Plasmodium Berghei ANKA Infection In Mice Research Articles

Antimalarial potential of Kerala Ayurvedic Water "Pathimugam".

The persistent threat of drug resistant malaria demands new cures. Low prevalence of malaria in the Indian state of Kerala compared with other proximal states led us to explore if there is any traditional practice in Kerala which may confer protection against malaria. In this context, our attention was drawn to "Pathimugam" i.e., Ceasalpinia sappan whose heartwood is used to prepare a red aqueous extract which is a uniquely popular drink all over Kerala. Aqueous and methanolic extracts of various organs of C. sappan were prepared and tested against Plasmodium falciparum grown in vitro culture using SYBR Green-I assay. The cytotoxicity of active extracts/fractions was studied using mammalian HeLa cell line. In vivo efficacy was determined using Plasmodium berghei ANKA infected mice. The highest antiplasmodial activities in the alcoholic and aqueous extracts were observed in leaf methanolic extract (IC 2 μg/ml) and heartwood aqueous extract (IC 12.5 μg/ml). C. sappan extracts were equipotent against both chloroquine sensitive Pf3D7 and resistant Pf INDO strains and showed suppression of % parasitemia in P. berghei infected mice. Activity guided chromatographic fractionation of aqueous wood extract led to the fortification of antiplasmodial activity (IC 5 μg/ml). Our results establish the antiplasmodial potential of C. sappan and suggest that its regular use might have prophylactic or curative actions that may assist in keeping check on malaria in the Indian state of Kerala.

In Vivo Antimalarial Activity of Cyperus rotundus and Its Combination with Dihydroartemisinin against Plasmodium berghei

Background. The increase in the number of drug-resistant Plasmodium species continues to be a serious public health concern. Therefore, identification of the potential novel antimalarial drugs derived from therapeutic plants could help solve this issue. This study investigated whether Cyperus rotundus aqueous crude extract (CRE) and its combination with dihydroartemisinin (DHA) were effective against Plasmodium berghei ANKA-infected mice. Methods. CRE was prepared from C. rotundus rhizomes and evaluated using acute and subacute toxicity tests on BALB/c mice. The antimalarial effectiveness of CRE was assessed at 100, 200, and 400 mg/kg in a 4-day suppressive test with curative and prophylactic testing and measurement of packed cell volume (PCV), body weight (BW), rectal temperature, and mean survival time (MST). Results. Following acute and subacute treatment, CRE caused no harmful effects or mortality in mice. When compared with that in the untreated control, infected mice administered with 400 mg/kg of CRE in a 4-day suppressive test exhibited the strongest antimalarial activity (55.30% inhibition) with prolonged MST. However, curative and prophylactic assays did not reveal CRE to have antimalarial activity. In comparison with that achieved with the single therapy, the combination of DHA and CRE at ED50/2 (1 and 200 mg/kg, respectively) produced considerable antimalarial activity at 90.08% inhibition with synergism (combination index = 0.21701). For the other parameters, CRE administration prevented malarial-induced changes in PCV, BW, and rectal temperature. Conclusions. CRE treatment significantly inhibited malaria in the 4-day suppressive test, and CRE combined with DHA had a synergistic antimalarial effect.

Resveratrol Reduce the Severity of Anemia and Thrombocytopenia in Plasmodium berghei ANKA-Infected Mice

Background: Malaria is an infectious disease with a high mortality rate. One of the complications of malaria is blood disorders. Hematological disorders such as anemia and thrombocytopenia are common in malaria infection. Resveratrol has been reported to have antimalarial activity. Objective: This study aimed to evaluate the effect of resveratrol in reducing the severity of anemia and thrombocytopenia in Plasmodum berghei-infected mice. Methods: The study began with parasite inoculation in mice. After the mice were infected, and randomly grouped into negative control, chloroquine 20 mg/kg (positive control), and resveratrol with doses 25, 50, and 100 mg/kg. The mice's blood profile was measured on day 0 and day 4 using a hematology analyzer. Results: The results showed that after administration of resveratrol at various doses, the number of RBC, hemoglobin, hematocrit and platelets was higher than the control. Conclusion: It can be concluded that resveratrol can reduce the severity of anemia and thrombocytopenia in mice infected with P. berghei.

