Evaluation of biocompatibility and mitochondria-targeted behaviors of Zn-NH-pyr. (A) Live/dead staining of macrophages treated with various concentrations of Zn-NH-pyr for 24 h (n = 3 independent samples). (B) Quantitative analysis of Calcein-AM and PI positive cells treated with various concentrations of Zn-NH-pyr for 24 h (mean ± SD, unpaired two-tailed Student's t-test, n = 3 independent samples). (C) CCK-8 assay of macrophages treated with various concentrations of Zn-NH-pyr for 24, 48 and 72 h (mean ± SD, n = 3 independent samples). (D) Cell cycle analysis of macrophages treated with various concentrations of Zn-NH-pyr for 24 h. (E) Hemolysis test of Zn-NH-pyr treated with various concentrations of Zn-NH-pyr for 2 h. (F) Confocal images show that similar to RB-TPP@Zn-NH-pyr, most RB@Zn-NH-pyr escaped from the lysosomes 3 h after addition to cells, despite partial colocalization observed (n = 3 independent samples). (G) Qualitative evaluation of colocalization of Zn-NH-pyr and endo/lysosome. (H) Confocal images show that RB@Zn-NH-pyr, as well as RB-TPP@Zn-NH-pyr, accumulated in mitochondria 3 h after addition to cells (n = 3 independent samples). (I) Qualitative evaluation of colocalization of Zn-NH-pyr and mitochondria. ** and ##
P < 0.01 compared to the 0 μg/mL group.