Journal: Cardiovascular research

Article Title: Non-enterobacterial endotoxins stimulate human coronary artery but not venous endothelial cell activation via Toll-like receptor 2.

doi: 10.1016/j.cardiores.2006.11.004

Figure Lengend Snippet: Fig. 3. Resting HCAECs but not HUVECs express TLR2. RT-PCR was performed on RNA from unstimulated HUVECs and HCAECs using primers specific for β-actin, TLR1, 2, 4, or 6 or CD14. Numbers beneath images show abundance of each transcript relative to β-actin as determined by real-time PCR. Dash indicates not detected within 40 cycles. (A). TLR2 mRNA expression was also measured in HUVECs cultured in medium alone, 1 μg/ml E. coli LPS, 10 ng/ml IFN-γ or LPS and IFN-γ combined for 18 h (B). Western blot for TLR2, TLR4, TLR6 and loading control GAPDH was performed on 10 μg of whole cell lysate from the TLR-deficient cell line HEK-293, HEK-293 cells transfected with TLR2 or TLR4/MD2, HUVECs, HCAECs or the human monocytic cell-line THP-1 (C).

Article Snippet: Western blot was performed on 10 μg of whole cell lysates obtained from unchallenged HEK-293, HEK293 cells transfected with TLR2 or TLR4/MD2, HUVEC, HCAEC and THP-1 cells using antibodies to TLR2 (sc10739, Santa-Cruz, US), TLR4 (sc10741, Santa-Cruz), TLR6 (3653, Prosci, CA, US) or loading control GAPDH (sc25778, Santa-Cruz).

Techniques: Reverse Transcription Polymerase Chain Reaction, Real-time Polymerase Chain Reaction, Expressing, Cell Culture, Western Blot, Control, Transfection

Journal: Cardiovascular research

Article Title: Non-enterobacterial endotoxins stimulate human coronary artery but not venous endothelial cell activation via Toll-like receptor 2.

doi: 10.1016/j.cardiores.2006.11.004

Figure Lengend Snippet: Fig. 4. NE-LPSs but not E. coli LPS stimulate TLR2-dependent signalling. TLR-deficient HEK-293 cells were transfected with CD14 and reporter construct alone (A), or with additional TLR4/MD2 (B) or TLR2 (C), then challenged with 0.1 to 1000 ng/ml of each type of LPS for 18 h. Data are expressed as fold induction of NF-κB dependent reporter expression normalised to internal transfection efficiency control vs cells cultured in medium alone, and are representative of at least four experiments. Ec = E. coli, Bf = B. fragilis, Pg = P. gingivalis, Pa = P. aeruginosa. (D) Phenol re-extraction removes all TLR2-stimulatory activity from a crude E. coli LPS preparation and neither TLR1 nor TLR6 is sufficient or required for responsiveness to NE-LPSs. HEK-293 cells were transfected with TLR1 and TLR6, TLR2 only, or the combination of TLRs 1, 2 and 6 together. Cells were then challenged with 1 μg/ml of crude E. coli LPS, phenol re-extracted E. coli LPS, or each of the NE-LPSs for 18 h. ⁎⁎Pb0.01 vs cells cultured in medium alone.

Article Snippet: Western blot was performed on 10 μg of whole cell lysates obtained from unchallenged HEK-293, HEK293 cells transfected with TLR2 or TLR4/MD2, HUVEC, HCAEC and THP-1 cells using antibodies to TLR2 (sc10739, Santa-Cruz, US), TLR4 (sc10741, Santa-Cruz), TLR6 (3653, Prosci, CA, US) or loading control GAPDH (sc25778, Santa-Cruz).

Techniques: Transfection, Construct, Expressing, Control, Cell Culture, Extraction, Activity Assay

Journal: Cardiovascular research

Article Title: Non-enterobacterial endotoxins stimulate human coronary artery but not venous endothelial cell activation via Toll-like receptor 2.

doi: 10.1016/j.cardiores.2006.11.004

Figure Lengend Snippet: Fig. 5. Whole non-enterobacterial organisms and lipid-As can stimulate TLR2-dependent signalling. TLR-deficient HEK-293 cells were transfected with CD14, or with additional TLR2 or TLR4/MD2 co-expression, as described in the legend of Fig. 4. Cells were then challenged with 107 whole heat-killed bacteria per ml (A), or 1 μg/ml of lipid-A (LA) of each organism, for 18 h (B). Responses to 10 ng/ml E. coli or P. gingivalis LPS are included as positive controls for TLR4 and TLR2 signalling respectively. Data are expressed as mean fold induction of NF-κB dependent reporter expression normalised to internal transfection efficiency control vs cells cultured in medium alone, and are representative of three experiments. Ec = E. coli, Bf = B. fragilis, Pg = P. gingivalis, Pa = P. aeruginosa. ⁎Pb0.05 vs medium alone.

Article Snippet: Western blot was performed on 10 μg of whole cell lysates obtained from unchallenged HEK-293, HEK293 cells transfected with TLR2 or TLR4/MD2, HUVEC, HCAEC and THP-1 cells using antibodies to TLR2 (sc10739, Santa-Cruz, US), TLR4 (sc10741, Santa-Cruz), TLR6 (3653, Prosci, CA, US) or loading control GAPDH (sc25778, Santa-Cruz).

