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. 2021 May 20;22(10):5386. doi: 10.3390/ijms22105386

Figure 6.

Figure 6

Mitochondria loaded DCs (Mito-DC) have higher mitochondrial function and immunogenicity after LPS stimulation compared to Veh-DCs. Seven-day old Veh-DC and were incubated with 10 µg/mL HEK293 mitochondria one day before treatment with 100 ng/mL LPS for additional 24 h or left unstimulated (Veh). (A) 24 h after addition of mitochondria, DCs were seeded in a Seahorse XF-24e analyzer, stimulated with and without LPS for 24 h, and oxygen consumption rate (OCR) was determined during sequential treatments with oligomycin (1), FCCP (2) and antimycin A/rotenone (3). (B) Quantification of basal OCR and ATP production. (C) mRNA levels of Pgc1a and Tfam in Veh-DC and Mito-DC treated with 100 ng/mL LPS or left unstimulated for 24 h. (D) Flow cytometry analysis of MHCII, co-stimulatory molecules (CD40/80/86) and PDL1 expression was determined with and without LPS stimulation. (E) ELISA of IL10, IL6, and TNFα from the Veh-DC and Mito-DC treated with 100 ng/mL LPS for 24 h. (F–H) mRNA levels of Tnfa, Il6, and Il10 in Veh-DC and Mito-DC treated with 100 ng/mL LPS or left unstimulated for 24 h. Data represent means ± SEM, * p ≤ 0.05, ** p ≤ 0.01, and *** p ≤ 0.001, one-way ANOVA followed by Tukey’s post-test. Data represent means ± SEM of triplicates.