Hemozoin Research Articles

Hematin anhydride (β-hematin): An analogue to malaria pigment hemozoin possesses nonlinearity

Hematin anhydride (β-hematin), the synthetic analogue of the malaria pigment, “hemozoin”, is a heme dimer produced by reciprocal covalent bonds among carboxylic acid groups on the protoporphyrin-IX ring and the iron atom present in the two adjacent heme molecules. Hemozoin is a disposal product formed from the digestion of hemoglobin present in the red blood cells infected with hematophagous malaria parasites. Besides, as the parasites invade red blood cells, hemozoin crystals are eventually released into the bloodstream, where they accumulate over time in tissues. Severe malaria infection leads to significant dysfunction in vital organs such as the liver, spleen, and brain in part due to the autoimmune response to the excessive accumulation of hemozoin in these tissues. Also, the amount of these crystals in the vasculature correlates with disease progression. Thus, hemozoin is a unique indicator of infection used as a malaria biomarker and hence, used as a target for the development of antimalarial drugs. Hence, exploring various properties of hemozoin is extremely useful in the direction of diagnosis and cure. The present study focuses on finding one of the unknown properties of β-hematin in physiological conditions by using the Z-scan technique, which is simple, sensitive, and economical. It is observed that hemozoin possesses one of the unique material properties, i.e., nonlinearity with a detection limit of ∼ 15 µM. The self-defocusing action causes β-hematin to exhibit negative refractive nonlinearity. The observed data is analyzed with a thermal lensing model. We strongly believe that our simple and reliable approach to probing the nonlinearity of β-hematin will provide fresh opportunities for malaria diagnostics & cure in the near future.

Soluble meso and deuteroporphyrin analogs of the malaria pigment hematin anhydride

Two soluble heme analogs of the insoluble malaria pigment hematin anhydride (HA, or β-hematin), [Fe(III)(protoporphyrin)]2, with either mesoporphyrin (MHA) or deuteroporphyrin (DHA) are characterized by elemental analysis, SEM, IR spectroscopy, electronic spectroscopy, paramagnetic 1H NMR spectroscopy and solution magnetic susceptibility. While prior single crystal and X-ray powder diffraction results indicate all three have a common propionate linked dimer motif, there is considerable solid state variation in the conformation. This is associated with enhanced solubility of MHA and DHA. As with HA, DHA undergoes thermally promoted reversible hydration/dehydration in the solid state. Solution 1H NMR studies of DHA suggest a high spin dimeric structure with the porphyrin methyls distributed between two isomers which are also present in the solid state. These soluble iron(III)porphyrin dimers allow for the first direct solution studies by NMR and UV–Vis spectroscopies of these key species. Taken together the results illustrate the importance and utility of varying the substituents on the periphery of the porphyrin for studying heme aggregation and malaria pigment formation.

Evaluation of in vitro antimalarial activity of <i>Citrus aurantifolia</i> (CHRISTM.) and <i>Ziziphus mauritiana</i> (LAM) using heme polymerization inhibition assay

Citrus aurantifolia and Ziziphus mauritiana (Lam) are plants that have numerous medicinal properties and are used extensively in traditional medicine for the treatment of several diseases. Malaria parasites metabolize haemoglobin and detoxify the resulting haem by polymerizing it to form haemozoin (malaria pigment). This study aimed at evaluating the antiplasmodial activity through in vitro cell-free heme polymerization inhibition assay (HPIA) of C. aurantifolia and Z. mauritiana. The analysis was conducted using a 96-well microtiter plate reader at an absorption of 400 nm, at different concentrations (0-2 mg/ml). The IC50 was determined by non-linear regression analysis using GraphPad Prism. The result indicated that both the root extracts of C. aurantifolia and Z. mauritiana showed significant difference (p˂0.0001) with IC50 of 0.81 mg/ml and 0.77 mg/ml respectively compared to positive control. Phytochemical screening of the ethanol root extracts showed the presence of saponins, alkaloids, terpenoids, glycosides, steroids, and anthraquinone, while tannins and reducing sugars were not detected in both samples used, flavonoid was detected in C. aurantifolia. Both the extracts exhibited potential for inhibiting heme polymerization in vitro.