Administration of soluble gp130Fc disrupts M-1 macrophage polarization, dendritic cell activation, MDSC expansion and Th-17 induction during experimental cerebral malaria

Regulatory effect of IL-6 on various immune cells plays a crucial role during experimental cerebral malaria pathogenesis. IL-6 neutralization can restore distorted ratios of myeloid dendritic cells and plasmacytoid dendritic cells as well as the balance between Th-17 and T-regulatory cells. IL-6 can also influence immune cells through classical and trans IL-6 signalling pathways. As trans IL-6 signalling is reportedly involved during malaria pathogenesis, we focused on studying the effects of trans IL-6 signalling blockade on various immune cell populations and how they regulate ECM progression. Results show that administration of sgp130Fc recombinant chimera protein lowers the parasitemia, increases the survivability of Plasmodium berghei ANKA infected mice, and restores the distorted ratios of M1/M2 macrophage, mDC/pDC, and Th-17/Treg. IL-6 trans signalling blockade has been found to affect both expansion of myeloid derived suppressor cells (MDSCs) and expression of inflammatory markers on them during Plasmodium berghei ANKA infection indicating that trans IL-6 signalling might regulate various immune cells and their function during ECM. In this work for the first time, we delineate the effect of sgp130Fc administration on influencing the immunological changes within the host secondary lymphoid organ during ECM induced by Plasmodium berghei ANKA infection.

Cell-specific MHC class I deletion on CD11c+ APCs abrogates CD8 T Cell infiltration into the brain

Abstract Cerebral malaria (CM) is a severe complication of P. falciparum infection and a leading cause of death, globally. Plasmodium berghei ANKA (PbA) infection of C57BL/6 mice serves as a murine model of CM called experimental cerebral malaria (ECM) due to the neurological and neuropathological features that simulate the infection of human CM. A putative mechanism of disease in human CM requires CD8 T cell to infiltration into the brain and blood-brain barrier disruption. The specific antigen-presenting cell type(s) that prime this CNS infiltrating CD8 T cell response capable of inducing lethal blood-brain barrier disruption has not been defined. To answer this, single class I expressing K bloxP transgenic mice were generated. Crossing these mice to class I deficient CD11c-cre or LysM-cre mice created the LysM-cre K bcKO and CD11c-cre K bcKO mice, respectively. PbA infected K bloxP and LysM-cre K bcKO were susceptible to ECM. In contrast, CD11c-cre K bcKO mice survived acute PbA infection and avoided neurological manifestations. We therefore employed spectral flow cytometry to analyze splenocytes and brain infiltrating immune cells during acute PbA infection in these genotypes. We observed a reduction in activated CD8 T cells in the spleens of PbA infected CD11c-cre K bcKO mice versus the K bloxP mice and the LysM-cre K bcKO. This reduced peripheral CD8 T cell activation in PbA infected CD11c-cre Kb cKO mice also corresponded with reduced brain infiltration of CD8 T cells. We conclude a CD11c+ APC is therefore required for priming and potentially recruitment of CD8 T cells to the brain during lethal ECM pathology. 5R01NS103212-06

Effect of ethanol extract of Hedyotis corymbosa (L.) Lamk against parasitemia and hepatomegaly in Plasmodium berghei ANKA-infected mice

Malaria is a parasitic infectious disease that remains one of the focuses of world health problems. The ethanol extract of Hedyotis corymbosa has biochemical compounds potentially become a new antimalarial drug. This study aimed to analyze the effect of this extract on parasitemia and hepatomegaly in mice infected with Plasmodium berghei ANKA. Twentyfive BALB/c mice were infected with P. berghei ANKA and grouped into 5 groups. Group 1-3 were treated with 250 mg/Kg BW (HC250), 300 mg/Kg BW (HC300), and 350 mg/Kg BW (HC350) of ethanol extract of H. corymbosa (EEHC), respectively. Group 4 was a positive control (POS) which was given dihydroartemisisnin-piperaquin (DHP) and Group 5 was a negative control (NEG) which was only given CMC Na 1%. Treatments were given orally once a day for four consecutive days. Parasitemia was observed daily on Giemsa-stained tail blood smear. On day 5 the mice were sacrificed, blood were collected by cardiac punctured, the livers were removed and the length, width, and weight were measured. There was no significant difference on parasitemia between Group 1, 2, 3 and NEG. However, the highest inhibition of parasite s growth was found in Group 3 (61.4%. Observation on hepatomegaly, showed that a significant difference on the length of the liver was found between Group 3 and NEG.

