Hypertension Increases Susceptibility to Experimental Malaria in Mice.

Global prevalence of hypertension is on the rise, burdening healthcare, especially in developing countries where infectious diseases, such as malaria, are also rampant. Whether hypertension could predispose or increase susceptibility to malaria, however, has not been extensively explored. Previously, we reported that hypertension is associated with abnormal red blood cell (RBC) physiology and anemia. Since RBC are target host cells for malarial parasite, Plasmodium, we hypothesized that hypertensive patients with abnormal RBC physiology are at greater risk or susceptibility to Plasmodium infection. To test this hypothesis, normotensive (BPN/3J) and hypertensive (BPH/2J) mice were characterized for their RBC physiology and subsequently infected with Plasmodium yoelii (P. yoelii), a murine-specific non-lethal strain. When compared to BPN mice, BPH mice displayed microcytic anemia with RBC highly resistant to osmotic hemolysis. Further, BPH RBC exhibited greater membrane rigidity and an altered lipid composition, as evidenced by higher levels of phospholipids and saturated fatty acid, such as stearate (C18:0), along with lower levels of polyunsaturated fatty acid like arachidonate (C20:4). Moreover, BPH mice had significantly greater circulating Ter119+ CD71+ reticulocytes, or immature RBC, prone to P. yoelii infection. Upon infection with P. yoelii, BPH mice experienced significant body weight loss accompanied by sustained parasitemia, indices of anemia, and substantial increase in systemic pro-inflammatory mediators, compared to BPN mice, indicating that BPH mice were incompetent to clear P. yoelii infection. Collectively, these data demonstrate that aberrant RBC physiology observed in hypertensive BPH mice contributes to an increased susceptibility to P. yoelii infection and malaria-associated pathology.

Extracellular vesicles derived from plasmodium-infected red blood cells alleviate cerebral malaria in plasmodium berghei ANKA-infected C57BL/6J mice.

RTS,S is the first malaria vaccine recommended for implementation among young children at risk. However, vaccine efficacy is modest and short-lived. To mitigate the risk of cerebral malaria (CM) among children under the age of 5, it is imperative to develop new vaccines. EVs are potential vaccine candidates as they obtain the ability of brain-targeted delivery and transfer plasmodium antigens and immunomodulators during infections. This study extracted EVs from BALB/c mice infected with Plasmodium yoelii 17XNL (P.y17XNL). C57BL/6J mice were intravenously immunized with EVs (EV-I.V. + CM group) or subcutaneously vaccinated with the combination of EVs and CpG ODN-1826 (EV + CPG ODN-S.C. + CM group) on days 0 and 20, followed by infection with Plasmodium berghei ANKA (P.bANKA) on day 20 post-second immunization. We monitored Parasitemia and survival rate. The integrity of the Blood-brain barrier (BBB) was examined using Evans blue staining.The levels of cytokines and adhesion molecules were evaluated using Luminex, RT-qPCR, and WB. Brain pathology was evaluated by hematoxylin and eosin and immunohistochemical staining. The serum levels of IgG, IgG1, and IgG2a were analyzed by enzyme-linked immunosorbent assay. Compared with those in the P.bANKA-infected group, parasitemia increased slowly, death was delayed (day 10 post-infection), and the survival rate reached 75 %-83.3 % in the EV-I.V. + ECM and EV + CPG ODN-S.C. + ECM groups. Meanwhile, compared with the EV + CPG ODN-S.C. + ECM group, although parasitemia was almost the same, the survival rate increased in the EV-I.V. + ECM group.Additionally, EVs immunization markedly downregulated inflammatory responses in the spleen and brain and ameliorated brain pathological changes, including BBB disruption and infected red blood cell (iRBC) sequestration. Furthermore, the EVs immunization group exhibited enhanced antibody responses (upregulation of IgG1 and IgG2a production) compared to the normal control group. EV immunization exerted protective effects, improving the integrity of the BBB, downregulating inflammation response of brain tissue, result in reduces the incidence of CM. The protective effects were determined by immunological pathways and brain targets elicited by EVs. Intravenous immunization exhibited better performance than subcutaneous immunization, which perhaps correlated with EVs, which can naturally cross BBB to play a better role in brain protection.

Plasmodium yoelii surface-related antigen (PySRA) modulates the host pro-inflammatory responses via binding to CD68 on macrophage membrane.

