fitc lta  (Vector Laboratories)

Journal: Neoplasia (New York, N.Y.)

Article Title: Microbe-associated molecular patterns differentially mediate carcinogenic alterations of the breast tissue in the context of obesity

doi: 10.1016/j.neo.2026.101284

Figure Lengend Snippet: LPS and flagellin activate the NF-κB pathway and induce the transcription of inflammatory cytokines. A,G,M. Western blotting analysis of the nuclear (N) and cytosolic (C) fractions of NF-kB (p65) after treatment of HMT-3522 S1 cells with vehicle (control), the MAMPs LPS (A), LTA (G) or flagellin (M), in the absence or presence of inhibitor for their cognate TLR. Quantification of the nuclear to cytosolic ratios of NF-kB is shown in the graphs (B, H, N). Expression of TNF-α and IL-8 was quantified by qPCR in breast acini 1 h and 72 h after treatment with LPS (C-F), LTA (I-L), and flagellin (O-R). ns, not significant; *P<0.05; **P<0.01.

Article Snippet: Briefly, plate wells (Greiner bio-one; 655061) were coated with 50 μl of 40 μg/ml LPS (Sigma-Aldrich; L2630) or 40 μg/ml LTA (InvivoGen; tlrl-pslta) or 1 μg/ml flagellin (InvivoGen; tlrl-epstfla-5) in 50mM carbonate/bicarbonate buffer (pH 9.6) overnight at 4°C.

Techniques: Western Blot, Control, Expressing

Journal: Neoplasia (New York, N.Y.)

Article Title: Microbe-associated molecular patterns differentially mediate carcinogenic alterations of the breast tissue in the context of obesity

doi: 10.1016/j.neo.2026.101284

Figure Lengend Snippet: LPS and flagellin induce DNA double-strand breaks in breast acini in a TLR-dependent manner. A. Representative comet images from neutral comet assays performed on 3D cultures of breast acini treated with vehicle (control), 1 μg/ml LPS, 1 μg/ml LTA, and 10 ng/ml flagellin for 72 h. Bleomycin (BLM; 20 mU/ml; 2h) was used as positive control. B. Comet assay quantification. n=300 acini from 3 biological replicates. C. Representative images for immunofluorescence staining of 53BP1 (red) in acini treated with MAMPs ± TLR inhibitors. Nuclei are counterstained with DAPI (blue). D-F. Quantification of 53BP1 foci numbers in acini treated with LPS ± TLR4 inhibitor (D), with LTA ± TLR2 inhibitor (E) or with flagellin ± TLR5 inhibitor (F). ns is not significant. *P<0.05, **P<0.01, ***P<0.001.

Article Snippet: Briefly, plate wells (Greiner bio-one; 655061) were coated with 50 μl of 40 μg/ml LPS (Sigma-Aldrich; L2630) or 40 μg/ml LTA (InvivoGen; tlrl-pslta) or 1 μg/ml flagellin (InvivoGen; tlrl-epstfla-5) in 50mM carbonate/bicarbonate buffer (pH 9.6) overnight at 4°C.

Techniques: Control, Positive Control, Single Cell Gel Electrophoresis, Immunofluorescence, Staining

Journal: Neoplasia (New York, N.Y.)

Article Title: Microbe-associated molecular patterns differentially mediate carcinogenic alterations of the breast tissue in the context of obesity

doi: 10.1016/j.neo.2026.101284

Figure Lengend Snippet: LPS causes nuclear oxidative stress and the formation of single-strand breaks. A-C. Analysis of nuclear H 2 O 2 with the HyPer7 probe in MCF10A cells treated with MAMPs, in the presence or absence of glutathione (GSH). Cells were treated with LPS (A), flagellin (B), or LTA (C). n≈50 nuclei from 3 biological replicates. *P<0.05, **P<0.01 compared to control. # P<0.05, ## P<0.01 compared to the GSH condition. D. Representative images of 8-OHdG staining (green) in MCF10A cells treated with LPS, LPS+GSH, and culture medium (control). Nuclei are counterstained with DAPI (blue). E-G. Quantification of nuclear 8-OHdG signals in MFC10A cells treated with control medium or with LPS (±GSH) (E), LTA (±GSH) (F), or flagellin (±GSH) (G). H. Representative images showing comets from alkaline comet assays performed on acini cultures treated with vehicle (control), LPS, LTA, flagellin or bleomycin (BLM; positive control). I. Comet assay quantification in acini treated with MAMPs. n=300 acini from 3 biological replicates. ns is not significant. *P<0.05, **P<0.01, ***P<0.001, ****P<0.0001.

