T. gondii excretory proteins promote the osteogenic differentiation of human bone mesenchymal stem cells via the BMP/Smad signaling pathway.

BACKGROUND: Bone defects, resulting from substantial bone loss that exceeds the natural self-healing capacity, pose significant challenges to current therapeutic approaches due to various limitations. In the quest for alternative therapeutic strategies, bone tissue engineering has emerged as a promising avenue. Notably, excretory proteins from Toxoplasma gondii (TgEP), recognized for their immunogenicity and broad spectrum of biological activities secreted or excreted during the parasite's lifecycle, have been identified as potential facilitators of osteogenic differentiation in human bone marrow mesenchymal stem cells (hBMSCs). Building on our previous findings that TgEP can enhance osteogenic differentiation, this study investigated the molecular mechanisms underlying this effect and assessed its therapeutic potential in vivo. METHODS: We determined the optimum concentration of TgEP through cell cytotoxicity and cell proliferation assays. Subsequently, hBMSCs were treated with the appropriate concentration of TgEP. We assessed osteogenic protein markers, including alkaline phosphatase (ALP), Runx2, and Osx, as well as components of the BMP/Smad signaling pathway using quantitative real-time PCR (qRT-PCR), siRNA interference of hBMSCs, Western blot analysis, and other methods. Furthermore, we created a bone defect model in Sprague-Dawley (SD) male rats and filled the defect areas with the GelMa hydrogel, with or without TgEP. Microcomputed tomography (micro-CT) was employed to analyze the bone parameters of defect sites. H&E, Masson and immunohistochemical staining were used to assess the repair conditions of the defect area. RESULTS: Our results indicate that TgEP promotes the expression of key osteogenic markers, including ALP, Runx2, and Osx, as well as the activation of Smad1, BMP2, and phosphorylated Smad1/5-crucial elements of the BMP/Smad signaling pathway. Furthermore, in vivo experiments using a bone defect model in rats demonstrated that TgEP markedly promoted bone defect repair. CONCLUSION: Our results provide compelling evidence that TgEP facilitates hBMSC osteogenic differentiation through the BMP/Smad signaling pathway, highlighting its potential as a therapeutic approach for bone tissue engineering for bone defect healing.

d-galactose causes embryonic development arrest and placental development disorders in mice by increasing ROS and inhibiting SIRT1/FOXO3a axis.

INTRODUCTION: Does an elevation in d-Galactose (D-Gal) levels within the body contribute to abnormal embryonic development and placental dysfunction during pregnancy? METHODS: Mouse embryos were cultivated to the blastocyst stage under varying concentrations of D-Gal. The blastocyst formation rate was measured, and the levels of reactive oxygen species (ROS), sirtuin 1 (SIRT1), and forkhead box O3a (FOXO3a) in blastocysts were assessed. Mice were intraperitoneally injected with either saline or D-Gal with or without SRT1720. On the 14th day of pregnancy, the fetal absorption rate and placental weight were recorded. Placental levels of superoxide dismutase (SOD) and malondialdehyde (MDA) were determined. The expression of senescence-related factors, such as senescence-associated ß-galactosidase (SA-ß-gal) in the placenta was examined, and the expression of placental SIRT1, FOXO3a and p21 was evaluated by immunohistochemistry and Western blotting. RESULTS: D-Gal adversely affects early embryonic development in vitro, resulting in a decreased blastocyst formation rate. Furthermore, D-Gal downregulates SIRT1 and FOXO3a while increasing ROS levels in blastocysts. Concurrently, D-Gal induces placental dysfunction, characterized by an elevated fetal absorption rate, reduced placental weight, diminished SOD activity, and increased MDA content. The senescence-related factor SA-ß-gal was detected in the placenta, along with altered expression of placental SIRT1, FOXO3a, and p21. The SIRT1 agonist SRT1720 mitigated this damage by increasing SIRT1 and FOXO3a expression. DISCUSSION: The inhibition of early embryonic development and placental dysfunction induced by D-Gal may be attributed to the dysregulation of SIRT1. Activating SIRT1 emerges as a potentially effective strategy for alleviating the adverse effects of D-Gal exposure.

Solvent-Induced In(III)-MOFs with Controllable Interpenetration Degree Performing High-Efficiency Separation of CO2/N2 and CO2/CH4.

