leukadherin 1 la 1  (MedChemExpress)

Journal: Frontiers in Immunology

Article Title: Mechanosensitivity of macrophage polarization: comparing small molecule leukadherin-1 to substrate stiffness

doi: 10.3389/fimmu.2025.1420325

Figure Lengend Snippet: Summary of experimental systems and plan. (A) Schematic depiction of in vitro cell culture model to test mechanotransduction by incubation of murine BMDMs on collagen-coated silica gels with corresponding stiffness (quantified as kilopascal, kPa) (B) Brightfield images from BMDMs cultured on gels of 0.2 kPa (soft) or 64 kPa (stiff). Scale bar = 10 µm. (C) Area of BMDMs measured from microscopic images shown in (B) . Each symbol represents area of one cell, line at median, p-value determined by Mann-Whitney, data from three independent experiments. (D) Flow cytometric analysis of CD11b expression on BMDMs cultured on gels of indicated stiffness and primed with LPS, primed with LPS and activated with ATP, or left unstimulated. Each symbol represents value from one sample, bar at median, p-values determined by two-way ANOVA indicated significant effect of mechanosensation (MS) and cytokine stimulation (CS), with follow-up pairwise comparisons (***p< 0.001, ****p< 0.0001). Data from 3 independent experiments. (E) Molecular structure of CD11b activator LA-1 . (F) Schematic showing hypothesized activation of integrins on stiff substrate to induce mechanotransduction, compared to LA-1 ligation and activation of CD11b. (G) Summary of experimental plan to test if exposure to LA-1 and incubation on stiffer substrates similarly regulate BMDM activation and polarization.

Article Snippet: For engaging CD11b integrin receptor, specific activator Leukadherin-1 (LA-1 (ADH-503 (GB1275)) from Selleck Chemicals (Cat.#SO525) was administrated in culture 30 min before inflammasome activation (during the last 30 min of LPS-mediated priming.

Techniques: In Vitro, Cell Culture, Incubation, MANN-WHITNEY, Expressing, Activation Assay, Ligation

Journal: Frontiers in Immunology

Article Title: Mechanosensitivity of macrophage polarization: comparing small molecule leukadherin-1 to substrate stiffness

doi: 10.3389/fimmu.2025.1420325

Figure Lengend Snippet: Stiffness of collagen-coated silica gels, but not LA1 treatment, regulates BMDM differentiation. (A) Bone-marrow cells from WT mice were incubated on collagen-coated silica gels or TC plastic in M-CSF (L929-cells supernatant) for three or seven days. Cells were analyzed by flow cytometry for Ly6C and F4/80 to identify populations of monocytes (Ly6C high F4/80 low ) and BMDMs (Ly6C low F4/80 high ). Percentage of BMDMs indicated in each flow plot. (B) Percentage of total cells that were identified as BMDMs. Each symbol represents value from one of three independent experiments, line at median, Kruskal-Wallis with follow-up pairwise comparison used to determine p-values, “P” = plastic. (C) Monocytes were labeled with CellTrace-BV421 at start of incubation. After 3 days in culture, dilution of CellTrace-BV421 was analyzed by flow cytometry. Cells were defined as monocytes or macrophages as shown. (D) Bone marrow cells from WT mice were labeled with CellTrace-BV421, then incubated with or without LA1 (5 μg/ml) for 3 or 7 days. Flow cytometric analysis was used to determine surface expression of CD11b, Ly6C, and F4/80 and dilution of CellTrace-BV421. Monocytes and BMDMs were determined as in (A) . Each symbol shows value for one sample, with each experiment performed with technical triplicates. Line at median. Results of three independent experiments are shown. (E) Dilution of CellTrace-BV421 dilution from cells in (D) . Representative samples from one of three independent experiments shown.

Article Snippet: For engaging CD11b integrin receptor, specific activator Leukadherin-1 (LA-1 (ADH-503 (GB1275)) from Selleck Chemicals (Cat.#SO525) was administrated in culture 30 min before inflammasome activation (during the last 30 min of LPS-mediated priming.