Malaria, one of the major infectious diseases in the world, is caused by the Plasmodium parasite. Plasmodium antigens could modulate the inflammatory response by binding to macrophage membrane receptors. As an export protein on the infected erythrocyte membrane, Plasmodium surface-related antigen (SRA) participates in the erythrocyte invasion and regulates the immune response of the host. This study found that the F2 segment of P. yoelii SRA activated downstream MAPK and NF-κB signaling pathways by binding to CD68 on the surface of the macrophage membrane and regulating the inflammatory response. The anti-PySRA-F2 antibody can protect mice against P. yoelii, and the pro-inflammatory responses such as IL-1ß, TNF-α, and IL-6 after infection with P. yoelii are attenuated. These findings will be helpful for understanding the involvement of the pathogenic mechanism of malaria with the exported protein SRA.

Novel ether derivatives of 11-azaartemisinins with high order antimalarial activity against multidrug-resistant Plasmodium yoelii in Swiss mice.

This study investigates cutting-edge synthetic chemistry approaches for designing and producing innovative antimalarial drugs with improved efficacy and fewer adverse effects. Novel amino (-NH2) and hydroxy (-OH) functionalized 11-azaartemisinins 9, 12, and 14 were synthesized along with their derivatives 11a, 13a-e, and 15a-b through ART and were tested for their AMA (antimalarial activity) against Plasmodium yoelii via intramuscular (i.m.) and oral routes in Swiss mice. Ether derivative 13c was the most active compound by i.m. route, it has shown 100 % protection at the dose of 12 mg/kg × 4 days and showed 100 % clearance of parasitaemia on day 4 at dose of 6 mg/kg. Amine 11a, ether derivatives 13d, 13e and ether 15a also showed promising antimalarial activity. ß-Arteether gave 100 % protection at the dose of 48 mg/kg × 4 days and 20 % protection at 24 mg/kg × 4 days dose by oral route, while it showed 100 % protection at 6 mg/kg × 4 days and no protection at 3 mg/kg × 4 days by i.m. route.

Mast cell-derived IL-10 protects intestinal barrier integrity during malaria in mice and regulates parasite transmission to Anopheles stephensi with a female-biased immune response.

Malaria is strongly predisposed to bacteremia, which is associated with increased gastrointestinal permeability and a poor clinical prognosis. We previously identified mast cells (MCs) as mediators of intestinal permeability in malaria and described multiple cytokines that rise with parasitemia, including interleukin (IL)-10, which could protect the host from an inflammatory response and alter parasite transmission to Anopheles mosquitoes. Here, we used the Cre-loxP system and non-lethal Plasmodium yoelii yoelii 17XNL to study the roles of MC-derived IL-10 in malaria immunity and transmission. Our data suggest a sex-biased and local inflammatory response mediated by MC-derived IL-10, supported by early increased number and activation of MCs in females relative to males. Increased parasitemia in female MC IL-10 (-) mice was associated with increased ileal levels of chemokines and plasma myeloperoxidase (MPO). We also observed increased intestinal permeability in female and male MC IL-10 (-) mice relative to MC IL-10 (+) mice but no differences in blood bacterial 16S DNA levels. Transmission success of P. yoelii to A. stephensi was higher in female relative to male mice and from female and male MC IL-10 (-) mice relative to MC IL-10 (+) mice. These patterns were associated with increased plasma levels of pro-inflammatory cytokines in female MC IL-10 (-) mice and increased plasma levels of chemokines and markers of neutrophil activation in male MC IL-10 (-) mice. Overall, these data suggest that MC-derived IL-10 protects intestinal barrier integrity, regulates parasite transmission, and controls local and systemic host immune responses during malaria, with a female bias.

The dCas9-based genome editing in Plasmodium yoelii.