Herein, three In(III)-based metal-organic frameworks (In-MOFs) with different degrees of interpenetration (DOI), namely In-MOF-1, In-MOF-2, and In-MOF-3, constructed by In3+ and Y-shaped ligands 4,4',4″-s-triazine-2,4,6-triyltribenzoate (H3TATB), are successfully synthesized through the ionothermal/solvothermal method. Subsequently, three novel In-MOFs, including noninterpenetration polycatenation, 2-fold interpenetrated, and 4-fold interpenetrated structure, are employed as the platform for systematically investigating the separation efficiency of CO2/N2, CO2/CH4, and CO2/CH4/N2 mixture gas system. Among them, In-MOF-2 shows the highest CO2 uptake capacities at 298 K and simultaneously possesses the low adsorption enthalpy of CO2 (26.4 kJ/mol at low coverage), a feature desirable for low-energy-cost adsorbent regeneration. The CO2/N2 (v: v = 15/85) selectivity of In-MOF-2 reaches 37.6 (at 298 K and 1 bar), also revealing outstanding selective separation ability from flue gases and purifying natural gas, affording a unique robust separation material as it has moderate DOI and pore size. In-MOF-2 shows exceptional stability and feasibility to achieve reproducibility. Aperture adjustment makes In-MOF-2 a versatile platform for selectively capturing CO2 from flue gases or purifying natural gas.

Arterial Stiffness and Obesity as Predictors of Diabetes: Longitudinal Cohort Study.

BACKGROUND: Previous studies have confirmed the separate effect of arterial stiffness and obesity on type 2 diabetes; however, the joint effect of arterial stiffness and obesity on diabetes onset remains unclear. OBJECTIVE: This study aimed to propose the concept of arterial stiffness obesity phenotype and explore the risk stratification capacity for diabetes. METHODS: This longitudinal cohort study used baseline data of 12,298 participants from Beijing Xiaotangshan Examination Center between 2008 and 2013 and then annually followed them until incident diabetes or 2019. BMI (waist circumference) and brachial-ankle pulse wave velocity were measured to define arterial stiffness abdominal obesity phenotype. The Cox proportional hazard model was used to estimate the hazard ratio (HR) and 95% CI. RESULTS: Of the 12,298 participants, the mean baseline age was 51.2 (SD 13.6) years, and 8448 (68.7%) were male. After a median follow-up of 5.0 (IQR 2.0-8.0) years, 1240 (10.1%) participants developed diabetes. Compared with the ideal vascular function and nonobese group, the highest risk of diabetes was observed in the elevated arterial stiffness and obese group (HR 1.94, 95% CI 1.60-2.35). Those with exclusive arterial stiffness or obesity exhibited a similar risk of diabetes, and the adjusted HRs were 1.63 (95% CI 1.37-1.94) and 1.64 (95% CI 1.32-2.04), respectively. Consistent results were observed in multiple sensitivity analyses, among subgroups of age and fasting glucose level, and alternatively using arterial stiffness abdominal obesity phenotype. CONCLUSIONS: This study proposed the concept of arterial stiffness abdominal obesity phenotype, which could improve the risk stratification and management of diabetes. The clinical significance of arterial stiffness abdominal obesity phenotype needs further validation for other cardiometabolic disorders.

Diabetes, glycemic control and arterial stiffness: a real-world cohort study in the context of predictive, preventive, and personalized medicine.

