scholarly journals iTRAQ-based Quantitative Proteomics Analysis Identifies Host Pathways Modulated during Toxoplasma gondii Infection in Swine

2020 ◽  
Vol 8 (4) ◽  
pp. 518 ◽  
Author(s):  
Jun-Jun He ◽  
Jun Ma ◽  
Jin-Lei Wang ◽  
Fu-Kai Zhang ◽  
Jie-Xi Li ◽  
...  
Keyword(s):  
Toxoplasma Gondii ◽  
Protein Expression ◽  
Absolute Quantification ◽  
Mesenteric Lymph Nodes ◽  
Proteomics Analysis ◽  
Liquid Chromatography Tandem Mass ◽  
Isobaric Tags ◽  
Toxoplasma Gondii Infection ◽  
The Brain

Toxoplasma gondii is a leading cause of foodborne illness and consumption of undercooked pig meat is a major risk factor for acquiring toxoplasmosis, which causes a substantial burden on society. Here, we used isobaric tags for relative and absolute quantification (iTRAQ) labelling coupled with liquid chromatography-tandem mass spectrometry (LC-MS/MS) to identify cellular proteins and pathways altered during T. gondii infection in pigs. We also used parallel reaction monitoring-based LC-MS/MS to verify the levels of protein expression of infected spleens and mesenteric lymph nodes (MLNs). At 6 days post-infection (dpi), 156, 391, 170, 292, and 200 differentially expressed proteins (DEPs) were detected in the brain, liver, lung, MLNs and spleen, respectively. At 18 dpi, 339, 351, 483, 388, and 303 DEPs were detected in the brain, liver, lung, MLNs and spleen, respectively. Although proteins involved in immune responses were upregulated in all infected tissues, protein expression signature in infected livers was dominated by downregulation of the metabolic processes. By weighted gene co-expression network analysis, we could further show that all proteins were clustered into 25 co-expression modules and that the pink module significantly correlated with the infection status. We also identified 163 potential anti-T. gondii proteins (PATPs) and provided evidence that two PATPs (HSP70.2 and PDIA3) can reduce T. gondii burden in porcine macrophages in vitro. This comprehensive proteomics analysis reveals new facets in the pathogenesis of T. gondii infection and identifies key proteins that may contribute to the pig’s defense against this infection.

Proteomics-Based Characterization of the Effects of MMP14 on the Protein Content of Exosomes from Corneal Fibroblasts

2020 ◽  
Vol 27 (10) ◽  
pp. 979-988
Author(s):  
Kyu-Yeon Han ◽  
Jin-Hong Chang ◽  
Dimitri T. Azar
Keyword(s):  
Protein Content ◽  
Catalytic Domain ◽  
Protein Composition ◽  
Proteomics Analysis ◽  
Knock Out ◽  
Corneal Fibroblast ◽  
Flow Cytometric ◽  
Corneal Fibroblasts ◽  
Liquid Chromatography Tandem Mass

Background: Exosomes secreted by corneal fibroblasts contain matrix metalloproteinase (MMP) 14, which is known to influence pro-MMP2 accumulation on exosomes. Accordingly, we hypothesized that the enzymatic activity of MMP14 may alter the protein content of corneal fibroblast- secreted exosomes. Objective: The aim of this study was to investigate the effects of MMP14 on the composition and biological activity of corneal fibroblast-derived exosomes. Methods: Knock out of the catalytic domain (ΔExon4) of MMP14 in corneal fibroblasts was used to determine the effect of MMP14 expression on the characteristics of fibroblast-secreted exosomes. The amount of secreted proteins and their size distribution were measured using Nano Tracking Analysis. Proteins within exosomes from wild-type (WT) and ΔExon4-deficient fibroblasts were identified by liquid chromatography-tandem mass spectrometry (MS/MS) proteomics analysis. The proteolytic effects of MMP14 were evaluated in vitro via MS identification of eliminated proteins. The biological functions of MMP14-carrying exosomes were investigated via fusion to endothelial cells and flow cytometric assays. Results: Exosomes isolated from WT and ΔExon4-deficient fibroblasts exhibited similar size distributions and morphologies, although WT fibroblasts secreted a greater amount of exosomes. The protein content, however, was higher in ΔExon4-deficient fibroblast-derived exosomes than in WT fibroblast-derived exosomes. Proteomics analysis revealed that WT-derived exosomes included proteins that regulated cell migration, and ΔExon4 fibroblast-derived exosomes contained additional proteins that were cleaved by MMP14. Conclusion: Our findings suggest that MMP14 expression influences the protein composition of exosomes secreted by corneal fibroblasts, and through those biological components, MMP14 in corneal fibroblasts derived-exosomes may regulate corneal angiogenesis.