The association of intraleucocytic malaria pigment and disease severity in Papua New Guinean children with severe P. falciparum malaria.

Plasmodium falciparum pigment-containing leucocytes (PCLs) are associated with adverse clinical manifestations of severe malaria in African children. However, limited data exist on the association of PCLs in settings outside of Africa. Thin films on peripheral blood slides obtained from children ages 6months-10y with severe malaria were examined for PCLs. The intraleucocytic pigment data were correlated with clinical phenotypic data such as severe anaemia, metabolic acidosis and coma to determine the association of PCLs with clinical phenotypes of severe malaria and outcome. Of the 169 children with severe P. falciparum malaria confirmed by microscopy, 76% (129/169) had PCLs. Compared with children without PCLs, the presence (adjusted odds ratio [AOR] 3.2 [95% confidence interval {CI} 1.5 to 6.9], p≤0.01) and quantity (AOR 1.0 [95% CI 1.0 to 1.1], p=0.04) of pigment-containing monocytes (PCMs) was significantly associated with severe anaemia, while the quantity of both PCMs (AOR 1.0 [95% CI 1.0 to 1.1], p≤0.01) and pigment-containing neutrophils (AOR 1.0 [95% CI 1.0 to 1.1], p=0.01) was significantly associated with metabolic acidosis. Plasma P. falciparum histidine-rich protein-2 level negatively correlated with the platelet count (r=-0.5, p≤0.01) in patients with PCLs and no PCLs. In Papua New Guinean children with severe P. falciparum malaria, the presence and quantity of PCLs are predictors of disease severity, severe anaemia and metabolic acidosis.

Posttranslational Modification of Human Cytochrome CYP4F11 by 4-Hydroxynonenal Impairs ω-Hydroxylation in Malaria Pigment Hemozoin-Fed Monocytes: The Role in Malaria Immunosuppression.

Malaria is a frequent parasitic infection becomes life threatening due to the disequilibrated immune responses of the host. Avid phagocytosis of malarial pigment hemozoin (HZ) and HZ-containing Plasmodium parasites incapacitates monocyte functions by bioactive lipoperoxidation products 4-hydroxynonenal (4-HNE) and hydroxyeicosatetraenoic acids (HETEs). CYP4F conjugation with 4-HNE is hypothesised to inhibit ω-hydroxylation of 15-HETE, leading to sustained monocyte dysfunction caused by 15-HETE accumulation. A combined immunochemical and mass-spectrometric approach identified 4-HNE-conjugated CYP4F11 in primary human HZ-laden and 4-HNE-treated monocytes. Six distinct 4-HNE-modified amino acid residues were revealed, of which C260 and H261 are localized in the substrate recognition site of CYP4F11. Functional consequences of enzyme modification were investigated on purified human CYP4F11. Palmitic acid, arachidonic acid, 12-HETE, and 15-HETE bound to unconjugated CYP4F11 with apparent dissociation constants of 52, 98, 38, and 73 µM, respectively, while in vitro conjugation with 4-HNE completely blocked substrate binding and enzymatic activity of CYP4F11. Gas chromatographic product profiles confirmed that unmodified CYP4F11 catalysed the ω-hydroxylation while 4-HNE-conjugated CYP4F11 did not. The 15-HETE dose dependently recapitulated the inhibition of the oxidative burst and dendritic cell differentiation by HZ. The inhibition of CYP4F11 by 4-HNE with consequent accumulation of 15-HETE is supposed to be a crucial step in immune suppression in monocytes and immune imbalance in malaria.

Neuroprotection by Alstonia boonei De Wild., Anacardium occidentale L., Azadirachta indica A.Juss. and Mangifera indica L.

