rat anti mouse tlr4  (R&D Systems)

Journal: Microbial pathogenesis

Article Title: Exploring the effects of lactulose on lung-intestinal tissue-associated factors and the TLR4/NF-κB signaling pathway in COPD rats based on lung-gut axis theory.

doi: 10.1016/j.micpath.2024.107245

Figure Lengend Snippet: Fig. 6. Protein band diagrams of MUC5AC, TLR4, and p-P65 in lung tissue among different rat groups.

Article Snippet: LPS and 2 % pentobarbital were both sourced from Beijing Ita Biotechnology Co., Ltd., with batch numbers YT1319 and 23NO213, respectively; Hardman brand cigarettes were provided by Shandong Zhongshan Cigarette Industry Co., Ltd., with tar content of 10 mg, nicotine content of 0.8 mg, and carbon monoxide content of 13 mg; phosphate buffer saline (PBS) buffer was supplied by Wuxi Aorui Dongyuan Biotechnology Co., Ltd., with lot number 21031203; 2.5 % glutaraldehyde was from Beijing Solepol Science and Technology Co., Ltd., with lot number P1126; anhydrous ethanol and acetone were both from Sinopharm Chemical Reagent Co., Ltd., with lot numbers 100092183 and 10000418, respectively; 812 embedding agent was sourced from SPI, with batch number 90529-77-4; eosin staining solution and hematoxylin staining solution were both from Zhuhai Beso Biotechnology Co. Ltd., with batch numbers BA4024 and BA4041, respectively; TNF-α ELISA kit and IL-1β ELISA kit were provided by Jiangsu Zeyu Bio-technology Co. Ltd., with batch numbers 2312226006R and 231226017R, respectively; reverse transcription kit was supplied by Thermo, USA, with lot number K1622; rabbit anti-rat mucin 5AC (MUC5AC) antibody was sourced from Invitrogen, with lot number PA5-115601; rabbit anti-rat p65 phosphorylated antibody, rabbit anti-rat p65 antibody, and mouse anti-rat TLR4 antibody were all provided by Proteintech, with batch numbers 82335-1-RR, 10745-1-AP, and 66350-1-Ig, respectively; goat anti-rabbit IgG antibody was bought from Affinity, with batch number S0001; and sheep anti-mouse IgG antibody was sourced from Affinity, with batch number S0002.

Techniques:

Journal: Microbial pathogenesis

Article Title: Exploring the effects of lactulose on lung-intestinal tissue-associated factors and the TLR4/NF-κB signaling pathway in COPD rats based on lung-gut axis theory.

doi: 10.1016/j.micpath.2024.107245

Figure Lengend Snippet: Fig. 8. Protein band diagrams of MUC2, TLR4, and p-P65 in colon tissue among different rat groups.

Article Snippet: LPS and 2 % pentobarbital were both sourced from Beijing Ita Biotechnology Co., Ltd., with batch numbers YT1319 and 23NO213, respectively; Hardman brand cigarettes were provided by Shandong Zhongshan Cigarette Industry Co., Ltd., with tar content of 10 mg, nicotine content of 0.8 mg, and carbon monoxide content of 13 mg; phosphate buffer saline (PBS) buffer was supplied by Wuxi Aorui Dongyuan Biotechnology Co., Ltd., with lot number 21031203; 2.5 % glutaraldehyde was from Beijing Solepol Science and Technology Co., Ltd., with lot number P1126; anhydrous ethanol and acetone were both from Sinopharm Chemical Reagent Co., Ltd., with lot numbers 100092183 and 10000418, respectively; 812 embedding agent was sourced from SPI, with batch number 90529-77-4; eosin staining solution and hematoxylin staining solution were both from Zhuhai Beso Biotechnology Co. Ltd., with batch numbers BA4024 and BA4041, respectively; TNF-α ELISA kit and IL-1β ELISA kit were provided by Jiangsu Zeyu Bio-technology Co. Ltd., with batch numbers 2312226006R and 231226017R, respectively; reverse transcription kit was supplied by Thermo, USA, with lot number K1622; rabbit anti-rat mucin 5AC (MUC5AC) antibody was sourced from Invitrogen, with lot number PA5-115601; rabbit anti-rat p65 phosphorylated antibody, rabbit anti-rat p65 antibody, and mouse anti-rat TLR4 antibody were all provided by Proteintech, with batch numbers 82335-1-RR, 10745-1-AP, and 66350-1-Ig, respectively; goat anti-rabbit IgG antibody was bought from Affinity, with batch number S0001; and sheep anti-mouse IgG antibody was sourced from Affinity, with batch number S0002.

