Co-administration of chloroquine and coenzyme Q10 improved treatment outcome during experimental cerebral malaria

Abstract

Development of cerebral malaria (CM) is driven by parasitemia levels, harmful inflammatory response, oxidative stress and consequent breach of the blood brain barrier. Use of adjunct therapy that utilizes an antioxidant and anti-inflammatory agent alongside chloroquine (CQ), may improve treatment outcome and shorten recovery from post-infection sequelae. Though withdrawn in some countries, CQ is still in use for prophylaxis and treatment of malaria in many countries. Current study investigated whether oral co-administration of 50 mg/kg CQ and 200 mg/kg of coenzyme Q₁₀ (CoQ₁₀) would improve treatment outcome against experimental cerebral malaria (ECM) and assuage the deleterious effects of oxidative stress and inflammation upon infection by Plasmodium berghei ANKA (PbA) in a C57BL/6 J mouse model. Treatment with CQ + CoQ₁₀ resulted in an improved parasite elimination; clearing the parasite one day early, when compared to mice on CQ alone. Remarkably, treatment with CQ and CoQ₁₀ separately or in combination, assuaged PbA induced elevation of serum levels of TNF-α and IFN-γ an indication of protection from ECM progression. Furthermore, CQ and CoQ₁₀-administration, blocked parasite-driven elevation of aspartate transaminase (AST), alanine transaminase (ALT) and bilirubin. In the presence of CQ and CoQ_10, severe PbA-induced systemic induction of oxidative stress and resultant GSH depletion was reduced in the brain, liver, spleen, and kidney. Overall, these findings demonstrate that administration of CQ and CoQ₁₀ ameliorates harmful parasite-driven oxidative stress and inflammation, while slowing the progression to full blown ECM and may improve treatment outcome in CM.

Keywords: Chloroquine; Coenzyme Q10; Experimental cerebral malaria; Inflammation; Oxidative stress.

Fig. 1

Survival curve and line graph of percentage blood cells infected with Plasmodium berghei ANKA (PbA). C57BL6 mice receiving CoQ10 received 200 mg/kg for 30 days prior to infection with PbA, parasitized RBC (pRBC) at 5 X 10⁴ live injected intraperitoneally at day 0 in all groups indicated to receive the parasite. Chloroquine (CQ) 50 mg/kg post infection was administered for 3 days starting from day 4 post infection. Microscopic parasitemia analysis was done on a daily basis and percentage. The survival rate was analyzed with Log-rank (Mantel-Cox) Test. Blood parasitaemia was compared by ANOVA, followed by a Tukey’s post hoc test (P ≤ .0.05). n = 5 mice per group

Fig. 2

CoQ₁₀ in combination with CQ abrogates PbA induced pro inflammatory cytokines (IFN-γ and TNF-α). Following pretreatment by CoQ10 for 30 days, mice were infected with PbA (5 X 10⁴ live, parasitized RBC (pRBC) i.p on day 0 followed by CoQ₁₀ (50 mg/kg p.o for three days). After 11 days of infection, serum from the blood were analyzed and the level of pro and anti-inflammatory cytokines determined. (a) levels of IFN-γ; (b) TNF-α; (c) IL-10; (d) IFN-γ: IL-10 ratio and (e) TNF-α: 1L-10 in the serum were determined by ELISA. Serum level of cytokines was compared by ANOVA, followed by a Tukey’s post hoc test (indicated level of significance: *P ≤ .0.05; **P ≤ .0.01;***P ≤ .0.001). n = 5 mice per group

Fig. 3

Oral administration of CoQ₁₀ and CQ restored glutathione levels in liver, brain, spleen and kidney. C57BL/6 mice were orally administered with 200 mg/kg CoQ₁₀ on a daily basis for 30 days followed by intraperitoneal injection of 5 X 10⁴ PbA pRBC. CQ was orally administered 4 d.p.i. After 11 days of infection, organs were harvested, and their homogenates were prepared to analyze the distribution of GSH endogenous antioxidant. (a) Cellular concentration levels of brain GSH; (b) Liver GSH (c) Spleen GSH and Kidney GSH were determined spectrophotometrically. Cellular level of GSH were compared by ANOVA, followed by a Tukey’s post hoc test (indicated level of significance: *P ≤ .0.05; **P ≤ .0.01;***P ≤ .0.001). n = 5 mice per group

Fig. 4

CoQ₁₀ in combination with CQ supplemented/treated mice show altered levels of liver transaminases enzymes upon PbA infection. Following pretreatment by CoQ10 for 30 days, mice were infected with PbA (5 X 10⁴ live, parasitized RBC (pRBC) i.p on day 0 followed by CoQ₁₀ (50 mg/kg p.o for three days). After 11 days of infection, serum from the blood were analyzed and the level of transaminases and bilirubin determined. (a) Levels of AST; (b) ALT; (c) AST:ALT ratio and (d) Bilirubin in the serum were determined spectrophotometrically. Serum level of transaminases and bilirubin was compared by ANOVA, followed by a Tukey’s post hoc test (indicated level of significance: *P ≤ .0.05; **P ≤ .0.01;***P ≤ .0.001). n = 5 mice per group

Fig. 5

Comparison of serum lipid levels among the groups supplemented with CoQ₁₀, and treated with chloroquine upon PbA infection. After 11 days of infection, serum from the blood were analyzed and the level of lipids determined. (a) Levels of HDL; (b) Cholesterol and (c) Triglycerides in the serum were determined spectrophotometrically. Serum level of lipids was compared by ANOVA, followed by a Tukey’s post hoc test (indicated level of significance: *P ≤ .0.05; **P ≤ .0.01;***P ≤ .0.001). n = 5 mice per group