Alstonia boonei De Wild. (stem bark), Anacardium occidentale L. (stem bark), Azadirachta indica A.Juss (leaves), Enantia chlorantha Oliv. (stem bark), Khaya senegalensis A.Juss (stem bark) Mangifera indica L. (stem bark), and Nauclea latifolia Sm. (stem bark) are used for treating malaria in southwest Nigeria. Surveys revealed that these plants are also employed for treating symptoms of malaria and cerebral malaria in the region. In this study, the effects of freeze-dried extracts of these plants were investigated on synthetic hemozoin (HZ)-induced neuroinflammation, neuronal damage, and increased permeability of brain microvascular endothelial cells. Effects of freeze-dried plant extracts were investigated on neuroinflammation by measuring levels of pro-inflammatory mediators in culture supernatants, while in-cell western assays were used to measure protein levels of iNOS and NLRP3. Effects on HZ-induced neurotoxicity and ROS generation was measured using MTT and DCFDA assays, respectively. HZ-induced permeability of hCMEC/D3 endothelial cells was determined using the in vitro vascular permeability assay kit. The extracts produced significant (p<0.05) reduction in TNFα, IL-6, IL-1β, MCP-1, RANTES and iNOS/NO production in HZ-stimulated BV-2 microglia. Pre-treatment with 50μg/mL of A. boonei, A. indica, A. occidentale, E. chlorantha and M. indica also resulted in the inhibition of NF-κB activation. Pre-treatment with A. indica produced, A. occidentale, M. indica and A. boonei reduced HZ-induced increased NLRP3 protein expression. HZ-induced increased caspase-1 activity was also reduced by A. boonei, A. occidentale, A. indica, E. chlorantha, and M. indica. Freeze-dried extracts of A. boonei, A. occidentale, A. indica and M. indica produced neuroprotective effect in HT-22 neuronal cells incubated with HZ by preventing HZ-induced neurotoxicity, ROS generation, DNA fragmentation and caspase 3/7 activity. Inhibition of HZ-induced increase in permeability of human hCMEC/D3 brain endothelial cells was also observed with A. boonei, A. occidentale, A. indica and M. indica, while reducing the release of TNFα and MMP-9. These results suggest that A. boonei, A. occidentale, A. indica and M. indica are neuroprotective through inhibition of neuroinflammation, neuronal damage and increased permeability of blood brain barrier. The outcome of the study provides pharmacological evidence for the potential benefits of plants as herbal treatments for cerebral malaria symptoms.

Vitamin C-rich juice co-administration with artemether-lumefantrine ameliorates oxido-inflammatory responses in Plasmodium berghei-infected mice.

This study investigated the effects of co-administration of a commercial juice rich in vitamin C (Vit C) on the antimalarial efficacy of artemether-lumefantrine (AL) in Plasmodium berghei-infected mice. Fifty Balb/c mice were infected with Plasmodium berghei NK65 strain from a donor mouse. Parasitemia was established after 72h. Animals were grouped into 6 (n = 10) and treated daily for 3days with normal saline, chloroquine, artemether-lumefantrine (AL), AL plus 50% commercial juice (CJ), and AL plus 50% Vit C supplementation in drinks ad libitum, respectively. Body weight, parasitemia levels, and mean survival time were determined. Tumor necrosis factor-alpha (TNF-α), interleukin 6 (IL-6), nitrite, malondialdehyde, reduced glutathione (GSH), catalase, and superoxide dismutase (SOD) were determined in the serum and liver tissues. Spleen histopathological changes were determined by H&E staining. Parasitemia was cleared by administration of AL and was not affected by Vit C and CJ supplementation. Vit C significantly prevented body weight reduction in AL-treated mice. CJ and Vit C supplementation to AL-treated mice significantly improved survival proportion compared with AL alone animals. Vit C and CJ supplementation significantly improved reduction of TNF-α, IL-6, and malondialdehyde, and increased GSH, CAT, and SOD in AL-treated mice. Spleen cell degeneration and presence of malaria pigment were reduced in AL-treated animals. The results suggest that ad libitum co-administration of commercial juice and vitamin C with artemether-lumefantrine does not impair its antimalarial efficacy but rather improved antioxidant and anti-inflammatory effects in mice.

