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Journal of Neuroinflammation May 2024Cerebral malaria (CM) is the most lethal complication of malaria, and survivors usually endure neurological sequelae. Notably, the cytotoxic effect of infiltrating...
BACKGROUND
Cerebral malaria (CM) is the most lethal complication of malaria, and survivors usually endure neurological sequelae. Notably, the cytotoxic effect of infiltrating Plasmodium-activated CD8 T cells on cerebral microvasculature endothelial cells is a prominent feature of the experimental CM (ECM) model with blood-brain barrier disruption. However, the damage effect of CD8 T cells infiltrating the brain parenchyma on neurons remains unclear. Based on the immunosuppressive effect of the PD-1/PD-L1 pathway on T cells, our previous study demonstrated that the systemic upregulation of PD-L1 to inhibit CD8 T cell function could effectively alleviate the symptoms of ECM mice. However, it has not been reported whether neurons can suppress the pathogenic effect of CD8 T cells through the PD-1/PD-L1 negative immunomodulatory pathway. As the important inflammatory factor of CM, interferons can induce the expression of PD-L1 via different molecular mechanisms according to the neuro-immune microenvironment. Therefore, this study aimed to investigate the direct interaction between CD8 T cells and neurons, as well as the mechanism of neurons to alleviate the pathogenic effect of CD8 T cells through up-regulating PD-L1 induced by IFNs.
METHODS
Using the ECM model of C57BL/6J mice infected with Plasmodium berghei ANKA (PbA), morphological observations were conducted in vivo by electron microscope and IF staining. The interaction between the ECM CD8 T cells (immune magnetic bead sorting from spleen of ECM mice) and primary cultured cortical neurons in vitro was observed by IF staining and time-lapse photography. RNA-seq was performed to analyze the signaling pathway of PD-L1 upregulation in neurons induced by IFNβ or IFNγ, and verified through q-PCR, WB, IF staining, and flow cytometry both in vitro and in vivo using IFNAR or IFNGR gene knockout mice. The protective effect of adenovirus-mediated PD-L1 IgGFc fusion protein expression was verified in ECM mice with brain stereotaxic injection in vivo and in primary cultured neurons via viral infection in vitro.
RESULTS
In vivo, ECM mice showed infiltration of activated CD8 T cells and neuronal injury in the brain parenchyma. In vitro, ECM CD8 T cells were in direct contact with neurons and induced axonal damage, as an active behavior. The PD-L1 protein level was elevated in neurons of ECM mice and in primary cultured neurons induced by IFNβ, IFNγ, or ECM CD8 T cells in vitro. Furthermore, the IFNβ or IFNγ induced neuronal expression of PD-L1 was mediated by increasing STAT1/IRF1 pathway via IFN receptors. The increase of PD-L1 expression in neurons during PbA infection was weakened after deleting the IFNAR or IFNGR. Increased PD-L1 expression by adenovirus partially protected neurons from CD8 T cell-mediated damage both in vitro and in vivo.
CONCLUSION
Our study demonstrates that both type I and type II IFNs can induce neurons to upregulate PD-L1 via the STAT1/IRF1 pathway mediated by IFN receptors to protect against activated CD8 T cell-mediated damage, providing a targeted pathway to alleviate neuroinflammation during ECM.
Topics: Animals; Mice; B7-H1 Antigen; CD8-Positive T-Lymphocytes; Interferon Regulatory Factor-1; Interferon-gamma; Malaria, Cerebral; Mice, Inbred C57BL; Mice, Knockout; Neurons; Plasmodium berghei; Signal Transduction; STAT1 Transcription Factor; Up-Regulation
PubMed: 38715061
DOI: 10.1186/s12974-024-03114-7 -
Acta Parasitologica Jun 2024Artemisinin combination therapies, the first-line antimalarials in Nigeria, have reportedly suffered multiple failures in malaria treatment, hence the search for novel...
PURPOSE
Artemisinin combination therapies, the first-line antimalarials in Nigeria, have reportedly suffered multiple failures in malaria treatment, hence the search for novel combination of other compounds. Methyl gallate and palmatine have been reported to exhibit antiplasmodial activities but the antimalarial activity of their combination has not been evaluated. Therefore, the evaluation of the combination of methyl gallate and palmatine for antimalarial activity in vitro and in vivo in the presence of piperine was carried out.
MATERIALS AND METHODS
The inhibitory potential of methyl gallate and palmatine combination on β-hematin (hemozoin) formation was studied in vitro. Also, the antimalarial activity of methyl gallate and palmatine combination with/without a bioenhancer (piperine) was evaluated in Plasmodium berghei NK65-infected mice.