Studying the Rationale of Fire Ant Sting Therapy Usage by the Tribal Natives of Bastar Revealed Ant Venom-Derived Peptides with Promising Anti-Malarial Activity.

Prevailing drug resistance in malaria imposes the major roadblock for the existing interventions necessitating the timely need to search for alternative therapies. Ants in Solenopsis spp, termed 'Fire ants', are well known for their aggressive behavior, which leads to the release of toxic venom. Notably, the tribal natives of the malaria-laden densely forested Bastar region, Chhattisgarh, India, use fire ant sting-based therapy to cure malaria-like high fever. Inspired by this, we have collected the fire ants from the forest of Bastar and extracted peptide and alkaloid fractions from ant venom using HPLC and analyzed them by LC/MS-based applications. Evaluation of the anti-malarial efficacy of these peptide fractions demonstrated a significant reduction in the growth of Plasmodium falciparum (Pf 3D7) in vitro, whereas the alkaloid fraction showed a negligible effect. in vitro hemolytic activity confirmed the venom peptide fraction to be non-hemolytic. Additionally, the venom peptide fraction is purely non-toxic to HepG2 cells. Anti-malarial efficiency of the same in Plasmodium berghei ANKA infected mice models showed a drastic reduction in parasitemia representing promising anti-malarial activity. Overall, our study has unraveled the scientific rationale underlying fire ant sting therapy used as a tribal naturotherapy for curing malaria-like fever, thus, introducing a way forward to develop nature-inspired anti-malarial chemotherapeutics.

Piperine Enhances the Antimalarial Activity of Curcumin in Plasmodium berghei ANKA-Infected Mice: A Novel Approach for Malaria Prophylaxis.

Malaria is a prevalent vector-borne infectious disease in tropical regions, particularly in the absence of effective vaccines and because of the emergence resistance of Plasmodium to available antimalarial drugs. An alternative strategy for malaria eradication could be the combination of existing compounds that possess antimalarial activity to target multiple stages of the parasite. This study evaluated the antimalarial activity of a combination of curcumin and piperine in mice. A total of 42 mice were assigned to six groups depending on the treatment administered: group I (normal group) with aquadest; group II (negative control) with 0.2 ml DMSO; group III received a standard malarial drug (artesunate 5 mg/kg BW); groups IV, V, and VI with curcumin 300 mg/kg BW, curcumin 300 mg/kg BW and piperine 20 mg/kg BW, and piperine 20 mg/kg BW, respectively. The antimalarial activity was evaluated using prophylactic assays in Plasmodium berghei ANKA-infected mice, including the percentage parasitemia, clinical signs, survival rate, serum biochemical analysis, parasitic load in the liver, and liver histopathology. All treatments showed significant (p < 0.05) antiplasmodial activity, with considerable parasite inhibition (>50%), curcumin 300 mg/kg BW (60.22%), curcumin 300 mg/kg BW, and piperine 20 mg/kg BW (77.94%) except for piperine 20 mg/kg BW (47.20%), eliciting greater inhibition relative to that of artesunate (51.18%). The delayed onset of clinical symptoms and prolonged survival rate were also significant (p < 0.05) in the combination of curcumin and piperine treated group. In addition, the low parasitic load in the liver and mild histopathological changes in the liver suggest that the combination of curcumin and piperine had synergistic or additive effects. These findings demonstrate the promising use of these combined compounds as a malarial prophylactic. Further studies were recommended to assess their clinical usefulness.