Genetic editing is a powerful tool for functional characterization of genes in various organisms. With its simplicity and specificity, the CRISPR-Cas9 technology has become a popular editing tool, which introduces site-specific DNA double-strand breaks (DSBs), and then leverages the endogenous repair pathway for DSB repair via homology-directed repair (HDR) or the more error-prone non-homologous end joining (NHEJ) pathways. However, in the Plasmodium parasites, the lack of a typical NHEJ pathway selects for DSB repair through the HDR pathway when a homologous DNA template is available. The AT-rich nature of the Plasmodium genome exacerbates this drawback by making it difficult to clone longer homologous repair DNA templates. To circumvent these challenges, we adopted the hybrid catalytically inactive Cas9 (dCas9)-microbial single-stranded annealing proteins (SSAP) editor to the Plasmodium genome. In Plasmodium yoelii, we demonstrated the use of the dCas9-SSAP, as the cleavage-free gene editor, by targeted gene deletion and gene tagging, even using shorter homologous DNA templates. This dCas9-SSAP method with a shorter DNA template, which did not require DSBs, independent of HDR and NHEJ, would be a great addition to the existing genetic toolbox and could be deployed for the functional characterization of genes in Plasmodium, contributing to improving the ability of the malaria research community in characterizing more than half of genes with unknown functions.IMPORTANCEMalaria caused by Plasmodium parasites infection remains a serious threat to human health, with an estimated 249 million malaria cases and 608,000 deaths worldwide in 2022, according to the latest report from the World Health Organization (WHO). Here, we demonstrated the use of dCas9-single-stranded annealing protein, as the cleavage-free gene editor in Plasmodium yoelii, by targeted deletion and gene tagging, even using shorter homologous DNA templates. This method with a shorter DNA template, which did not require DSBs, independent of HDR and NHEJ, showing the potential significance in greatly improving our ability to elucidate gene functions, would contribute to assisting the malaria research community in deciphering more than half of genes with unknown functions to identify new drug and vaccine targets.

Deaggregation of mutant Plasmodium yoelii de-ubiquitinase UBP1 alters MDR1 localization to confer multidrug resistance.

Mutations in a Plasmodium de-ubiquitinase UBP1 have been linked to antimalarial drug resistance. However, the UBP1-mediated drug-resistant mechanism remains unknown. Through drug selection, genetic mapping, allelic exchange, and functional characterization, here we show that simultaneous mutations of two amino acids (I1560N and P2874T) in the Plasmodium yoelii UBP1 can mediate high-level resistance to mefloquine, lumefantrine, and piperaquine. Mechanistically, the double mutations are shown to impair UBP1 cytoplasmic aggregation and de-ubiquitinating activity, leading to increased ubiquitination levels and altered protein localization, from the parasite digestive vacuole to the plasma membrane, of the P. yoelii multidrug resistance transporter 1 (MDR1). The MDR1 on the plasma membrane enhances the efflux of substrates/drugs out of the parasite cytoplasm to confer multidrug resistance, which can be reversed by inhibition of MDR1 transport. This study reveals a previously unknown drug-resistant mechanism mediated by UBP1 through altered MDR1 localization and substrate transport direction in a mouse model, providing a new malaria treatment strategy.

Novel amino- and hydroxy-functionalized 1,2,4-trioxanes and their antimalarial activity against multidrug-resistant Plasmodium yoelii nigeriensis in Swiss mice via intramuscular and oral route.

Following the economic and social state of humanity, Malaria is categorized as one of the life-threatening illness epidemics in under developed countries. For the eradication of the same, 1,2,4-trioxanes 17a1-a2, 17b1-b2, 17c1-c2 15a-c, 18 and 19 have been synthesized continuing the creation of a novel series. Additionally, these novel compounds were tested for their effectiveness against the multidrug-resistant Plasmodium yoelii nigeriensis in mice model using both oral and intramuscular (im) administration routes. The two most potent compounds of the series, 17a1 and 17a2, demonstrated 100 % protection at 48 mg/kg x 4 days via oral route, which is twice as potent as artemisinin. In this model artemisinin provided 100 % protection at a dose of 48 mg/kg × 4 days and 80 % protection at 24 mg/kg × 4 days via im route.

TLR7 enhancing follicular helper T (Tfh) cells response in C57BL/6 mice infected with Plasmodium yoelii NSM TLR7 mediated Tfh cells in P. yoelii infected mice.