RESULTS
Methyl gallate and palmatine in the ratio 3:2 acted synergistically in vitro and had the highest inhibitory effect (IC = 0.73 µg/mL) on β-hematin (hemozoin) formation. The 3:2 combination of methyl gallate and palmatine exhibited no antimalarial activity in vivo in the absence of piperine but caused reduction in parasitemia that exceeded 40% in the presence of piperine at the dose of 25 mg/kg body weight on days 6 and 8 post-inoculation in mice.
CONCLUSION
The 3:2 combination of methyl gallate and palmatine in the presence of piperine exhibited antimalarial activity in vivo, possibly by synergistic inhibition of hemozoin formation which may cause accumulation of haem within the food vacuole of Plasmodium spp. and its death.
Topics: Animals; Polyunsaturated Alkamides; Antimalarials; Benzodioxoles; Piperidines; Malaria; Mice; Gallic Acid; Alkaloids; Plasmodium berghei; Drug Synergism; Berberine Alkaloids; Parasitemia; Inhibitory Concentration 50; Hemeproteins
PubMed: 38705947
DOI: 10.1007/s11686-024-00850-x -
Biochemical Pharmacology Jul 2024The spread of malarial parasites resistant to first-line treatments such as artemisinin combination therapies is a global health concern. Differentiation-inducing factor...
The spread of malarial parasites resistant to first-line treatments such as artemisinin combination therapies is a global health concern. Differentiation-inducing factor 1 (DIF-1) is a chlorinated alkylphenone (1-(3,5-dichloro-2,6-dihydroxy-4-methoxyphenyl) hexan-1-one) originally found in the cellular slime mould Dictyostelium discoideum. We previously showed that some derivatives of DIF-1, particularly DIF-1(+2) (1-(3,5-dichloro-2,6-dihydroxy-4-methoxyphenyl) octan-1-one), exert potent antimalarial activities. In this study, we synthesised DIF-1(+3) (1-(3,5-dichloro-2,6-dihydroxy-4-methoxyphenyl) nonan-1-one). We then evaluated the effects of DIF-1(+3) in vitro on Plasmodium falciparum and in vivo over 7 days (50-100 mg/kg/day) in a mouse model of Plasmodium berghei. DIF-1(+3) exhibited a half-maximal inhibitory concentration of approximately 20-30 % of DIF-1(+2) in three laboratory strains with a selectivity index > 263, including in strains resistant to chloroquine and artemisinin. Parasite growth and multiplication were almost completely suppressed by treatment with 100 mg/kg DIF-1(+3). The survival time of infected mice was significantly increased (P = 0.006) with no apparent adverse effects. In summary, addition of an acyl group to DIF-1(+2) to prepare DIF-1(+3) substantially enhanced antimalarial activity, even in drug-resistant malaria, indicating the potential of applying DIF-1(+3) for malaria treatment.
Topics: Antimalarials; Animals; Mice; Hexanones; Plasmodium falciparum; Plasmodium berghei; Malaria; Dictyostelium; Acylation; Female; Hydrocarbons, Chlorinated
PubMed: 38697310
DOI: 10.1016/j.bcp.2024.116243 -
Computational and Structural... Dec 2024Malaria, a significant global health challenge, is caused by parasites. The liver stage plays a pivotal role in the establishment of the infection. This study focuses...
Malaria, a significant global health challenge, is caused by parasites. The liver stage plays a pivotal role in the establishment of the infection. This study focuses on the liver stage development of the model organism Plasmodium berghei, employing fluorescent microscopy imaging and convolutional neural networks (CNNs) for analysis. Convolutional neural networks have been recently proposed as a viable option for tasks such as malaria detection, prediction of host-pathogen interactions, or drug discovery. Our research aimed to predict the transition of Plasmodium-infected liver cells to the merozoite stage, a key development phase, 15 hours in advance. We collected and analyzed hourly imaging data over a span of at least 38 hours from 400 sequences, encompassing 502 parasites. Our method was compared to human annotations to validate its efficacy. Performance metrics, including the area under the receiver operating characteristic curve (AUC), sensitivity, and specificity, were evaluated on an independent test dataset. The outcomes revealed an AUC of 0.873, a sensitivity of 84.6%, and a specificity of 83.3%, underscoring the potential of our CNN-based framework to predict liver stage development of . These findings not only demonstrate the feasibility of our methodology but also could potentially contribute to the broader understanding of parasite biology.
PubMed: 38690550
DOI: 10.1016/j.csbj.2024.04.029 -
Journal of Ethnopharmacology Sep 2024Andrographis paniculata (AP) ((Burm f.) Wall. ex Nees) is a medicinal plant, documented for its folkloric use in the treatment of malaria.