Background: Arterial stiffness is a major contributor to morbidity and mortality worldwide. Although several metabolic markers associated with arterial stiffness have been developed, there is limited data regarding whether glycemic control modifies the association between diabetes and arterial stiffness. For these reasons, identification of traits around diabetes will directly contribute to arterial stiffness and atherosclerosis management in the context of predictive, preventive, and personalized medicine (PPPM). Thus, this study aimed to explore the relationship of diabetes and glycemic control status with arterial stiffness in a real-world setting. Methods: Data of participants from Beijing Xiaotangshan Examination Center (BXEC) with at least two surveys between 2008 and 2019 were used. Cumulative hazards were presented by inverse probability of treatment weighted (IPTW) Kaplan-Meier curves. Cox models were used to estimate the hazard ratio (HR) and 95% confidence interval (CI). Arterial stiffness was defined as brachial-ankle pulse wave velocity (baPWV) ≥1400 cm/s. Results: Of 5837 participants, the mean baseline age was 46.5±9.3 years, including 3791 (64.9%) males. During a median follow-up of 4.0 years, 1928 (33.0%) cases of incident arterial stiffness were observed. People with diabetes at baseline had a 48.4% (HR: 1.484, 95% CI: 1.250-1.761) excessive risk of arterial stiffness. Adherence to good glycemic control attenuated the relationship between diabetes and arterial stiffness (HR: 1.264, 95% CI: 0.950-1.681); while uncontrolled diabetes was associated with the highest risk of arterial stiffness (HR: 1.629, 95% CI: 1.323-2.005). Results were consistent using IPTW algorithm and multiple imputed data. Conclusion: Our study quantified that diabetes status is closely associated with an increased risk of arterial stiffness and supported that adherence to good glycemic control could attenuate the adverse effect of diabetes on arterial stiffness. Therefore, glucose monitoring and control is a cost-effective strategy for the predictive diagnostics, targeted prevention, patient stratification, and personalization of medical services in early vascular damages and arterial stiffness. Supplementary Information: The online version contains supplementary material available at 10.1007/s13167-023-00347-z.

Toxoplasma gondii Induces Pyroptosis in Human Placental Trophoblast and Amniotic Cells by Inducing ROS Production and Activation of Cathepsin B and NLRP1/NLRP3/NLRC4/AIM2 Inflammasome.

Toxoplasma gondii infection in pregnant women may cause fetal anomalies; however, the underlying mechanisms remain unclear. The current study investigated whether T. gondii induces pyroptosis in human placental cells and the underlying mechanisms. Human placental trophoblast (BeWo and HTR-8/SVneo) and amniotic (WISH) cells were infected with T. gondii, and then reactive oxygen species (ROS) production, cathepsin B (CatB) release, inflammasome activation, and pyroptosis induction were evaluated. The molecular mechanisms of these effects were investigated by treating the cells with ROS scavengers, a CatB inhibitor, or inflammasome-specific siRNA. T. gondii infection induced ROS generation and CatB release into the cytosol in placental cells but decreased mitochondrial membrane potential. T. gondii-infected human placental cells and villi exhibited NLRP1, NLRP3, NLRC4, and AIM2 inflammasome activation and subsequent pyroptosis induction, as evidenced by increased expression of ASC, cleaved caspase-1, and mature IL-1ß and gasdermin D cleavage. In addition to inflammasome activation and pyroptosis induction, adverse pregnancy outcome was shown in a T. gondii-infected pregnant mouse model. Administration of ROS scavengers, CatB inhibitor, or inflammasome-specific siRNA into T. gondii-infected cells reversed these effects. Collectively, these findings show that T. gondii induces NLRP1/NLRP3/NLRC4/AIM2 inflammasome-dependent caspase-1-mediated pyroptosis via induction of ROS production and CatB activation in placental cells. This mechanism may play an important role in inducing cell injury in congenital toxoplasmosis.

Toxoplasma gondii excretory/secretory proteins promotes osteogenic differentiation of bone marrow mesenchymal stem cells via aerobic glycolysis mediated by Wnt/ß­catenin signaling pathway.

Toxoplasma gondii excretory/secretory proteins (TgESPs) are a group of proteins secreted by the parasite and have an important role in the interaction between the host and Toxoplasma gondii (T. gondii). They can participate in various biological processes in different cells and regulate cellular energy metabolism. However, the effect of TgESPs on energy metabolism and osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs) has remained elusive. In the present study, TgESPs were extracted from the T. gondii RH strain and used to treat BMSCs to observe the effect of TgESPs on energy metabolism and osteogenic differentiation of BMSCs and to explore the molecular mechanisms involved. The osteogenic differentiation and energy metabolism of BMSCs were evaluated using Alizarin Red S staining, qRT-PCR, western blot, immunofluorescence and Seahorse extracellular flux assays. The results indicated that TgESPs activated the Wnt/ß­catenin signaling pathway to enhance glycolysis and lactate production in BMSCs, and promoted cell mineralization and expression of osteogenic markers. In conclusion, the present study uncovered the potential mechanism by which TgESPs regulate BMSCs, which will provide a theoretical reference for the study of the function of TgESPs in the future.