Interleukin-1b and Interferon-g are Associated with Malaria-Induced Hypoinsulinemic Hypoglycemia in Plasmodium berghei Anka-Infected Mice

Malaria-induced hypoglycemia is recognized as a serious complication of malaria and has one of the strongest associations with mortality in children. It has been speculated that oxidative stress and pro-inflammatory response during parasite infection were involved in its pathophysiology. Hence, this study aimed to investigate the development of malaria-induced hypoglycemia during Plasmodium berghei ANKA (PbANKA) infection with particular attention to the involvement of c-peptide, interleukin-1b (IL-1b), and interferon-g (IFN-g). ICR mice were infected with 1×107 parasitized erythrocytes of PbANKA, and parasitemia was monitored, and the development of hypoglycemia was assessed by measuring plasma glucose levels. The change of c-peptide level was evaluated. The pro-inflammatory response of IL-1b and IFN-g were also quantified in plasma. It was found that PbANKA infection resulted in hypoglycemia as indicated by a significantly (P &lt; 0.05) decrease in plasma glucose levels on day 4 post-infection and associated with parasitemia. The c-peptide was slightly increased at day 2 post-infection, and then significantly (P &lt; 0.05) decreased since day 4. Furthermore, we observed a significantly (P &lt; 0.05) increased IL-1b, firstly responded, at day 2 post-infection followed by increasing the IFN-g level at day 4 in PbANKA-induced hypoglycemia. Our findings support the idea that hypoinsulinemic hypoglycemia in the PbANKA infected mice may be involved in the high IL-1b and IFN-g against the parasite infection.

IL-6 dependent expansion of inflammatory MDSCs (CD11b+ Gr-1+) promote Th-17 mediated immune response during experimental cerebral malaria.

Myeloid derived suppressor cells (MDSCs) are a group of heterogeneous cell populations that can suppress T cell responses. Various aspects of MDSCs in regulating immune responses in several cancer and infectious diseases have been reported till date. But the role and regulation of MDSCs have not been systematically studied in the context of malaria. This study depicts the phenotypic and functional characteristics of splenic MDSCs and how they regulate Th-17 mediated immune response during Experimental Cerebral Malaria (ECM). Flow cytometric analysis reveals that MDSCs in the spleen and bone marrow expand at 8 dpi during ECM. Among subtypes of MDSCs, PMN-MDSCs show significant expansion in the spleen but M-MDSCs remain unaltered. Functional analysis of sorted MDSCs from spleens of Plasmodium berghei ANKA (PbA) infected mice shows suppressive nature of these cells and high production of Nitric oxide (NO). Besides, MDSCs were also found to express various inflammatory markers during ECM suggesting the M1 type phenotype of these cells. In-vivo depletion of MDSCs by the use of Anti Gr-1 increases mice survival but doesn't significantly alter the parasitemia. Previously, it has been reported that Treg/Th-17 balance in the spleen is skewed towards Th-17 during ECM. Depletion of MDSCs was found to regulate Th-17 percentages to homeostatic levels and subvert various inflammatory changes in the spleen. Among different factors, IL-6 was found to play an important role in the expansion of MDSCs and expression of inflammatory markers on MDSCs in a STAT3-dependent manner. These findings provide a unique insight into the role of IL-6 in the expansion of the MDSC population which causes inflammatory changes and increased Th-17 responses during ECM.

Effect of Allicin and Artesunate Combination Treatment on Experimental Mice Infected with Plasmodium berghei.

Malaria is still a significant health problem in endemic countries and increases Plasmodium resistance to the available antimalarial drugs. Hence, this study aimed to investigate the antimalarial activity of allicin and its combination with artesunate (ART) against rodent malaria Plasmodium berghei ANKA (PbANKA) infected mice. Allicin was prepared in 20% Tween-80. Balb/c mice were inoculated intraperitoneally with 1×107 PbANKA-infected erythrocytes and orally given by gavage with the chosen doses of 1, 10, 50, and 100 mg/kg of allicin and 1, 5, 10, and 20 mg/kg of ART once a day for 4 consecutive days. Effective dose 50 (ED50) of allicin and ART was subsequently investigated. Moreover, the combination (1 : 1) of allicin and ART at the doses of their respective ED50, ED50 1/2, ED50 1/4, and ED50 1/8 was also carried out. The untreated control was given 20% Tween-80. The results showed that allicin presented a dose-dependent antimalarial activity with significance (p < 0.05). The ED50 values of allicin and ART were about 14 and 5 mg/kg, respectively. For combination, allicin and ART showed a synergistic effect at the combination doses of ED50, ED50 1/2, and ED50 1/4 with significantly (p < 0.01) prevented reduction of packed cell volume, bodyweight loss, rapid dropping of rectal temperature, and markedly prolonged mean survival time, compared with the untreated control and single treatment. It can be concluded that allicin exerted potential antimalarial activity in single and its combination with ART.

