Enhancing the therapeutic potential of P29 protein-targeted monoclonal antibodies in the management of alveolar echinococcosis through CDC-mediated mechanisms.

Alveolar echinococcosis (AE) is a highly lethal helminth infection. Current chemotherapeutic strategies for AE primarily involve the use of benzimidazoles (BZs) such as mebendazole (MDZ) and albendazole (ABZ), which exhibit limited efficacy. In a previous study, the vaccine of recombinant Echinococcus granulosus P29 (rEgP29) showed significant immunoprotection against E. granulosus in both mice and sheep. In the current study, we utilized hybridoma technology to generate five monoclonal antibodies (mAbs) against P29, among which 4G10F4 mAb exhibited the highest antigen-specific binding capacity. This mAb was selected for further investigation of anti-AE therapy, both in vivo and in vitro. In vitro, 4G10F4 inhibited a noteworthy inhibition of E. multilocularis protoscoleces and primary cells viability through complement-dependent cytotoxicity (CDC) mechanism. In vivo, two experiments were conducted. In the first experiment, mice were intraperitoneally injected with Em protoscoleces, and subsequently treated with 4G10F4 mAb (2.5/5/10 mg/kg) at 12 weeks postinfection once per week for 8 times via tail vein injection. Mice that were treated with 4G10F4 mAb only in dosage of 5mg/kg exhibited a significant lower mean parasite burden (0.89±0.97 g) compared to isotype mAb treated control mice (2.21±1.30 g). In the second experiment, mice were infected through hepatic portal vein and treated with 4G10F4 mAb (5mg/kg) at one week after surgery once per week for 8 times. The numbers of hepatic metacestode lesions of the 4G10F4 treatment group were significantly lower in comparison to the isotype control group. Pathological analysis revealed severe disruption of the inner structure of the metacestode in both experiments, particularly affecting the germinal and laminated layers, resulting in the transformation into infertile vesicles after treatment with 4G10F4. In addition, the safety of 4G10F4 for AE treatment was confirmed through assessment of mouse weight and evaluation of liver and kidney function. This study presents antigen-specific monoclonal antibody immunotherapy as a promising therapeutic approach against E. multilocularis induced AE.

The Acute Inflammatory Potential of Particles From the Echinococcus granulosus Laminated Layer Is Moderated by Its Calcium Inositol Hexakisphosphate Component.

Cystic echinococcosis is caused by the tissue-dwelling larva (hydatid) of Echinococcus granulosus sensu lato. A salient feature is that this larva is protected by the acellular laminated layer (LL). As the parasite grows, the LL sheds abundant particles that can accumulate in the parasite's vicinity. The potential of LL particles to induce inflammation in vivo has not been specifically analysed. It is not known how each of its two major components, namely highly glycosylated mucins and calcium inositol hexakisphosphate (InsP6) deposits, impacts inflammation induced by the LL as a whole. In this work, we show that LL particles injected intraperitoneally cause infiltration of eosinophils, neutrophils and monocytes/macrophages as well as the disappearance of resident (large peritoneal) macrophages. Strikingly, the absence of calcium InsP6 enhanced the recruitment of all the inflammatory cell types analysed. In contrast, oxidation of the mucin carbohydrates caused decreased recruitment of neutrophils. The carbohydrate-oxidised particles caused cell influx nonetheless, which may be explained by possible receptor-independent effects of LL particles on innate immune cells, as suggested by previous works from our group. In summary, LL particles can induce acute inflammatory cell recruitment partly dependent on its mucin glycans, and this recruitment is attenuated by the calcium InsP6 component.

Effects of annexin B18 from Echinococcus granulosus sensu lato on mouse macrophages.