Article Snippet: Briefly, plate wells (Greiner bio-one; 655061) were coated with 50 μl of 40 μg/ml LPS (Sigma-Aldrich; L2630) or 40 μg/ml LTA (InvivoGen; tlrl-pslta) or 1 μg/ml flagellin (InvivoGen; tlrl-epstfla-5) in 50mM carbonate/bicarbonate buffer (pH 9.6) overnight at 4°C.

Techniques: Control, Staining, Positive Control, Single Cell Gel Electrophoresis

Journal: Neoplasia (New York, N.Y.)

Article Title: Microbe-associated molecular patterns differentially mediate carcinogenic alterations of the breast tissue in the context of obesity

doi: 10.1016/j.neo.2026.101284

Figure Lengend Snippet: LPS and flagellin cause DNA damage and inflammation in mice mammary glands. A. Schematic of the MAMPs injection experiment. B-D. Representative images for 3, 3′-diaminobenzidine (DAB) immunohistochemistry against LPS (B), LTA (C), and flagellin (D) in mice mammary glands. Hematoxylin was used as a counterstain. E. Representative comet images from the neutral comet assay performed on cells extracted from the mammary glands of mice treated with LPS, LTA, Flagellin, or control medium as indicated in the schematic. F. Quantification of comet assay results. G. Schematic of the obesogenic diet experiment. H. Plasma LPS concentrations (pg/ml) in the dietary groups. I. Comet assay analysis showing Spearman’s correlation between DNA damage levels (tail moments) and LPS concentrations (pg/ml) in the mice dietary groups. AU is Arbitrary units and ns is not significant. *P<0.05.

Article Snippet: Briefly, plate wells (Greiner bio-one; 655061) were coated with 50 μl of 40 μg/ml LPS (Sigma-Aldrich; L2630) or 40 μg/ml LTA (InvivoGen; tlrl-pslta) or 1 μg/ml flagellin (InvivoGen; tlrl-epstfla-5) in 50mM carbonate/bicarbonate buffer (pH 9.6) overnight at 4°C.

Techniques: Injection, Immunohistochemistry, Neutral Comet Assay, Control, Single Cell Gel Electrophoresis, Clinical Proteomics

Journal: Neoplasia (New York, N.Y.)

Article Title: Microbe-associated molecular patterns differentially mediate carcinogenic alterations of the breast tissue in the context of obesity

doi: 10.1016/j.neo.2026.101284

Figure Lengend Snippet: Proteobacteria is associated with higher DNA damage levels in breast tissues. A. 16S rRNA sequencing results showing relative abundance of LPS-containing bacteria, LTA-containing bacteria, and bacteria with unknown status in each of the study subjects. B. Bar graph showing the relative abundance of bacterial phyla in breast tissue samples from each subject. C-D. Spearman’s correlation between BMI and the relative abundance of LPS-containing bacteria (C) or Proteobacteria (D). E. Spearman’s correlation between BMI and breast tissue levels of anti-MAMPs IgA, IgM, and IgG. F-G. Spearman’s correlation between DNA damage and the relative abundance of LPS-containing bacteria (F) or Proteobacteria (G). H. Representative neutral comet assay images from breast tissues samples. I-J. Spearman’s correlation between the relative abundance of LPS-containing bacteria (I) or Proteobacteria (J) and 8-OHdG concentrations in breast tissue.

Article Snippet: Briefly, plate wells (Greiner bio-one; 655061) were coated with 50 μl of 40 μg/ml LPS (Sigma-Aldrich; L2630) or 40 μg/ml LTA (InvivoGen; tlrl-pslta) or 1 μg/ml flagellin (InvivoGen; tlrl-epstfla-5) in 50mM carbonate/bicarbonate buffer (pH 9.6) overnight at 4°C.

Techniques: Sequencing, Bacteria, Neutral Comet Assay