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It is known that premature elimination of non-parasitized RBCs (nRBCs) plays an important role in the pathogenesis of malarial anemia, in which suicidal death process (eryptosis) of nRBCs has been suggested to be involved. To check this possibility, we investigate eryptosis during infection of P. berghei ANKA in Wistar rats, a malaria experimental model that, similar to human malaria, the infection courses with low parasitemia and acute anemia. As expected, P. berghei ANKA infection was marked by low parasite burdens that reached a mean peak of 3% between days six and nine post-infection and solved spontaneously. A significant reduction of the hemoglobin levels (~ 30%) was also observed on days subsequent to the peak of parasitemia, persisting until day 16 post-infection. In eryptosis assays, it was observed a significant increase in the levels of PS-exposing nRBC, which coincided with the reduction of hemoglobin levels and was positively related to anemia. In addition to PS externalization, eryptosis of nRBC induced by P. berghei infection was characterized by cytoplasm calcium influx, but not caspases activity. These results confirm our previous studies evidencing a pro-eryptotic effect of malaria infection on nRBCs and show that a caspase-independent eryptotic process is implicated in anemia induced by P. berghei ANKA infection in Wistar rats.

Background: Glucose homeostasis derangement is a common pathophysiology of malaria whose aetiology is still controversial. The Plasmodium parasite, immunological and inflammatory responses, as well as chemotherapeutics currently used cause hypoglycaemia in malaria. Anti-parasitic and anti-disease drugs are required to combat malaria while ameliorating the pathophysiology of the infection. Asiatic acid has anti-hyperglycaemic, antioxidant, pro-oxidant properties useful in glucose homeostasis but its influence in malaria is yet to be reported. Here we present findings on the influence of asiatic acid on glucose metabolism in vivo using P. berghei -infected Sprague Dawley rats., Materials and Methods: Acute as well as sub-chronic studies were carried out in vivo where physicochemical properties and glucose homeostasis were monitored after administration of asiatic acid (10mg/kg) in both non-infected and infected animals. Glucose metabolism associated biochemical changes in malaria were also investigated., Results: In acute studies, asiatic acid improved oral glucose response while in the sub-chronic state it maintained food and water intake and suppressed parasitaemia. Normoglycaemic control was maintained in infected animals through insulin suppression and increasing glucagon secretion, in both acute and chronic studies. Asiatic acid administration curtailed lactate concentration towards normal., Conclusion: Per oral post-infection asiatic acid administration preserved drinking and eating habits, inhibited sickness behaviour while suppressing parasitaemia. Reciprocal relationship between insulin and glucagon concentrations was maintained influencing glucose homeostasis positively and inhibition of hyperlactaemia in malaria. Abbreviations: ip -intraperitoneal, po -per oral, ig -intragastric, AA-Asciatic acid, OGTT-oral glucose tolerance test, OS-oxidative stress, ROS-reactive oxygen species, NO-nitric oxide, ONOO - - peroxynitrite, BRU-Biomedical Research Unit, SD-Sprague Dawley., Competing Interests: Conflict of interest The authors declare no conflict of in interest in this work.