Cystic echinococcosis (CE) is a zoonotic disease, caused by Echinococcus granulosus sensu lato (E. granulosus s. l.), which posed significant public health concern globally. E. granulosus s. l. annexin B18 (EgANXB18) acts as a secretory protein, exerting a crucial influence in mediating host-parasite interactions. Recombinant annexin B18 (rEgANXB18) was expressed by Escherichia coli and the immunoreactivity was assessed by western blotting. The binding affinity between rEgANXB18 and total protein of RAW264.7 cells was assessed by ELISA. The impact of rEgANXB18 on the metabolic activity of RAW264.7 cells was assayed by Cell Counting Kit-8 assay. The mRNA levels of polarization markers (inducible nitrous oxide synthase (iNOS) and arginase 1 (Arg1)) and key cellular factors (IL-1ß，IL-6，IL-10 and TNFα) were evaluated by qRT-PCR. rEgANXB18 was successfully expressed and recognized by E. granulosus s.l. infected canine sera, as well as could bind to the total protein of RAW264.7 cells. Additionally, rEgANXB18 could promote metabolic activity at 5, 10, 20, and 40 µg/mL while no significant impact on metabolic activity was observed at 80 µg/mL. Co-culture RAW264.7 cells with rEgANXB18 resulted in significantly upregulation of the transcript levels of polarization markers iNOS and Arg1. Moreover, rEgANXB18 significantly upregulated the transcript levels of IL-1ß, IL-6, TNFα, and IL-10, while dose-effect relationship was observed in IL-1ß, IL-6, and IL-10. Our results indicated that EgANXB18 showed the potential to regulate immune response of macrophages by shifting the cell polarization and cytokine profile, thereby promoting the parasitism of CE.

A pilot study of microRNAs expression profile in plasma of patients with hydatid disease: potential immunomodulation of hydatid disease.

Echinococcosis is a zoonotic disease, which seriously endangers human health. The immune game between parasite and host is not fully understood. Exosomes are thought to be one of the ways of information communication between parasite and host. In this study, we attempted to explore the communication between Echinococcus granulosus and its host through the medium of exosomes. We collected plasma from E. granulosus patients (CE-EXO) and healthy donors (HD-EXO) and extracted exosomes from the plasma. The expression profile of miRNA in plasma was determined by second generation sequencing. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) were used to annotate the function of target genes of differential miRNAs. Meanwhile, we co-cultured plasma exosomes from healthy donors and plasma exosomes from E. granulosus patients with Jurkat T cells with or without phytohaemagglutinin (PHA) stimulation. The expression of CD69 on Jurkat T cells was detected by flow cytometry. The results showed that the miRNA of exosomes between healthy donors and E. granulosus patients was significantly different. GO and KEGG were used to annotate the function of target genes of differential miRNAs. The results indicate that many important pathways are involved in inflammation, metabolism, and immune response after parasite infection, such as p53 signaling pathway, PI3K-Akt signaling pathway, and glycolysis/gluconeogenesis. Flow cytometry showed that CE-EXO reduced the expression of CD69 + on Jurkat T cells. Our present results suggest that these differentially expressed miRNAs may be important regulators of parasite-host interactions. Meanwhile, functional prediction of its target genes provides valuable information for understanding the mechanism of host-parasite interactions. These results provide clues for future studies on E. granulosus escape from host immune attack, which could help control E. granulosus infection.

Involvement of TIGIT in Natural Killer Cell Exhaustion and Immune Escape in Patients and Mouse Model With Liver Echinococcus multilocularis Infection.

BACKGROUND AND AIMS: Alveolar echinococcosis (AE) is a lethal helminthic liver disease caused by persistent infection with Echinococcus multilocularis. Although more attention has been paid to the immunotolerance of T cells caused by E. multilocularis infection, the role of natural killer (NK) cell, a critical player in liver immunity, is seldom studied. APPROACH AND RESULTS: Here, we observed that NK cells from the blood and closed liver tissue (CLT) of AE patients expressed a higher level of inhibitory receptor TIGIT and were functionally exhausted with a lower expression of granzyme B, perforin, interferon-gamma (IFN-γ), and TNF-α. Addition of anti-TIGIT (T-cell immunoreceptor with immunoglobulin and immunoreceptor tyrosine-based inhibitory motif domain) monoclonal antibody into AE patients' peripheral blood mononuclear cell culture significantly enhanced the synthesis of IFN-γ and TNF-α by NK cells, indicating the reversion of exhausted NK cells by TIGIT blockade. In the mouse model of E. multilocularis infection, liver and splenic TIGIT+ NK cells progressively increased dependent of infection dosage and timing and were less activated and less degranulated with lower cytokine secretion. Furthermore, TIGIT deficiency or blockade in vivo inhibited liver metacestode growth, reduced liver injury, and increased the level of IFN-γ produced by liver NK cells. Interestingly, NK cells from mice with persistent chronic infection expressed a higher level of TIGIT compared to self-healing mice. To look further into the mechanisms, more regulatory CD56bright and murine CD49a+ NK cells with higher TIGIT expression existed in livers of AE patients and mice infected with E. multilocularis, respectively. They coexpressed higher surface programmed death ligand 1 and secreted more IL-10, two strong inducers to mediate the functional exhaustion of NK cells. CONCLUSIONS: Our results indicate that inhibitory receptor TIGIT is involved in NK cell exhaustion and immune escape from E. multilocularis infection.

