Biology subjects

Introini, V.

Publications and source records attributed to Introini, V..

9 recordsLinked to original sources

bioRxiv2025

Febrile temperature enhances Plasmodium falciparum cytoadhesion by disrupting the endothelial glycocalyx

Fever, a universal host defense response in infection and inflammation, paradoxically contributes to neurological complications in malaria. Febrile temperatures are known to enhance parasite virulence protein expression, but direct effects on the human endothelium remain unknown. We found that a 1-hour exposure to 40 {degrees}C, representative of fever in children with cerebral malaria, increased adhesion of Plasmodium falciparum-infected red blood cells and neutrophils to 3D brain microvascular models displaying a wide wall shear stress gradient. Mechanistically, this brief hyperthermia triggered rapid endothelial glycocalyx shedding, exposing endothelial receptors for binding. This response was more pronounced in brain than in pulmonary microvessels, revealing a greater vulnerability of the cerebral vasculature to fever. Pharmacological inhibition of matrix metalloproteinase activity preserved glycocalyx integrity and abolished the temperature-induced increase in adhesion. These findings identify fever as a host-specific amplifier of malaria-associated microvascular pathology, highlighting the importance of antipyretic strategies to mitigate disease severity.

microbiology↗

bioRxiv2025

Microfluidic Platform for Automatic Quantification of Malaria Parasite Invasion Under Physiological Flow Conditions

Understanding the impact of forces generated by blood flow on biological processes in the circulatory system, such as the invasion of human red blood cells by malaria parasites, is currently limited by the lack of experimental systems that integrate them. Recent systematic quantification of the growth of Plasmodium falciparum, the species that causes the majority of malaria mortality, under a range of shaking conditions has shown that parasite invasion of erythrocytes is affected by the shear stress to which the interacting P. falciparum merozoites and their target red blood cells are exposed. Blood flow could similarly impact shear stress and therefore invasion in vivo, but there is currently no method to test the impact of flow-induced forces on parasite invasion. We have developed a microfluidic device with four channels, each with dimensions similar to those of a post-capillary venule, but with different flow velocities. Highly synchronised P. falciparum parasites are injected into the device, and parasite egress and invasion rates are quantified using newly developed custom video analysis, which fully automates cell type identification and trajectory tracking. The device was tested with both wild-type P. falciparum lines and lines in which genes encoding proteins involved in parasite invasion had been deleted. Deletion of Erythrocyte Binding Antigen 175 (PfEBA175) has a significant impact on invasion under flow, but not in static culture. These findings establish for the first time that flow conditions can critically affect parasite invasion in a genotype-dependent manner. The method can be applied to other biological processes affected by fluid motion, such as cell adhesion, migration, and mechanotransduction.

microbiology↗

bioRxiv2025

Invasion preferences suggest a possible role for Plasmodium falciparum parasites in the expansion of Duffy negativity in West and Central Africa

Duffy antigen receptor for chemokines (DARC) is the primary red blood cell (RBC) receptor for invasion of human RBCs by Plasmodium vivax and knowlesi parasites. By contrast, Plasmodium falciparum parasites use multiple RBC receptors for invasion. Whether DARC is one of these receptors has never been systematically explored. We used flow cytometry and microscopy-based approaches to investigate whether P. falciparum parasites preferentially invade specific Duffy RBC phenotypes and explored two potential explanations for invasion preference - differences in RBC biophysical properties and surface protein composition. P. falciparum parasites showed a consistent preference for Duffy-positive RBCs, and some biophysical properties and surface protein expression varied between Duffy-positive and Duffy-negative RBCs. We then used our in vitro invasion data to parametrise an evolutionary- epidemiological model of the relationship between P. falciparum and the FYBES allele. Our model accounts for immunity against P. falciparum virulence, gained through exposure, and thus mutations that impede infection are not always advantageous. The inhibition of P. falciparum invasion that we observed in vitro leads to FYBES frequencies increasing at low levels of P. falciparum transmission but decreasing at high levels of transmission. The impact of P. falciparum on the prevalence of Duffy negativity may therefore be most apparent in lower transmission settings. Our findings are the first to show a link between Duffy negativity and P. falciparum and suggest that DARC may direct or indirectly be involved in P. falciparum invasion of human RBCs which could, together with P. vivax, explain the distribution of Duffy negativity in sub-Saharan Africa.

