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. 2023 Oct 4;33(1):78–90. doi: 10.1093/hmg/ddad162

Figure 5.

Figure 5

DYRK1A inhibitor, harmine reduces EGR1 mRNA expression in T21-EC via PPARG pathway and restores angiogenesis and mitochondrial respiratory function. (A) Relative mRNA expressions of DYRK1A in T21- and T21’-ECs, evaluated by quantitative PCR. Approximately 1.5-fold higher expression is observed both in T21- and T21’-ECs as compared with cDi21-ECs. (B) Relative mRNA expression of PPARG in cDi21-ECs, T21-, and T21’-ECs treated with DMSO or harmine; the latter significantly increases PPARG expression in T21- and T21’-ECs, but not in cDi21-ECs. (C) Relative mRNA expression of EGR1 treated with DMSO or harmine; the latter significantly reduces EGR1 expression in T21- and T21’-ECs. (D) Representative phase contrast images of tube formation assays in each cell line and condition. Scale bar: 500 μm. (E) Quantitative analyses of tube formation assays reveal that angiogenesis is significantly recovered in harmine-treated T21- and T21’-ECs as same level as that of cDi21-ECs. (F) OCR of each cell line and condition at baseline and after sequential treatment of oligomycin (Oligo), FCCP, and Rotenone/Antimycin A (R/AA). (G) Quantitative analyses of basal respiration and maximal respiration show significant improvement of OCR in harmine-treated T21- and T21’-ECs. (H) Quantitative analyses of annexin V-positive and PI-negative cell populations show that apoptotic cells are significantly decreased in harmine-treated T21- and T21’-ECs, similar to cDi21-ECs. (I) Quantitative measurements of MitoSOX fluorescence intensity show that the level of mitochondrial superoxide production is significantly decreased in harmine-treated T21- and T21’-ECs, which is similar to the level in cDi21-ECs. Three to four independent experiments were conducted in each. Data are presented as mean ± SEM. Data were analyzed by Tukey–Kramer multiple comparison test. ***P < 0.001, **P < 0.01, *P < 0.05.