Human hydatid cyst fluid-induced therapeutic anti-cancer immune responses via NK1.1+ cell activation in mice.

Echinococcus granulosus is a cestode parasite which causes cystic echinococcosis disease. Previously we observed that vaccination with E. granulosus antigens from human hydatid cyst fluid (HCF) significantly inhibits colon cancer growth. In the present work, we evaluate the anti-tumor immune response induced by human HCF against LL/2 lung cancer in mice. HCF vaccination protected from tumor growth, both in prophylactic and therapeutic settings, and significantly increased mouse survival compared to control mice. Considering that tumor-associated carbohydrate antigens are expressed in E. granulosus, we oxidized terminal carbohydrates in HCF with sodium periodate. This treatment abrogates the anti-tumor activity induced by HCF vaccination. We found that HCF vaccination-induced IgG antibodies that recognize LL/2 tumor cells by flow cytometry. An antigen-specific immune response is induced with HCF vaccination in the tumor-draining lymph nodes and spleen characterized by the production of IL-5 and, in less extent, IFNÉ£. In the tumor microenvironment, we found that NK1.1 positive cells from HCF-treated mice showed higher expression of CD69 than control mice ones, indicating a higher level of activation. When we depleted these cells by administrating the NK-specific antibody NK1.1, a significantly decreased survival was observed in HCF-induced mice, suggesting that NK1.1+ cells mediate the anti-tumor protection induced by HCF. These results suggest that HCF can evoke an integrated anti-tumor immune response involving both, the innate and adaptive components, and provide novel insights into the understanding of the intricate relationship between HCF vaccination and tumor growth.

Immune Exhaustion of T Cells in Alveolar Echinococcosis Patients and Its Reversal by Blocking Checkpoint Receptor TIGIT in a Murine Model.

BACKGROUND AND AIMS: The cestode Echinococcus multilocularis infection, a serious health problem worldwide, causes alveolar echinococcosis (AE), a tumor-like disease predominantly located in the liver and able to spread to any organs. Until now, there have been few studies that explore how T-cell exhaustion contributes to the parasite's escape from immune attack and how it might be reversed. APPROACH AND RESULTS: In this study, we found that liver T-cell immunoreceptor with immunoglobulin and immunoreceptor tyrosine-based inhibitory motif domain (TIGIT) expression was significantly enhanced and positively correlated with lesion activity in AE patients. High TIGIT expression in both liver-infiltrating and blood T cells was associated with their functional exhaustion, and its ligand CD155 was highly expressed by hepatocytes surrounding the infiltrating lymphocytes. In co-culture experiments using human blood T cells and hepatic cell line HL-7702, CD155 induced functional impairment of TIGIT+ T cells, and in vitro blockade with TIGIT antibody restored the function of AE patients' T cells. Similar TIGIT-related functional exhaustion of hepatic T cells and an abundant CD155 expression on hepatocytes were observed in E. multilocularis-infected mice. Importantly, in vivo blocking TIGIT prevented T-cell exhaustion and inhibited disease progression in E. multilocularis-infected mice. Mechanistically, CD4+ T cells were totally and CD8+ T cells partially required for anti-TIGIT-induced regression of parasite growth in mice. CONCLUSIONS: This study demonstrates that E. multilocularis can induce T-cell exhaustion through inhibitory receptor TIGIT, and that blocking this checkpoint may reverse the functional impairment of T cells and represent a possible approach to immunotherapy against AE.

Expression analysis of circulating miR-146a and miR-155 as novel biomarkers related to effective immune responses in human cystic echinococcosis.

