Comparative assessment of embryotoxicity of 2,4,6-triiodophenol to mouse blastoid and pre-implantation embryo models.

Embryonic developmental effects of disinfection by-products, which are generated during drinking water treatment and widely detected in environment, have gained more and more attention nowadays, calling for construction of in vitro research models which can mimic early embryonic development to evaluate the embryotoxicity. The embryonic stem cell test offers a promising assay to predict embryotoxicity of environmental pollutions. However, it is not appropriate for the toxicological study of preimplantation embryos. Here, we used mouse extended stem cells (mEPS) to reconstruct embryo-like structures (blastoid), furtherly attempting to evaluate the reliability of this model for the prediction of possible developmental toxicity of 2,4,6-triiodophenol (TIP, 5-50 µM), a novel halogenated disinfection byproduct widely detected in water and even drinking water, to mammalian preimplantation embryo. To verify this, we treated mouse embryo derived from in vitro fertilization (IVF-embryo) as reference. The results showed that mEPS-blastoid was like natural blastocyst in morphology, cell composition, and could recapitulate key developmental events happened during mouse preimplantation stage. When blastoid and IVF-embryo models were separately exposed to TIP, their final blastocyst formation rates were not impaired, according to morphological features, meanwhile that TIP exposure caused slight cell apoptosis. Besides, TIP induced an ICM cell bias in cell fate decision, resulting in cell proportion change, which implied abnormal developmental potential. Though we could not evaluate TIP's embryotoxicity before 8-cell stage using blastoid model, its viability as a novel and high-throughput assessment platform for increasing environmental pollutants was still recognized.

Effects of virtual reality simulation on medical students' learning and motivation in human parasitology instruction: a quasi-experimental study.

BACKGROUND: Despite the proven effectiveness of simulation-based learning activities, its adoption in medical education remains limited, and the influence of simulation on student motivation, particularly subjective task values, is seldom explored. This study aimed to investigate the impact of a simulation-based learning activity on student learning and subjective task values in a medical morphology-related course of Human Parasitology. METHODS: A quasi-experimental study was conducted with 113 Chinese undergraduate medical students who participated in a Human Parasitology course during April to May 2022. Students were divided into two groups: Simulation Group (n = 55), where students used the simulation, and Lecture Group (n = 58), where students attended an online lecture. Students' learning was measured prior to the intervention, immediately after the intervention, and three weeks later to assess knowledge retention. The subjective task values questionnaire was administered before and after the interventions. Data were analyzed using one-way ANCOVA and MANOVA. RESULTS: Students in the Simulation Group exhibited significantly higher knowledge gain compared to the Lecture Group [F (1,110) = 23.69, p < 0.01]. Additionally, the Simulation Group retained knowledge significantly better than the Lecture Group [F (1,101) = 10.05, p < 0.005]. Furthermore, students in the Simulation Group experienced a significant increase in subjective task values after the intervention [F (3, 52) = 3.57, p < 0.05, ηp2 = 0.17], while students in the Lecture Group reported a significant decrease in subjective task values [F (3, 55) = 2.96, p < 0.05, ηp2 = 0.14]. CONCLUSIONS: Simulation-based learning not only leads to superior learning but also enhances students' subjective task values. These findings offer valuable insights into designing effective simulation-based learning experiences in medical education and have significant practical implications for educators and medical professionals.

Peritoneal GATA6+ macrophage drives hepatic immunopathogenesis and maintains the Treg cell niche in the liver.

The source of macrophages that contribute to human liver disease remains poorly understood. The purpose of this study is to investigate the functional mechanism of peritoneal macrophages in the development of hepatic immunopathology. By performing the natural infection with the blood fluke Schistosoma japonicum (S. japonicum) and the chemically carbon tetrachloride (CCl4 )-induced liver injured mouse model, we identified the peritoneal cavity as an essential source of hepatic macrophages. Here, we show that a large number of F4/80+ macrophages was accumulated in the peritoneal cavity during liver injury. An unknown source population of macrophages, which highly expressed GATA6 that is specific to peritoneal macrophages, was found to exist in the injured livers. Peritoneal macrophage deletion by injection with clodronate-containing liposomes led to an attenuated hepatic pathology and the inflammatory microenvironment, while adoptive transfer of macrophages into the abdominal cavity, by contrast, results in restoring liver pathology. Importantly, there are set genes of monocyte chemoattractant protein (MCP)-1, -2, and -3 that are highly related to recruit GATA6+ macrophages during S. japonicum infection, while administration of bindarit, a selective inhibitor of MCPs synthesis, dramatically decreased the hepatic expression of GATA6+ macrophages and thus attenuated hepatic pathology. Furthermore, in vivo study showed that peritoneal macrophages promote hepatic immunopathology is dependent on the accumulation of regulatory T cells (Tregs) in the liver. Altogether, these data provide the first clear evidence that GATA6+ peritoneal macrophages play critical roles in both the formation of hepatic immunopathology and the accumulation of Tregs cells.