Toll-like receptors (TLRs) play an important role in inducing innate and acquired immune responses against infection. However, the effect of Toll-like receptor 7 (TLR7) on follicular helper T (Tfh) cells in mice infected with Plasmodium is still not clear. The results showed that the splenic CD4+ CXCR5+ PD-1+ Tfh cells were accumulated after Plasmodium yoelii NSM infection, the content of splenic Tfh cells was correlated to parasitemia and/or the red blood cells (RBCs) counts in the blood. Moreover, the expression of TLR7 was found higher than TLR2, TLR3 and TLR4 in splenic Tfh cells of the WT mice. TLR7 agonist R848 and the lysate of red blood cells of infected mice (iRBCs) could induce the activation and differentiation of splenic Tfh cells. Knockout of TLR7 leads to a decrease in the proportion of Tfh cells, down-regulated expression of functional molecules CD40L, IFN-γ, IL-21 and IL-10 in Tfh cells; decreased the proportion of plasma cells and antibody production and reduces the expression of STAT3 and Ikzf2 in Tfh cells. Administration of R848 could inhibit parasitemia, enhance splenic Tfh cell activation and increase STAT3 and Ikzf2 expression in Tfh cells. In summary, this study shows that TLR7 could regulate the function of Tfh cells, affecting the immune response in the spleen of Plasmodium yoelii NSM-infected mice.

Plasmodium yoelii iron transporter PyDMT1 interacts with host ferritin and is required in full activity for malarial pathogenesis.

BACKGROUND: The rapid reproduction of malaria parasites requires proper iron uptake. However, the process of iron absorption by parasites is rarely studied. Divalent metal transporter (DMT1) is a critical iron transporter responsible for uptaking iron. A homolog of human DMT1 exists in the malaria parasite genome, which in Plasmodium yoelii is hereafter named PyDMT1. RESULTS: PyDMT1 knockout appears to be lethal. Surprisingly, despite dwelling in an iron-rich environment, the parasite cannot afford to lose even partial expression of PyDMT1; PyDMT1 hypomorphs were associated with severe growth defects and quick loss of pathogenicity. Iron supplementation could completely suppress the defect of the PyDMT1 hypomorph during in vitro culturing. Genetic manipulation through host ferritin (Fth1) knockout to increase intracellular iron levels enforced significant growth inhibition in vivo on the normal parasites but not the mutant. In vitro culturing with isolated ferritin knockout mouse erythrocytes completely rescued PyDMT1-hypomorph parasites. CONCLUSION: A critical iron requirement of malaria parasites at the blood stage as mediated by this newly identified iron importer PyDMT1, and the iron homeostasis in malarial parasites is finely tuned. Tipping the iron balance between the parasite and host will efficiently kill the pathogenicity of the parasite. Lastly, PyDMT1 hypomorph parasites were less sensitive to the action of artemisinin.

Heterogeneity in killing efficacy of individual effector CD8+ T cells against Plasmodium liver stages.

Vaccination strategies in mice inducing high numbers of memory CD8+ T cells specific to a single epitope are able to provide sterilizing protection against infection with Plasmodium sporozoites. We have recently found that Plasmodium-specific CD8+ T cells cluster around sporozoite-infected hepatocytes but whether such clusters are important in elimination of the parasite remains incompletely understood. Here, we used our previously generated data in which we employed intravital microscopy to longitudinally image 32 green fluorescent protein (GFP)-expressing Plasmodium yoelii parasites in livers of mice that had received activated Plasmodium-specific CD8+ T cells after sporozoite infection. We found significant heterogeneity in the dynamics of the normalized GFP signal from the parasites (termed 'vitality index' or VI) that was weakly correlated with the number of T cells near the parasite. We also found that a simple model assuming mass-action, additive killing by T cells well describes the VI dynamics for most parasites and predicts a highly variable killing efficacy by individual T cells. Given our estimated median per capita kill rate of k = 0.031/h we predict that a single T cell is typically incapable of killing a parasite within the 48 h lifespan of the liver stage in mice. Stochastic simulations of T cell clustering and killing of the liver stage also suggested that: (i) three or more T cells per infected hepatocyte are required to ensure sterilizing protection; (ii) both variability in killing efficacy of individual T cells and resistance to killing by individual parasites may contribute to the observed variability in VI decline, and (iii) the stable VI of some clustered parasites cannot be explained by measurement noise. Taken together, our analysis for the first time provides estimates of efficiency at which individual CD8+ T cells eliminate intracellular parasitic infection in vivo.

Evaluating the stability of host-reference gene expression and simultaneously quantifying parasite burden and host immune responses in murine malaria.

