c3a  (R&D Systems)

Journal: Journal of Neuroinflammation

Article Title: Microglia undergo sex-dimorphic transcriptional and metabolic rewiring during aging

doi: 10.1186/s12974-024-03130-7

Figure Lengend Snippet: Microglial C3a – C3aR signaling promotes glycolysis and phagocytosis. Young microglia (postnatal day 0–2) were treated with 10 nM recombinant mouse C3a. ( a - d ) p-AKT (Ser473), p-mTOR (Ser2448) and HIF1α were assessed by Western blotting at the indicated timepoints (0–24 h), and expression was normalized to total AKT, total mTOR and β-actin, respectively. n = 5 replicates. Data are presented as mean (SEM). * p < 0.05, ** p < 0.01, *** p < 0.001 (one-way ANOVA). ( e - g ) Seahorse assays were used to evaluate real-time glycolytic rate in young microglia (postnatal day 0–2, pooled male and female) after 18 h in vitro treatment with 10 nM recombinant mouse C3a. Stimuli were added as indicated ( e ), and basal glycolysis ( f ) and compensatory glycolysis ( g ) were determined by calculating the glycolytic Proton Efflux Rate (e; glycoPER). n = 5/group. # p < 0.05 (unpaired t-test). ( h , i ) Flow cytometry analysis was performed to assess phagocytosis of FITC-fAβ 1−42 . Rapamycin (50 µM) or 2-DG (5 mM) were pre- (1 h) and co-treated (18 h) with C3a. Proportion of phagocytic cells ( h ) was assessed by evaluating FITC-positive microglia, and FITC MFI was evaluated in total live microglia ( i ). Data are presented as mean (SEM). * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001 (one-way ANOVA)

Article Snippet: Primary microglia were stimulated with vehicle (PBS) or recombinant mouse C3a (10 nM; Peprotech) for the indicated time as in each figure legend.

Techniques: Recombinant, Western Blot, Expressing, In Vitro, Flow Cytometry

Journal: Journal of Inflammation Research

Article Title: Macrophage-Myofibroblast Transition as a Potential Origin for Skeletal Muscle Fibrosis After Injury via Complement System Activation

doi: 10.2147/JIR.S450599

Figure Lengend Snippet: MMT was induced in BMDMs with the stimulation of C3a. ( A ) The representative immunofluorescence image demonstrated the location and relative expression of F4/80. Scale bar = 100μm. ( B ) The representative immunofluorescence image demonstrated the relative expression of Col 1. Scale bar = 50μm. ( C ) Quantification of relative fluorescence intensity of Col 1. (n=6) ***p<0.001. ( D ) α-SMA and Col 1 expression level was determined by Western blot after different treatment. ( E and F ) Quantification of relative protein expression. (n=6) Data was presented as mean ± SD. *p<0.05, **p<0.01.

Article Snippet: For C3a (novoprotein, No.: CM99) stimulation, 10 ng/mL recombinant mouse C3a was added into the medium for 72h and then collected for measurements.

Techniques: Immunofluorescence, Expressing, Fluorescence, Western Blot