Individuals in natural populations may be infected with multiple different parasites at a time. These parasites may interact with each other or act independently in the host, and this may result to varying outcomes on host health and survival. This study therefore aimed at investigating the health impact of co-infection of mice with Plasmodium berghei and Trypanosoma brucei. Forty Swiss albino mice (14-17g) were divided into four groups of ten. Mice in groups A and B received 10(6)P. berghei and groups B and C 10(5)T. brucei, while group D were uninfected. The co-infected mice had higher P. berghei and T. brucei parasitaemia, compared with the mono-infected mice. The co-infected mice had significantly (p<0.05) lower survival rate compared with the mono-infected mice. Co-infection of mice with P. berghei and T. brucei resulted in rapid P. berghei and T. brucei development and increased parasitaemia. The leukocyte numbers significantly (p<0.05) reduced on days 12 and 15 post infection among P. berghei infected mice, in the presence or absence of T. brucei. Anaemia and hypoglycaemia was more severe in the co-infected mice. Therefore, co-infection of mice with P. berghei and T. brucei may increase pathologic impact to the host by increasing parasitaemia., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Background: The objective of this study was to explore effects of khat (Catha edulis) on outcome of rodent malaria infection and its anti-plasmodial activities on Plasmodium berghei ANKA (PbA)., Methods: Female Swiss albino mice were orally treated with crude khat (Catha edulis) extracts (100, 200 and 300 mg/kg) on a daily basis for 4 weeks prior to PbA infection. Physical, clinical, hematological, biochemical and histo-pathological features of the mice were assessed. In addition, in vivo anti-plasmodial activities of khat were evaluated., Results: The finding of this study showed that khat use was strongly associated with increment of levels of liver and kidney biomarkers, leucopenia, severe anemia, rise in level of inflammation biomarkers: C-reactive protein (CRP), uric acid (UA), increased monocyte-lymphocyte count ratio (MLCR), manifestation of cerebral malaria symptoms such as ataxia, paralysis and deviation of the head but with no pulmonary edema. Significantly lower level of parasitemia (P<0.05), rectal temperature, but, high level of hemoglobin were observed at the early stage of the PbA infection in khat treated mice than the control. With extension of the treatment period, however, drastic increments were observed in parasite load and rectal temperature although there was reduction in hemoglobin (Hb) level. Moreover, khat showed poor anti-plasmodial activity with <10% parasite suppression activity and lack protection against major malaria symptoms. The significant reduction (P<0.01) of hematological parameters during PbA infection strengthen the notion that hematological parameters could be good predictors of severe malaria complications in human., Conclusions: In mice model treated with khat prior to infection with the rodent malaria parasite, khat was found to worsen manifestation of most malaria complications. Furthermore, the same plant showed poor in vivo anti-plasmodial activity and protection against major malaria symptoms.

Degenerative and demyelinating diseases are known to alter the mechanical properties of brain tissue. While few studies have characterized these biomechanical changes, it is clear that accurate characterization of the mechanical properties of diseased brain tissue could be a substantial asset to neuronavigation and surgery simulation through haptic devices. In this study, samples of brain tissue from rats infected with Plasmodium berghei ANKA, an African murine malaria parasite, are evaluated using a uniaxial tensile test machine. Infected brains having different levels of parasitemia are mounted on the testing machine and extended until failure of the tissue. The stress-strain curve of each sample is obtained and compared to healthy rat brain tissue. Young's modulus of each sample is extracted from the Hookean part of the stress-strain diagram. Young's modulus of rats' brain shows considerable difference among the samples having various levels of parasitemia compared with the controls. For instance, the brains with 0% (control), 1.5%, and 9% parasitemia showed a Young's modulus of 46.15, 54.54, and 266.67 kPa, respectively. This suggests sequestration of the stiffened and less deformable parasitized red blood cells in the brain microvasculature.

Cerebral malaria (CM) is a severe neurologic complication that arises predominantly in children and non-immune adults infected with Plasmodium falciparum. In the current study, the dynamics of CD4(+)CD25(+)Foxp3(+) regulatory T cells (Tregs) and pro- and anti-inflammatory cytokines were analyzed in P. berghei ANKA (P.bANKA)-infected C57BL/6, BALB/c, and DBA/2 mice. We showed that C57BL/6 mice were susceptible to CM, while BALB/c and DBA/2 mice were resistant to CM and succumbed to hyperparasitemia and severe anemia. The proportion and absolute numbers of Tregs in BALB/c and DBA/2 mice were significantly higher than in C57BL/6 mice. The levels of pro-inflammatory cytokines, such as IFN-gamma, TNF-alpha, IL-6, IL-17 and NO in CM-susceptible C57BL/6 mice were obviously higher than in CM-resistant BALB/c and DBA/2 mice, while the level of the anti-inflammatory cytokine IL-10 was the opposite to that of pro-inflammatory cytokines, confirming that an appropriate balance between pro- and anti-inflammatory immune responses is essential to control the pathogenesis of severe malaria, and Tregs are important regulators if this balance is to be maintained. In vivo depletion of Tregs significantly protected C57BL/6 mice from experimental CM and the production of pro- and anti-inflammatory cytokines was reversed, indicating that this cell population contributes to pathogenesis by modulating the balance of pro- and anti-inflammatory responses. Our data demonstrate that Tregs mediate the incidence and outcome of CM in P.bANKA-infected mice by modifying the pro-inflammatory response.