The efficacy of pre-erythrocytic stage malaria antigens or vaccine platforms is routinely assessed in murine models challenged with Plasmodium sporozoites. Relative liver-stage parasite burden is quantified using reverse transcription quantitative PCR (RTqPCR), which relies on constitutively expressed endogenous control reference genes. However, the stability of host-reference gene expression for RTqPCR analysis following Plasmodium challenge and immunization has not been systematically evaluated. Herein, we evaluated the stability of expression of twelve common RTqPCR reference genes in a murine model of Plasmodium yoelii sporozoite challenge and DNA-adenovirus IV 'Prime-Target' immunization. Significant changes in expression for six of twelve reference genes were shown by one-way ANOVA, when comparing gene expression levels among challenge, immunized, and naïve mice groups. These changes were attributed to parasite challenge or immunization when comparing group means using post-hoc Bonferroni corrected multiple comparison testing. Succinate dehydrogenase (SDHA) and TATA-binding protein (TBP) were identified as stable host-reference genes suitable for relative RTqPCR data normalisation, using the RefFinder package. We defined a robust threshold of 'partial-protection' with these genes and developed a strategy to simultaneously quantify matched host parasite burden and cytokine responses following immunisation or challenge. This is the first report systematically identifying reliable host reference genes for RTqPCR analysis following Plasmodium sporozoite challenge. A robust RTqPCR protocol incorporating reliable reference genes which enables simultaneous analysis of host whole-liver cytokine responses and parasite burden will significantly standardise and enhance results between international malaria vaccine efficacy studies.

Gut Bacteroides act in a microbial consortium to cause susceptibility to severe malaria.

Malaria is caused by Plasmodium species and remains a significant cause of morbidity and mortality globally. Gut bacteria can influence the severity of malaria, but the contribution of specific bacteria to the risk of severe malaria is unknown. Here, multiomics approaches demonstrate that specific species of Bacteroides are causally linked to the risk of severe malaria. Plasmodium yoelii hyperparasitemia-resistant mice gavaged with murine-isolated Bacteroides fragilis develop P. yoelii hyperparasitemia. Moreover, Bacteroides are significantly more abundant in Ugandan children with severe malarial anemia than with asymptomatic P. falciparum infection. Human isolates of Bacteroides caccae, Bacteroides uniformis, and Bacteroides ovatus were able to cause susceptibility to severe malaria in mice. While monocolonization of germ-free mice with Bacteroides alone is insufficient to cause susceptibility to hyperparasitemia, meta-analysis across multiple studies support a main role for Bacteroides in susceptibility to severe malaria. Approaches that target gut Bacteroides present an opportunity to prevent severe malaria and associated deaths.

Ultrastructural alterations due to sodium metavanadate treatment in the blood stages of Plasmodium yoelii yoelii.

Malaria is a potentially mortal disease caused by parasites of the genus Plasmodium spp. It has a wide distribution in the world and unfortunately there are several factors that make its control difficult; among which the development of pharmacological resistance to the different drugs used to treat this disease stands out, which makes it necessary to design new compounds that have an antimalarial effect. Previous studies have shown that vanadium has a broad antiparasitic spectrum and is also safe for the host, so the objective of this research was to evaluate the antimalarial potential of sodium metavanadate (SM) and to analyze the ultrastructural changes in parasites exposed. The method consisted of inoculating CD-1 male mice with Plasmodium yoelii yoelii and administering a 10 mg/kg/day dose of SM orally for 4 days. On the fifth day, whole blood samples were obtained, processed for ultrastructural analysis, and the changes in the different parasite stages were compared against the control. Our results showed that SM decreased parasitemia compared to the group that did not receive treatment and modified the ultrastructure in all parasitic stages because it damaged the membranes, causing alterations mainly in the nucleus and in the mitochondria as well as the loss of cellular organization, which could affect the integrity of these parasites and decrease its viability.

Bhlhe40 limits early IL-10 production from CD4+ T cells during Plasmodium yoelii 17X infection.