evolutionary biology↗

bioRxiv2024

Optical tweezers reveal that PfEBA and PfRH ligands, not PfMSP1, play a central role in Plasmodium-falciparum merozoite-erythrocyte attachment

Malaria pathogenesis and parasite multiplication both depend on the ability of Plasmodium falciparum merozoites to invade human erythrocytes. Invasion is a complex multi-step process that is known to involve multiple P. falciparum proteins but dissecting the precise role of individual proteins has to date been limited by the availability of quantifiable phenotypic assays. In this study, we apply a new approach to assigning function to invasion proteins by using optical tweezers to directly manipulate recently egressed merozoites and erythrocytes and quantify the strength of attachment between them, as well as the frequency with which such attachments occur. Using a range of inhibitors, antibodies, and genetically modified P. falciparum strains, we quantitated the contribution of individual P. falciparum proteins to these merozoite-erythrocyte attachment phenotypes for the first time. Most of the interactions investigated did not affect the force needed to pull merozoites and erythrocytes apart, including loss of the major P. falciparum merozoite surface protein PfMSP1 and PfGAP45, part of the glideosome actinomyosin motor complex. The only factors that significantly reduced the strength of merozoite-erythrocyte attachment were ones that disrupted the function of members of the EBA-175 like Antigen (PfEBA) family and Reticulocyte Binding Protein Homologue (PfRH) invasion ligand families. While these assays also reinforced the known redundancy within these families, with the deletion of some ligands not impacting detachment force, it appears that the PfEBA/PfRH families play a central role in merozoite attachment, not the major merozoite surface protein PfMSP1. Author summaryMalaria is a devastating disease caused by a parasitic infection. The deadliest species is Plasmodium falciparum, which causes more than 600,000 deaths annually. The parasites life cycle is complex, but all the symptoms of malaria are caused when the parasites replicate in human red blood cells. Replication depends on the invasion of the red blood cells by the parasites which is a complex process involving multiple molecular interactions and multiple steps. Invasion begins with the attachment of the parasite to the red blood cell, making this step of particular interest in the development of new therapeutics. We assessed which interactions are key to the strength of attachment using an optical tweezer assay, which allowed us to directly measure the binding force between individual parasites and red blood cells whilst using a range of molecular and genetic tools that target specific interactions known to have a role in invasion. This showed that loss of a protein commonly thought to be critical to the early stages of binding (PfMSP1) had no effect on attachment strength, whereas disruptions of several members from two families of proteins (the Erythrocyte Binding Like protein family and the reticulocyte binding-like protein family) affect attachment strength.

molecular biology↗

bioRxiv2024

Broadly inhibitory antibodies against severe malaria virulence proteins

Plasmodium falciparum pathology is driven by the accumulation of parasite-infected erythrocytes in microvessels. This process is mediated by the parasites polymorphic erythrocyte membrane protein 1 (PfEMP1) adhesion proteins. A subset of PfEMP1 variants that bind human endothelial protein C receptor (EPCR) through their CIDR1 domains is responsible for severe malaria pathogenesis. A longstanding question is whether individual antibodies can recognize the large repertoire of circulating PfEMP1 variants. Here, we describe two broadly reactive and binding-inhibitory human monoclonal antibodies against CIDR1. The antibodies isolated from two different individuals exhibited a similar and consistent EPCR-binding inhibition of 34 CIDR1 domains, representing five of the six subclasses of CIDR1. Both antibodies inhibited EPCR binding of both recombinant full-length and native PfEMP1 proteins as well as parasite sequestration in bioengineered 3D brain microvessels under physiologically relevant flow conditions. Structural analyses of the two antibodies in complex with two different CIDR1 antigen variants reveal similar binding mechanisms that depend on interactions with three highly conserved amino acid residues of the EPCR-binding site in CIDR1. These broadly reactive antibodies likely represent a common mechanism of acquired immunity to severe malaria and offer novel insights for the design of a vaccine or treatment targeting severe malaria.

immunology↗

bioRxiv2022

The structure of a Plasmodium vivax Tryptophan Rich Antigen suggests a lipid binding function for a pan-Plasmodium multi-gene family