Oral Administration of Piperine as Curative and Prophylaxis Reduces Parasitaemia in Plasmodium berghei ANKA-Infected Mice.

Malaria remains a public health problem and a leading cause of death worldwide. Consequently, the discovery of novel agents, including substances from medicinal plants, is urgently needed. Piper nigrum has long been used by the community in the treatment of the symptoms of malaria. In a previous study, Piper nigrum was demonstrated to exhibit promising antiplasmodial activity against Plasmodium falciparum 3D7 and INDO strains. The aim of this study was to further investigate the antimalarial activity (curative and prophylactic) of piperine (a major isolated constituent of Piper nigrum) in Plasmodium berghei ANKA-infected mice. Piperine 10, 20, and 40 mg/kg body weight (bw), artesunate 5 mg/kg bw, and DMSO were administered orally for four days to different groups of Swiss Webster mice. Then, mice were monitored for parasitaemia, body weight, rectal temperature, survival rate, and clinical parameters. Piperine 40 mg/kg bw in curative and prophylactic tests had the maximum parasitaemia chemosuppression of 79.21% and 58.8% (p < 0.05), respectively, with a significant effect on the survival rate compared with control animals. In the curative test, piperine 40 mg/kg bw reduced the mean clinical score compared with the control group. Additionally, piperine showed an ability to protect organs (lungs, liver, spleen, and kidneys) from some damage in a dose-dependent manner. This study can be used as a basis for further discovery of novel chemotherapeutic or chemoprophylactic compounds.

Antimalarial Activity of Ethanol Extract of Kelakai Leaves (Stenochlaena palustris) to Parasitemia and Splenomegaly in BALB/c Mice Infected with Plasmodium berghei ANKA

Introduction: Malaria is one of global health problems. Splenomegaly is one of malaria symptoms. Antimalarial drug resistance had been reported. Alternative treatment is by using traditional medicinal plants such as kelakai (Stenochlaena palustris). Kelakai contains alkaloid and flavonoid which had been reported to have antimalarial activity. The aim of this study was to discover antimalarial activity of ethanol extract of kelakai leaves to parasitemia and splenomegaly of Plasmodium berghei ANKA in infected BALB/c mice.Methods: This study was based on a modified Peter test using BALB/c mice infected with P. berghei ANKA treated with ethanol extract of kelakai leaves, with chloroquine diphosphate as a positive control. The negative control was P. berghei ANKA infected mice without any additional treatment. Administration of ethanol extract of kelakai leaves was performed for 4 days with a serial doses of 100, 10, and 1 mg/kg body weight. The positive control was given chloroquine diphosphate 20 mg/kg body weight. Parasitemia was observed daily prior to the calculation of the percentage of parasite growth and parasite growth inhibition. At the end of the test, the mice were sacrificed and spleens were isolated to measure their sizes. Probit analysis was performed to obtain ED50 to find the effect of extract in parasite killing by 50%. Spearman test was performed to analyze the correlation of doses of extract and splenomegaly.Results: Parasitemia growth inhibition was directly proportional to the dose. Higher parasitemia inhibition was obtained at higher doses and vice versa. Result of probit analysis showed an ED50 was 77.05 mg/kg body weight. Statistical analysis resulted in insignificant correlation between doses and splenomegaly p = 1.0 (significancy &lt; 0.05).Conclusion: Ethanol extract of kelakai leaves possessed good antimalarial activity and there was no correlation between extract doses and splenomegaly in Plasmodium berghei ANKA-infected mice.