The cytokine IL-10 suppresses T-cell-mediated immunity, which is required to control infection with Plasmodium yoelii. Consequently, IL-10 can delay the time needed to resolve this infection, leading to a higher parasite burden. While the pathways that lead to IL-10 production by CD4+ T cells are well defined, much less is known about the mediators that suppress the expression of this potent anti-inflammatory cytokine. Here, we show that the transcription factor basic helix-loop-helix family member e40 (Bhlhe40) contributes to controlling parasite burden in response to P. yoelii infection in mice. Loss of Bhlhe40 expression in mice results in higher Il10 expression, higher peak parasitemia, and a delay in parasite clearance. The observed phenotype was not due to defects in T-cell activation and proliferation or the humoral response. Nor was it due to changes in regulatory T-cell numbers. However, blocking IL-10 signaling reversed the outcome in Bhlhe40-/ - mice, suggesting that excess IL-10 production limits their ability to control the infection properly. In addition to suppressing Il10 expression in CD4+ T cells, Bhlhe40 can promote Ifng expression. Indeed, IFN-γ production by CD4+ T cells isolated from the liver was significantly affected by the loss of Bhlhe40. Lastly, Bhlhe40 deletion in T cells resulted in a phenotype similar to that observed in the Bhlhe40-/ - mice, indicating that Bhlhe40 expression in T cells contributes to the ability of mice to control infection with P. yoelii.

Thymic atrophy induced by Plasmodium berghei ANKA and Plasmodium yoelii 17XL infection.

The thymus is the anatomical site where T cells undergo a complex process of differentiation, proliferation, selection, and elimination of autorreactive cells which involves molecular signals in different intrathymic environment. However, the immunological functions of the thymus can be compromised upon exposure to different infections, affecting thymocyte populations. In this work, we investigated the impact of malaria parasites on the thymus by using C57BL/6 mice infected with Plasmodium berghei ANKA and Plasmodium yoelii 17XL; these lethal infection models represent the most severe complications, cerebral malaria, and anemia respectively. Data showed a reduction in the thymic weight and cellularity involving different T cell maturation stages, mainly CD4-CD8- and CD4+CD8+ thymocytes, as well as an increased presence of apoptotic cells, leading to significant thymic cortex reduction. Thymus atrophy showed no association with elevated serum cytokines levels, although increased glucocorticoid levels did. The severity of thymic damage in both models reached the same extend although it occurs at different stages of infection, showing that thymic atrophy does not depend on parasitemia level but on the specific host-parasite interaction.

Dysfunction of CD169+ macrophages and blockage of erythrocyte maturation as a mechanism of anemia in Plasmodium yoelii infection.

Plasmodium parasites cause malaria with disease outcomes ranging from mild illness to deadly complications such as severe malarial anemia (SMA), pulmonary edema, acute renal failure, and cerebral malaria. In young children, SMA often requires blood transfusion and is a major cause of hospitalization. Malaria parasite infection leads to the destruction of infected and noninfected erythrocytes as well as dyserythropoiesis; however, the mechanism of dyserythropoiesis accompanied by splenomegaly is not completely understood. Using Plasmodium yoelii yoelii 17XNL as a model, we show that both a defect in erythroblastic island (EBI) macrophages in supporting red blood cell (RBC) maturation and the destruction of reticulocytes/RBCs by the parasites contribute to SMA and splenomegaly. After malaria parasite infection, the destruction of both infected and noninfected RBCs stimulates extramedullary erythropoiesis in mice. The continuous decline of RBCs stimulates active erythropoiesis and drives the expansion of EBIs in the spleen, contributing to splenomegaly. Phagocytosis of malaria parasites by macrophages in the bone marrow and spleen may alter their functional properties and abilities to support erythropoiesis, including reduced expression of the adherence molecule CD169 and inability to support erythroblast differentiation, particularly RBC maturation in vitro and in vivo. Therefore, macrophage dysfunction is a key mechanism contributing to SMA. Mitigating and/or alleviating the inhibition of RBC maturation may provide a treatment strategy for SMA.

Immunological characteristics of CD103+CD8+ Tc cells in the liver of C57BL/6 mouse infected with plasmodium NSM.