Techniques:

Journal: The Journal of Experimental Medicine

Article Title: Intercrypt sentinel macrophages tune antibacterial NF-κB responses in gut epithelial cells via TNF

doi: 10.1084/jem.20210862

Figure Lengend Snippet: Primary antibodies

Article Snippet: TLR4 , BioRad , MCA2154T.

Techniques:

Journal: The Journal of Experimental Medicine

Article Title: Intercrypt sentinel macrophages tune antibacterial NF-κB responses in gut epithelial cells via TNF

doi: 10.1084/jem.20210862

Figure Lengend Snippet: TLR4 + immune cells induce epithelial NF-κB signaling in the cecal mucosa upon LPS exposure. Mice were i.v. injected with LPS. Cecal explants were imaged at 1 h.p.inj. by two-photon microscopy. (A–C) Representative images and quantification of epithelial NF-κB activation in the indicated mice (A, n = 4–7) or BMCs (B, n = 5; C, n = 5). Each circle represents one mouse. Black line: median. **, P ≤ 0.01 by Mann–Whitney U test. (D) Small intestinal organoids were treated with 5, 50, or 500 ng/ml LPS and imaged for ∼1 h. Representative images of one organoid over time (top) and quantification of NF-κB activation (bottom; relative change). Each circle represents one organoid at the given time (minutes after start of the treatment, n = 7). Lines connect data points from the same organoid. Red dashed line: 50% activation threshold. Black dotted line: no change. Scale bars: 50 µm. Combined data of two (C and D), three (A), or four (B) independent experiments.

Article Snippet: TLR4 , BioRad , MCA2154T.

Techniques: Injection, Microscopy, Activation Assay, MANN-WHITNEY

Journal: The Journal of Experimental Medicine

Article Title: Intercrypt sentinel macrophages tune antibacterial NF-κB responses in gut epithelial cells via TNF

doi: 10.1084/jem.20210862

Figure Lengend Snippet: TLR4 + immune cells induce epithelial NF-κB activation in the small intestine and colon. (A) Schematic drawing of the two-photon imaging (left). The intestinal mucosa is imaged from the luminal side (black arrowhead), resulting in images in horizontal plane of the mucosa (right; part of the image shown in ). White dashed line/E, epithelium; L, lumen; white asterisks, epithelial nuclei. (B) Fold changes in expression of A20 , Cxcl2 , and Tnf in the cecal mucosa of mice depicted in ( n = 5). (C) Fold changes in expression of A20 , Cxcl2 , and Tnf in the cecal mucosa of mice depicted in in comparison to PBS-injected mice in ( n = 5). (D and E) Two-photon microscopy images and quantification of epithelial NF-κB activation in the (D; n = 3–5) small intestine and (E; n = 3–5) colon of LPS-injected BMCs, and (F) in small intestine and colon of Myd88 −/− > p65 GFP-FL x Tlr4 −/− , Ticam1 −/− > p65 GFP-FL x Tlr4 −/− BMCs. Each circle represents one mouse. Black line: median. Statistical analysis: Mann–Whitney U test. *, P ≤ 0.05; **, ≤ 0.01. Scale bars: 50 µm. Combined data of three (B), four (C), five (D), six (E), or seven (F) independent experiments. Each circle represents one mouse. Black line: median. Scale bars: 50 µm.

Article Snippet: TLR4 , BioRad , MCA2154T.

Techniques: Activation Assay, Imaging, Expressing, Comparison, Injection, Microscopy, MANN-WHITNEY

Journal: The Journal of Experimental Medicine

Article Title: Intercrypt sentinel macrophages tune antibacterial NF-κB responses in gut epithelial cells via TNF