Unusual blood smear with multiple stages of Plasmodium falciparum infection and intraleukocytic malaria pigments in an expatriate with severe malaria and delayed clearance of parasites.

A French expatriate in Ethiopia presented with severe Plasmodium falciparum infection. The blood smear was remarkable associating multiple stages of parasites including circulating schizonts with a high rate of intraleukocytic malaria pigments. Under artesunate treatment, without polymorphism in PfK13 gene, delayed clearance of parasites occurred, probably following the massive merogony.

Plasmodium berghei Purified Hemozoin Associated with DNA Strongly Inhibits P. berghei Liver-Stage Development in BALB/c Mice after Intravenous Inoculation.

In the acidic lysosome-like digestive vacuole, Plasmodium parasites crystallize heme from hemoglobin into hemozoin, or malaria pigment. Upon release of progeny merozoites, the residual hemozoin is phagocytized by macrophages principally in the liver and spleen where the heme crystals can persist for months to years, as heme oxygenase does not readily degrade the crystal. Previous studies demonstrated hemozoin modulation of monocytes and macrophages. Hemozoin modulates immune function activity of monocytes/macrophages. Here, we used purified/washed hemozoin (W-Hz) isolated from murine Plasmodium berghei infections and intravenously (i.v.) injected it back into naive mice. We characterized the modulating effect of W-Hz on liver-stage replication. Purified washed hemozoin decreases P. berghei liver levels both at 1 week and 1 month after i.v. injection in a dose and time dependent fashion. The injected hemozoin fully protected in nine out of 10 mice given a 50 sporozoite inoculum, and in 10 out of 10 mice against 2,000 sporozoites when they were infected an hour or a day after hemozoin inoculation. DNase treatment at the hemozoin reversed the observed liver load reduction. The liver load reduction was similar in mature B- and T-cell-deficient RAG-1 knockout (KO) mice suggesting an innate immune protection mechanism. This work indicates a role for residual hemozoin in down modulation of Plasmodium liver stages.

The prognostic and diagnostic value of intraleukocytic malaria pigment in patients with severe falciparum malaria

Severe falciparum malaria is a major cause of death in tropical countries, particularly in African children. Rapid and accurate diagnosis and prognostic assessment are critical to clinical management. In 6027 prospectively studied patients diagnosed with severe malaria we assess the prognostic value of peripheral blood film counts of malaria pigment containing polymorphonuclear leukocytes (PMNs) and monocytes. We combine these results with previously published data and show, in an individual patient data meta-analysis (n = 32,035), that the proportion of pigment containing PMNs is predictive of in-hospital mortality. In African children the proportion of pigment containing PMNs helps distinguish severe malaria from other life-threatening febrile illnesses, and it adds to the prognostic assessment from simple bedside examination, and to the conventional malaria parasite count. Microscopy assessment of pigment containing PMNs is simple and rapid, and should be performed in all patients hospitalised with suspected severe malaria.