2,4,6-triiodophenol exhibits embryotoxicity to pre-implantation mouse embryos in an in vitro exposure model.

2,4,6-triiodophenol (TIP), a novel type of halophenolic disinfection byproducts, has been widely detected in water bodies, even in drinking water. Recently, TIP has drawn increasing concerns on account of considerable developmental toxicity towards lower organisms and cytotoxicity for mammalian cells. However, it remains unknown about its toxicity on mammalian pre-implantation embryos. Here, by exposing mouse zygotes derived in vitro fertilization to TIP, which ranged from 5 to 50 µM, we found that TIP impaired the quality of pre-implantation mouse embryos in a dose-dependent manner, inducing decline of both total and trophectoderm cell numbers, enhancing caspase 3/7 activity and reactive oxygen species generation, though it did not decrease blastocyst formation efficiency. For the sake that only high qualified embryos are able to implant in endometrium and generate health body finally, we applied a previously modified in vitro culture system to assess TIP-exposed blastocysts' further developmental potency beyond pre-implantation stage. Surprisingly, although the exposed dose was only 5 µM and TIP was removed as soon as the zygotes reached blastocyst stage, these blastocysts still nearly lost their implantation and egg cylinder formation ability, exhibiting abnormal embryonic lineage differentiation pattern as well. Therefore, our study not only entirely shows TIP embryonic toxicity on mouse pre-implantation embryos, but also proposes a model to evaluate embryotoxicity from the zygote to egg cylinder stage.

Oral Exposure to 1,4-Dioxane Induces Hepatic Inflammation in Mice: The Potential Promoting Effect of the Gut Microbiome.

1,4-Dioxane is a widely used industrial solvent that has been frequently detected in aquatic environments. However, the hepatotoxicity of long-term dioxane exposure at environmentally relevant concentrations and underlying mechanisms of liver damage remain unclear. In this study, male mice were exposed to dioxane at concentrations of 0.5, 5, 50, and 500 ppm for 12 weeks, followed by histopathological examination of liver sections and multiomics investigation of the hepatic transcriptome, serum metabolome, and gut microbiome. Results showed that dioxane exposure at environmentally relevant concentrations induced hepatic inflammation and caused changes in the hepatic transcriptome and serum metabolic profiles. However, no inflammatory response was observed after in vitro exposure to all concentrations of dioxane and its in vivo metabolites. The gut microbiome was considered to be contributing to this apparently contradictory response. Increased levels of lipopolysaccharide (LPS) may be produced by some gut microbiota, such as Porphyromonadaceae and Helicobacteraceae, after in vivo 500 ppm of dioxane exposure. LPS may enter the blood circulation through an impaired intestinal wall and aggravate hepatic inflammation in mice. This study provides novel insight into the underlying mechanisms of hepatic inflammation induced by dioxane and highlights the need for concerns about environmentally relevant concentrations of dioxane exposure.

Research on the effect and mechanism of antimicrobial peptides HPRP-A1/A2 work against Toxoplasma gondii infection.