Tryptophan Rich Antigens (TRAgs) are encoded by a multi-gene family in all Plasmodium species, significantly expanded in P. vivax, but their function is not currently known. We show that multiple P. vivax TRAgs are expressed on the merozoite surface and that one, PVP01_0000100 binds red blood cells with a strong preference for reticulocytes. Solving the structure of the C-terminal tryptophan rich domain that defines the TRAg family revealed a three-helical bundle that is conserved across Plasmodium and has homology with lipid-binding BAR domains involved in membrane remodelling. Biochemical assays confirmed that this domain has lipid binding activity with preference for sulfatide, a glycosphingolipid present in the outer leaflet of plasma membranes. Deletion of the putative orthologue in P. knowlesi, PKNH_1300500, impacts invasion in reticulocytes, suggesting a role for membrane remodelling during this essential process. Together, this work suggests a molecular function for the TRAg family for the first time.

microbiology↗

bioRxiv2022

The erythrocyte membrane properties of beta thalassaemia heterozygotes and their consequences for Plasmodium falciparum invasion

Malaria parasites such as Plasmodium falciparum have exerted formidable selective pressures on the human genome. Of the human genetic variants associated with malaria protection, beta thalassaemia (a haemoglobinopathy) was the earliest to be associated with malaria prevalence. However, the malaria protective properties of beta thalassaemic erythrocytes remain unclear. Here we studied the mechanics and surface protein expression of beta thalassaemia heterozygous erythrocytes, measured their susceptibility to P. falciparum invasion, and calculated the energy required for merozoites to invade them. We found invasion-relevant differences in beta thalassaemic cells versus matched controls, specifically: elevated membrane tension, reduced bending modulus, and higher levels of expression of the major invasion receptor basigin. However, these differences acted in opposition to each other with respect to their likely impact on invasion, and overall we did not observe beta thalassaemic cells to have lower P. falciparum invasion efficiency for any of the strains tested.

microbiology↗

bioRxiv2021

Centripetal nuclear shape fluctuations associate with chromatin condensation towards mitosis

The cell nucleus plays a central role in several key cellular processes, including chromosome organisation, replication and transcription. Recent work intriguingly suggests an association between nuclear mechanics and cell-cycle progression, but many aspects of this connection remain unexplored. Here, by monitoring nuclear shape fluctuations at different cell cycle stages, we uncover increasing inward fluctuations in late G2 and early mitosis, which are initially transient, but develop into instabilities that culminate into nuclear-envelope breakdown in mitosis. Perturbation experiments and correlation analysis reveal an association of these processes with chromatin condensation. We propose that the contrasting forces between an extensile stress and centripetal pulling from chromatin condensation could link mechanically chromosome condensation and nuclear- envelope breakdown, the two main nuclear processes during mitosis. Significance StatementThe nucleus was recently shown to exhibit shape fluctuations that vary with cell-cycle stage, but we know very little about the possible links between nuclear mechanics and cell cycle- progression. Through flickering analysis, this study monitors radius and nuclear envelope fluctuations across the cell cycle. The authors discover that as the cell cycle progresses towards mitosis, localised inward invaginations of the nuclear shape form initially transiently and gradually increasing their amplitude, in association with chromatin condensation. This phenomenon develops into nuclear envelope breakdown, suggesting a novel link between cell cycle, chromatin mechanics and nuclear shape fluctuations.

biophysics↗

bioRxiv2018

Red blood cell tension controls Plasmodium falciparum invasion and protects against severe malaria in the Dantu blood group

Malaria has had a major effect on the human genome, with many protective polymorphisms such as sickle cell trait having been selected to high frequencies in malaria endemic regions1, 2. Recently, we showed that a novel blood group variant, Dantu, provides 74% protection against all forms of severe malaria in homozygous individuals3-5. This is a similar degree of protection to sickle cell trait and considerably greater than the most advanced malaria vaccine, but until now the mechanism of protection has been unknown. In the current study, we demonstrate a significant impact of Dantu on the ability of Plasmodium falciparum merozoites to invade RBCs. The Dantu variant was associated with extensive changes to the RBC surface protein repertoire, but unexpectedly the malaria protective effect did not correlate with specific RBC-parasite receptor-ligand interactions. By following invasion using video microscopy, we found a strong link between RBC tension and parasite invasion and, even in non-Dantu RBCs, identified a tension threshold above which RBC invasion did not occur. Dantu RBCs had higher average tension, meaning that a higher proportion of Dantu RBCs could not be invaded. These findings not only provide an explanation for the protective effect of Dantu against severe malaria, but also provide fresh insights into the essential process of P. falciparum parasite invasion, and how invasion efficiency varies across the heterogenous populations of RBCs that are present both within and between individuals.

genetics↗