Cell viability of A375 cells with TF treatment at 24, 48, and 72 h (A), cell viability of HFF-1, A375, and A875 cells with TF treatment at 24 h (B), and morphology of A375 cells with theaflavin (TF) treatment at 24 h (C). Data were mean ± SD (n = 5). By means of Fisher’s least significant difference (LSD) multiple comparisons, data (mean ± SD) with same lowercase letter (b vs. bc; bc vs. cd; c vs. c; cd vs. d; d vs. d) indicate no significant difference between each other, while data with different letters (a vs. b vs. c vs. b vs. e vs. f vs. g) indicate significant difference with each other. Scale bar = 200 μm.

DAPI staining observation (A) and flow cytometry analysis (B) on A375 cells with theaflavin (TF) treatment. Statistical analysis of apoptotic cell rate (C), statistical analysis of early apoptotic rate (D) and statistical analysis of late apoptotic rate (E). By means of Fisher’s least significant difference (LSD) multiple comparisons, data (mean ± SD) with same lowercase letter (a vs. ab; ab vs. b; b vs. b; c vs. c) indicate no significant difference between each other, while data with different letters (a vs. b vs. c) indicate significant difference with each other. Scale bar = 100 μm.

Mortality and adverse events of larval zebrafishes induced by theaflavin (TF) and observation of larval zebrafishes xenotransplanted with A375 cells with treatment of TF or cisplatin (A) as well as the fluorescence intensity and inhibitory rates of TF (B). Fluorescent area in red represents the A375 cell mass. By means of LSD multiple comparisons, data (mean ± SD) with same lowercase letter (a vs. a; c vs. c) indicate no significant difference between each other, while data with different letters (a vs. b vs. c) indicate significant difference with each other.

Relative mRNA expression of theaflavin (TF)-targeted genes in A375 cells after 24 h treatment. By means of Fisher’s least significant difference (LSD) multiple comparisons, data (mean ± SD) with same lowercase letter (a vs. a; b vs. b; c vs. c) indicate no significant difference between each other, while data with different letters (a vs. b vs. c vs. d) indicate significant difference with each other.

Expression and phosphorylation of theaflavin (TF)-targeted proteins in A375 cells after 24 h treatment. Data (mean ± SD) with different lowercase letters are significantly different with each other at Fisher’s least significant difference (LSD) multiple comparisons.