Fig. 4: In vitro characterization of HMGB-1 stimulated macrophages.
From: The immune checkpoint TIM-3/HMGB-1 axis in myocardial infarction
A In vitro model of macrophage differentiation from monocytes to unstimulated macrophage (Mφ) and then to pro-inflammatory (M1) or anti-inflammatory (M2) macrophages. B Gene expression of classical M1 (IL6 and CXCL10) and M2 (FN1) markers. C Expression of SPP1 and CCL18 showing M1 and M2 mimicking of transcriptomics macrophage clusters. D Schematic of HMGB-1 and receptor blockade experiments. E Expression of M1 and M2 macrophage markers in macrophages stimulated with 5 µg/mL HMGB-1 for 48 hours. F TIM-3, RAGE and TLR4 expression in macrophages stimulated with 5 µg/mL HMGB-1 for 48 hours. G Quantification of TIM-3 and TLR-4 expression in macrophages stimulated with HMGB-1 compared to RAGE based on the 2-ΔCt. H Expression of IL6 upon stimulation with 5 µg/mL HMGB-1 for 48 hours with or without preincubation of 10 µg/mL neutralizing antibodies against TIM-3, RAGE or TLR4. I Expression of inflammasome pathway markers CASP1, IL1B and IL18. qPCR expression was normalized by RSP9 and data is presented as mean ± standard error of the mean (SEM). HMGB-1 High mobility group box-1; TIM-3 T-cell immunoglobulin and mucin-domain containing protein-3; RAGE Receptor for advanced glycation endproducts; TLR4 toll-like receptor 4.