Cystic echinococcosis, an important zoonotic disease, is caused by Echinococcus granulosus. MicroRNAs are a small group of single-stranded noncoding RNAs, which play an effective role in biological processes. This study aimed at comparing the expression levels of miR-146a and miR-155 in the plasma of patients with hydatidosis and healthy individuals. A group of 20 patients with hydatid cyst formed a study group and 20 healthy individuals with no known chronic diseases formed a control group. Plasma samples were collected from hydatidosis patients as well as sex- and age-matched healthy volunteers. After that, RNA extraction and cDNA synthesis were done and the expression levels of miR-146a and miR-155 were determined by quantitative real-time polymerase chain reaction (PCR) for both groups. The results indicated that the level of miR-146a increased in all patients with hydatidosis compared to the control group. Also, the level of miR-155 increased in all hydatidosis patients, but no correlation was observed in the level of miR-155 between the two groups. The results also revealed that miR-146a and miR-155 upregulation in the plasma leads to the development of novel biomarkers for echinococcosis. One of the reasons for the increase of miRNAs in hydatidosis may be their role in modulating the immune system. These miRNAs are likely to be considered as one of the most important biomarkers in determining the severity of hydatidosis.

MicroRNA profile in immune response of alveolar and cystic echinococcosis patients.

It is known that miRNAs are effective in immune response in the diagnosis and treatment of many infectious diseases. However, the miRNAs profile is unknown in Alveolar and Cystic Echinococcosis which can be fatal if left untreated. The miRNAs profile that activates the T and B cells forming the immune system in Alveolar and Cystic Echinococcosis patients was investigated in this study. A total of 50 liver tissue samples were obtained from Alveolar and Cystic Echinococcosis patients in Kilis State Hospital Pathology Laboratory in southeast of Turkey. The circulating cell-free miRNAs were evaluated by a quantitative real-time polymerase chain reaction, statistically calculated within ΔΔCt values and fold changes were evaluated by Welch T test, in which P < .05 was considered to be significant. Twenty-five microRNAs, including let-7a-5p, let-7c, let-7e-5p, miR-15b-5p, miR16, miR-17-5p, miR-23a-5p, miR-24-3p, miR-25-3p, miR-26a-3p, miR-26b-3p, miR-29b-3p, miR-29c-3p, miR-30a-5p, miR-30b-5p, miR-30c-5p, miR-30d-5p, miR-30e-5p, miR-98-5p, miR-101-3p, miR-106b-5p, miR-125b-5p, miR-142-5p, miR-222-3p and miR-223-3p, were found as down-regulated in Alveolar and Cystic Echinococcosis patients than control groups. Twelve miRNAs, including miR-15a-5p, miR-21-5p, miR-27a-3p, miR-29a-3p, miR-146a-5p, miR-181a-5p, miR-181b-5p, miR-181d, miR-181c-5p, miR-195-5p, miR-214-3p and miR-365-3p, were found as up-regulated in Alveolar and Cystic Echinococcosis patients than healthy person. It has been shown that T- and B-cell activities are related in the progressive of both Alveolar and Cystic Echinococcosis in this study. The miRNA panel activated by T and B cells may be important for exploring the mechanisms underlying early development in Alveolar and Cystic Echinococcosis providing novel information that may be used to discover new therapeutics for these diseases.

Soluble programmed death-1 (sPD-1) as predictor of early surgical outcomes of paediatric cystic echinococcosis.

AIMS: Following treatment, cystic echinococcosis (CE) exhibits a relatively high relapse rate. Here, we evaluated the value of soluble programmed death-1 (sPD-1), sPD-1 ligand (sPD-L1) and anti-recP29 antibody concentrations, as predictors of early surgical treatment outcomes in young CE-affected patients. METHODS AND RESULTS: This prospective study included 59 Tunisian children (177 plasmas), where CE was surgically treated and monitored for 3 post-operative years. Based on CE post-surgical development, patients were clustered into a 'No relapsed' CE (NRCE; n = 39) and a 'Relapsed' CE (RCE; n = 20) group. Plasma levels of sPD-1, sPD-L1 and anti-recP29 IgG were measured using ELISA. In the NRCE group, sPD-1, sPD-L1 and anti-recP29 IgG concentrations were significantly lower at D365 than at D30. By contrast, in the RCE group, no significant difference was observed between D0, D30 and D365. When considering individual variations, the probability to be 'relapse-free' was 67% and 73% when anti-recP29 IgG and sPD-L1 level, respectively, decreased between D30 and D365. The probability to be 'relapse-free' was 86% when the sPD-1 level decreased between D30 and D365 (P = .003; chi-square test). CONCLUSION: sPD-1 may be a useful biomaker for the early evaluation of surgical procedure efficacy in paediatric CE cases.

Polyreactive antibodies as potential humoral biomarkers of host resistance to cystic echinococcosis.