With increasing antibiotic resistance and drug safety concerns, novel therapeutics are urgently needed. Antimicrobial peptides are promising candidates that could address the spread of multidrug-resistant pathogens. HPRP-A1/A2 are known to display antimicrobial activity against gram-negative bacteria, gram-positive bacteria and some pathogenic fungi, but whether HPRP-A1/A2 work on Toxoplasma gondii (T gondii) is unknown. In this study, we found that the viability of tachyzoites that received HPRP-A1/A2 treatment was significantly decreased, and there was a reduction in the adhesion to and invasion of macrophages by tachyzoites after HPRP-A1/A2 treatment. HPRP-A1/A2 damaged the integrity of tachyzoite membranes, as characterized by membrane disorganization in and cytoplasm outflow from tachyzoites. In addition, in vivo injection with HPRP-A1/A2 resulted in a significantly decreased number of tachyzoites and an accelerated Th1/Tc1 response, and elicited pro-inflammatory cytokines in T gondii-infected mice. Furthermore, HPRP-A1/A2-treated splenocytes exhibited a significantly increased Tc1/Th1 response, and HPRP-A1/A2-stimulated macrophages inhibited the growth of carboxyfluorescein succinimidyl amino ester (CFSE)-labelled tachyzoites, which had higher TNF-α/IL-12 mRNA levels. Altogether, these results imply that HPRP-A1/A2 are effective against T gondii through damaging the structure of tachyzoites and inducing a protective immune response, which could offer an alternative approach against T gondii infection.

A comparative evaluation of the immunotoxicity and immunomodulatory effects on macrophages exposed to aromatic trihalogenated DBPs.

Objective: 2,4,6-trichlorophenol (TCP), 2,4,6-tribromophenol (TBP), and 2,4,6-triiodophenol (TIP) are three aromatic halogenated disinfection byproducts (DBPs) identified in chlorinated saline effluents. This study aimed to evaluate and compare their immunotoxicity and immunomodulatory effects on macrophages. Materials and methods: CCK-8 assay was used to evaluate cytotoxicity of TCP, TBP, and TIP in mouse macrophage RAW264.7 cells. A light microscope and digital camera were used to record the morphological changes of RAW264.7 cells. qRT-PCR was used to measure the mRNA levels of polarization markers and secreted cytokines. Cytokine production was also detected by ELISA. Flow cytometry was performed to analyze the expression of M1 and M2 markers on macrophages. Results: TCP, TBP, and TIP had different cytotoxic effects on macrophages. The rank order of cytotoxicity was TIP > TBP > TCP. Furthermore, the three halogenated DBPs displayed different preferences for macrophage polarization. Intriguingly, 200 µM TIP remarkably induced the M2-dominant polarization of macrophages, while 200 µM TCP induced an M1-dominant polarization of macrophages. TBP has a moderate ability in inducing M1 and M2 polarization compared with TCP and TIP. Conclusions: TIP displayed higher cytotoxicity against macrophages than TBP and TCP, its brominated and chlorinated analogs. Since M1 and M2 macrophages facilitate the inflammatory and anti-inflammatory responses, respectively, the discrepancy of TCP, TBP, and TIP in inducing macrophage polarization may lead to distinct immunomodulatory and toxicological outcomes, thus giving rise to different types of diseases. This finding may provide novel insights into evaluating the toxicity of environmental pollutants on the immune system.

Evaluation of factors influencing the guide to read biomedical English literature course for Chinese new medical postgraduates-a multiple regression analysis.

BACKGROUND: There is a dearth of published literature that demonstrates the impact of a Guide to Reading Biomedical English Literature course on new Chinese medical postgraduates. Keeping this gap in mind, the objectives of this study were to assess the factors associated with course effectiveness using the teacher, postgraduate and organizational factors. METHODS: This study was conducted at Nanjing Medical University from December 2014 to December 2015. The participants were 440 new graduate students from different medical specialties. At baseline, each student was assessed for teacher factors, individual factors and organizational factors using a self-administered structured scored anonymous questionnaire. After that, Pearson chi-square analysis was conducted to evaluate the factors that impact teacher factors (knowledge level, teaching style, individualized teaching, logical teaching, heuristic teaching, literature difficulty, bilingual teaching), individual factors (gender, attitude toward studying, previewing literature, English literacy level) and course management (such as teaching objectives and assessment system) on this course. Furthermore, multiple logistic regression analyses were performed to determine the impact of the above factors on our outcome variables (knowledge level, teaching style, individualized teaching, heuristic teaching, study attitude, previewing literature, management). RESULTS: Nearly all of the participants (420 of 440, 95.5%) thought this course was helpful for learning to read scientific literature and understanding scientific research design. Multivariate logistic regression analyses showed that the participants perception of the course as effective was associated with teachers' high knowledge level (Adjusted Odds Ratio, AOR = 49.673; 95% confidence interval, 95% CI = 4.28, 575.90). In addition, heuristic teaching was found to be significantly associated with a positive teaching effect of teaching (AOR = 12.76; 95% CI = 1.78, 91.64). Furthermore, the participants perception of the course as effective was associated with positive attitude toward studying (AOR = 25.004; 95% CI = 2.51, 249.09). Previewing literature was also associated with course effectiveness (AOR = 0.02; 95% CI = 0.04, 0.11). CONCLUSIONS: This study indicated that the course effectiveness of the Guide for Reading Biomedical English Literature was associated with i) teachers' knowledge, ii) heuristic teaching, iii) students' positive attitude, and iv) students' previewing literature.