Background: The relationship between malaria and obesity are largely unknown. This is partly due to the fact that malaria occurs mainly in tropical areas where, until recently, obesity was not prevalent. It now appears, however, that obesity is emerging as a problem in developing countries. To investigate the possible role of obesity on the host-parasite response to malarial infection, this study applied a murine model, which uses the existence of genetically well characterized obese mice., Methods: The receptivity of obese homozygous ob/ob mice was compared to the receptivity of control heterozygous ob/+ lean mice after a single injection of Plasmodium berghei ANKA sporozoites. Both parasitaemia and mortality in response to infection were recorded., Results: The control mice developed the expected rapid neurological syndromes associated with the ANKA strain, leading to death after six days, in absence of high parasitaemia. The obese mice, on the other hand, did not develop cerebral malaria and responded with increasing parasitaemia, which produced severe anemia leading to death 18-25 days after injection., Conclusion: The observed major differences in outward symptoms for malarial infection in obese versus control mice indicate a link between obesity and resistance to the infection which could be addressed by malariologists studying human malaria.

Background and Aim: Previous studies using isolated perfused rat liver in vivo have suggested that during the erythrocytic phase of malaria infection, overall phagocytosis by Kupffer cells is enhanced. The aim of the present study was to further investigate the individual phagocytic capacity and prostaglandin E(2) (PGE(2)) secretion of isolated Kupffer cells in vitro, and the immunohistochemical characteristics of Kupffer cells in vivo., Methods: Malaria was induced in male Sprague-Dawley rats (n = 12) by inoculation with parasitized red cells containing Plasmodium berghei. Kupffer cells were isolated by centrifugal elutriation., Results: A significantly increased yield of Kupffer cells was obtained from malaria-infected livers compared to controls (36.7 +/- 4.5 vs 11.8 +/- 1.1 x10(6) cells, P < 0.0001, n = 12). There was an increased internalization by phagocytosis of [(3)H]-BSA latex microspheres after 60 min in malaria-infected Kupffer cells compared to controls (65.05 +/- 1.5 vs 48.6 +/- 0.7, P < 0.001, n = 12). PGE(2) secretion into the cell culture medium was significantly suppressed in malaria-infected Kupffer cells compared to controls (1167 +/- 88 vs 4537 +/- 383 pg per 10(6) cells, P < 0.001, n = 5). Staining of ED1, ED2 and PCNA was greater in malaria-infected livers compared to control., Conclusion: The results indicate that the number of Kupffer cells is significantly increased and their phagocytic activity on a cell-by-cell basis is enhanced during the erythrocytic stage of malaria.

Objective: The effect of castration and subsequent replacement of dehydroepiandrosterone (DHEA) or estradiol on parasitemia, mortality and incidence of cerebral malaria (CM) was evaluated in CBA mice infected with Plasmodium berghei ANKA., Methods: Female mice were castrated, and groups of 12-15 animals received daily injections of DHEA, estradiol or saline. Four days after the start of treatment, mice were inoculated with 1 x 10(6)P. berghei ANKA-parasitized erythrocytes. DHEA treatment was continued during the 5 days after infection, and estradiol was administered during the follow-up. Parasitemia was evaluated daily in Giemsa-stained blood smears. Signs of CM were determined by the manifestation of coma, limb paralysis and/or convulsions. Plasma TNF-alpha levels were evaluated by sandwich ELISA. Nitric oxide synthase (NOS) activity in the brain of moribund mice was measured by the method of Bredt and Snyder., Results: In non-castrated infected mice, the incidence of CM was 50%, and plasma TNF-alpha increased and brain NOS activity decreased compared to non-infected controls. Castration had no major effect on the parameters analyzed (parasitemia, mortality, CM incidence, TNF-alpha levels or NOS activity). Estradiol replacement caused a decrease in parasitemia but resulted in higher CM incidence and faster mortality, with an increase in NOS activity., Conclusions: Estradiol modulated the immune response of P. berghei ANKA-infected CBA mice, decreasing parasitemia and increasing NOS activity, and impacted negatively on survival and CM incidence, showing that neuroimmunoendocrine interactions are important in the physiopathogenesis of malaria infections.