Techniques: Transfection, Expressing, Bacteria, Control, Cell Culture

Journal: Cardiovascular research

Article Title: Non-enterobacterial endotoxins stimulate human coronary artery but not venous endothelial cell activation via Toll-like receptor 2.

doi: 10.1016/j.cardiores.2006.11.004

Figure Lengend Snippet: Fig. 6. TLR2-neutralizing antibody blunts HCAEC responses to NE — but not E. coli LPS. HCAECs were incubated with 10 μg/ml of a TLR2- neutralizing monoclonal antibody, TL2.5 (Anti-TLR2) or isotype matched antibody control, TLR3.7 (Irr. Ab) for 30 min prior to challenge with 1 μg/ ml of each LPS or 100 ng/ml Pam3CSK4 for 4 h (A). Alternatively, HCAECs were preincubated with 10 μg/ml of TLR6-neutralizing polyclonal antibody (Anti-TLR6) or irrelevant antibody before challenge with 1 μg/ml of each LPS or 10 ng/ml of the synthetic diacyl lipopeptide FSL-1 for 4 h (B). Cell- surface ELISA was then performed to measure relative expression of E- selectin. Results are representative of three experiments and are presented as mean+/−S.D. ⁎Pb0.05 by Student's t-test.

Article Snippet: Western blot was performed on 10 μg of whole cell lysates obtained from unchallenged HEK-293, HEK293 cells transfected with TLR2 or TLR4/MD2, HUVEC, HCAEC and THP-1 cells using antibodies to TLR2 (sc10739, Santa-Cruz, US), TLR4 (sc10741, Santa-Cruz), TLR6 (3653, Prosci, CA, US) or loading control GAPDH (sc25778, Santa-Cruz).

Techniques: Incubation, Control, Enzyme-linked Immunosorbent Assay, Expressing

Journal: Frontiers in Immunology

Article Title: Nocardia rubra cell wall skeleton-induced MARCO expression: implications for improved phagocytosis and cytokine secretion in tumor-associated macrophages

doi: 10.3389/fimmu.2026.1611476

Figure Lengend Snippet: Nr-CWS affects MARCO expression through TLR4. The expression of TLR4 significantly increased after Nr-CWS treatment [ (D-G) , scale bars = 100 μm], while the expression TLR2 was unchanged (A-C) . Subsequent inhibition of TLR4 expression resulted in decreased levels of MARCO [ (H-J) , scale bars = 100 μm]. n = 3, * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: The following commercial antibodies (vendor, catalog number and dilution) utilized for western blot and immunohistochemical staining were employed in accordance with the manufacturers’ guidelines: mouse anti-CD68 antibody (Abcam, ab201340, 1:200), rabbit anti-MARCO antibody (Abcam, ab231046, 1:1000 for western blot and 1:100 for immunohistochemistry), rabbit anti-CD163 antibody (Abways Technology, CY6845, 1:500), mouse anti-CD86 antibody (Proteintech, 68674-2-Ig, 1:5000), mouse anti-TLR4 antibody (Santa Cruz Biotechnology, sc-293072, 1:1000 for western blot and 1:200 for immunohistochemistry), rabbit anti-TLR2 antibody(Abways Technology, CY5102, 1:2000), mouse anti-GAPDH antibody (Proteintech, 6004-1-Ig, 1:50000), HRP-goat anti-mouse recombinant secondary antibody (H+L)(Proteintech, RGAM001, 1:5000), goat anti-mouse IgG (H+L) Alexa Fluor 594 (Abways Technology, AB0152, 1:300), HRP-goat anti-rabbit recombinant secondary antibody (H+L) (Proteintech, RGAR001, 1:5000), goat anti-rabbit IgG (H+L) secondary antibody DyLightTM 594 (Report Biotech, S7002, 1:500), goat anti-mouse IgG (H+L) secondary antibody DyLightTM 488 (Report Biotech, S6001, 1:500), goat anti-mouse IgG (H+L) secondary antibody DyLightTM 594 (Abways Technology, AB0152, 1:500).

Techniques: Expressing, Inhibition

Journal: The Journal of Veterinary Medical Science

Article Title: Immune-Stimulatory Effects of a Bacteria-Based Probiotic on Peripheral Leukocyte Subpopulations and Cytokine mRNA Expression Levels in Scouring Holstein Calves

doi: 10.1292/jvms.13-0534

Figure Lengend Snippet: Monoclonal antibodies used for flow cytometry

Article Snippet: CD282 , TLR2 , Monocytes , HCA151F , IgG , AbD-Serotec.

Techniques: Bioprocessing

Journal: Journal of Inflammation Research

Article Title: Therapeutic Effect of C-C Chemokine Receptor Type 1 (CCR1) Antagonist BX471 on Allergic Rhinitis

doi: 10.2147/JIR.S254717

Figure Lengend Snippet: Relative quantification of NF-kB p65, TLR4, and TLR2 protein expression levels. ( A ) Western blot analysis for NF-kB p65, TLR4, and TLR2 relative expression from groups indicated. ( B ) summary graph for relative expression of NF-kB (n=8 per group). ( C ) summary graph for relative expression of TLR4 (n=8 per group). ( D ) summary graph for relative expression of TLR2 (n=8 per group). All values are represented as mean ± SEM. * p<0.05; ** p<0.01; and *** p<0.001 versus the vehicle control group.

Article Snippet: Samples were subsequently treated with anti β-actin antibody (BM3873, Boster Biological Technology, China), NF-kB p65 monoclonal antibody (A10609, Abclonal, USA.), TLR4 antibody (BA1717, Boster Biological Technology, China), or TLR2 antibody (BM4001, Boster Biological Technology, China), and were incubated overnight at 4°C.

Techniques: Quantitative Proteomics, Expressing, Western Blot, Control