Polyreactive antibodies (pAb) bind to a broad range of unrelated structures, providing hosts with functional components able to rapidly recognize and protect against different pathogens. However, their roles against helminth parasites are still unexplored. Here, pAb profiles were analysed in cystic echinococcosis (CE), a zoonosis caused by the cestode Echinococcus granulosus sensu lato. Levels of anti-DNP (2,4-dinitrophenyl-hapten) antibodies were measured as a surrogate parameter of pAb in different biological settings. Firstly, levels of serum and peritoneal pAb were measured during early experimental secondary CE, using both high (Balb/c) and low (C57Bl/6) susceptible mouse strains. Serum pAb mostly differed in normal mice, being pAb levels of IgG subclasses with poor anti-parasite activities predominant in Balb/c animals. Conversely, peritoneal pAb isotypes/subclasses with efficient anti-parasite activities predominated in normal and infected C57Bl/6 mice. Secondly, sera from potentially resistant patients, susceptible individuals and healthy donors were analysed, showing higher pAb levels of the IgA and IgG-particularly IgG1-isotypes in potentially resistant individuals compared to control groups. Finally, since remarkable differences were observed in pAb profiles according to the intrinsic host susceptibility to the infection, we proposed here that pAb might be considered as potential humoral biomarkers for host resistance to CE.

Agranulocytosis leads to intestinal Echinococcus multilocularis oncosphere invasion and hepatic metacestode development in naturally resistant Wistar rats.

Susceptibility to Echinococcus multilocularis infection considerably varies among intermediate (mostly rodents) and dead-end host species (e.g. humans and pig), in particular regarding intestinal oncosphere invasion and subsequent hepatic metacestode development. Wistar rats are highly resistant to infection and subsequent diseases upon oral inoculation with E. multilocularis eggs, however, after immunosuppressive treatment with dexamethasone, rats become susceptible. To address the role of the cellular innate immunity, Wistar rats were individually or combined depleted of natural killer (NK) cells, macrophages (MΦ) and granulocytes (polymorphonuclear cells, PMN) prior to E. multilocularis egg inoculation. Although NK cell and MΦ depletion did not alter the resistance status of rats, the majority of PMN-depleted animals developed liver metacestodes within 10 weeks, indicating that PMN are key players in preventing oncosphere migration and/or development in Wistar rats. In vitro studies indicated that resistance is not caused by neutrophil reactive oxygen species or NETosis. Also, light microscopical examinations of the small intestine showed that oral inoculation of E. multilocularis eggs does not elicit a mucosal neutrophil response, suggesting that the interaction of oncospheres and neutrophils may occur after the former have entered the peripheral blood. We suggest to consider granulocytes as mediators of resistance in more resistant species, such as humans.

Effects of Allium sativum on IFN-γ and IL4 concentrations in mice with cystic echinococcosis.

The aim of the current investigation was to assess the impacts of methanolic extract of Allium sativum (MEAS) on IL-4 (a cytokine derived from Th2 cells) and IFN-É£ (a cytokine derived from Th1 cells) levels in mice infected with Echinococcus granulosus. Sixty healthy BALB/c female mice were used in this study. Each animal was intraperitoneally injected with 1500 protoscoleces. The infected animals were randomly divided into six groups: albendazole (100 mg/kg), MEAS 10 (10 mg/kg), MEAS 20 (20 mg/kg), MEAS 40 (40 mg/kg), MEAS 80 (80 mg/kg) and control group with no treatment. The studied animals received albendazole and/or MEAS through drinking water for 30 days. Serum IFN-γ concentration significantly increased in the MEAS 20 and 80 groups in comparison to the control, albendazole and MEAS 10 groups (P < 0.05). The serum IL-4 level showed no significant difference between the trial groups. The findings of this study showed that MEAS at 20 and 80 mg/kg concentrations enhanced Th1 cell response in mice with cystic echinococcosis.

The role of regulatory B cells in Echinococcus granulosus-infected mice.