Insights into the molecular basis of host behaviour manipulation by Toxoplasma gondii infection

2017 ◽  
Vol 1 (6) ◽  
pp. 563-572 ◽  
Author(s):  
Pierre-Mehdi Hammoudi ◽  
Dominique Soldati-Favre
Keyword(s):  
Toxoplasma Gondii ◽  
Stress Responses ◽  
Molecular Mechanisms ◽  
Brain Regions ◽  
Intermediate Hosts ◽  
Toxoplasma Gondii Infection ◽  
Host Parasite ◽  
Host Behaviour ◽  
The Brain ◽  
Brain Homeostasis

Typically illustrating the ‘manipulation hypothesis’, Toxoplasma gondii is widely known to trigger sustainable behavioural changes during chronic infection of intermediate hosts to enhance transmission to its feline definitive hosts, ensuring survival and dissemination. During the chronic stage of infection in rodents, a variety of neurological dysfunctions have been unravelled and correlated with the loss of cat fear, among other phenotypic impacts. However, the underlying neurological alteration(s) driving these behavioural modifications is only partially understood, which makes it difficult to draw more than a correlation between T. gondii infection and changes in brain homeostasis. Moreover, it is barely known which among the brain regions governing fear and stress responses are preferentially affected during T. gondii infection. Studies aiming at an in-depth dissection of underlying molecular mechanisms occurring at the host and parasite levels will be discussed in this review. Addressing this reminiscent topic in the light of recent technical progress and new discoveries regarding fear response, olfaction and neuromodulator mechanisms could contribute to a better understanding of this complex host–parasite interaction.

scholarly journals iTRAQ-Based Proteomics Analysis of Autophagy-Mediated Responses against MeJA in Laticifers of Euphorbia kansui L.

2019 ◽  
Vol 20 (15) ◽  
pp. 3770
Author(s):  
Fang ◽  
Yao ◽  
Zhang ◽  
Tian ◽  
Wang ◽  
...  
Keyword(s):  
Developmental Process ◽  
Cysteine Proteinase ◽  
Defense Responses ◽  
Absolute Quantification ◽  
Pcr Analysis ◽  
Control Group ◽  
Proteomics Analysis ◽  
Meja Treatment ◽  
Euphorbia Kansui ◽  
Isobaric Tags

Autophagy is a well-defined catabolic mechanism whereby cytoplasmic materials are engulfed into a structure termed the autophagosome. Methyl jasmonate (MeJA), a plant hormone, mediates diverse developmental process and defense responses which induce a variety of metabolites. In plants, little is known about autophagy-mediated responses against MeJA. In this study, we used high-throughput comparative proteomics to identify proteins of latex in the laticifers. The isobaric tags for relative and absolute quantification (iTRAQ) MS/MS proteomics were performed, and 298 proteins among MeJA treated groups and the control group of Euphorbia kansui were identified. It is interesting to note that 29 significant differentially expressed proteins were identified and their associations with autophagy and ROS pathway were verified for several selected proteins as follows: α-L-fucosidase, β-galactosidase, cysteine proteinase, and Cu/Zn superoxide dismutase. Quantitative real-time PCR analysis of the selected genes confirmed the fact that MeJA might enhance the expression of some genes related to autophagy. The western blotting and immunofluorescence results of ATG8 and ATG18a which are two important proteins for the formation of autophagosomes also demonstrated that MeJA could promote autophagy at the protein level. Using the electron microscope, we observed an increase in autophagosomes after MeJA treatment. These results indicated that MeJA might promote autophagy in E. kansui laticifers; and it was speculated that MeJA mediated autophagy through two possible ways: the increase of ROS induces ATG8 accumulation and then aotophagosome formation, and MeJA promotes ATG18 accumulation and then autophagosome formation. Taken together, our results provide several novel insights for understanding the mechanism between autophagy and MeJA treatment. However, the specific mechanism remains to be further studied in the future.