Age-modified factors associated with placental malaria in rural Burkina Faso

BackgroundMalaria in pregnancy can result in placental infection with fetal implications. This study aimed at assessing placental malaria (PM) prevalence and its associated factors in a cohort of pregnant women with peripheral malaria and their offspring.MethodThe data were collected in the framework of a clinical trial on treatments for malaria in pregnant women . Placental malaria (PM) was diagnosed by histopathological detection of parasites and/or malaria pigment on placenta biopsies taken at delivery. Factors associated with PM were assessed using logistic regression.ResultsOut of 745 biopsies examined, PM was diagnosed in 86.8 % of women. Acute, chronic and past PM were retrieved in 11 (1.5 %), 170 (22.8 %), and 466 (62.6 %) women, respectively. A modifying effect was observed in the association of gravidity or anemia at the study start with pooled PM (presence of parasites and/or malaria pigment). In women under 30, gravidity ≤ 2 was associated with an increased prevalence of pooled PM but in women aged 30 years or more, gravidity was no more associated with pooled PM (OR 6.81, 95 % CI 3.18 – 14.60; and OR 0.52, 95 % CI 0.10 – 2.76, respectively). Anemia was associated with pooled PM in women under 30 (OR 1.96, 95 % CI 1.03 – 3.72) but not in women aged 30 years or more (OR 0.68, 95 % CI 0.31 – 1.49). Similarly, the association of gravidity with past-chronic PM depended also on age. A higher prevalence of active PM was observed in women under 30 presenting with symptomatic malaria (OR 3.79, 95 % CI 1.55 – 9.27), while there was no significant increase in the prevalence of active PM (presence of parasites only) in women with symptomatic malaria when aged 30 years or more (OR 0.42, 95 % CI 0.10 – 1.75). In women with chronic PM, the prevalence of low birth weight or prematurity was the highest (31.2 %) as compared with past PM or no PM.ConclusionDespite the rapid diagnosis and efficacious treatment of peripheral infection, the prevalence of placental malaria remained high in women with P. falciparum peripheral infection in Nanoro, especially in younger women This underlines the importance of preventive measures in this specific group.

Imported malaria in adults: about a case of cerebral malaria

BackgroundMalaria is the first parasitic infection endemic in the world caused by parasites species of Plasmodium. Cerebral malaria (CM) is a rapidly progressive and severe form of Plasmodium falciparum infection, characterized by a greater accumulation of red blood cells parasitized by Plasmodium falciparum in the brain. The diagnosis of malaria is usually made in living patients from a blood sample taken in the course of a fever on return from an endemic country, whereas CM, often associated with fatal outcomes even in treated subjects, is usually diagnosed at autopsy.Case presentationWe present the case of a 36-year-old man who died a few days after returning from a business trip to the Ivory Coast. As a result of an unclear cause of death, a medicolegal autopsy was ordered. Autopsy findings revealed massive congestion and edema of the brain with no other macroscopic abnormalities at organ gross examination. Histology and laboratory tests were conducted revealing a Plasmodium falciparum infection, with numerous parasitized erythrocytes containing dots of hemozoin pigment (malaria pigment) in all examined brain sections and all other organs. Death was attributed to cerebral malaria with multiple organ failure.ConclusionsThis report summarizes several features for the diagnosis of malaria and how postmortem investigations, as well as histology and laboratory diagnosis, may lead to a retrospective diagnosis of a fatal complicated form with cerebral involvement.

Malaria Pigment Hemozoin Impairs GM-CSF Receptor Expression and Function by 4-Hydroxynonenal.

Malarial pigment hemozoin (HZ) generates the lipoperoxidation product 4-hydroxynonenal (4-HNE), which is known to cause dysregulation of the immune response in malaria. The inhibition of granulocyte macrophage colony-stimulating factor (GM-CSF)-dependent differentiation of dendritic cells (DC) by HZ and 4-HNE was previously described in vitro, and the GM-CSF receptor (GM-CSF R) was hypothesised to be a primary target of 4-HNE in monocytes. In this study, we show the functional impact of HZ on GM-CSF R in monocytes and monocyte-derived DC by (i) impairing GM-CSF binding by 50 ± 9% and 65 ± 14%, respectively (n = 3 for both cell types); (ii) decreasing the expression of GM-CSF R functional subunit (CD116) on monocyte’s surface by 36 ± 11% (n = 6) and in cell lysate by 58 ± 16% (n = 3); and (iii) binding of 4-HNE to distinct amino acid residues on CD116. The data suggest that defective DC differentiation in malaria is caused by GM-CSF R dysregulation and GM-CSF R modification by lipoperoxidation product 4-HNE via direct interaction with its CD116 subunit.

Hemozoin: a Complex Molecule with Complex Activities.