Praziquantel Targets M1 Macrophages and Ameliorates Splenomegaly in Chronic Schistosomiasis.

Splenomegaly is a common feature of many infectious diseases, including schistosomiasis japonica. However, the immunopathogenesis and the treatment of splenomegaly due to schistosomiasis have been largely neglected. Praziquantel (PZQ), a classical schistosomicide, has been demonstrated by us and others to have antifibrotic and anti-inflammatory activities against schistosomiasis. In this study, we investigated the effect of PZQ on alleviating the splenomegaly caused by Schistosoma japonicum infection in mice. The results showed that the number of macrophages, especially the number of M1 macrophages, was significantly increased in the enlarged spleens of infected mice (P < 0.001). After PZQ treatment for 4 weeks, the number of splenic macrophages, especially the number of M1 macrophages, was significantly reduced (P < 0.001) by the way of apoptosis, and another schistosomicide, mefloquine, had no effect either on the splenomegaly or on reducing the number of macrophages. Furthermore, by using the murine macrophage line RAW 264.7, we found that PZQ could inhibit the formation of the NLRP3 inflammasome and attenuate phagocytic activity in M1 macrophages. Thus, our studies suggest that PZQ plays a powerful role in ameliorating the splenomegaly caused by S. japonicum infection, which presents a new strategy for the therapy of splenomegaly resulting from other pathological conditions.

MHC II-, but not MHC II+, hepatic Stellate cells contribute to liver fibrosis of mice in infection with Schistosoma japonicum.

Hepatic stellate cells (HSCs) are considered as the main effector cells in vitamin A metabolism and liver fibrosis, as well as in hepatic immune regulation. Recently, researches have revealed that HSCs have plasticity and heterogeneity, which depend on their lobular location and whether liver is normal or injured. This research aimed to explore the biological characteristics and heterogeneity of HSCs in mice with Schistosoma japonicum (S. japonicum) infection, and determine the subpopulation of HSCs in pathogenesis of hepatic fibrosis caused by S. japonicum infection. Results revealed that HSCs significantly increased the expressions of MHC II and fibrogenic genes after S. japonicum infection, and could be classified into MHC II+ HSCs and MHC II- HSCs subsets. Both two HSCs populations suppressed the proliferation of activated CD4+T cells, whereas only MHC II- HSCs displayed a myofibroblast-like phenotype. In response to IFN-γ, HSCs up-regulated the expressions of MHC II and CIITA, while down-regulated the expression of fibrogenic gene Col1. In addition, praziquantel treatment decreased the expressions of fibrogenic genes in MHC II- HSCs. These results confirmed that HSCs from S. japonicum-infected mice have heterogeneity. The MHC II- α-SMA+ HSCs were major subsets of HSCs contributing to liver fibrosis and could be considered as a potential target of praziquantel anti-fibrosis treatment.

Vascular endothelial growth factor promotes the activation of hepatic stellate cells in chronic schistosomiasis.

The activation of hepatic stellate cells (HSCs) is a key event in fibrotic pathogenesis. However, the mechanism involving activation of HSCs in chronic schistosomiasis is not entirely clear. Human HSC LX-2 and human umbilical vein endothelial cells (ECs) were cultured with Schistosoma japonicum antigens (SA) in vitro. Fibrosis-associated genes and cell proliferation were analyzed. HSCs were isolated from mice of chronic schistosomiasis with or without praziquantel (PZQ) treatment, followed by the microarray analysis for the liver fibrosis-associated pathways. Although SA inhibited the activation and proliferation of HSCs, it induced the EC proliferation and vascular endothelial growth factor-a (VEGF) production. VEGF significantly increased the proliferation of HSCs and upregulated the expression of collagen and α-smooth muscle actin. For in vivo study, we found that several fibrosis-associated pathways were involved in the HSCs during the reversal of liver fibrosis caused by schistosomiasis, including VEGF, platelet-derived growth factor, tumor necrosis factor and endothelin-1 pathways. The Ingenuity Pathway Analysis showed that VEGF directly regulated several pro-fibrotic and immune cytokine genes in HSCs, including integrin, fibronectin, interferon-γ, interleukin (IL)-6 and IL-10. Our data indicated the critical role of VEGF signaling in HSC activation in chronic schistosomiasis and highlighted several promising genes and pathways in HSCs as potential targets for therapeutic treatment of liver fibrosis.