scholarly journals DDRE-14. HEMP DERIVED EXTRACELLULAR VESICLES (EVS): A POTENTIAL ANTI-GLIOMA THERAPY

2020 ◽  
Vol 22 (Supplement_2) ◽  
pp. ii64-ii64
Author(s):  
Hassan Azari ◽  
Nasser Nassiri Koopaei ◽  
Mohammad-Zaman Nouri ◽  
Jesse D Hall ◽  
Nancy D Denslow ◽  
...  
Keyword(s):  
Tumor Progression ◽  
Extracellular Vesicles ◽  
Chromatography Tandem Mass Spectrometry ◽  
Anti Cancer ◽  
Liquid Chromatography Tandem Mass ◽  
Median Diameter ◽  
The Impact ◽  
The Brain ◽  
Plant Sources

Abstract INTRODUCTION Extracellular vesicles (EVs) have been harvested from many plant sources, some of which have anti-cancer effects and some could be used as therapeutic nanodelivery vectors. Hemp plant is a natural source of cannabinoids, of which delta 9-tetrahydroxicannabinol (THC) and cannabidiol (CBD) have proven anti-cancer proprieties. HYPOTHESIS We hypothesized that hemp EVs are enriched in cannabinoids and their application will reduce glioblastoma (GBM) tumor progression. APPROACH EVs were isolated from the hemp plant using ultracentrifugation. Nanotracking analysis, electron microscopy and liquid chromatography tandem mass spectrometry (LC-MS/MS) were utilized to characterize EVs. GBM cell lines were cultured in the neuropshere assay to evaluate hemp EVs anti-glioma effects. Fluorescent-labelled EVs were used to evaluate their brain tissue distribution in orthotopic patient-derived GBM xenografts. RESULTS Hemp EVs have a median diameter of 112.6nm with a typical lipid-bilayer structure. LC-MS/MS have shown that while cannabidiolic, cannabigerolic, and tetrahydroxicannabinolic acids represent 69.1 ± 2.1%, 19.1 ± 1.6%, 6.5 ± 0.54% of the total cannabinoids in hemp EVs, CBD and THC only make 4.75 ± 0.26%, and 0.5 ± 0.3%. Hemp EVs are potent anti-glioma agents with a 7-day LD-50 of 1.04µM and 2.4µM [based on EVs total cannabinoid content] for KR-158 and L0 GBM lines, respectively. Compared to the vehicle, overnight incubation of L0 cells with 1µM hemp EVs significantly reduced GBM cell migration (630.3 ± 61.43 vs 143.7 ± 8.7). Intranasal administration of hemp EVs led to a widespread distribution in tumor bearing brain including GBM tumor core. CONCLUSION Based on these results, hemp EVs with enriched cannabinoid content exert antiglioma effect in-vitro and when delivered intranasally, are widely distributed throughout the brain and within the tumor of PDX animals. Further experiments are ongoing to address the impact of nasally-delivered hemp EVs on tumor progression and compare to the application of purified acidic cannabinoids.