In order to study the effects of acclimatization of Plasmodium in beta-thalassaemic mice, we used a mouse model of beta-thalassaemia (DBA/2J/beta-thal/beta-thal), similar to that observed in humans. We acclimatized 3 rodent malarias (P. berghei, P. chabaudi and P. yoelii) in DBA/2J and DBA/2J/beta-thal mice lines, by 4 intraperitoneal serial transfers. All 3 rodent malarias developed in red blood cells of beta-thalassaemic mice without losing their virulence. There was no delay in infection and peaks of parasitaemia were similar in beta-thalassaemic and normal mice. The mortality occurred earlier in beta-thalassaemic mice than in control mice for P. berghei and P. chabaudi. The difference was more pronounced for P. yoelii NS where normal mice did not die. These results could be explained by a failure of erythropoiesis in beta-thalassaemic mice, which are unable to compensate for the destruction of red blood cells by the parasites, and the mice died of anaemia. Ultrastructural examination of the rodent malaria parasites in beta-thalassaemic RBC showed a normal development even in the presence of Heinz bodies. In conclusion, no effective protection against malaria was provided by the beta-thalassaemia in this mouse model.

Cerebral malaria-susceptible (C57BL/6) mice infected with Plasmodium berghei ANKA (PbA) developed low parasitemia and died from typical neurological symptoms between 8 to 10 days after infection. In contrast, nonsusceptible (BALB/c) mice developed high peripheral blood parasitemia and died 22-24 days later without neurological implications. Daily injections of fatty acids (FA) during the first 3 days after infection protected C57BL/6 mice from cerebral symptoms but had no effect on BALB/c mice. Thus, treated C57BL/6 mice developed hyperparasitemia and died 25 days after infection, like BALB/c mice. Red blood cells from C57BL/6 control mice were found to be more resistant to lysis by linoleic acid than those of BALB/c mice. Three days following infection with PbA, these differences disappeared. Treatment with FA prevented these changes. We concluded that the host's cells were altered soon after infection and that the nature and degree of alterations depended on the mouse strain, thus determining the eventual outcome of the infection. Likewise, the effects of FA might not be directed against the parasite but rather seem to act early after infection on these parasite-induced modifications of host cells.

Glutathione (GSH) plays a critical role in the detoxication and the protection of cells against oxidative stress. In the present study we examined the relationship between the intracellular GSH levels as well as glutathione S-transferase (GST), glutathione reductase (GR), and glutathione peroxidase (GPx) activities and how they relate to Plasmodium berghei resistance to chloroquine. Resistant strains (CQR30 and CQR60) were selected in vivo from a sensitive strain (NK65). Marked increases in GSH levels and GST activity within resistant parasites were observed, compared to sensitive parasites. On the other hand, GR and GPx activities were similar in sensitive and resistant parasites. Treatment with chloroquine did not influence the intracellular level of GSH, but it was found to significantly decrease GR activity. Intracellular depletion of GSH, by a nontoxic concentration of buthionine sulfoximine (BSO), significantly sensitized the resistant parasites to chloroquine. These results suggest that the P. berghei resistance results from altered GSH and GST levels and activity, respectively, which enable the detoxification of chloroquine in resistant parasites.

Mefloquine is an antimalarial drug with schizonticidal activity on blood-stage parasites. Studies of the role of mefloquine on the development of Plasmodium berghei ANKA in anopheles stephensi have been carried out that showed a dose-dependent effect on the sporogonic cycle of these parasites, with changes in the numbers of oocysts and the extent of sporozoite invasion of salivary glands. In this study, we show that mefloquine-resistant P. berghei ANKA blood stage parasites could be selected through drug pressure during continuous cyclical transmission of Anopheles gambiae s.l.