Activation of the hypothalamic-pituitary-adrenal (HPA) axis contributes to the immunosuppression of mice infected with Angiostrongylus cantonensis.

BACKGROUND: Immunosuppression has been described as a consequence of brain injury and infection by different mechanisms. Angiostrongylus cantonensis can cause injury to the central nervous system and eosinophilic meningitis to human. Both T cell and B cell immunity play an essential role in the resistance of the infection. However, whether brain injury caused by A. cantonensis infection can lead to immunosuppression is not clear. Therefore, the present study sought to observe the alteration of immune responses in mice infected with A. cantonensis. METHODS: Mice were infected with 20 third-stage A. cantonensis larvae. The messenger RNA (mRNA) expression of inflammatory mediators in brain tissues was observed by qRT-PCR. Cell surface markers including CD3, CD4, CD8, CD19, B220, 7-AAD, annexin-V, IgM, AA4.1, and CD23 were evaluated by using flow cytometry. The immune functions of T and B lymphocytes were detected upon stimulation by ConA and antibody responses to a nonself antigen OVA, respectively. Activation of the hypothalamic-pituitary-adrenal axis was evaluated by analyzing the concentration of plasma corticosterone and levels of mRNA for corticotropin-releasing hormone, tyrosine hydroxylase, and c-fos. RESULTS: A. cantonensis infection results in obvious immunosuppression evidenced as progressive spleen and thymus atrophy and significant decrease in the number of lymphocyte subsets including B cells, CD3+ T cells, CD4+ T cells, and CD8+ T cells, as well as reduced T cell proliferation at 21 days post-infection and antibody reaction to exogenous protein after infection. However, the sharp decrease of splenic and thymic cells was not due to cell apoptosis but to B cell genesis cessation and impairing thymocyte development. In addition, helminthicide treatment with albendazole on infected mice at 7 days post-infection could prevent immunosuppressive symptoms. Importantly, infected mice displayed hypothalamic-pituitary-adrenal axis activation, with peak responses occurring at 16 days post-infection, and glucocorticoid receptor antagonist could partially restore the infection-induced cessation of B cell genesis. CONCLUSIONS: Brain injury caused by A. cantonensis infection, like that of brain stroke and trauma, enhanced endogenous corticosteroid activity, resulting in peripheral immunosuppression.

[Interferon-γ Induced by Toxoplasma gondii Excreted/Secreted Antigens Promotes Apoptosis of CD4+CD25+ Regulatory T Cells].