doi: 10.1084/jem.20210862

Figure Lengend Snippet: TNF produced by CD11c + cells induces local epithelial NF-κB activation in the intestinal mucosa. (A and B) p65 GFP-FL intestinal epithelial organoids established from the indicated regions were treated with 5, 50, and 500 ng/ml or 5 µg/ml LPS (+ LBP and CD14, if indicated) and imaged for 1 h (A; n = 3–17), or analyzed by qPCR at 3 h of treatment (B and C; n = 6 or 7). (C) Colon organoids from p65 GFP-FL x Tlr4 −/− mice ( n = 4). (D) Representative two-photon microscopy overview image of the cecal mucosa of mice described in at 1 h.p.inj. of LPS ( n = 6). Red squares indicate RFP + ( Tlr4 +/+ ) cells. White lines indicate IEC NF-κB activation zones (defined as areas with continuous epithelial NF-κB activation). (E) Quantification of epithelial NF-κB activation in Il18 −/− > p65 GFP-FL x Tlr4 −/− , Il18r −/− > p65 GFP-FL x Tlr4 −/− , and Il1ab −/− > p65 GFP-FL x Tlr4 −/− BMCs at 1 h.p.inj. of LPS ( n = 7 or 8). (F) p65 GFP-FL intestinal epithelial organoids from cecum (left) or colon (right) were treated with 5, 50, and 500 ng/ml TNF and imaged for 1 h ( n = 3–17). (G) Quantification of epithelial NF-κB activation in mice as described in . Mice pretreated with DTX were injected with PBS or TNF ( n = 2–6). Cecae were imaged at 1 h.p.inj. Data of LPS-injected mice are replotted from for comparison. Black line: median (B, C, E, and G). Dashed line: detection limit (C and G) or error range (A and F). Each circle represents one organoid sample (B and C), one mouse (E and G), or the median (A and F). Statistical analysis: one-way ANOVA with Dunett’s correction (B and C) or Mann–Whitney U test (E and G). *, P ≤ 0.05; **, P ≤ 0.01. Scale bars: 50 µm. Combined data of two (A, small intestine; B, C, D, and F, cecum), three (A, cecum), four (F, colon), six (B and E), or eight (A, colon) independent experiments.

Article Snippet: TLR4 , BioRad , MCA2154T.

Techniques: Produced, Activation Assay, Microscopy, Injection, Comparison, MANN-WHITNEY

Journal: The Journal of Experimental Medicine

Article Title: Intercrypt sentinel macrophages tune antibacterial NF-κB responses in gut epithelial cells via TNF

doi: 10.1084/jem.20210862

Figure Lengend Snippet: CD11c + cells induce local epithelial NF-κB activation via TNF. Mice were i.v. injected with LPS and cecal explants imaged at 1 h.p.inj. by two-photon microscopy (representative image and quantification) if not indicated otherwise. (A) Cecum mucosa from p65 GFP-FL x Tlr4 −/− mice reconstituted with a 1:10 mix of ActRFP (10%, Tlr4 +/+ ) and p65 GFP-FL x Tlr4 −/− (90%) BM. Analysis of RFP + cells within an epithelial NF-κB activation zone (see , n = 10–18). (B – D) Cecal epithelium NF-κB activation of the indicated BMCs or p65 GFP-FL mice pretreated with isotype control/anti-TNF antibody or i.v. injected with TNF and analyzed at the indicated time points ( n = 5 or 6). (E) TNF-treated small-intestinal epithelial organoids. Representative image and quantification of NF-κB activation kinetics with 5, 50, or 500 ng/ml TNF ( n = 9–17). Lines connect data points from the same organoid. Red dashed line: 50% activation threshold. Black dotted line: no change. (F) Representative images of the cecal epithelium and quantification of epithelial NF-κB activation of p65 GFP-FL x Tlr4 −/− mice reconstituted with a 1:20 mix of CD11c-DTR and TNFa −/− BM, pretreated with DTX ( n = 5–8). (B–D and F) Black line: median. *, P ≤ 0.05; **, P ≤ 0.01 by Mann–Whitney U test. Each circle represents one mouse or one organoid (E). Combined data of two (A and B), three (D and E), four (F), or six (C) independent experiments. Scale bars: 50 µm.

Article Snippet: TLR4 , BioRad , MCA2154T.

Techniques: Activation Assay, Injection, Microscopy, Control, MANN-WHITNEY

Journal: The Journal of Experimental Medicine

Article Title: Intercrypt sentinel macrophages tune antibacterial NF-κB responses in gut epithelial cells via TNF