Nephroprotective effect of virgin coconut oil in Plasmodium berghei ANKA infected Balb/c mice

AbstractMalaria is a parasitic infectious disease caused by Plasmodium, which remains a world health problem with an estimated 219 million cases worldwide. In severe malaria infection, several organs of the body can be affected, including the kidneys. One of the pathophysiology associated with the worsening of this disease is oxidative stress. The use of antioxidants is expected to prevent this, and one product that has a high antioxidant content is virgin coconut oil (VCO). This study aimed to analyze the effect of VCO on the kidney in Plasmodium berghei ANKA-infected mice. This study was an in vivo laboratory experimental study with a randomized post-test only control group design using 35 BALB/c mice infected with P. berghei ANKA, weighing 20-30 grams. VCO with the Javara® brand is used with doses of 1, 5, and 10 ml/kg body weight (kgBW)/ day. The parameter assessed were levels of BUN, creatinine, and renal histopathological changes. The administration of VCO on the treated group shows minimal tubular necrosis and glomerulonephritis compared to the negative control group. The BUN and creatinine levels in the treated group were also lower than the negative control group. The results showed that VCO has a nephroprotective effect against P. berghei ANKA infection in mice.Keywords: malaria, kidney, virgin coconut oil

Assessment of in vitro and in vivo antimalarial efficacy and GC-fingerprints of selected medicinal plant extracts

A hallmark of mortality and morbidity, malaria is affecting nearly half of the world's population. Emergence of drug-resistant strains of malarial parasite prompts identification and evaluation of medicinal plants and their constituents that may hold the key to a new and effective anti-malarial drug. In this context, nineteen methanolic extracts from seventeen medicinal plants were evaluated for anti-plasmodial potential against Plasmodium falciparum strain 3D7 (Chloroquine (CQ) sensitive) and INDO (CQ resistant) using fluorescence based SYBR-Green assay and for cytotoxic effects against mammalian cell lines. Leaf extract of two plants showed promising in vitro anti-malarial activity (Pf3D7 IC50 ≤ 10 μg/ml); one plant extract showed good activity (Pf3D7 IC50 = 10.1–20 μg/ml); seven were moderately active (IC50 = 20.1–50 μg/ml), four plant extracts showed poor activity (PfD7 IC50 = 50.1–100 μg/ml) and five extracts showed no activity up to IC50 = 100 μg/ml. Further, six extracts were found equipotent to PfINDO (resistance index ranging 0.4–2) and relatively nontoxic to mammalian cell lines HEK293 (cytotoxicity index ranging 1.4–12.5). Based on good resistance and selectivity indices, three extracts were evaluated for in vivo activity in Plasmodium berghei ANKA infected mice at a dose of 500 mg/kg and they showed significant suppression of P. berghei parasitemia. Further, these active plant extracts were fractionated using silica-gel chromatography and their fractions were evaluated for anti-plasmodial action. Obtained fractions showed enrichment in antimalarial activity. Active fractions were analyzed by gas chromatography and mass-spectrometery. Results suggests that the three active plant extracts could serve as potent source of anti-malarial agent and therefore require further analysis.

In vivo Antimalarial Activities of Five Ugandan Medicinal Plants on Plasmodium berghei in Mice

Aim: Medicinal plants have played an important role in the treatment of different ailments including malaria in developing countries particularly in Africa. This study has evaluated the antimalarial activities of Azadirachta indica A. Juss (Meliaceae), Cymbopogon citratus Stapf. (Poaceae), Moringa oleifera Lam. (Moringaceae), Tithonia diversifolia (Hemsl) A. Grey (Asteraceae) and Vernonia amygdalina Del. (Asteraceae) which are commonly-used for malaria treatment in Uganda.&#x0D; Study Design: This is an experimental laboratory report on antimalarial activities of some Ugandan medicinal plants for subsequent profiling in an herbal pharmacopoeia and eventual drug development.&#x0D; Place and Duration of Study: The Animal Research Facility and the Clinical and Research Laboratory, Faculty of Medicine, Mbarara University of Science and Technology, Uganda, between July 2019 and March 2020.&#x0D; Methodology: The antimalarial activity of the hot infusion of each leaf was evaluated on chloroquine-sensitive Plasmodium berghei ANKA-infected mice using 4-day test at 100 – 400 mg/kg with chloroquine (10 mg/kg) and artemether-lumefantrine (4 mg/kg) as positive controls; and distilled water as negative control. The observed haematological responses of the animals were determined with an automated haematometer.&#x0D; Results: The results showed dose-dependent activities in the animals treated with the extract of each plant leaf in varying degrees. Thus, V. amygdalina and T. diversifolia showed the highest antimalarial activities with the chemosuppression values of 75% and 66% at 400 mg/kg, respectively. The results of V. amygdalina, T. diversifolia and M. oleifera, extracts gave the lowest ED50 of 141, 195 and 231 mg/kg, respectively being significantly different from A. indica (ED50 319 mg/kg) and C. citratus (ED50 346 mg/kg) with V. amygdalina as the most potent extract among the five plant leaves.&#x0D; Conclusion: The observed activities of the five plants have therefore supported their folkloric uses as antimalarial remedies by the Ugandan traditional medicine practitioners with obvious potentials for drug development.

