Supplementary material for the article: Manzano, J. I.; Konstantinović, J.; Scaccabarozzi, D.; Perea, A.; Pavić, A.; Cavicchini, L.; Basilico, N.; Gamarro, F.; Šolaja, B. A. 4-Aminoquinoline-Based Compounds as Antileishmanial Agents That Inhibit the Energy Metabolism of Leishmania. European Journal of Medicinal Chemistry 2019, 180, 28–40. https://doi.org/10.1016/j.ejmech.2019.07.010

Supplementary material for: [https://doi.org/10.1016/j.ejmech.2019.07.010]Related to published version: [http://cherry.chem.bg.ac.rs/handle/123456789/3287

Supplementary data for the article: Konstantinović, J.; Videnović, M.; Orsini, S.; Bogojević, K.; D’Alessandro, S.; Scaccabarozzi, D.; Terzić Jovanović, N.; Gradoni, L.; Basilico, N.; Šolaja, B. A. Novel Aminoquinoline Derivatives Significantly Reduce Parasite Load in Leishmania Infantum Infected Mice. ACS Medicinal Chemistry Letters 2018, 9 (7), 629–634. https://doi.org/10.1021/acsmedchemlett.8b00053

Supplementary material for: [https://pubs.acs.org/doi/10.1021/acsmedchemlett.8b00053]Related to published version: [http://cherry.chem.bg.ac.rs/handle/123456789/2209]Related to accepted version: [http://cherry.chem.bg.ac.rs/handle/123456789/2948

Interplay between Plasmodium falciparum haemozoin and l-arginine: implication for nitric oxide production

Abstract Background Plasmodium falciparum haemozoin, a detoxification product of digested haemoglobin from infected erythrocytes, is released into the bloodstream upon schizont rupture and accumulates in leukocytes. High levels of haemozoin correlate with disease severity. Some studies have shown that concentrations of the substrate of inducible nitric oxide synthase (iNOS), l-arginine, as well as nitric oxide are low in patients infected with P. falciparum malaria. The present study investigates, in vitro, the role of P. falciparum haemozoin on nitric oxide production, iNOS expression in macrophages, and the possible interaction between l-arginine and haemozoin. Methods Plasmodium falciparum haemozoin was obtained from in vitro cultures through magnetic isolation. Phagocytosis of haemozoin by immortalized bone marrow derived macrophages was detected by confocal reflection combined with fluorescence microscopy. Nitrite concentrations in the supernatants was evaluated by Griess assay as a standard indication of nitric oxide production, while iNOS expression was detected on cell extracts by western blotting. Detection of l-arginine in haemozoin-treated or untreated media was achieved by liquid chromatography–tandem mass spectrometry (LC–MS/MS). Results Haemozoin synergizes in vitro with interferon-gamma to produce nitric oxide. However, when mouse macrophages were stimulated with haemozoin, a proportional increase of nitric oxide was observed up to 25 μM of haemozoin, followed by a decrease with doses up to 100 μM, when nitric oxide release was completely abrogated. This was not due to reactive oxygen species production, nor to an effect on iNOS activity. Interestingly, when at 24 h, haemozoin-treated macrophages were washed and incubated in fresh medium for further 24 h, the nitric oxide production was restored in a dose–response manner. Similar results were seen when l-arginine-enriched media was used in the stimulation. Moreover, muramyldipeptide, a strong nitric oxide inducer, was unable to activate macrophages to release nitric oxide in the presence of haemozoin-treated medium. By LC–MS/MS a complete depletion of l-arginine was observed in this haemozoin-treated, conditioned medium. Conclusions It is proposed that haemozoin interacts with l-arginine reducing its availability for iNOS, and thus decreasing nitric oxide production. The clinical (or pathological) implications of these results are discussed