CD103 is an important marker of tissue-resident memory T cells (TRM) which play important roles in fighting against infection. However, the immunological characteristics of CD103+ T cells are not thoroughly elucidated in the liver of mouse infected with Plasmodium. Six- to eight-week-old C57BL/6 mice were infected with Plasmodium yoelii nigeriensis NSM. Mice were sacrificed on 12-16 days after infection and the livers were picked out. Sections of the livers were stained, and serum aspartate aminotransferase (AST) and alanine transaminase (ALT) levels were measured. Moreover, lymphocytes in the liver were isolated, and the expression of CD103 was determined by using qPCR. The percentage of CD103 on different immune cell populations was dynamically observed by using flow cytometry (FCM). In addition, the phenotype and cytokine production characteristics of CD103+CD8+ Tc cell were analyzed by using flow cytometry, respectively. Erythrocyte stage plasmodium infection could result in severe hepatic damage, a widespread inflammatory response and the decrease of CD103 expression on hepatic immune cells. Only CD8+ Tc and γδT cells expressed higher levels of CD103 in the uninfected state.CD103 expression in CD8+ Tc cells significantly decreased after infection. Compared to that of CD103- CD8+ Tc cells, CD103+ CD8+ Tc cells from the infected mice expressed lower level of CD69, higher level of CD62L, and secreted more IL-4, IL-10, IL-17, and secreted less IFN-γ. CD103+CD8+ Tc cells might mediate the hepatic immune response by secreting IL-4, IL-10, and IL-17 except IFN-γ in the mice infected with the erythrocytic phase plasmodium, which could be involved in the pathogenesis of severe liver damage resulted from the erythrocytic phase plasmodium yoelii nigeriensis NSM infection.

[The number of TIGIT+CD8+ T cells increases but their cytokine secretion decreases in the lungs of Plasmodium yoelii infected mice].

Objective To investigate the effect of T cell immunoreceptor with Ig and ITIM domains (TIGIT) on the function of CD8+ T cells in the lungs of Plasmodium infected mice. Methods The lungs of the mice infected with Plasmodium yoelii were isolated, weighed and photographed after 12 days' infection. After dissolution, lung lymphocytes were isolated, counted and stained, and then the contents of CD8+ and TIGIT+CD8+ T cells were detected by flow cytometry. The expressions of L selectin (CD62L), CD69, programmed death 1 (PD-1), CD25, and C-X3-C motif chemokine receptor 1 (CX3CR1) on TIGIT+CD8+ T cells were detected by flow cytometry. After stimulation with phorbol 12-myristate 13-acetate (PMA) and ionomycin, the ability of TIGIT+CD8+T cells to secrete interferon γ(IFN-γ), interleukin 21 (IL-21), IL-4, IL-17, and IL-10 was detected. Results The body mass of mice with Plasmodium infection was reduced. The lungs became darker, and the ratio of the lung mass to body mass was significantly increased. Compared with the normal mice, the percentages and absolute quantity of CD8+ and TIGIT+CD8+ T cells in the lungs of the infected mice were significantly increased. The percentage of TIGIT+CD8+ T cells expressing CD62L in the infected group was significantly lower, while the percentage of the CD69, PD-1, and CX3CR1 cells were significantly higher than that of TIGIT+CD8+ T cells from the normal mice. The percentages of TIGIT+CD8+ T cells secreting IL-21, IL-4, IL-17 and IL-10 cells in the infected group were significantly lower. Conclusion The lung lesions from mice with Plasmodium infection are obvious, the numbers of TIGIT+CD8+ T cells increase, and these cells express a variety of activation-related molecules, but the ability to secrete cytokines is reduced.

Why is Babesia not killed by artemisinin like Plasmodium?

Babesia spp. are intraerythrocytic apicomplexans that digest and utilize red blood cells in a similar way to intraerythrocytic Plasmodium spp., but unlike the latter, are not sensitive to artemisinin. A comparison of Babesia and Plasmodium genomes revealed that Babesia genomes, which are smaller than those of Plasmodium, lack numerous genes, and especially haem synthesis-related genes, that are found in the latter. Single-cell sequencing analysis showed that the different treatment groups of Babesia microti with expressed pentose phosphate pathway-related, DNA replication-related, antioxidation-related, glycolysis-related, and glutathione-related genes were not as sensitive to artemether as Plasmodium yoelii 17XNL. In particular, pentose phosphate pathway-related, DNA replication-related, and glutathione-related genes, which were actively expressed in P. yoelii 17XNL, were not actively expressed in B. microti. Supplying iron in vivo can promote the reproduction of B. microti. These results suggest that Babesia spp. lack a similar mechanism to that of malaria parasites through which the haem or iron in hemoglobin is utilized, and that this likely leads to their insensitivity to artemisinin.