doi: 10.1084/jem.20210862

Figure Lengend Snippet: Receptor expression in IECs. (A–D) Confocal microscopy images of (A) the cecal patch, and (B) a mucosa-associated lymphoid follicle in fixed cecae of p65 GFP-FL mice i.v. injected with LPS at 1 h.p.inj. Boxes in overview images indicate insets. Arrowheads indicate p65 + nuclei. Arrows indicate p65 − nuclei. Scale bars: 50 µm (overview images) or 10 µm (insets). TLR4 staining in small intestine (C) and colon (D) of WT mice. Arrowheads indicate MHCII + cells. Arrows indicate IECs. Scale bars: 20 µm. Representative images of mice from two experiments. (E) Heat map depicting expression levels of Tlr2, Tlr4, Tlr6, Tlr11 , Tnfrsf1a (TNFR1), and Tnfrsf1b (TNFR2) in untreated or TNF-treated (5 ng/ml, 8 h) small intestinal epithelial organoids derived from SPF (SPF1, SPF2) or germ-free (GF) mice, m-IC c12 cells, and mouse embryonic fibroblasts (MEFs; reanalysis of a previously published transcriptome dataset, all detectable Tlrs depicted; ). (F) Cecal mucosa stained for TNFR1 at 1 h.p.inj. of LPS. Scale bars: 50 µm. Representative images of mice from three independent experiments ( n = 4–7).

Article Snippet: TLR4 , BioRad , MCA2154T.

Techniques: Expressing, Confocal Microscopy, Injection, Staining, Derivative Assay

Journal: The Journal of Experimental Medicine

Article Title: Intercrypt sentinel macrophages tune antibacterial NF-κB responses in gut epithelial cells via TNF

doi: 10.1084/jem.20210862

Figure Lengend Snippet: Tissue resident, monocyte-derived macrophages secrete TNF to induce local epithelial NF-κB activation. (A) ELISA measurements of TNF concentrations in the cecal mucosa of LPS injected WT mice ( n = 5 or 6). Dashed line: detection limit. y axis in log 10 scale. (B) Percentage of TNF + DCs or macrophages (gating as shown in ) in the cecum, small intestine, and colon of LPS-treated WT mice (1 h.p.inj.) and PBS-treated controls ( n = 5–7). (C) Representative images of the cecal epithelium and quantification of epithelial NF-κB activation of p65 GFP-FL mice pretreated with anti-CSF1R or isotype control, injected with LPS, and imaged 1 h.p.inj. ( n = 7). Depletion efficiency of macrophages and DCs in anti-CSF1R treated mice. (D) Normalized marker expression of TNF + compared with TNF − macrophages in the cecum, small intestine, and colon of LPS-injected WT mice ( n = 4–7). (E) Percentage of CD4 +/− Tim4 +/− cells among TNF − and TNF + macrophages in the cecum of LPS-injected WT mice ( n = 4). (F) TNF-PLA analysis of cecae from p65 GFP-FL x Tlr4 −/− mice reconstituted with a 1:40 mix of ActRFP (2.5%, Tlr4 +/+ ) and p65 GFP-FL x Tlr4 −/− (97.5%) BM. Representative confocal microscopy image of fixed cecal tissue at 40 min.p.inj. (left) and quantification of PLA for TNF in crypts without (−) or with (+) epithelial NF-κB activation at 1 h.p.inj. ( n = 11–13). Scale bar: 10 µm. Black line: median. Statistical analysis: one-way ANOVA with Dunett’s correction (A), two-way ANOVA with Sidak’s multiple comparison test (B), or Mann–Whitney U test (C, E, and F). *, P ≤ 0.05; **, P ≤ 0.01. Each circle represents one mouse (A–E) or one crypt (F; five mice analyzed). Combined data of two (D), three (B and C), four (F), or six (A) independent experiments, or exemplary data of two (E) independent experiments.

Article Snippet: TLR4 , BioRad , MCA2154T.

Techniques: Derivative Assay, Activation Assay, Enzyme-linked Immunosorbent Assay, Injection, Control, Marker, Expressing, Confocal Microscopy, Comparison, MANN-WHITNEY

Journal: The Journal of Experimental Medicine

Article Title: Intercrypt sentinel macrophages tune antibacterial NF-κB responses in gut epithelial cells via TNF

