docking, Molecular Dynamics Simulation, and In Vitro Validation

doi: 10.3390/ijms27052179

Figure Lengend Snippet: Representative images of Western blotting of EGT (0.1, 0.5, 1.0 mM) influencing protein expression of TLR4, p-NF-κB/NF-κB, and MyD88. ( A ) Western blotting of TLR4, p-NF-κB/NF-κB, and MyD88. ( B ) Quantification of ratio of p-NF-κB/NF-κB. ( C ) Quantification of ratio of MyD88/β-actin. ( D ) Quantification of ratio of TLR4/β-actin. Data are presented as mean ± SEM (n = 3). Values with different letters differ significantly ( p < 0.05).

Article Snippet: The primary antibodies include β-actin (1:5000; Proteintech; Cat. No. 66009-1-lg), NF-κB p65 (1:1000; Proteintech; Cat. No. 10268-1-AP), p-NF-κB p65 (Ser536) (1:1000; CST; Cat. No. 3033), MyD88 (23230-1-AP), TLR4 (19811-1-AP), PI3 Kinase (PI3K) (1:5000; Proteintech; Cat. No. 60225-1-AP), p-PI3K (1:1000; CST; Cat. No. 4228), AKT (1:2000; Proteintech; Cat. No. 10176-1-AP), p-AKT (Ser473) (1:1000; CST; Cat. No. 8326), mTOR (1:5000; Proteintech; Cat. No. 66888-1-AP), and p-mTOR (Ser2448) (1:1000; CST; Cat. No. 5536).

Techniques: Western Blot, Expressing