Journal: PLOS Pathogens
Article Title: Glutamine alleviates Staphylococcus aureus -induced mastitis by modulating macrophage polarisation
doi: 10.1371/journal.ppat.1014053
Figure Lengend Snippet: Mice were orally administered αKG prior to establishment of a S. aureus -induced mastitis model. Mammary gland tissues were subsequently collected for analysis (n = 7 per group). A: Mitochondrial superoxide levels in frozen mammary gland tissue sections were assessed using MitoSOX Red staining, with green fluorescence indicating ROS. B: ATP levels in mammary gland tissues from Control, S. au, and αKG + S. au . C-D: OCR of mammary gland tissue cells in the three groups. E: Expression of LC3, ATG5, and p62 in protein lysates of mammary tissues from the three groups. G-H: RAW264.7 cells cultured with αKG were treated with oligomycin A and stimulated with S. aureus . Inflammatory cytokine levels and NF-κB signaling pathway activity of Control, S. au, αKG + S. au, αKG + S. au +oligomycin A were measured. I-P: qPCR analysis of M2 and M1 marker gene mRNA expression in four experimental groups of RAW264.7 cells. The data are presented as mean ± SD. One-way analysis of variance was performed for statistical analysis. * P < 0.05, ** P < 0.01 and *** P < 0.001 indicate significance from each group. ns, no significance.
Article Snippet: The specific primary antibodies used in the experiment included p65 (#8242, Cell Signaling Technology, USA), p-p65 (#3033, Cell Signaling Technology, America), IκB (AF5002, Affinity Biosciences, Jiangsu, China), p-IκB (AF2002, Affinity Biosciences, Jiangsu, China), ZO-1 (AF5145, Affinity Biosciences, Jiangsu, China), Occludin (DF7504, Affinity Biosciences, Jiangsu, China), Claudin-3 (AF0129, Affinity Biosciences, Jiangsu, China), ATG5 (bsm-52596R, Bioss, Woburn, MA, USA), LC3-I/LC3-II (AF5402, Affinity Biosciences, Jiangsu, China), p62 (ab91526, Abcam plc, Cam bridge, UK), SOCS3 (bs-0580R, Bioss, Woburn, MA, USA), JAK1 (bs-1439R , Bioss, Woburn, MA, USA), p-JAK1 (bs-3238R, Bioss, Woburn, MA, USA), STAT3 (AF6294, Affinity Biosciences, Jiangsu, China), p-STAT3 (AF3293, Affinity Biosciences, Jiangsu, China) and β-actin (T0022, Affinity Biosciences, Jiangsu, China).
Techniques: Staining, Fluorescence, Control, Expressing, Cell Culture, Activity Assay, Marker
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