Identification and evaluation of circulating exosomal miRNAs for the diagnosis of postmenopausal osteoporosis.

BACKGROUND: Postmenopausal osteoporosis (PMOP) is a common condition that leads to a loss of bone density and an increased risk of fractures in women. Recent evidence suggests that exosomal miRNAs are involved in regulating bone development and osteogenesis. However, exosomal miRNAs as biomarkers for PMOP diagnosis have not been systematically evaluated. In this study, we aim to identify PMOP-associated circulating exosomal miRNAs and evaluate their diagnostic performance. METHODS: We performed next-generation sequencing and bioinformatics analysis of plasma exosomal miRNAs from 12 PMOP patients and 12 non-osteoporosis controls to identify PMOP-associated exosomal miRNAs, and then validated them in an independent natural community cohort with 26 PMOP patients and 21 non-osteoporosis controls. Exosomes were isolated with the size exclusion chromatography method from the plasma of elder postmenopausal women. The plasma exosomal miRNA profiles were characterized in PMOP paired with controls with next-generation sequencing. Potential plasma exosomal miRNAs were validated by qRT-PCR in the validation cohort, and their performance in diagnosing PMOP was systematically evaluated with the receiver operating characteristic curve. RESULTS: Twenty-seven miRNAs were identified as differentially expressed in PMOP versus controls in sequencing data, of which six exosomal miRNAs (miR-196-5p, miR-224-5p, miR320d, miR-34a-5p, miR-9-5p, and miR-98-5p) were confirmed to be differentially expressed in PMOP patients by qRT-PCR in the validation cohort. The three miRNAs combination (miR-34a-5p + miR-9-5p + miR-98-5p) demonstrated the best diagnostic performance, with an AUC = 0.734. In addition, the number of pregnancies was found to be an independent risk factor that can improve the performance of exosomal miRNAs in diagnosing PMOP. CONCLUSIONS: These results suggested that the plasma exosomal miRNAs had the potential to serve as noninvasive diagnostic biomarkers for PMOP.

The impact of high background particle concentration on the spatiotemporal distribution of Serratia marcescens bioaerosol.

Airborne transmission is a well-established mode of dissemination for infectious diseases, particularly in closed environments. However, previous research has often overlooked the potential impact of background particle concentration on bioaerosol characteristics. We compared the spatial and temporal distributions of bioaerosols under two levels of background particle concentration: heavily polluted (150-250 µg/m3) and excellent (0-35 µg/m3) in a typical ward. Serratia marcescens bioaerosol was adopted as a bioaerosol tracer, and the bioaerosol concentrations were quantified using six-stage Andersen cascade impactors. The results showed a significant reduction (over at least 62.9%) in bioaerosol concentration under heavily polluted levels compared to excellent levels at all sampling points. The temporal analysis also revealed that the decay rate of bioaerosols was higher (at least 0.654 min-1) under heavily polluted levels compared to excellent levels. These findings suggest that background particles can facilitate bioaerosol removal, contradicting the assumption made in previous research that background particle has no effect on bioaerosol characteristics. Furthermore, we observed differences in the size distribution of bioaerosols between the two levels of background particle concentration. The average bioaerosols size under heavily polluted levels was found to be higher than that under excellent levels, and the average particle size under heavily polluted levels gradually increased with time. In conclusion, these results highlight the importance of considering background particle concentration in future research on bioaerosol characteristics.

Effects of phthalates on human chorionic trophoblast cells and mouse embryonic development.

Phthalate exposure is associated with reproductive health, but the mechanism is unclear. This study used human chorionic trophoblast epithelial cells (HTR8/Svneo cells) and mouse embryos as objects aims to explore the effects of phthalate plasticizers on germ cells and fertility and the possible signalling pathways. In the present study, high concentrations of MEHP for 24 h significantly inhibited the proliferation and viability of HTR8/SVneo cells. Compared with the negative control (NC) group, the MEHP medium and high concentration groups promoted the apoptosis of HTR8/SVneo cells and inhibited the cell cycle, HTR8/SVneo cells were blocked in G1/G0 phase and could not enter S phase, and cell meiosis was inhibited. Western blot experiments showed that there was no difference in the protein expression of wnt inhibitory factor 1 (WIF1) and ß-catenin in HTR8/SVneo cells between the MEHP exposure groups and the NC groups. In vitro embryo culture experiments found that there was no difference in blastocyst formation rate among groups after exposure to DEHP for 2 h. Immunofluorescence showed that the expression of WIF1 decreased in the low concentration group, and there was no difference in the medium and high concentration groups, while the expression of ß-catenin was increased in both the low concentration group and the high concentration group. Our data suggest that exposure to phthalate plasticizers can affect the viability, cell cycle and apoptosis of trophoblast cells, resulting in abnormal expression of the embryonic WIF1/ß-catenin signalling pathway and impaired fertility.

