Parasite Burdens In Bone Marrow Research Articles

Resistance ofLeishmania donovanito Sodium Stibogluconate Is Related to the Expression of Host and Parasite γ-Glutamylcysteine Synthetase

Sequencing studies showed that the gamma-glutamylcysteine synthetase (gamma-GCS) heavy chain genes from sodium stibogluconate (SSG)-resistant (SSG-R) and SSG-susceptible (SSG-S) Leishmania donovani strains were identical, indicating that SSG resistance was related to quantitative differences in gamma-GCS expression rather than gene interstrain polymorphisms. In vitro infection of murine macrophages with the SSG-R strain, but not the SSG-S strain, down regulated expression of host gamma-GCS, which would result in a reduction in intramacrophage glutathione (GSH) levels and promote an oxidative intramacrophage environment. This would inhibit, or minimize, the reduction of SSG pentavalent antimony to its more toxic trivalent form. Macrophage studies showed that the SSG-R strain expressed higher levels of gamma-GCS compared to the SSG-S strain, which would result in higher GSH levels, giving increased protection against oxidative stress and facilitating SSG efflux. However a similar differential effect on host and parasite gamma-GCS expression was not obtained when using tissues from infected mice. In this case gamma-GCS expression was organ and strain dependent for both the host and the parasite, indicating that environmental conditions have a profound effect on gamma-GCS expression. Consistent with the proposed mechanism from in vitro studies, increasing tissue GSH levels in the presence of SSG by cotreatment of L. donovani-infected mice with SSG solution and GSH incorporated into nonionic surfactant vesicles was more effective in reducing liver, spleen, and bone marrow parasite burdens than monotherapy with SSG. Together, these results indicate that SSG resistance is associated with manipulation of both host and parasite GSH levels by L. donovani.

Immune responses of Leishmania donovani infected BALB/c mice following treatment with free and vesicular sodium stibogluconate formulations

The anti-parasitic efficacy of free and a non-ionic surfactant vesicular (NIV) sodium stibogluconate (SSG) formulation of the drug, and their effect on the immune responses of Leishmania donovani infected BALB/c mice, was compared. The SSG NIV formulation maintained a significant suppression of splenic, hepatic and bone marrow parasite burdens ( P<0.005) compared to control values throughout the study. Infected controls and drug treated animals had high levels of L. donovani specific antibodies by day 14 of the study and the titre of these antibodies increased throughout the study for infected controls and free SSG treated animals. Initially SSG NIV treated animals had significantly higher specific IgG2a levels ( P < 0.01, day 16) compared with infected controls and free SSG treated mice, but by day 31 the levels of this isotype and other antibodies (IgG1, IgG3 and IgM) were significantly lower ( P<0.05) than values for the other two groups. There was no difference in the proliferative responses of spleen cells taken from infected controls and drug treated animals to both specific and non-specific stimulation at day 16 of the study. On day 31, only spleen cells taken from infected mice given SSG NIV displayed a significant proliferative response to parasite antigen preparations and Concanavalin A stimulation ( P<0.01). No IL4 was detected in supernatants from in vitro spleen cells cultures. Significant levels of IFN-γ was induced by stimulation of cells from vesicular drug treated animals with a frozen parasite preparation compared with medium controls ( P < 0.05) on day 49 but not day 16. Similar stimulation did not induce IFN γ production in spleen cells from infected controls or free drug treated animals. Only SSG NIV treated animals gave a significant positive DTH response to an L. donovani parasite preparation ( P<0.05) given on day 31. The results of this study indicate that there was a formulation dependant qualitative difference in the post-treatment immune responses of L. donovani infected animals, with SSG NIV animals displaying immune responses expected for a cured phenotype.

Visceral Leishmaniasis in the BALB/c Mouse: A Comparison of thein VivoActivity of Five Non-Ionic Surfactant Vesicle Preparations of Sodium Stibogluconate

Five non-ionic surfactants (Surfactants V-IX) were screened for their ability to produce vesicles for the delivery of sodium stibogluconate. Mean vesicle diameter and antimony content were determined prior to in vivo assessment of antiparasitic activity in a mouse model of acute visceral leishmaniasis. V/D suspensions (i.e. stibogluconate loaded vesicles kept in the hydrating drug solution) were more effective against spleen, liver and bone marrow parasites than drug loaded vesicle suspensions that had unentrapped drug removed. A Surfactant IX V/D suspension was the most active antileishmanial preparation causing 74 +/- 10%, 99 +/- 1% and 38 +/- 8% suppression of liver, spleen and bone marrow parasite burdens respectively. Contrary to previous findings, a reduction in splenic and bone marrow parasite burdens was achieved using large vesicles (mean diameter > 800nm). The significance of these results is discussed.

A direct comparison of sodium stibogluconate treatment in two animal models of human visceral leishmaniasis, mouse and hamster

The effect of sodium stibogluconate therapy, in the free or liposomal form, on the spleen, liver and bone marrow parasite burdens of L. donovani infected mice (BALB/c and B10) and hamsters was studied. Animals were infected i.v. (tail vein in mice, jugular vein in hamsters) and treated on either days 7 and 8 postinfection (acute model) or on days 31 and 32 postinfection or later (chronic model) with free (44.4 mg Sb (v)/kg/day) or liposomal (7.1 mg Sb (v)/kg/day) drug. Six days after the second drug dose animals were killed and parasite burdens assessed. Comparison of the parasite burdens in untreated L. donovani infected mice (B10 and BALB/c) and hamsters showed that higher burdens were obtained in hamsters in all 3 infection sites when the data were expressed as Leishman-Donovan units. However, if the data were expressed as the number of parasites/1000 host cell nuclei, liver burdens were similar in the two species although the parasite burdens remained higher in the spleen and bone marrow of hamsters. In the acute model, both free and liposomal drug treatment significantly reduced liver and spleen parasite burdens in BALB/c mice compared with controls, the liposomal form being significantly more suppressive at a sixth of the free drug dose. The two treatments had little effect on murine bone marrow burdens. In the hamster, the relative efficacies of the free and liposomal forms of the drug in the liver were similar to that in the mouse but in the spleen and bone marrow both treatments markedly reduced parasite numbers. The parasites in the hamster were therefore much more susceptible to drug treatment and this species-dependent effect was most marked in the bone marrow. In mice suffering from the chronic infection the effectiveness of free and liposomal stibogluconate decreased with the length of infection. A similar effect was also demonstrated in hamsters.

Visceral Leishmaniasis: Drug Carrier System Characteristics and the Ability to Clear Parasites from the Liver, Spleen and Bone Marrow in Leishmania donovani Infected BALB/c Mice

The efficacy of various sodium stibogluconate formulations against Leishmania donovani has been investigated using a BALB/c mouse model of visceral leishmaniasis. Only one therapy, multiple dosing with drug loaded sonicated vesicles, liposomes or niosomes, was found to be effective against parasites in the liver, spleen and bone marrow. Other treatments significantly reduced parasite liver burdens but either failed to effect spleen and bone marrow parasites, or were effective but toxic. Prophylactic treatment with sodium stibogluconate preparations, six days before infection, reduced parasite multiplication in the liver (free, niosomal and liposomal drug) and the spleen (sonicated, drug loaded niosomes only), but had no suppressive effect on bone marrow parasite burdens compared with controls. These results indicate that in-vivo sodium stibogluconate persists in some compartments at parasiticidal concentrations and that failure to reach this concentration at some sites of infection such as bone marrow, is the cause of treatment failure and relapse.