To investigate the phenotypic changes of the expression level of regulatory B cells and related molecules during the continuous infection of Echinococcus granulosus (E. granulosus) in mice and its relationship with E. granulosus infection and its immune effect. Experimental group mice were inoculated with protoscoleces suspension via intraperitoneally injection to prepare a mouse model of E. granulosus infection. Flow cytometry was used to detect the expression of regulatory B cells CD1dhiCD5+CD19hi cells and CD1dhiCD5+CD19hi IL-10+ cells in spleen and peripheral blood of mice. The expressions of IL-10 and TGF-ß1 in mouse serum were detected via ELISA. The liver pathological changes in mice were observed by H&E staining; Moreover, the expressions and distribution of IL-10 and TGF-ß1 in mice liver were measured through immunohistochemistry. The ELISA test results showed no significant changes in serum IL-10 and TGF-ß1 levels in early infected mice. However, at the middle and late stages of infection, the levels of IL-10 and TGF-ß1 in the serum of mice increased significantly (P < 0.05). The proportion of CD1dhiCD5+CD19hiBreg cells and the proportion of CD1dhiCD5+CD19hiIL-10+Breg cells in the spleen of mice infected with E. granulosus were increased at 90 days after infection, which indicating that Breg cells proliferated in the late stage of infection. CD1dhiCD5+CD19hi regulatory B cells may be one of the causes of immunosuppression of E. granulosus infection. It is speculated that Bregs inhibitory effect may play a role by regulating the expression of cytokines and inducing the secretion of inhibitory cytokines IL-10 and TGF-ß1.

Screening, construction, and serological identification of the diagnostic antigen molecule EG-06283 for the diagnosis of Echinococcus granulosus.

Several strategies exist to prevent and control echinococcosis, a global parasitic disease. However, most treatments are ineffective and adverse effects are common. Therefore, we aimed to screen protoscolex antigen molecules of Echinococcus granulosus to identify a diagnostic biomarker for hydatid disease. Published E. granulosus transcriptome sequencing data were analyzed to screen for antigen molecules that are highly expressed in protoscoleces but not in oncospheres. The membrane protein EG-06283 (annotated as Frizzled-4) was selected from 16 antigens, and its gene fragment was subjected to codon optimization and synthesis. rEG-06283 expression was induced in the pET-24a/EG-06283/BL21 strain; subsequently, the protein was purified and subcutaneously injected into ICR mice at weeks 0, 2, 4, and 6. Blood sampling occurred periodically to quantify serum immunoglobulin G (IgG) levels via enzyme-linked immunosorbent assays (ELISA). Immunogenicity was determined by western blot assays using sera from normal mice and mice with secondary hydatid infections. The antigen's immune reactivity and diagnostic value were validated using sera of patients with hydatid disease. ELISA results confirmed that the antigen molecule induced specific IgG production in mice, resulting in significantly higher levels than those in the adjuvant and control groups (P < 0.05). The western blot results indicated that the protein was recognized by antibodies in the sera of mice with hydatid infection and the antisera of immunized mice. Quantification of protein levels in the sera of patients with hydatid disease significantly differed from levels in healthy participants (P < 0.05). These results indicate that rEG-06283 is a potential diagnostic antigen for E. granulosus infections.

Echinococcus multilocularis vesicular fluid induces the expression of immune checkpoint proteins in vitro.

AIMS: Alveolar echinococcosis is a severe chronic helminthic infection that mimics a tumour-like disease. This study aimed at investigating in vitro interactions between Echinococcus multilocularis vesicular fluid (VF) and different immune checkpoints (PD-1/PD-L1, CTLA-4, LAG-3 and TIM-3). METHODS AND RESULTS: Peripheral blood mononuclear cells (PBMC) from healthy blood donors were isolated by Ficoll. Natural killer (NK) cells were selected. Each type of cell was stimulated individually with E. multilocularis-VF. Expression of the different immune checkpoints was measured by flow cytometry on day 3 and day 6; all supernatants were used for immunoassays. Cells and supernatants from 22 healthy donors were analysed. A significant increase of PD-1, PD-L1, LAG-3 and TIM-3 was observed upon E. multilocularis-VF exposure for NK cells on day 3 (P < .05, Wilcoxon signed-rank test). A significant increase of PD-L1 and CTLA-4 was observed upon E. multilocularis-VF exposure for T cells on day 6 (P < .05, Wilcoxon signed-rank test), which was associated with increased levels of Th1 and Th2 cytokines P < .05, Wilcoxon signed-rank test). CONCLUSION: These preliminary data suggest that immune checkpoints could be a way for E. multilocularis to modulate the host immune response during alveolar echinococcosis.

The correlations between Th1 and Th2 cytokines in human alveolar echinococcosis.