A Superhydrophobic Trifluoromethyl-Containing Covalent Organic Framework Membrane for Efficient Oil/Water Separation.

Oily water caused in the process of industry leads to not only the waste of resources, but also environmental pollution. Membrane separation, as a facile and efficient separation technology, has attracted widespread attention in the field of oil/water separation. The development of membrane materials with high separation performance is one of the key elements to improve separation efficiency. In this work, a superhydrophobic membrane composited with a trifluoromethyl-containing covalent organic framework (COF) is prepared, which exhibits excellent performance on separations of oil/water mixtures and water-in-oil emulsions. For different composition of oil/water mixtures, the highest flux of oil is up to 32 000 L m-2  h-1 and oil/water separation efficiency is above 99%. Moreover, the high oil/water separation efficiency remains unchanged after successive cycles. This work provides a feasible scheme for the design of high-efficiency oil/water separation membranes.

A New perspective: How Pathogens Manipulate Phagocytosis?

The immune system is a complicated, closely regulated mechanism that evolved to keep people healthy from infectious pathogens. Phagocytosis is important for both innate and acquired immunity, which is a critical process for microbial pathogens and apoptotic cells to be consumed and eliminated. However, several pathogens have evolved different strategies to escape detection and killing by phagocytosis. Recently, with the increase in infectious diseases and antibiotic resistance, it is significant for people to have a deep understanding of immune evasion, which may become an opportunity to explore new treatments and vaccination. Additionally, researchers mostly study immune evasion of a single pathogen but rarely summarize pathogens from the perspective of immune mechanisms. Here, we present the current understanding of phagocytosis and give a brief discussion of how pathogens control phagocytosis at different stages.

Chondrogenic differentiation of mesenchymal stem cells through cartilage matrix-inspired surface coatings.

The stem cell niche comprises soluble molecules and extracellular matrix components which provide chemical and mechanical cues that determine the differentiation of stem cells. Here, the effect of polyelectrolyte multilayer (PEM) composition and terminal layer fabricated with layer-by-layer technique (LBL) pairing either hyaluronan [in its native (nHA) and oxidized form (oHA)] or chondroitin sulfate (CS) with type I collagen (Col I) is investigated on chondrogenic differentiation of human umbilical mesenchymal stem cells (hUC-MSCs). Physical studies performed to investigate the establishment and structure of the surface coatings show that PEM composed of HA and Col I show a dominance of nHA or oHA with considerably lesser organization of Col I fibrils. In contrast, distinguished fibrilized Col I is found in nCS-containing PEM. Generally, Col I-terminated PEM promote the adhesion, migration, and growth of hUC-MSCs more than GAG-terminated surfaces due to the presence of fibrillar Col I but show a lower degree of differentiation towards the chondrogenic lineage. Notably, the Col I/nHA PEM not only supports adhesion and growth of hUC-MSCs but also significantly promotes cartilage-associated gene and protein expression as found by histochemical and molecular biology studies, which is not seen on the Col I/oHA PEM. This is related to ligation of HA to the cell receptor CD44 followed by activation of ERK/Sox9 and noncanonical TGF-ß signaling-p38 pathways that depends on the molecular weight of HA as found by immune histochemical and western blotting. Hence, surface coatings on scaffolds and other implants by PEM composed of nHA and Col I may be useful for programming MSC towards cartilage regeneration.

Platelet Rich Plasma in the Repair of Articular Cartilage Injury: A Narrative Review.