Malaria is a disease caused by parasites that reside in host red blood cells and use hemoglobin as a nutrient source. Heme released by hemoglobin catabolism is modified by the parasite to produce hemozoin (HZ), which has toxic effects on the host. Experimentation aiming to elucidate how HZ contributes to malaria pathogenesis has utilized different preparations of this molecule, complicating interpretation and comparison of findings. We examine natural synthesis and isolation of HZ and highlight studies that have used multiple preparations, including synthetic forms, in a comparative fashion. Recent work utilizing sophisticated imaging and detection techniques reveals important molecular characteristics of HZ synthesis and biochemistry. Other recent studies further refine understanding of contributions of HZ to malaria pathogenesis yet highlight the continuing need to characterize HZ preparations and contextualize experimental conditions in the in vivo infection milieu. This review highlights the necessity of collectively determining what is physiologically relevant HZ. Characterization of isolated natural HZ and use of multiple preparations in each study are recommended with application of in vivo studies whenever possible. Adoption of such practices is expected to improve reproducibility of results and elucidate the myriad of ways that HZ participates in malaria pathogenesis.

Hemozoin is a potential threat in cerebral malaria pathology through the induction of RBC-EC cytoadherence

Cerebral malaria is an outcome of multifaceted and complicated condition. Cytoadherence is one critical factor in cerebral malaria pathology as high order cytoadherence complexes result in vascular congestion and cell apoptosis. Morphological abnormalities in uninfected RBCs can be a contributing factor to aggravate cytoadherence. Malaria pigment hemozoin is a potential bioactive molecule and the role of this pigment in cerebral malaria pathology is not completely understood. To understand this, primarily we investigated the impact of hemozoin pigment on uninfected RBCs. Secondarily, we investigated the role of this pigment in formation of endothelial cells-RBCs (EC-RBC) cytoadherence complex. We first observed that a dose dependent hemozoin exposure to uninfected RBCs induced structural abnormalities. Differential counting of these abnormal RBCs indicated population of acanthocytes, spherocytes and microcytes. The formation of abnormal RBCs was observed with phosphatidylserine externalization. Lipid peroxidation, reduced glutathione and reactive oxygen species (ROS) levels indicated an increase in hemozoin exposure mediated oxidative stress. Our in-vitro cytoadherence assay indicated formation of endothelial EC-RBC cytoadherence complex. The dose dependent hemozoin exposure to uninfected RBCs resulted in oxidative stress mediated high order cytoadherence complex formation. This effect was reversed in presence of antioxidant molecules. The inhibitory effect of antioxidant molecules indicates that oxidative stress can be a regulatory factor to control cerebral malaria pathology. Being the first report to highlight the impact of malaria pigment hemozoin on uninfected RBCs, this study brings attention to the role of abnormal RBCs in worsening of cerebral malaria pathology.

Hemozoin Promotes Lung Inflammation via Host Epithelial Activation.

Respiratory distress in severe malaria is associated with high mortality, but its pathogenesis remains unclear. The malaria pigment hemozoin (HZ) is abundant in target organs of severe malaria, including the lungs, and is known to be a potent innate immune activator of phagocytes. We hypothesized that HZ might also stimulate lung epithelial activation and thereby potentiate lung inflammation. We show here that airway epithelium stimulated with HZ undergoes global transcriptional reprogramming and changes in cell surface protein expression that comprise an epithelial activation phenotype. Proinflammatory signaling is induced, and key cytoadherence molecules are upregulated, including several associated with severe malaria, such as CD36 and ICAM1. Epithelial and extracellular matrix remodeling pathways are transformed, including induction of key metalloproteases and modulation of epithelial junctions. The overall program induced by HZ serves to promote inflammation and neutrophil transmigration, and is recapitulated in a murine model of HZ-induced acute pneumonitis. Together, our data demonstrate a direct role for hemozoin in stimulating epithelial activation that could potentiate lung inflammation in malaria.IMPORTANCE Respiratory distress (RD) is a complication of severe malaria associated with a particularly high risk for death in African children infected with the parasite Plasmodium falciparum The pathophysiology underlying RD remains poorly understood, and the condition is managed supportively. The parasite-derived factor HZ accumulates in target organs of severe malaria, including the lungs, and is a potent stimulator of immune cells. Our findings demonstrate that HZ causes global activation of lung epithelial cells, a response that directly promotes lung inflammation. HZ stimulates expression of key proinflammatory and cell surface molecules, alters signaling pathways involved in epithelial-matrix remodeling, and promotes neutrophil transmigration and airway inflammation. The lung epithelial activation induced by HZ mimics patterns seen in malarial lung injury and provides new insights into the molecular pathogenesis of RD.

