growth factor β1  (MedChemExpress)

Journal: Advanced Science

Article Title: Neutrophil Mobilization Triggers Microglial Functional Change to Exacerbate Cerebral Ischemia‐Reperfusion Injury

doi: 10.1002/advs.202503722

Figure Lengend Snippet: MRP14 inhibits microglia to phagocytize neutrophils and promotes microglial pyroptosis after ischemic stroke. A,B) Immunofluorescence staining showing the percentage of neutrophils (green) touching or surrounded by microglia (red) in the ischemic brain of WT and MRP14 KO groups (n = 6/group). The white arrow indicates neutrophils touching or surrounded by microglia. C) The neutrophil numbers per field in the ischemic brain of WT and MRP14 KO groups (n = 6/group). D,E) The flow cytometry analysis showing the percentage of Ly6G + microglia in the ischemic brain of the tMCAO mice from the WT and MRP14 KO groups (n = 6/group). IgG was used as a negative control. F,G) Immunofluorescence staining showing the expression of CD68 (green) in the microglia (red) in the ischemic brain of WT and MRP14 KO groups (n = 6/group). H‐L) Western blot analysis of the expression of GSDMD, NLRP3, pro‐caspase 1, caspase 1, pro‐IL‐1 β and IL‐1 β in the ischemic brain of WT or MRP14 KO mice 24 h after tMCAO (n = 6/group). M,N) Immunofluorescence staining showing the expression of GSDMD (green) in the microglia (red) in the ischemic brain of WT or MRP14 KO mice 24 h after tMCAO (n = 6/group). Data are presented as means ± SEM; Mann‐Whitney test, unpaired t‐test, one‐way ANOVA. **P < 0.01. Scale bar: 25 µm (A and F); 50 µm (M). tMCAO, transient middle cerebral artery occlusion; WT, wild type; KO, knockout; veh, vehicle; paq, paquinimod; GSDMD, gasdermin D; NLRP3, NOD‐like receptor family, pyrin domain containing 3; IL‐1 β , interleukin‐1 β ; Casp1, caspase 1.

Article Snippet: Peripheral neutrophils were treated with recombined IL‐1 β (MedChemExpress, USA) at a concentration of 10 ng mL −1 for 4 h, and anakinra (10 μg mL −1 , MedChemExpress, USA) or PD 169316 (10 μM, MedChemExpress, USA) was added 30 min before IL‐1 β treatment.

Techniques: Immunofluorescence, Staining, Flow Cytometry, Negative Control, Expressing, Western Blot, MANN-WHITNEY, Knock-Out

Journal: Advanced Science

Article Title: Neutrophil Mobilization Triggers Microglial Functional Change to Exacerbate Cerebral Ischemia‐Reperfusion Injury

doi: 10.1002/advs.202503722

Figure Lengend Snippet: Conditional Knockout of microglial TLR4 alleviates MRP14‐induced impaired phagocytosis and NLRP3 inflammasome‐associated pyroptosis after ischemic stroke. A,B) The schematic diagram showing the breeding strategy for generating mice which were specifically lacking TLR4 expression in microglia (TLR4 flox/flox ; CX3CR1‐CreERT). C,D) Verification of conditionally knockout of TLR4 expression in TLR4 flox/flox ; CX3CR1‐CreERT mice (n = 6/group). E,F) Immunofluorescence staining showing the percentage of neutrophils (green) touching or surrounded by microglia (red) in the ischemic brain of TLR4 flox/flox or TLR4 flox/flox ; CX3CR1‐CreERT mice treated with vehicle or recombined MRP14 protein (0.6 mg kg −1 ) 24 h after tMCAO (n = 6/group). G) The neutrophil numbers per high‐power field in the ischemic brain of TLR4 flox/flox or TLR4 flox/flox ; CX3CR1‐CreERT mice treated with vehicle or recombined MRP14 protein (0.6 mg kg −1 ) 24 h after tMCAO (n = 6/group). H‐L) Western blot analysis of the expression of GSDMD, NLRP3, pro‐caspase 1, caspase 1, pro‐IL‐1 β and IL‐1 β in the ischemic brain of TLR4 flox/flox or TLR4 flox/flox ; CX3CR1‐CreERT mice treated with vehicle or recombined MRP14 protein (0.6 mg kg −1 ) 24 h after tMCAO (n = 6/group). M,N) Immunofluorescence staining showing the expression of GSDMD (green) in the microglia (red) in the ischemic brain of TLR4 flox/flox or TLR4 flox/flox ; CX3CR1‐CreERT mice treated with vehicle or recombined MRP14 protein (0.6 mg kg −1 ) 24 h after tMCAO (n = 6/group). Data are presented as means ± SEM; unpaired t‐test, one‐way ANOVA. *P < 0.05, **P < 0.01. Scale bar, 50 µm. TLR4, Toll‐like receptor 4; veh, vehicle; TLR4 f/f , TLR4 flox/flox ; GSDMD, gasdermin D; NLRP3, NOD‐like receptor family, pyrin domain containing 3; IL‐1 β , interleukin‐1 β ; Casp1, caspase 1.

Article Snippet: Peripheral neutrophils were treated with recombined IL‐1 β (MedChemExpress, USA) at a concentration of 10 ng mL −1 for 4 h, and anakinra (10 μg mL −1 , MedChemExpress, USA) or PD 169316 (10 μM, MedChemExpress, USA) was added 30 min before IL‐1 β treatment.

Techniques: Knock-Out, Expressing, Immunofluorescence, Staining, Western Blot