Mechanism of antimalarial action and mitigation of infection-mediated mitochondrial dysfunction by phyto-constituents of Andrographis paniculata ((Burm f.) Wall. ex Nees) in Plasmodium berghei-infected mice.
ETHNOPHARMACOLOGICAL RELEVANCE
Andrographis paniculata (AP) ((Burm f.) Wall. ex Nees) is a medicinal plant, documented for its folkloric use in the treatment of malaria.
AIM
This study was designed to determine the potency of extract and fractions of A. paniculata (AP) as a curative, both for susceptible and resistant malaria and to also determine the plant's mechanism of action. This study was also designed to determine whether AP extract and its most potent fraction will mitigate infection-mediated mitochondrial dysfunction, and to assess the phytochemical constituents of the most potent fraction.
MATERIALS AND METHODS
n-Hexane, dichloromethane, ethylacetate and methanol were used to partition the methanol extract of A. paniculata. Graded doses of these extract and fractions were used to treat mice infected with chloroquine-sensitive strain of P. berghei in a curative model. The most potent fraction was used to treat mice infected with resistant (ANKA strain) P. berghei. Inhibition of hemozoin formation, reversal of mitochondrial dysfunction and antiinflammatory potentials were determined. A combination of ultraperformance liquid chromatography-quadrupole time of flight-mass spectrometry and nuclear magnetic resonance spectroscopy were used for chemical analysis.
RESULTS
Microscopy revealed that the dichloromethane fraction decreased the parasite burden the most, and inhibition of the hemozoin formation is one of its mechanisms of action. The dichloromethane fraction reversed parasite-induced mitochondrial pore opening in the host, enzyme-dependent ATP hydrolysis and peroxidation of host mitochondrial membrane phospholipids as well as its antiinflammatory potentials. The UPLC-qTOF-MS report and NMR fingerprints of the dichloromethane fraction of A. paniculata yielded fourteen compounds of which sibiricinone C was identified from the plant for the first time.
CONCLUSION
Fractions of A. paniculata possess antiplasmodial effects with the dichloromethane fraction having the highest potency. The potent effect of this fraction may be attributed to the phytochemicals present because it contains terpenes implicated with antimalarial and antiinflammatory activities.
Topics: Animals; Plasmodium berghei; Antimalarials; Malaria; Plant Extracts; Mice; Andrographis; Mitochondria; Male; Hemeproteins; Anti-Inflammatory Agents; Female
PubMed: 38670400
DOI: 10.1016/j.jep.2024.118241 -
Microbes and Infection Apr 2024Hemozoin is a crystal synthesized by Plasmodium parasites during hemoglobin digestion in the erythrocytic stage. The hemozoin released when the parasites egress from the...
Hemozoin is a crystal synthesized by Plasmodium parasites during hemoglobin digestion in the erythrocytic stage. The hemozoin released when the parasites egress from the red blood cell, which is complexed with parasite DNA, is cleared from the circulation by circulating and tissue-resident monocytes and macrophages, respectively. Recently, we reported that intravenous administration of purified hemozoin complexed with Plasmodium berghei DNA (Hz) resulted in an innate immune response that blocked liver stage development of sporozoites that was dose-dependent and time-limited. Here, we further characterize the organismal, cellular, and molecular events associated with this protective innate response in the liver and report that a large proportion of the IV administered Hz localized to F4/80 cells in the liver and that the rapid and strong protection against liver-stage development waned quickly such that by 1 week post-Hz treatment animals were fully susceptible to infection. RNAseq of the liver after IV administration of Hz demonstrated that the rapid and robust induction of genes associated with the acute phase response, innate immune activation, cellular recruitment, and IFN-γ signaling observed at day 1 was largely absent at day 7. RNAseq analysis implicated NK cells as the major cellular source of IFN-γ. In vivo cell depletion and IFN-γ neutralization experiments supported the hypothesis that tissue-resident macrophages and NK cells are major contributors to the protective response and the NK cell-derived IFN-γ is key to induction of the mechanisms that block sporozoite development in the liver. These findings advance our understanding of the innate immune responses that prevent liver stage malaria infection.
PubMed: 38670216
DOI: 10.1016/j.micinf.2024.105343 -
Malaria Journal Apr 2024The use of fluorescent proteins (FPs) in Plasmodium parasites has been key to understand the biology of this obligate intracellular protozoon. FPs like the green... (Review)
Review
The use of fluorescent proteins (FPs) in Plasmodium parasites has been key to understand the biology of this obligate intracellular protozoon. FPs like the green fluorescent protein (GFP) enabled to explore protein localization, promoter activity as well as dynamic processes like protein export and endocytosis. Furthermore, FP biosensors have provided detailed information on physiological parameters at the subcellular level, and fluorescent reporter lines greatly extended the malariology toolbox. Still, in order to achieve optimal results, it is crucial to know exactly the properties of the FP of choice and the genetic scenario in which it will be used. This review highlights advantages and disadvantages of available landing sites and promoters that have been successfully applied for the ectopic expression of FPs in Plasmodium berghei and Plasmodium falciparum. Furthermore, the properties of newly developed FPs beyond DsRed and EGFP, in the visualization of cells and cellular structures as well as in the sensing of small molecules are discussed.