The Relationship Between Fetal Weight with Sequestration of Infected Erythrocyte, Monocyte Infiltration, and Malaria Pigment Deposition in Placenta of Mother Giving Birth Suffering from Plasmodium Vivax Infection.

Background:Malaria in pregnancy can cause fatal complications by parasite sequestration mechanism, which can cause monocyte infiltration in the intervillous space. P. vivax infection was significantly associated with malaria pigment in the placenta, indicating past subclinical infectionsObjective:This study aimed to determine the mechanism of P. vivax in the pathogenesis of placental malaria and its relationship with LBW.Methods:This study was observational analytic with a cross-sectional approach. Placental tissue samples were obtained from pregnant women with LBW babies during delivery in Maumere, Nusa Tenggara Timur. The samples used in this study were confirmed by a polymerase chain reaction and consisted of 25 samples with 12 positive and 13 negative samples. Placental tissue samples were made with Hematoxylin-Eosin staining and observed under 1000x magnification at 100 fields using a light microscope. Parasite density, monocyte infiltration, and parasite pigments deposition were calculated.Results:Microscopic observation revealed that there was a significant difference in infected erythrocytes sequestration between groups. Interestingly, monocyte and malaria pigments accumulation were found in malaria-positive and -negative groups, and no significant difference between groups. The correlation test showed no significant relationship between monocyte infiltration and LBW in the malaria-positive and -negative group and between parasite pigments and LBW in both groups. Moreover, there was no significant correlation between parasite density and LBW in the positive and negative groups.Conclusion:P. vivax infection causes acute, sub-acute, and chronic placental malaria in subclinical infected pregnant women in Maumere, Nusa Tenggara Timur that might cause an LBW baby.

Description, measurement, and automatic classification of the Plasmodium berghei oocyst morphology during early differentiation in vitro

After colonization of the mosquito midgut by the malaria parasite, Plasmodium differentiates from an invasive, motile ookinete to a multiplicative, sessile oocyst. Despite their importance in establishing the infection and increasing its population, relatively little is known about the early morphological transformation associated with these changes in function. Oocyst differentiation begins with the formation of a spherical protrusion near the center of the crescent-shaped ookinete. As this protuberance grows, it engulfs the content of the two distal ends, thus rounding the cell. In this work, scrutinized observations of the overall changes in shape, coupled with the migration of the malaria pigment granules and the nucleus into the protuberance, revealed that the movement of the cell content happens in an anteroposterior manner. The resulting data, formalized as morphometric measurements, led to the identification of 5 transitional stages and to the development of a computer training algorithm that automatically classifies them. Since cell differentiation has been associated with redox fluctuations, the classification algorithm was tested with parasites stained with a glutathione-specific fluorescent probe. This revealed changes in the glutathione content during differentiation that are suggestive of a redox modulation during transformation. • Plasmodium ookinetes differentiate into reproductive oocysts after mosquito invasion. • Visual scrutiny of the transformation led to the recognition of 5 transitional stages. • Transformation and organelle migration happens in an anteroposterior manner. • A machine learning algorithm was developed to automatically classify these stages. • The discretization into stages allowed the determination of GSH oscillations.