Objective: To investigate the effect of excreted/secreted antigens (ESAs) from Toxoplasma gondii RH strain and TgCtwh3 strain on apoptosis of CD4+CD25+ regulatory T cells and interferon-γ (IFN-γ) secretion. Methods: ESAs of Toxoplasma gondii RH strain and TgCtwh3 strain were prepared. Splenic mononuclear cells were isolated from C57BL/6 mice and randomly divided into RH ESA group（2×106 cells/well with addition of 10 µg/ml RH ESA）, TgCtwh3 ESA group （2×106 cells/well with addition of 10 µg/ml TgCtwh3 ESA） and control group（2×106 cells/well with addition of 10 µg/ml ovalbumin）. Flow cytometry was performed to examine the early apoptosis of CD4+CD25+ regulatory T cells after treatment for 48 h and 72 h. ELISA was conducted to determine the level of IFN-γ in the supernatant after treatment for 72 h. In another experiment, the 3 groups of splenic mononuclear cells were added with 10 µg/ml anti-IFN-γ antibody for 72 h and flow cytometry was performed to examine the early apoptosis of CD4+CD25+ regulatory T cells. Meanwhile, splenic mononuclear cells from IFN-γ knockout and wild-type C57BL/6 mice were also divided into the above-described 3 groups, and flow cytometry was performed to examine the early apoptosis of CD4+CD25+ regulatory T cells after treatment for 72 h. Results: The concentrations of RH ESA and TgCtwh3 ESA were 0.54 mg/ml and 2.14 mg/ml, respectively. Flow cytometry showed that the early apoptosis rate of CD4+CD25+ regulatory T cells in the RH ESA group and the TgCtwh3 ESA group after 48 h treatment was （12.90±1.26）% and （9.71±1.04）%, respectively （P＜0.05）, both significantly higher than that in control group （4.48±0.48）% （P<0.01） . The early apoptosis rate of CD4+CD25+ regulatory T cells after 72 h in the RH ESA group was（15.21±1.11）%, significantly higher than that in the TgCtwh3 ESA group[（11.02±0.92）%] （P<0.05） and the control group[（10.10±1.49）%]（P<0.01）. ELISA showed that the level of interferon-γ in the RH ESA group and the TgCtwh3 ESA group after 72 h was（4 764.0±118.7） pg/ml and （3 629.0±33.6） pg/ml, respectively （P＜0.01）, both significantly higher than that in the control［（679.4±30.6） pg/ml］（P<0.01）. Flow cytometry revealed lower early apoptosis rate of CD4+CD25+ regulatory T cells in the RH ESA group added with anti-IFN-γ antibody［（10.44±1.44）%ï¼½ compared with that without the addition of the antibody［（14.96±0.83）］（P<0.05）. But this difference was not observed for the TgCtwh3 ESA group. Moreover, the RH ESA-induced apoptosis rate of regulatory T cells from IFN-γ knockout mice［（10.64±0.55）%ï¼½ was significantly lower than that from the wild-type mice ［（15.21±1.11）%］（P<0.01）. But this difference was not found for the TgCtwh3 ESA treatment. Conclusion: T. gondii RH ESA induces apoptosis of CD4+CD25+ regulatory T cells and IFN-γ secretion, and these effects are stronger than those of TgCtwh3 ESA. The T. gondii ESA-induced IFN-γ stimulates generation of anti-Toxoplasma immunity during acute Toxoplasma infection through mediation of regulatory T cell apoptosis.

Development of a magnetic solid-phase extraction coupled with gas chromatography and mass spectrometry method for the analysis of semivolatile organic compounds.

Semivolatile organic compounds are a category of organic micropollutants including phthalate esters, polycyclic aromatic hydrocarbons and so on, which are commonly analyzed by solid-phase extraction followed by gas chromatography with mass spectrometry. In this work, a highly sensitive and feasible method of magnetic solid-phase extraction combined with gas chromatography with mass spectrometry was established for the determination of semivolatile organic compounds in water. The novel method was based on a permanent magnetic resin with uniform particle size and high surface area (1154.3 m2 /g). The results demonstrated that the extraction efficiency of the resin was superior to that of a C18 cartridge. The method was proved to be of satisfactory recoveries (75-115.7%) and limits of detection and quantification (0.063-6.524 and 0.212-21.745 µg/L, respectively). The method was applied to the analysis of semivolatile organic compounds in the midstream Huai River. It was observed that polychlorinated biphenyls exceeded current water standards. To further illustrate the potential effects on human health, health risk assessment was conducted based on the obtained data. The existence of health risk was proved, with hexachlorobenzene and 2,2',4,4'-tetrachlorobiphenyl as the major causes. The method possesses the characteristics of high efficiency and rapid analysis, offering a good prospect of applications in large quantities of practical water.

[Research Advances on Toxoplasma gondii Proteomics].

The proteomic techniques have been widely used in Toxoplasma gondii research since the past decade, providing proteomic data that facilitate understanding of T. gondii activities. Currently, the global proteomic studies of T. gondii are mainly confined to the tachyzoite and the oocyst stages. The subproteomic research involves some important antigens, such as the soluble tachyzoite antigen, glycoproteins, and immunoproteins, etc. The differential proteomics of T. gondii is mainly focused on identifying differentially-expressed proteins in different T. gondii strains. This review summarizes the current status of proteomic research on T. gondii, with specific focuses on global proteomic, subproteomic, and differential proteomic findings. In addition, this review gives an overview on web-based resources that provide proteomic data and support for studies on T. gondii, and finally discusses future prospects of T. gondii proteomics applications.

The quantitative and functional changes of NK cells in mice infected with Angiostrongylus cantonensis.