Supplementary data for the article: Konstantinović, J.; Videnović, M.; Orsini, S.; Bogojević, K.; D’Alessandro, S.; Scaccabarozzi, D.; Terzić Jovanović, N.; Gradoni, L.; Basilico, N.; Šolaja, B. A. Novel Aminoquinoline Derivatives Significantly Reduce Parasite Load in Leishmania Infantum Infected Mice. ACS Medicinal Chemistry Letters 2018, 9 (7), 629–634. https://doi.org/10.1021/acsmedchemlett.8b00053

Supplementary material for: [https://pubs.acs.org/doi/10.1021/acsmedchemlett.8b00053]Related to published version: [http://cherry.chem.bg.ac.rs/handle/123456789/2209]Related to accepted version: [http://cherry.chem.bg.ac.rs/handle/123456789/2948

Altered Lipid Composition of Surfactant and Lung Tissue in Murine Experimental Malaria-Associated Acute Respiratory Distress Syndrome.

Malaria-associated acute lung injury (MA-ALI) and its more severe form malaria-associated acute respiratory distress syndrome (MA-ARDS) are common, often fatal complications of severe malaria infections. However, little is known about their pathogenesis. In this study, biochemical alterations of the lipid composition of the lungs were investigated as possible contributing factors to the severity of murine MA-ALI/ARDS. C57BL/6J mice were infected with Plasmodium berghei NK65 to induce lethal MA-ARDS, or with Plasmodium chabaudi AS, a parasite strain that does not induce lung pathology. The lipid profile of the lung tissue from mice infected with Plasmodium berghei NK65 developing MA-ALI/ARDS, but not that from mice without lung pathology or controls, was characterized by high levels of phospholipids -mainly phosphatidylcholine- and esterified cholesterol. The high levels of polyunsaturated fatty acids and the linoleic/oleic fatty acid ratio of the latter reflect the fatty acid composition of plasma cholesterol esters. In spite of the increased total polyunsaturated fatty acid pool, which augments the relative oxidability of the lung membranes, and the presence of hemozoin, a known pro-oxidant, no excess oxidative stress was detected in the lungs of Plasmodium berghei NK65 infected mice. The bronchoalveolar lavage (BAL) fluid of Plasmodium berghei NK65 infected mice was characterized by high levels of plasma proteins. The phospholipid profile of BAL large and small aggregate fractions was also different from uninfected controls, with a significant increase in the amounts of sphingomyelin and lysophosphatidylcholine and the decrease in phosphatidylglycerol. Both the increase of proteins and lysophosphatidylcholine are known to decrease the intrinsic surface activity of surfactant. Together, these data indicate that an altered lipid composition of lung tissue and BAL fluid, partially ascribed to oedema and lipoprotein infiltration, is a characteristic feature of murine MA-ALI/ARDS and possibly contribute to lung dysfunction

Differential induction of malaria liver pathology in mice infected with Plasmodium chabaudi AS or Plasmodium berghei NK65

Cerebral malaria and severe anaemia are the most common deadly complications of malaria, and are often associated, both in paediatric and adult patients, with hepatopathy, whose pathogenesis is not well characterized, and sometimes also with acute respiratory distress syndrome (ARDS). Here, two species of murine malaria, the lethal Plasmodium berghei strain NK65 and self-healing Plasmodium chabaudi strain AS which differ in their ability to cause hepatopathy and/or ARDS were used to investigate the lipid alterations, oxidative damage and host immune response during the infection in relation to parasite load and accumulation of parasite products, such as haemozoin.status: publishe

MOESM1 of Differential induction of malaria liver pathology in mice infected with Plasmodium chabaudi AS or Plasmodium berghei NK65

Additional file 1. ALT and AST determination in mice infected with P. berghei NK65 or P. chabaudi AS. C57BL/6J mice were injected intraperitoneally with 104 erythrocytes infected with P. berghei NK65 or P. chabaudi AS. Serum levels of AST (panel A), ALT (panel B) and the AST/ALT ratio (panel C) were determined at day 8 and 10 post infection according to manufacturer’s protocol (Teco Diagnostics, California, USA). n = 3-6 mice for each time point and strain, additional data can be found in [16]. *p < 0.05; **p < 0.01 versus control