doi: 10.1084/jem.20210862

Figure Lengend Snippet: Intestinal macrophages secrete TNF to induce local epithelial NF-κB activation. (A) Gating strategy for intestinal MP subsets in the cecal mucosa of mice depicted in B and C. (B and C) Flow cytometry analysis of cecal MPs from PBS- or LPS-injected (B) KappaBle mice for assessment of NF-κB activation (gating as shown in A; n = 3–5) or (C) WT mice for identification of TNF-producing MP subsets (gating as shown in A; n = 3–6). (D) Updated gating strategy for differentiation of intestinal DCs and macrophages as shown in . Lineage = NK1.1, CD3, B220. (E) For scRNAseq, CD45 + live MHCII + lineage (NK1.1, CD3, B220) − cells were sorted from the cecal mucosa of 40 min LPS-injected mice or PBS-treated controls ( n = 4 mice) and subsequently analyzed by scRNAseq (10X Genomics). T-distributed stochastic neighbor embedding plots showing the distribution of the analyzed cells indicated by cluster (left) or treatment (right). (F) Expression levels of intestinal MP markers: this analysis revealed two clear macrophages clusters (7 and 10). CD11b − CD103 + Xcr1 + DCs were represented in clusters 1–4, out of which cluster 2 mainly consisted of cells from LPS-treated mice, indicating that this might represent an activated state. This is in line with the secondary TNF production of this subset at later time points after injection, as detected by flow cytometry (C). Clusters 0 and 5 were positive for Sirpa and contained CD11b + CD103 + DCs (cluster 0) and CD11b + CD103 − DCs (cluster 5). While clusters 12 and 13 were positive for a number of monocyte/macrophage markers and therefore likely represent maturing macrophages, the assignment of clusters 6, 8, and 9 was challenging due to overlapping marker expression. These clusters, together with cluster 11 (mast cells), likely contained precursors (macrophage/DC, cluster 8) or contaminating cells (T cells, cluster 6 and 8; plasma cells, cluster 9). (G) Expression analysis of TLR4 signaling–associated genes. (H) Gene set enrichment analysis for macrophage clusters (7 and 10). (I) Frequency of DCs and macrophages in the cecum, small intestine, and colon of naive WT mice ( n = 5). Each circle represents one mouse (B, C, and I) or one cell (E). Black line: median. Combined data of two (I), three (B), or nine (C) independent experiments. Statistical analysis: one-way ANOVA with Tukey’s correction (C) or Mann–Whitney U test (B). *, P ≤ 0.05; **, P ≤ 0.01.

Article Snippet: TLR4 , BioRad , MCA2154T.

Techniques: Activation Assay, Flow Cytometry, Injection, Expressing, Marker, Clinical Proteomics, MANN-WHITNEY

Journal: The Journal of Experimental Medicine

Article Title: Intercrypt sentinel macrophages tune antibacterial NF-κB responses in gut epithelial cells via TNF

doi: 10.1084/jem.20210862

Figure Lengend Snippet: TNF-mediated epithelial NF-κB activation occurs upon bacterial infection and induces an antibacterial response. (A) Representative two-photon microscopy images of cecal explants of streptomycin-pretreated p65 GFP-FL mice infected with S . Tm for 8–13.5 h ( n = 28). Categories for scoring of epithelial NF-κB activation status: “no activation” (green); “patchy activation” (blue); “full activation” (orange); “inflammation” (red; tissue distortion evident); “unspecified” (gray; was excluded from further analysis). Scale bars: 50 µm. (B) Distribution of the analyzed 28 samples of A among the four epithelial NF-κB activation categories (bottom). For simplification, the blue, orange, and red categories were summarized as “NF-κB signaling” (yellow, top). (C) Tnf transcript levels in the cecal mucosa of mice described in A and naive p65 GFP-FL mice, grouped according to the epithelial NF-κB activation status of the respective mice and depicted as 2 -ΔCT . Expression levels were normalized to Actb ( n = 33). (D) TNFa +/− or TNFa −/− > p65 GFP-FL x Tlr4 −/− BMCs were analyzed as described in A and B. (E) Log 2 ratios of selected genes in a transcriptome analysis of TNF-treated (8 h, 5 ng/ml) compared with untreated small intestinal epithelial organoids . FDR, false discovery rate. (F) C3 transcript levels in untreated and TNF-treated (5 ng/ml, 4 h) small intestinal organoids depicted as 2 -ΔCT . Expression levels were normalized to Actb . (G) Streptomycin-pretreated TNFa −/− mice and heterozygous littermates were orally infected with S . Tm for 36h. S . Tm in the cecal lumen (gated on O12 + cells, see ) were stained for surface C3 to assess coating of luminal bacteria by flow cytometry (C3 + population). MFI, median fluorescence intensity. Statistical analysis: Mann–Whitney U test (C, F, and G) or χ 2 test (D). *, P ≤ 0.05; **, P ≤ 0.01. Each circle represents one mouse (C and G) or one experiment (average; F). y axis in log 10 scale (C, F, and G). Combined data of three (G), four (F), five (A–C), or six (D) independent experiments.

Article Snippet: TLR4 , BioRad , MCA2154T.