Angiostrongylus cantonensis is a neurotropic parasite which can cause injury to central nervous system and eosinophilic meningitis to human. Natural killer (NK) cells are specialized innate lymphocytes important in early defense against pathogens as in a variety of intracellular bacterial, viral, and protozoan infections. However, the number and function of NK cells in extracellular parasitic infection of A. cantonensis are unclear. In this study, on A. cantonensis infected mice which may mimic the human's infection, we found that the percentage of splenic NK cells and the absolute number of peripheral blood NK cells were decreased at 21-day post infection compared with that of controls. When administrating with albendazole treatment at early stage of the infection, the changes of NK cells could be avoided. Further analysis confirmed that the reduction of NK cells was due to their apoptosis manifested as increased expressions of annexin V and activated caspase-3 after 16-day post infection. Moreover, both activated and inhibitory receptors such as CD16, CD69, NKG2D, and Ly49a on NK cells were down-regulated after 16-day post infection. Interestingly, NK cells isolated from mice of 21-day post infection showed enhanced IFN-γ production when stimulated with IL-12 for 24 h and cytotoxicity to YAC-1 cells, as well as elevated CD107a expression. It is evident that NK cell population and its function were changed in A. cantonensis infected mice, suggesting their involvement in pathogenesis of the infection.

Identify organic contaminants of high-concern based on non-targeted toxicity testing and non-targeted LC-HRMS analysis in tap water and source water along the Yangtze River.

Many organic pollutants were detected in tap water (TW) and source water (SW) along the Yangtze River. However, the potential toxic effects and the high-concern organics (HCOs) which drive the effect are still unknown. Here, a non-targeted toxicity testing method based on the concentration-dependent transcriptome and non-targeted LC-HRMS analysis combining tiered filtering were used to reveal the overall biological effects and chemical information. Subsequently, we developed a qualitative pathway-structure relationship (QPSR) model to effectively match the biological and chemical information and successfully identified HCOs in TW and SW along the Yangtze River by potential substructures of HCOs. Non-targeted toxicity testing found that the biological potency of both TW and SW was stronger in the downstream of the Yangtze River, and disruption of the endocrine system and cancer were the main drivers of the effect. In addition, non-targeted LC-HRMS analysis combined with retention time prediction results identified 3220 and 631 high-confidence compound structures in positive and negative ion modes, respectively. Then, QPSR model was further implied and identified a total of 103 HCOs, containing 35 industrial chemicals, 30 PPCPs, 26 pesticides, and 12 hormones in TW and SW, respectively. Among them, the neuroactive and hormonal compounds oxoamide, 8-iso-16-cyclohexyl-tetranor prostaglandin E2, E Keppra, and Tocris-0788 showed the highest frequency of detection, which were identified in more than 1/3 of the samples. The strategy of combining non-targeted toxicity testing and non-targeted LC-HRMS analysis will support comprehensive biological effect assessment, identification of HCOs, and risk control of mixtures.

Toxoplasma gondii microneme protein MIC3 induces macrophage TNF-α production and Ly6C expression via TLR11/MyD88 pathway.

Toxoplasma gondii is the most successful parasite worldwide. It is of great interest to understand how T. gondii induce different immune responses in different hosts. In this study, we found that a peptide of T. gondii microneme protein MIC3 induced TNF-α production, NF-κB phosphorylation, iNOS transcription and Ly6C expression in mouse macrophage RAW264.7 cells. MyD88 inhibition, small interfering RNA against Tlr11 and CRISPR/Cas9-mediated knock-out of Tlr11 all reduced MIC3-induced TNF-α production, NF-κB phosphorylation, iNOS transcription and Ly6C expression. Additionally, we determined the location of MIC3 peptide in mouse macrophages using immunofluorescence. MIC3 could both adhere to the cell membrane of mouse macrophages and enter the cells. These results suggest that MIC3 triggered the immune responses in mouse macrophages via TLR11/MyD88/NF-κB pathway. It is known that human macrophages lacking TLR11. We predicted that the immune responses induced by MIC3 in human macrophages were significantly different from those in mouse macrophages. As expected, MIC3 peptide failed to induce TNF-α expression, iNOS expression and NF-κB phosphorylation in human THP-1 derived macrophages. MIC3 induced macrophage immune responses via TLR11. Intriguingly, the amino acid sequence of MIC3 is completely different from the well-known TLR11 ligand profilin, which generates a potent IL-12p40, TNF-α and IL-6 response. In marked contrast to profilin, MIC3 could not induce IL-12p40 expression in both mouse RAW264.7 cells and human THP-1 derived macrophages. Furthermore, the simulated tertiary structure of MIC3 peptide shows poor similarity with the crystal structure of profilin, suggesting that MIC3 might be a different ligand from profilin. These findings about MIC3 and TLR11 will provide us with important insights into the pathogenesis of toxoplasmosis and coevolution during host-parasite interaction.