Factors associated with sub-microscopic placental malaria and its association with adverse pregnancy outcomes among HIV-negative women in Dar es Salaam, Tanzania: a cohort study

BackgroundMalaria infection during pregnancy has negative health consequences for both mothers and offspring. Sub-microscopic malaria infection during pregnancy is common in most African countries. We sought to identify factors associated with sub-microscopic placental malaria, and its association with adverse pregnancy outcomes among HIV-negative pregnant women in Dar es Salaam, Tanzania.MethodsWe recruited a cohort of pregnant women during their first trimester and assessed for the occurrence of placental malaria and pregnancy outcomes. The follow-up was done monthly from recruitment until delivery. Histopathology placental malaria positive results were defined as the presence of malaria pigment or parasitized erythrocytes on the slide (histology-positive (HP)), and the sub-microscopic placental infection was defined as positive Plasmodium falciparum DNA by polymerase chain reaction (DNA PCR) amplification in a negative histopathology test. Adverse pregnancy outcomes investigated included low birth weight (birth weight below 2.5 kg), prematurity (live birth below 37 weeks), and small-for-gestational-age (SGA) (live born with a birth weight below 10th percentile for gestational age and sex). Weighted baseline category logit, log-binomial, and log-Poisson models were used to assess factors associated with placental malaria, and its association with adverse pregnancy outcomes.ResultsAmong 1115 women who had histopathology and DNA PCR performed, 93 (8%) had HP placental infection, and 136 (12%) had the sub-microscopic placental infection. The risk of sub-microscopic placental malaria was greater in women who did not use mosquito prevention methods such as bed nets, fumigation, or mosquito coils (odds ratio (OR) = 1.75; 95% confidence interval (CI): 1.05–2.92; P = 0.03) and in women who were anemic (OR = 1.59; 95% CI: 1.20–2.11; P = 0.001). Women who were underweight had reduced odds of sub-microscopic placental malaria infection (OR = 0.33; 95% CI: 0.17–0.62; P = 0.001). Women who were overweight/obese had 1.48 times higher the odds of HP placental malaria compared to normal weight (OR = 1.48; 95% CI: 1.03–2.11; P = 0.03). HP placental malaria infection was associated with an increased risk of SGA births (RR = 1.30, 95% CI: 0.98–1.72, P = 0.07). In contrast, the sub-microscopic infection was associated with a reduced risk of SGA births (RR = 0.61, 95% CI: 0.43–0.88, P = 0.01). Placental malaria was not associated with low birth weight or prematurity.ConclusionMalaria prevention methods and maternal nutrition status during early pregnancy were important predictors of sub-microscopic placental malaria. More research is needed to understand sub-microscopic placental malaria and the possible mechanisms mediating the association between placental malaria and SGA.

Plasmodium infection inhibits tumor angiogenesis through effects on tumor-associated macrophages in a murine implanted hepatoma model

BackgroundHepatocellular carcinoma (HCC) is one of the leading causes of cancer-related death in China. The lack of an effective treatment for this disease results in a high recurrence rate in patients who undergo radical tumor resection, and the 5-year survival rate of these patients remains low. Our previous studies demonstrated that Plasmodium infection provides a potent antitumor effect by inducing innate and adaptive immunity in a murine Lewis lung carcinoma (LLC) model.MethodsThis study aimed to investigate the inhibitory effect of Plasmodium infection on hepatocellular carcinoma in mice, and various techniques for gene expression analysis were used to identify possible signal regulation mechanisms.ResultsWe found that Plasmodium infection efficiently inhibited tumor progression and prolonged survival in tumor-bearing mice, which served as a murine implanted hepatoma model. The inhibition of tumor progression by Plasmodium infection was related to suppression of tumor angiogenesis within the tumor tissue and decreased infiltration of tumor-associated macrophages (TAMs). Further study demonstrated that matrix metalloprotease 9 (MMP-9) produced by TAMs contributed to tumor angiogenesis in the tumor tissue and that the parasite-induced reduction in MMP-9 expression in TAMs resulted in the suppression of tumor angiogenesis. A mechanistic study revealed that the Plasmodium-derived hemozoin (HZ) that accumulated in TAMs inhibited IGF-1 signaling through the PI3-K and MAPK signaling pathways and thereby decreased the expression of MMP-9 in TAMs.ConclusionsOur study suggests that this novel approach of inhibiting tumor angiogenesis by Plasmodium infection is of high importance for the development of new therapies for cancer patients.DcTAkd_JguhWjRG6hMrBFrVideo abstract