OBJECTIVE: This paper reviews the research of platelet-rich plasma (PRP) in articular cartilage injury repair, to assess the mechanism, utilization, and efficacy of PRP in the treatment of articular cartilage injury, hoping to provide a theoretical basis for the clinical application of PRP in the future. MATERIALS AND METHODS: A comprehensive database search on PRP applications in cartilage repair was performed. Among them, the retrieval time range of PRP in clinical trials of repairing knee cartilage injury was from January 1, 2021 to January 1, 2022. Non-clinical trials and studies unrelated to cartilage injury were excluded. RESULT: PRP can affect inflammation, angiogenesis, cartilage protection, and cellular proliferation and differentiation after articular cartilage injury through different pathways. In all, 13 clinical trials were included in the analysis. CONCLUSION: PRP is an emergent therapeutic approach in tissue engineering. Most studies reported that PRP has a positive effect on cartilage injury, improving the joint function, meanwhile there is a lack of standardized standards. The technology of PRP in the repair and treatment of articular cartilage injury is worthy of further research.

SRT1720 plays a role in oxidative stress and the senescence of human trophoblast HTR8/SVneo cells induced by D-galactose through the SIRT1/FOXO3a/ROS signalling pathway.

D-galactose (D-gal) is a reducing sugar widely distributed in food. In a pregnant animal model exposed to D-gal, D-gal was found to have toxic effects on both the mother and foetus through oxidative stress. However, little is known about the effect of D-gal exposure on the placenta and its underlying mechanism. In this study, we evaluated the effects of D-gal on HTR8/SVneo cells and the mechanisms in vitro. In the present study, the activity of HTR8/SVneo human trophoblasts decreased in a time- and concentration-dependent manner after exposure to D-gal. D-gal resulted in premature senescence of HTR8/SVneo cells, as confirmed by assessing ß-galactosidase (SA-ß-gal) activity and the expression of senescence-related factor p21. We also verified the damage of oxidative stress induced by D-gal by measuring the expression of reactive oxygen species (ROS), sirtuin 1 (SIRT1) and forkhead box O (FOXO) 3a. SRT1720, as a SIRT1 activator, mitigated D-gal-induced oxidative stress and senescence by upregulating SIRT1 and FOXO3a expression and reducing ROS production. Our data suggest that D-gal may induce HTR8/SVneo premature ageing through the SIRT1/FOXO3a/ROS signalling pathway mediated by oxidative stress and that SIRT1 protects cells from this damage.

Silver nanoparticles induce apoptosis via NOX4-derived mitochondrial reactive oxygen species and endoplasmic reticulum stress in colorectal cancer cells.

Aim: To investigate the anticancer mechanisms of silver nanoparticles (AgNPs) in colorectal cancer. Methods: Anticancer effects of AgNPs were determined in colorectal cancer HCT116 cells and xenograft mice using cellular and molecular methods. Results: AgNPs induced mitochondrial reactive oxygen species production, mitochondrial dysfunction and endoplasmic reticulum (ER) stress responses through NOX4 and led to HCT116 cell apoptosis. Pretreatment with DPI or 4-PBA significantly inhibited mitochondrial reactive oxygen species production, apoptosis, ER stress response, NOX4 expression and mitochondrial dysfunction in AgNP-treated HCT116 cells. AgNPs also significantly suppressed HCT116 cell-based xenograft tumor growth in nude mice by inducing apoptosis and ER stress responses. Conclusion: AgNPs exert anticancer effects against colorectal cancer via ROS- and ER stress-related mitochondrial apoptosis pathways.

Expression profiles of NOD-like receptors and regulation of NLRP3 inflammasome activation in Toxoplasma gondii-infected human small intestinal epithelial cells.

