Bioimmunotherapy of rodent malaria: co-treatment with recombinant mouse granulocyte-macrophage colony-stimulating factor and an enkephalin fragment peptide Tyr–Gly–Gly

We have earlier shown that recombinant mouse granulocyte-macrophage colony-stimulating factor (rmGM-CSF) and methionine-enkephalin co-treatment can protect mice from malaria. We now report the bioimmunotherapeutic effect of rmGM-CSF and a synthetic enkephalin fragment peptide Tyr–Gly–Gly (TGG) co-treatment on blood-induced Plasmodium berghei infection in Swiss mice. Mice were completely aparasitimic following co-treatment with rmGM-CSF (10.0 μg/kg) and TGG (2.0 mg/kg × 3 per day, intraperitoneally (i.p.)) starting from day −1 to day +4; however, in monotherapy, neither of these agents showed any detectable bioimmunotherapeutic effect. Curiously, similar co-treatment with rmGM-CSF (10.0 μg/kg) and higher doses of TGG (10.0 mg/kg) did not protect the mice. The combined bioimmunotherapeutic effect of these agents was abrogated by the separate administration each of rabbit neutralizing anti-rmGM-CSF antibody, non-selective opioid receptor antagonist naltrexone (10.0 mg/kg × 6 per day, i.p.), and silica (3.0 mg per mouse, intravenously (i.v.)). The peritoneal and splenic macrophages from the protected mice showed a significant (P<0.05) increase in their pool-size and the phagocytic activity, ex vivo. Furthermore, the protected mice, as compared to the unprotected ones, showed a significant (P<0.05) maximum increase in their serum nitrate and nitrite, interferon-γ (IFN-γ), and tumor necrosis factor-α (TNF-α) levels in their splenic homogenates, on the day before the beginning of the resolution of parasitaemia. Selective inhibitors of both inducible (aminoguanidine) and all forms ( -NG-monomethyl arginine) of nitric oxide (NO) synthase, significantly (P<0.05) augmented the mortality of co-treated mice, suggesting the role of NO in protection. These data show that, in P. berghei-infected mice, co-treatment with rmGM-CSF and conditional doses of TGG can impart protection, apparently through partly NO-dependent and macrophage-mediated mechanism(s).