Figure 6
Combined effect of histamine and doxorubicin on triple-negative breast tumors induced in nude mice. (A) Relative tumor volume of the control group versus doxorubicin (Dox, 2 mg/kg) or the combination of Dox (2 mg/kg) and histamine (HA, 5 mg/kg). (6 mice per group, **P<0.01 versus Control; T-Test). (B) Median tumor doubling time of each group is depicted numerically (*P<0.05, ***P<0.001 versus Control; ##P<0.01 versus Dox. T-Test). (C) Histopathological and immunohistochemical analyses of tumor tissues. (a–c) Representative H&E-stained sections are shown. (a) Untreated tumors presented undifferentiated adenocarcinoma cells with marked anisokaryosis and anisocytosis. (b) Dox increased necrosis and the nuclear optical density. (c) The combined treatment Dox+HA reduced tumor cellularity, with evident nuclear hyperchromasia, that was replaced by extracellular matrix. Representative immunohistochemical images of (d–f) TUNEL, (g–i) PCNA and (j–l) 8-dOHG in paraffin-embedded tumor tissues. x630 Original magnification. Scale bar, 20 μm. (D) The number of tumor cells and the percentage of TUNEL, PCNA and 8-OHdG-positive stained cells were quantified by counting 10 random fields. (*P<0.05, ***P<0.001 versus Control; ##P<0.01, ###P<0.001 versus Dox). (E) Representative H&E stained sections of heart and liver are shown. (a) Normal histological appearance of untreated heart. (b) Heart of Dox-treated animals displaying severe myocytolysis (red arrow), areas of necrosis (black arrow) and reduced striated muscle bands. (c) Heart of Dox+HA-treated animals showing preservation of the structure with reduced myocytolysis, nuclei and muscle bands with normal appearance. (d) Normal histological appearance of untreated liver. (e) Liver of Dox-treated animals displaying focal necrosis (black arrow), sinusoidal atrophy (white arrow), inflammatory infiltrates, and fibrosis (red arrow). (f) Liver of Dox+HA-treated animals showing reduced sinusoidal disarrangement, displaying similar characteristics of the untreated liver. x1000 Original magnification. Scale bar, 20 μm.