BACKGROUND: Toxoplasma gondii is a parasite that primarily infects through the oral route. Nucleotide-binding oligomerization domain (NOD)-like receptors (NLRs) play crucial roles in the immune responses generated during parasitic infection and also drive the inflammatory response against invading parasites. However, little is known about the regulation of NLRs and inflammasome activation in T. gondii-infected human small intestinal epithelial (FHs 74 Int) cells. METHODS: FHs 74 Int cells infected with T. gondii were subsequently evaluated for morphological changes, cytotoxicity, expression profiles of NLRs, inflammasome components, caspase-cleaved interleukins (ILs), and the mechanisms of NLRP3 and NLRP6 inflammasome activation. Immunocytochemistry, lactate dehydrogenase assay, reverse transcription polymerase chain reaction (RT-PCR), real-time quantitative RT-PCR, and western blotting techniques were utilized for analysis. RESULTS: Under normal and T. gondii-infected conditions, members of the NLRs, inflammasome components and caspase-cleaved ILs were expressed in the FHs Int 74 cells, except for NLRC3, NLRP5, and NLRP9. Among the NLRs, mRNA expression of NOD2, NLRP3, NLRP6, and NAIP1 was significantly increased in T. gondii-infected cells, whereas that of NLRP2, NLRP7, and CIITA mRNAs decreased significantly in a time-dependent manner. In addition, T. gondii infection induced NLRP3, NLRP6 and NLRC4 inflammasome activation and production of IL-1ß, IL-18, and IL-33 in FHs 74 Int cells. T. gondii-induced NLRP3 inflammasome activation was strongly associated with the phosphorylation of p38 MAPK; however, JNK1/2 had a weak effect. NLRP6 inflammasome activation was not related to the MAPK pathway in FHs 74 Int cells. CONCLUSIONS: This study highlighted the expression profiles of NLRs and unraveled the underlying mechanisms of NLRP3 inflammasome activation in T. gondii-infected FHs 74 Int cells. These findings may contribute to understanding of the mucosal and innate immune responses induced by the NLRs and inflammasomes during T. gondii infection in FHs 74 Int cells.

Toxoplasma gondii Induces Apoptosis via Endoplasmic Reticulum Stress-Derived Mitochondrial Pathway in Human Small Intestinal Epithelial Cell-Line.

Toxoplasma gondii, an intracellular protozoan parasite that infects one-third of the world's population, has been reported to hijack host cell apoptotic machinery and promote either an anti- or proapoptotic program depending on the parasite virulence and load and the host cell type. However, little is known about the regulation of human FHs 74 small intestinal epithelial cell viability in response to T. gondii infection. Here we show that T. gondii RH strain tachyzoite infection or ESP treatment of FHs 74 Int cells induced apoptosis, mitochondrial dysfunction and ER stress in host cells. Pretreatment with 4-PBA inhibited the expression or activation of key molecules involved in ER stress. In addition, both T. gondii and ESP challenge-induced mitochondrial dysfunction and cell death were dramatically suppressed in 4-PBA pretreated cells. Our study indicates that T. gondii infection induced ER stress in FHs 74 Int cells, which induced mitochondrial dysfunction followed by apoptosis. This may constitute a potential molecular mechanism responsible for the foodborne parasitic disease caused by T. gondii.

Adenosine A3 Receptor Mediates ERK1/2- and JNK-Dependent TNF-α Production in Toxoplasma gondii-Infected HTR8/SVneo Human Extravillous Trophoblast Cells.

Toxoplasma gondii is an intracellular parasite that causes severe disease when the infection occurs during pregnancy. Adenosine is a purine nucleoside involved in numerous physiological processes; however, the role of adenosine receptors in T. gondii-induced trophoblast cell function has not been investigated until now. The goal of the present study was to evaluate the intracellular signaling pathways regulated by adenosine receptors using a HTR-8/SVneo trophoblast cell model of T. gondii infection. HTR8/SVneo human extravillous trophoblast cells were infected with or without T. gondii and then evaluated for cell morphology, intracellular proliferation of the parasite, adenosine receptor expression, TNF-α production and mitogen-activated protein (MAP) kinase signaling pathways triggered by adenosine A3 receptor (A3AR). HTR8/SVneo cells infected with T. gondii exhibited an altered cytoskeletal changes, an increased infection rate and reduced viability in an infection time-dependent manner. T. gondii significantly promoted increased TNF-α production, A3AR protein levels and p38, ERK1/2 and JNK phosphorylation compared to those observed in uninfected control cells. Moreover, the inhibition of A3AR by A3AR siRNA transfection apparently suppressed the T. gondii infection-mediated upregulation of TNF-α, A3AR production and MAPK activation. In addition, T. gondii-promoted TNF-α secretion was dramatically attenuated by pretreatment with PD098059 or SP600125. These results indicate that A3AR-mediated activation of ERK1/2 and JNK positively regulates TNF-α secretion in T. gondii-infected HTR8/SVneo cells.