Techniques: Incubation, Flow Cytometry, Comparison, Labeling, Expressing

Journal: Frontiers in Immunology

Article Title: Mechanosensitivity of macrophage polarization: comparing small molecule leukadherin-1 to substrate stiffness

doi: 10.3389/fimmu.2025.1420325

Figure Lengend Snippet: Macrophage surface markers are differentially regulated by integration of mechanical and cytokine cues. BMDMs were incubated on collagen-coated silica gels (open circles, light grey bars: 0.2 kPa; closed circles, dark grey bars: 64 kPa) with LPS+IFNγ, IL-4+IL-13, or without specific cytokine stimulation (none) for 24 h. Expression of indicated surface markers were analyzed by flow cytometry and quantified as median fluorescence intensity (MFI) of all BMDMs gated (CD11b pos F4/80 pos ). Each symbol represents value from one sample; bar shows median value; 95% confidence interval shown. Data were combined from three independent experiments, enabled by consistent fluorophore staining and identical cytometer settings for independent acquisitions. Two-way ANOVA was used to determine if substrate stiffness (MS, mechanosensation) and/or cytokine stimulation (CS) significantly altered marker expression, and to determine if there was a significant interaction of mechanosensation and cytokine stimulation on MFI. No specific pairwise comparisons were tested.

Article Snippet: For engaging CD11b integrin receptor, specific activator Leukadherin-1 (LA-1 (ADH-503 (GB1275)) from Selleck Chemicals (Cat.#SO525) was administrated in culture 30 min before inflammasome activation (during the last 30 min of LPS-mediated priming.

Techniques: Incubation, Expressing, Flow Cytometry, Fluorescence, Staining, Cytometry, Marker

Journal: Frontiers in Immunology

Article Title: Mechanosensitivity of macrophage polarization: comparing small molecule leukadherin-1 to substrate stiffness

doi: 10.3389/fimmu.2025.1420325

Figure Lengend Snippet: Macrophage surface markers are differentially regulated by LA1 treatment and cytokine cues. BMDMs were incubated with (closed circles) or without (gray circles) LA1 (5 µg/ml) and with LPS+IFNγ, IL-4+IL-13, or without specific cytokine stimulation (none) for 24 h. Expression of indicated surface markers were analyzed by flow cytometry and quantified as median fluorescence intensity (MFI) of all BMDMs gated (CD11b pos F4/80 pos ). Each symbol represents value from one sample; bar shows median value. Two-way ANOVA was used to determine if LA1 exposure (LA1) and/or cytokine stimulation (CS) significantly altered marker expression, and to determine if there was a significant interaction of LA1 exposure and cytokine stimulation on MFI. No specific pairwise comparisons were tested. Results from one representative experiment of two or three independent experiments shown; variability across independent acquisitions precluded combining data from multiple experiments. Results from other experiments are shown in Supplementary Figure 2 .

Article Snippet: For engaging CD11b integrin receptor, specific activator Leukadherin-1 (LA-1 (ADH-503 (GB1275)) from Selleck Chemicals (Cat.#SO525) was administrated in culture 30 min before inflammasome activation (during the last 30 min of LPS-mediated priming.

Techniques: Incubation, Expressing, Flow Cytometry, Fluorescence, Marker

Journal: Frontiers in Immunology

Article Title: Mechanosensitivity of macrophage polarization: comparing small molecule leukadherin-1 to substrate stiffness

doi: 10.3389/fimmu.2025.1420325

Figure Lengend Snippet: Interrogation of candidate tyrosine kinases downstream of LA1-mediated CD11b activation. (A) Immunoblot of lysates derived from BMDMs incubated with or without LA1 during final 30 min of LPS priming (4 h) and activation with nigericin (30 min). Densitometry of probed proteins shown below corresponding bands. (B–E) Quantification of (B) phospho-JNK1, (C) phospho-p65 (NF-κB) (D) phospho-ERK1/2, and (E) phospho-p38 normalized to total substrate (JNK1, p65, ERK1/2, or p38, respectively), then actin. Each symbol represents value from one of the 3 independent experiments, line at median, p-values determined using Mann-Whitney. Whole immunoblots with molecular weight markers shown in Supplementary Figure 7 .

Article Snippet: For engaging CD11b integrin receptor, specific activator Leukadherin-1 (LA-1 (ADH-503 (GB1275)) from Selleck Chemicals (Cat.#SO525) was administrated in culture 30 min before inflammasome activation (during the last 30 min of LPS-mediated priming.

Techniques: Activation Assay, Western Blot, Derivative Assay, Incubation, MANN-WHITNEY, Molecular Weight