RAGE modulatory effects on cytokines network and histopathological conditions in malarial mice

This study was aimed at investigating the involvement of Receptor for Advanced Glycation End Products (RAGE) during malaria infection and the effects of modulating RAGE on the inflammatory cytokines release and histopathological conditions of affected organs in malarial animal model. Plasmodium berghei (P. berghei) ANKA-infected ICR mice were treated with mRAGE/pAb and rmRAGE/Fc Chimera drugs from day 1 to day 4 post infection. Survival and parasitaemia levels were monitored daily. On day 5 post infection, mice were sacrificed, blood were drawn for cytokines analysis and major organs including kidney, spleen, liver, brain and lungs were extracted for histopathological analysis. RAGE levels were increased systemically during malaria infection. Positive correlation between RAGE plasma concentration and parasitaemia development was observed. Treatment with RAGE related drugs did not improve survival of malaria-infected mice. However, significant reduction on the parasitaemia levels were recorded. On the other hand, inhibition and neutralization of RAGE production during the infection significantly increased the plasma levels of interleukin (IL-4, IL-17A, IL-10 and IL-2) and reduced interferon (IFN)-γ secretion. Histopathological analysis revealed that all treated malarial mice showed a better outcome in histological assessment of affected organs (brain, liver, spleen, lungs and kidney). RAGE is involved in malaria pathogenesis and targeting RAGE could be beneficial in malaria infected host in which RAGE inhibition or neutralization increased the release of anti-inflammatory cytokines (IL-10 and IL-4) and reduce pro-inflammatory cytokine (IFNγ) which may help alleviate tissue injury and improve histopathological conditions of affected organs during the infection.

Dimorphic effect of 17β-oestradiol on pathology and oxidative stress in experimental malaria.

Malaria is the parasitic disease with the highest mortality worldwide; males exhibit higher mortality and more severe symptomatology than females, suggesting the participation of sexual hormones in protection and pathology. We have documented that gonadectomy modifies oxidative stress in Plasmodium berghei ANKA-infected mice in a dimorphic manner. However, gonadectomy decreases all sexual steroids levels, making it difficult to determine the contribution of each hormone to the results. This study aimed to explore the participation of 17β-oestradiol (E2) in oxidative stress in the blood, spleen, liver and brain of P. berghei-infected female and male mice. E2 was administered to intact or gonadectomized (GX) male and female mice to assess their effects on parasitaemia, body weight loss and hypothermia. We also measured the effect of E2 on the specific activities of superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx) and on malondialdehyde (MDA) levels in the blood, spleen, liver and brain of CBA/Ca male and female mice infected with P. berghei ANKA. We detected the effects of E2 and sexual dimorphism on all tissues and variables analysed. Administration of E2 increased parasitaemia in intact mice. However, reconstitution of GX female mice with E2 decreased parasitaemia. E2 decreased body weight and differentially modulated oxidative stress depending on the sex, infection and tissue analysed. Low antioxidant activity was detected in the brain, suggesting additional protective antioxidant mechanisms in the brain independent of antioxidant enzymes. Our results explained, at least in part, the sexual dimorphism in this experimental model of malaria.

