Hepatotoxicity of pentavalent antimonial drug: possible role of residual Sb(III) and protective effect of ascorbic acid

Abstract

Pentavalent antimonial drugs such as meglumine antimoniate (Glucantime [Glu; Sanofi-Aventis, São Paulo, Brazil]) produce severe side effects, including cardiotoxicity and hepatotoxicity, during the treatment of leishmaniasis. We evaluated the role of residual Sb(III) in the hepatotoxicity of meglumine antimoniate, as well as the protective effect of the antioxidant ascorbic acid (AA) during antimonial chemotherapy in a murine model of visceral leishmaniasis. BALB/c mice infected with Leishmania infantum were treated intraperitoneally at 80 mg of Sb/kg/day with commercial meglumine antimoniate (Glu) or a synthetic meglumine antimoniate with lower Sb(III) level (MA), in association or not with AA (15 mg/kg/day), for a 20-day period. Control groups received saline or saline plus AA. Livers were evaluated for hepatocytes histological alterations, peroxidase activity, and apoptosis. Increased proportions of swollen and apoptotic hepatocytes were observed in animals treated with Glu compared to animals treated with saline or MA. The peroxidase activity was also enhanced in the liver of animals that received Glu. Cotreatment with AA reduced the extent of histological changes, the apoptotic index, and the peroxidase activity to levels corresponding to the control group. Moreover, the association with AA did not affect the hepatic uptake of Sb and the ability of Glu to reduce the liver and spleen parasite loads in infected mice. In conclusion, our data supports the use of pentavalent antimonials with low residue of Sb(III) and the association of pentavalent antimonials with AA, as effective strategies to reduce side effects in antimonial therapy.

FIG 1

Parasite loads in the livers (A) and spleens (B) of BALB/c mice infected with L. infantum after 20 days of treatment with commercial (Glu) or synthetic meglumine antimoniate (MA) (80 mg of Sb/kg/day) or saline by the i.p. route. The data are shown as dot plots, and lines correspond to the median of each group (n = 6 to 7). *, P < 0.05 (Kruskal-Wallis, followed by Dunn's multiple-comparison post test).

FIG 2

Histopathology (D to F) and apoptosis (A to C) analysis in the livers of L. infantum-infected BALB/c mice treated as follows: A and D, Glu, given i.p. at 80 mg of Sb/kg/day; B and E, MA, given i.p. at 80 mg of Sb/kg/day; or C and F, saline, given i.p. Histology was evaluated by H&E staining. Apoptosis was assessed by TUNEL. Cells were considered positive when they showed a strong nuclear staining recognized by the presence of dark brown lumps and morphological changes characteristic of apoptosis (see Materials and Methods for details). Bar, 20 μm.

FIG 3

Effect of treatment of L. infantum-infected BALB/c mice with commercial (Glu) or synthetic (MA) meglumine antimoniate using different liver parameters: A, lucent area in hepatocytes; B, apoptotic index of hepatocytes; and C, peroxidase activity. Mice were treated with Glu or MA (80 mg of Sb/kg/day) or saline for 20 days by the i.p. route. Noninfected mice receiving saline were also used as controls. The data are means ± the standard errors of the mean (SEM; n = 3 to 7). ***, P < 0.001 (one-way ANOVA with Bonferroni post test).

FIG 4

Influence of AA on the liver parameters of L. infantum-infected BALB/c mice treated or not to with commercial meglumine antimoniate (Glu). The liver parameters measured included the percentage of lucent area in the hepatocytes (A), the apoptotic index of the hepatocytes (B), and the peroxidase activity (C). Mice were treated with Glu (80 mg/kg/day), AA (15 mg/kg/day), Glu+AA, or saline alone for 20 days by the i.p. route. Data are means ± the SEM (n = 3 to 7). ***, P < 0.001; **, P < 0.01 (one-way ANOVA with Bonferroni post test).

FIG 5

Parasite loads in the liver (A) and spleen (B) in BALB/c mice infected with L. infantum after 20 days of treatment with Glu (80 mg of Sb/kg/day) in the presence or absence of AA (15 mg/kg/day) by the i.p. route. The data are shown as dot plots, and lines correspond to the median of each group (n = 6 to 7). **, P < 0.01 (Mann-Whitney test).