Topics: Green Fluorescent Proteins; Plasmodium berghei; Promoter Regions, Genetic; Plasmodium falciparum; Protein Transport
PubMed: 38643106
DOI: 10.1186/s12936-024-04936-9 -
Antimicrobial Agents and Chemotherapy Apr 2024The development of novel antiplasmodial compounds with broad-spectrum activity against different stages of parasites is crucial to prevent malaria disease and parasite...
The development of novel antiplasmodial compounds with broad-spectrum activity against different stages of parasites is crucial to prevent malaria disease and parasite transmission. This study evaluated the antiplasmodial activity of seven novel hydrazone compounds (referred to as CB compounds: CB-27, CB-41, CB-50, CB-53, CB-58, CB-59, and CB-61) against multiple stages of parasites. All CB compounds inhibited blood stage proliferation of drug-resistant or sensitive strains of in the low micromolar to nanomolar range. Interestingly, CB-41 exhibited prophylactic activity against hypnozoites and liver schizonts in , a primate model for . Four CB compounds (CB-27, CB-41, CB-53, and CB-61) inhibited oocyst formation in mosquitoes and five CB compounds (CB-27, CB-41, CB-53, CB-58, and CB-61) hindered the development of ookinetes. The CB compounds did not inhibit the activation of female and male gametocytes . Isobologram assays demonstrated synergistic interactions between CB-61 and the FDA-approved antimalarial drugs, clindamycin and halofantrine. Testing of six CB compounds showed no inhibition of glutathione S-transferase as a putative target and no cytotoxicity in HepG2 liver cells. CB compounds are promising candidates for further development as antimalarial drugs against multidrug-resistant parasites, which could also prevent malaria transmission.
PubMed: 38639491
DOI: 10.1128/aac.01643-23 -
Journal of Vector Borne Diseases Apr 2024The persistent threat of drug resistant malaria demands new cures. Low prevalence of malaria in the Indian state of Kerala compared with other proximal states made us...
BACKGROUND OBJECTIVES
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 made us 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 in Kerala.
METHODS
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 P. berghei ANKA infected mice.
RESULTS
The highest antiplasmodial activities in the alcoholic and aqueous extracts were observed in leaf methanolic extract (IC50 2 μg/ml) and heartwood aqueous extract (IC50 12.5 μg/ml). Ceasalpinia sappan extracts were equipotent against both chloroquine-sensitive Pf3D7 and resistant PfINDO strains and showed suppression of percentage parasitemia in P. berghei infected mice. Activity- guided chromatographic fractionation of aqueous wood extract led to the fortification of antiplasmodial activity (IC50 5 μg/ml).
INTERPRETATION CONCLUSION
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.
PubMed: 38634464
DOI: 10.4103/JVBD.JVBD_18_24 -
ACS Medicinal Chemistry Letters Apr 2024Toward addressing the cardiotoxicity liability associated with the antimalarial drug astemizole (AST, hERG IC = 0.0042 μM) and its derivatives, we designed and...
Toward addressing the cardiotoxicity liability associated with the antimalarial drug astemizole (AST, hERG IC = 0.0042 μM) and its derivatives, we designed and synthesized analogues based on compound ( NF54 IC = 0.012 μM; hERG IC = 0.63 μM), our previously identified 3-trifluoromethyl-1,2,4-oxadiazole AST analogue. Compound retained multistage antiplasmodium activity (ABS NF54 IC = 0.017 μM; gametocytes iGc/LGc IC = 1.24/1.39 μM, and liver-stage HepG2 IC = 2.30 μM), good microsomal metabolic stability (MLM CL < 11 μL·min·mg, < 0.33), and solubility (150 μM). It shows a ∼6-fold and >6000-fold higher selectivity against human ether-á-go-go-related gene higher selectively potential over hERG relative to and AST, respectively. Despite the excellent antiplasmodium activity profile, efficacy in the mouse infection model was diminished, attributable to suboptimal oral bioavailability ( = 14.9%) at 10 mg·kg resulting from poor permeability (log = -0.82). No cross-resistance was observed against 44 common mutant lines, suggesting activity via a novel mechanism of action.
PubMed: 38628794
DOI: 10.1021/acsmedchemlett.3c00496