Tim-3 signaling blockade with \u03b1-lactose induces compensatory TIGIT expression in Plasmodium berghei ANKA-infected mice

BackgroundMalaria, one of the largest health burdens worldwide, is caused by Plasmodium spp. infection. Upon infection, the host’s immune system begins to clear the parasites. However, Plasmodium species have evolved to escape the host’s immune clearance. T-cell immunoglobulin and mucin domain 3 (Tim-3), a surface molecule on most immune cells, is often referred to as an exhaustion marker. Galectin (Gal)-9 is a Tim-3 ligand and the T helper (Th) 1 cell response is inhibited when Gal-9 binds to Tim-3. In the present study, dynamic expression of Tim-3 on key populations of lymphocytes during infection periods of Plasmodium berghei and its significance in disease resistance and pathogenesis were explored.MethodsTim-3 expression on critical lymphocyte populations and the proportion of these cells, as well as the levels of cytokines in the sera of infected mice, were detected by flow cytometry. Further, in vitro anti-Tim-3 assay using an anti-Tim-3 antibody and in vivo Tim-3-Gal-9 signaling blockade assays using α-lactose (an antagonist of Gal-9) were conducted. An Annexin V Apoptosis Detection Kit with propidium iodide was used to detect apoptosis. In addition, proteins associated with apoptosis in lung and spleen tissues were confirmed by Western blotting assays.ResultsIncreased Tim-3 expression on splenic CD8+ and splenic CD4+, and circulatory CD4+ T cells was associated with a reduction in the proportion of these cells. Furthermore, the levels of interleukin (IL)-2, IL-4, IL-6, IL-22, and interferon (IFN)-γ, but not that of tumor necrosis factor alpha (TNF-α), IL-10, and IL-9, increased to their highest levels at day 4 post-infection and decreased thereafter. Blocking Tim-3 signaling in vitro inhibited lymphocyte apoptosis. Tim-3-Gal-9 signaling blockade in vivo did not protect the mice, but induced the expression of the immunosuppressive molecule, T cell immunoreceptor with Ig and ITIM domains (TIGIT), in Plasmodium berghei ANKA-infected mice.ConclusionsTim-3 on lymphocytes negatively regulates cell-mediated immunity against Plasmodium infection, and blocking Tim-3-galectin 9 signaling using α-lactose did not significantly protect the mice; however, it induced the compensatory expression of TIGIT. Further investigations are required to identify whether combined blockade of Tim-3 and TIGIT signaling could achieve a better protective effect.

Pre-clinical study of iron oxide nanoparticles fortified artesunate for efficient targeting of malarial parasite.

BackgroundArtesunate the most potent antimalarial is widely used for the treatment of multidrug-resistant malaria. The antimalarial cytotoxicity of artesunate has been mainly attributed to its selective, irreversible and iron- radical-mediated damage of parasite biomolecules. In the present research, iron oxide nanoparticle fortified artesunate was tested in P. falciparum and in an experimental malaria mouse model for enhancement in the selectivity and toxicity of artesunate towards parasite. Artesunate was fortified with nontoxic biocompatible surface modified iron oxide nanoparticle which is specially designed and synthesized for the sustained pH-dependent release of Fe2+ within the parasitic food vacuole for enhanced ROS spurt.MethodsAntimalarial efficacy of Iron oxide nanoparticle fortified artesunate was evaluated in wild type and artemisinin-resistant Plasmodium falciparum (R539T) grown in O + ve human blood and in Plasmodium berghei ANKA infected swiss albino mice. Internalization of nanoparticles, the pH-dependent release of Fe2+, production of reactive oxygen species and parasite biomolecule damage by iron oxide nanoparticle fortified artesunate was studied using various biochemical, biophysical, ultra-structural and fluorescence microscopy. For determining the efficacy of ATA-IONP+ART on resistant parasite ring survival assay was performed.ResultsThe nanoparticle fortified artesunate was highly efficient in the 1/8th concentration of artesunate IC50 and led to retarded growth of P. falciparum with significant damage to macromolecules mediated via enhanced ROS production. Similarly, preclinical In vivo studies also signified a radical reduction in parasitemia with ~8–10-fold reduced dosage of artesunate when fortified with iron oxide nanoparticles. Importantly, the ATA-IONP combination was efficacious against artemisinin-resistant parasites.InterpretationSurface coated iron-oxide nanoparticle fortified artesunate can be developed into a potent therapeutic agent towards multidrug-resistant and artemisinin-resistant malaria in humans.FundThis study is supported by the Centre for Study of Complex Malaria in India funded by the National Institute of Health, USA.