Techniques: Activation Assay, Infection, Microscopy, Expressing, Staining, Bacteria, Flow Cytometry, Fluorescence, MANN-WHITNEY

Journal: The Journal of Experimental Medicine

Article Title: Intercrypt sentinel macrophages tune antibacterial NF-κB responses in gut epithelial cells via TNF

doi: 10.1084/jem.20210862

Figure Lengend Snippet: Primary antibodies

Article Snippet: TLR4 , BioRad , MCA2154T.

Techniques:

Journal: bioRxiv

Article Title: Gram-negative bacterial infection increases lung cancer metastasis via Toll-like receptor activation and increased cancer cell proliferation post-tumor adhesion

doi: 10.1101/2021.09.15.460508

Figure Lengend Snippet: A . Dot plot of number of gross hepatic metastases in WT and TLR4 -/- mice injected with 30,000 H59-GFP cells followed by CLP or sham surgery 24hrs later and sacrificed 2 weeks after tumor cell injection with means and standard deviations. * p = 0.0082; ** p = 0.0006; *** p = 0.0005. B . Representative images of the livers from the 4 groups of mice in (A). C . Representative images of Ki67 staining in tumor island and liver sinusoids of livers from sham and CLP mice two weeks following tumor injection of H59-GFP cells followed by CLP after 24hrs using a 20X microscope objective. D . Dot plot of number of ki67 tumor islands seen in 10 random fields (± SEM) from the 4 groups of mice in (A). * p = 0.0203; ** p < 0.0001. E . Representative H&E images of the livers of the 4 groups of mice in (A) at 20X magnification and inset of micrometastases of the micrometastases seen in the CLP group at 40X magnification. F . Dot plot of number of micrometastases seen in 10 random fields (± SEM) from the 4 groups of mice in (A). * p = 0.117; ** p < 0.0001. For all panels, n=7-14 mice/group, n.s.=not significant.

Article Snippet: Fluorescein-conjugated monoclonal mouse anti-mouse TLR5 IgG2a and fluorescein-conjugated monoclonal rat anti-mouse TLR4 IgG2b (both from Novus Biologicals (Oakville, ON)) were used at 500 μg/mL.

Techniques: Injection, Staining, Microscopy

Journal: bioRxiv

Article Title: Gram-negative bacterial infection increases lung cancer metastasis via Toll-like receptor activation and increased cancer cell proliferation post-tumor adhesion

doi: 10.1101/2021.09.15.460508

Figure Lengend Snippet: A . Dot plot number of % confluence of GFP+ cells (average ± SEM) from cultures of H59-GFP cells stimulated with either heat inactivated E. coli (ECOLI) or a combination of heat inactivated E. coli and Eritoran (ERIT) at 24, 48 and 72 hrs post-stimulation. Non-treated (NT) cells were used as controls. n=9, * p<0.05. B . Representative images of panel A. C . Cell surface expression of H59-GFP by flow cytometry for TLR2, TLR4 and TLR5. For all 3 panels, non-treated cells are in green, isotype control in red and antibody against TLR in pink. 10,000 cells were counted for each sample. D . Bar graph of average (± SEM) number of H59 cells following stimulation with heat-inactivated E. Coli in the presence or absence of blockade of TLR2, TLR5 and TLR2, 4 and 5 combination along with their isotype controls. n≥3, * p<0.05. E . Representative images of panel D. F . Bar graph of MTT assay 48 hours post-stimulation with conditioned media from untreated BEAS-2B cells (NT), BEAS-2B cells stimulated with LPS (LPS), BEAS-2B cells stimulated with LPS following treatment with small molecule inhibitor Eritoran (LPS + Eritoran). n≥3, n.s.=not significant, * p<0.05 increase from NT, ** p<0.05 decrease from LPS. G . Bar graph of MTT assay 48 hours post-stimulation with conditioned media from untreated BEAS-2B cells (NT), BEAS-2B cells stimulated with LPS (LPS) and BEAS-2B cells stimulated with LPS following treatment with small molecule inhibitors PD184352, BIRB0796 and PI103 (LPS + “inhibitor”) or inhibitor alone. Data are presented as mean ± standard deviation. n≥3, * p<0.05 increase NT and ** p<0.05 decrease from LPS

Article Snippet: Fluorescein-conjugated monoclonal mouse anti-mouse TLR5 IgG2a and fluorescein-conjugated monoclonal rat anti-mouse TLR4 IgG2b (both from Novus Biologicals (Oakville, ON)) were used at 500 μg/mL.

Techniques: Expressing, Flow Cytometry, Control, MTT Assay, Standard Deviation