BACKGROUND: Alveolar echinococcosis (AE) is a zoonotic parasitic disease caused by Echinococcus multilocularis larval tapeworm infections in humans that severely impairs the health of affected patients in the northern hemisphere. METHODS: The expression levels of 20 cytokines associated with AE infection were measured by enzyme-linked immunosorbent assay, and the correlations between these cytokines were analysed in the R programming language. RESULTS: Serum cytokine levels differed among individuals in both the AE patient and healthy control groups. The results of the correlations among the cytokines showed obvious differences between the two groups. In the AE patients group, Th1 and Th2 cytokines formed a more complicated network than that in the healthy control group. CONCLUSIONS: The altered correlations between Th1 and Th2 cytokines may be closely associated with AE infection, which may provide a new explanation for the essential differences between AE patients and healthy individuals.

Myeloid-derived suppressor cells exert immunosuppressive function on the T helper 2 in mice infected with Echinococcus granulosus.

Cystic echinococcosis (CE) is a worldwide hazardous zoonotic parasitosis caused by Echinococcus granulosus. CE development involves complex immunological mechanisms, including participation of multiple immune cells and effector molecules. Myeloid-derived suppressor cells (MDSCs) are known to be involved in chronic and acute inflammatory conditions. In this study, we aimed to characterize the immune function of MDSCs in CE to improve the understanding, prevention and treatment of CE. Our results indicated that MDSCs overexpressing Ly6C and Ly6G inhibit the formation and activity of T helper 2 cells in a NO-dependent manner during E. granulosus infection.

Diagnostic performance of Echinococcus granulosus protoscolices antigens in the serodiagnosis of human cystic echinococcosis.

The development of suitable serological tests for the diagnosis of CE is still necessary. This study aimed to evaluate the efficacy of ELISA in the diagnosis of human cystic echinococcosis (CE), using parasite protoscolices antigens. Liver hydatid cysts were isolated from sheep infected with hydatid cysts and the protoscolices were isolated from the hydatid cyst fluid. Protoscolices crude antigen was prepared by mechanical disruption, plus freeze-thawing and sonication methods. Thirty sera samples of confirmed hydatid cyst patients, 30 samples of healthy individuals, and 30 samples of people with other infections were collected and the samples were evaluated in an ELISA system, using the crude protoscolices antigen. The sera samples were also simultaneously evaluated by antigen B-ELISA. The estimated value of sensitivity and specificity for the ELISA, using the crude protoscolices antigens, was 93.3% (95% CI: 76.4-98.8%) and 90% (95% CI: 78.8-95.8%), respectively. These values were 86.6 (95% CI: 68.3-95.6) and 91 (95% CI: 80.81-96.9) for the antigen-B based ELISA. Antigens prepared from protoscolices of hydatid cyst are suitable candidates for the serologic diagnosis of human CE. Further studies are needed to identify a single specific antigen among the protoscolices antigens to improve the diagnostic performance of these antigens.

Serodiagnosis of human cystic echinococcosis based on recombinant antigens B8/1 and B8/2 of Echinococcus granulosus.

BACKGROUND: Cystic echinococcosis (CE) is a widespread parasitic disease caused by the larval stage of Echinococcus granulosus. Since current methods for the diagnosis of CE are not efficient enough, rapid, and reliable tests are required for the acceleration of CE diagnosis. The present study aimed to produce recombinant B8/1 and B8/2 antigens of E. granulosus and evaluate their sensitivities and specificities separately and simultaneously for the diagnosis of CE. METHODS: The recombinant B8/1 and B8/2 antigens were produced and used in an ELISA system for the diagnosis of CE. The sera specimens including 30 sera from pathologically confirmed CE patients, 30 from other non-CE patients, and 30 from healthy controls, were evaluated by the ELISA, using AgB8/1 and AgB8/2. RESULTS: The results showed a sensitivity of 93.33%, 90%, and 96.7% for AgB8/1, AgB8/2, and their combination, respectively. The specificities were 91.7%, 93.33%, and 93.33% for AgB8/1, AgB8/2, and their combination, respectively. CONCLUSION: Simultaneous usage of AgB8/1 and AgB8/2 increased the test sensitivity for the diagnosis of CE. Furthermore, the specificity of AgB8/1 and AgB8/2 combination was more than AgB8/1 and equal to AgB8/2 alone. The findings revealed that the simultaneous usage of AgB8/1 and AgB8/2 could be a suitable approach for the diagnosis of CE.