sa pgn  (InvivoGen)

Journal: Frontiers in Immunology

Article Title: A feedback loop sustaining neutrophil extracellular trap formation involves S100 proteins, histones, TLR2 and RAGE, and is restrained by albumin

doi: 10.3389/fimmu.2026.1774475

Figure Lengend Snippet: Sequential contribution of RAGE and TLR2 to the belated action of endogenous mediators driving NET formation. (A) Human neutrophils adherent to poly-L-lysine-coated coverslips were incubated for 4 h in the absence (“unstim”) or presence of 1 μg/mL ultra-pure peptidoglycan (“PGN”), 100 U/mL TNFα, 1 nM GM-CSF, or 100 nM fMLP. After 90 min of stimulation, a TLR2 receptor antagonist (20 nM CU-CPT22, “TLR2i +90’”) or its diluent was added to the cells. NET formation was then assessed by microscopy. Mean ± s.e.m. of standardized NET indices from 3 independent experiments. (B) Human neutrophils adherent to poly-L-lysine-coated coverslips were incubated for 4 h in the absence (“unstim”) or presence of 100 U/mL TNFα. The receptors, TLR2 and RAGE, were antagonized either prior to (-15 min) or after (+30, +60, +120, +180 min) stimulation using 20 nM CU-CPT22 (“TLR2i”) or 1 µM FPS-ZM1 (“RAGEi”). NET formation was then assessed by microscopy. Mean ± s.e.m. of standardized NET indices from 3 independent experiments. *, p < 0.05; **, p < 0.01; ***, p < 0.001 vs the positive control; ns, not significantly different from the positive control; using Student’s paired t test.

Article Snippet: Among neutrophil stimuli, monosodium urate (MSU) (tlrl-msu) and ultra-pure peptidoglycan (PGN) were from Invivogen (#tlrl-pgnb3); TNFα (#210-TA) and GM-CSF (#7954-GM) were from R&D Systems; and N-formyl-methionyl-phenylalanine (fMLP) was from Millipore Sigma (#F3506).

Techniques: Incubation, Microscopy, Positive Control

Journal: bioRxiv

Article Title: Liver sinusoidal endothelial cells integrate metabolic and immune signals for MAPK-dependent BMP6 regulation and hepcidin induction

doi: 10.64898/2026.05.07.723498

Figure Lengend Snippet: (A) Bmp6 mRNA expression in primary LSECs treated with 5 ng/mL LPS or 2.5 μM heme for 6 h in the presence or absence of hepatocyte-conditioned medium. (B) Bmp6 mRNA expression in primary LSECs treated with various TLR ligands: Pam3CSK4 (TLR1/2), PGN (TLR2), Poly I:C (TLR3), LPS (TLR4), FLA-ST (TLR5), FSL1 (TLR2/6), R848 (TLR7/8), and ODN (TLR9) or vehicle control (NT, non-treated) for 6 h. (C-D) Bmp6 mRNA expression in primary LSECs treated with increasing concentrations of LPS for 6 h or with 5 ng/mL LPS for 2, 4, and 6 h. (E) Bmp6 mRNA expression in LSECs treated with 2.5 μM heme or protoporphyrin IX (PPIX) for 6 h. (F-G) Bmp6 mRNA expression in primary LSECs treated with increasing concentrations of heme for 6 h or treated with 2.5 μM heme for 2, 4, or 6 h. (H) Bmp6 mRNA expression in primary LSECs pre-treated with TAK242 (5 μM) or DMSO for 1 h, followed by 5 ng/mL LPS or 2.5 μM heme treatment for 6 h. (I) Transcription factor activity analysis by RNA-seq of LSECs treated with heme with respect to untreated controls (contrast-wise), in presence of hepatocyte-conditioned medium. (J) Bmp6 mRNA expression in primary LSECs pre-treated with CHX (5 μM) or DMSO for 1 h, followed by 5 ng/mL LPS treatment for 6 h. Cell culture experiments, except those in panel A, were always conducted in the presence of hepatocyte-conditioned medium. Gene expression levels were assessed by RT-qPCR, normalized to the housekeeping gene Rpl19 , and expressed as fold change relative to vehicle-treated controls. The dashed line (ut) represents the mRNA expression of LSECs treated with the conditions shown, in the absence of LPS or heme. Data are obtained from three or four independent experiments and displayed as mean ± SD. Statistical significance: *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001, one-way ANOVA or Student’s t-test. CM, conditioned medium; PPIX, protoporphyrin IX; CHX, cycloheximide.

Article Snippet: The TLR ligands Pam3CSK4 (TLR2:1) (#tlrl-pms), PGN-SA (TLR2) (#tlrl-pgns2), Poly I:C (TLR3) (#tlrl-picw), FLA-ST (TLR5) (#tlrl-stfla), FSL1 (TLR2:TLR6) (tlrl-fsl), R848 (TLR7:8) (#tlrl-r848-1), ODN (TLR9) (#tlrl-1826) and the MAPK inhibitor SP600125 (#tlrl-sp60) were purchased form Invivogen and diluted in PBS (TLR ligands) or DMSO (SP600125).

Techniques: Expressing, Control, Activity Assay, RNA Sequencing, Cell Culture, Gene Expression, Quantitative RT-PCR

Journal: EMBO Molecular Medicine

Article Title: A skin colonizer disrupts inflammatory and humoral immune defenses in hidradenitis suppurativa

doi: 10.1038/s44321-026-00407-7

Figure Lengend Snippet: ( A ) IgG (left) and IgA (right) reactivities of HC and HS3 serum pools towards healthy skin sections are shown. Scale bar: 100 µm. The white dotted line indicates the separation between epidermis (Epi) and dermis. ( B ) Same as in ( A ) towards healthy donor-derived KCs, with quantification of IgG signals normalized to Dapi. Scale bar: 50 µm. Mann–Whitney Statistical test, n = 3. ( C ) Dot plots illustrating the reactivity of HC and HS3 sera ( n = 11) against peptidoglycan (PGN), Keyhole limpet hemocyanin (KLH), MAPK14, lipopolysaccharide (LPS), double-stranded DNA (dsDNA), tubulin (TB), and lysozyme (LZ). Data were area under the curve (AUC) values determined by ELISA with serially diluted sera. Mann–Whitney statistical test. ( D ) Methodology for depleting HC and HS3 serum pools on bacteria (top). Panels below show the differential reactivity of HC and HS3 sera pools against LZ after depletion on P. uenonis or P. bivia , expressed as AUC values from ELISA with serially diluted sera, and as the mean percentage of IgG loss in bacteria-depleted pools ( n = 3) compared to non-depleted sera, analyzed by two-way ANOVA with Sidak’s multiple comparisons test. .

Article Snippet: Peptidoglycan from B. subtilis , Invivogen , Cat# tlrl-pgnb3.

Techniques: Derivative Assay, MANN-WHITNEY, Enzyme-linked Immunosorbent Assay, Bacteria