Immunotoxicity Evaluation of Trihalophenolic Disinfection By-Products in Mouse and Human Mononuclear Macrophage Systems: The Role of RNA Epitranscriptomic Modification in Mammalian Immunity.

BACKGROUND: 2,4,6-Trichlorophenol (TCP), 2,4,6-tribromophenol (TBP) and 2,4,6-triiodophenol (TIP) are three widely detected trihalophenolic disinfection by-products (DBPs). Previous studies have mainly focused on the carcinogenic risk and developmental toxicity of 2,4,6-trihalophenols. Very little is known about their immunotoxicity in mammals. OBJECTIVES: We investigated the effects of 2,4,6-trihalophenols on mammalian immunity using a mouse macrophage model infected with bacteria or intracellular parasites and aimed to elucidate the underlying mechanisms from an epitranscriptomic perspective. The identified mechanisms were further validated in human peripheral blood mononuclear cells (PBMCs). METHODS: The mouse macrophage cell line RAW264.7 and primary mouse peritoneal macrophages were exposed to different concentrations of TCP, TBP, and TIP. The pro-inflammatory marker Ly6C, the survival of the bacterium Escherichia coli (E. coli), and the parasite burden of Toxoplasma gondii (T. gondii) were assessed. Furthermore, the global gene expression profiling of macrophages following exposure to 2,4,6-trihalophenols was obtained through RNA-sequencing (RNA-seq). The effects of 2,4,6-trihalophenols on RNA N6-methyladenosine (m6A) methyltransferases and total RNA m6A levels were evaluated using Western blotting and dot blot, respectively. Transcriptome-wide m6A methylome was analyzed by m6A-seq. In addition, expression of m6A regulators and total RNA m6A levels in human PBMCs exposed to 2,4,6-trihalophenols were detected using quantitative reverse transcriptase polymerase chain reaction and dot blot, respectively. RESULTS: Mouse macrophages exposed to TCP, TBP, or TIP had lower expression of the pro-inflammatory marker Ly6C, with a greater difference from control observed for TIP-exposed cells. Consistently, macrophages exposed to such DBPs, especially TIP, were susceptible to infection with the bacterium E. coli and the intracellular parasite T. gondii, indicating a compromised ability of macrophages to defend against pathogens. Intriguingly, macrophages exposed to TIP had significantly greater m6A levels, which correlated with the greater expression levels of m6A methyltransferases. Macrophages exposed to each of the three 2,4,6-trihalophenols exhibited transcriptome-wide redistribution of m6A. In particular, the m6A peaks in genes associated with immune-related pathways were altered after exposure. In addition, differences in m6A were also observed in human PBMCs after exposure to 2,4,6-trihalophenols. DISCUSSION: These findings suggest that 2,4,6-trihalophenol exposure impaired the ability of macrophages to defend against pathogens. This response might be associated with notable differences in m6A after exposure. To the best of our knowledge, this study presents the first m6A landscape across the transcriptome of immune cells exposed to pollutants. However, significant challenges remain in elucidating the mechanisms by which m6A mediates immune dysregulation in infected macrophages after 2,4,6-trihalophenol exposure. https://doi.org/10.1289/EHP11329.

[Research advances in actinomycete ATP-binding cassette transporters--a review].

Actinomycetes can produce numerous secondary metabolites with novel structures and unique bioactivities, which are significant for pharmaceutical industry, agriculture and environmental protection. Whole-genome sequencing data demonstrate that actinomycetes contain plenty genes coding for transporters with ATP-binding cassette (ABC), which play important roles in nutrient uptake, secondary metabolite export, xenogenous toxin detoxification, and so on. In this review, the structures and mechanisms of the ABC transporters were described. We also comprehensively discussed research advances including ours on actinomycete ABC transporters, with emphasis on ABC exporters responsible for the secretion of secondary metabolites. Finally, research hotspots and application prospect of actinomycete ABC transporters were also addressed.