Tissue cyst tropism in Toxoplasma gondii: a comparison of tissue cyst formation in organs of cats, and rodents fed oocysts

Parasitology ◽  
1997 ◽  
Vol 115 (1) ◽  
pp. 15-20 ◽  
Author(s):  
J. P. DUBEY
Keyword(s):  
Toxoplasma Gondii ◽  
Cyst Formation ◽  
Tissue Cyst ◽  
Intermediate Hosts ◽  
Mesenteric Lymph Nodes ◽  
Marked Contrast ◽  
Mesenteric Lymph ◽  
Time Period ◽  
Rats And Mice ◽  
The Brain

The persistence of Toxoplasma gondii tissue cysts in organs of cats (definitive host) and rodents (intermediate hosts) was studied. Nine cats, 12 rats, and 12 mice were fed T. gondii oocysts and their organs were digested in pepsin and then bioassayed for bradyzoites in mice. Of 9 cats killed 37 or 51 days after feeding 102 (2 cats), 103 (3 cats) or 104 (4 cats) oocysts of the VEG strain, tissue cysts were found in each cat; in the tongue of 9, in the heart of 5, in the brain of 4, and in the eyes of 1 cat. The dose had no effect on the distribution of tissue cysts in cats. Twelve rats were each fed 105 oocysts of the VEG strain of T. gondii and killed 21, 29, 64 or 237 days later. At each time-period, 11 tissues of 3 rats were pooled and bioassayed in mice. Tissue cysts were found in the brain, skeletal muscle, heart and kidneys of rats at each killing time; in the lungs, intestines, and mesenteric lymph nodes in 3 of 4 instances; in the tongue, liver, and eyes in 2 instances and in the spleen in 1 instance. Also, using the same procedures and sampling the same 11 tissues as used for rats, tissue cysts were seen in all organs except in the tongue and liver of 3 mice killed on day 82 after feeding the VEG strain. In 9 mice (3 with each strain) fed oocysts of the ME-49, GT-1, or P89 T. gondii strain and killed 62–130 days later, tissue cysts were found consistently only in the brain. Thus, in rats and mice, most tissue cysts were found in the brain and rarely in the tongue. This was in marked contrast to the distribution of tissue cysts in cats.

scholarly journals GZFLW Induces Apoptosis of Ectopic Endometrial Stromal Cells via Promoting VPS53 Protein Stability

2018 ◽  
Vol 2018 ◽  
pp. 1-10
Author(s):  
Ziyu Zhang ◽  
Faying Liu ◽  
YunYun Xu ◽  
Huang Huang ◽  
Yang Zou ◽  
...  
Keyword(s):  
Molecular Mechanism ◽  
Stromal Cells ◽  
In Vitro Study ◽  
Absolute Quantification ◽  
Endometrial Stromal Cells ◽  
Gynecological Diseases ◽  
Isobaric Tags ◽  
Differential Expression Protein ◽  
Vacuolar Protein

Endometriosis is still a major problem in obstetrics and gynecology. While GZFLW (Gui Zhi Fu Ling Wan) has been originally used for treating gynecological diseases, however, the molecular mechanism that GZFLW acts on endometriosis is not clear. To investigate the molecular mechanism that GZFLW plays role on endometriosis, iTRAQ (isobaric tags for relative and absolute quantification) proteomics and human endometrial stromal cells (Y14) obtained from a patient with endometriosis were used in in vitro study. Our results demonstrated that GZFLW decreased Y14 cells proliferation while increased cells apoptosis. The differential expression protein VPS53 (Vacuolar protein sorting 53 homolog) was predicted by iTRAQ coupled LC-MS/MS and further identified by western blot. Besides, GZFLW induced VPS53 protein level by promoting its stabilization. Our findings highlight a novel role for VPS53 in gynecology and provide a potent therapeutic strategy against endometriosis.

scholarly journals Effects of Toxoplasma gondii Infection on the Brain

2007 ◽  
Vol 33 (3) ◽  
pp. 745-751 ◽  
Author(s):  
V. B. Carruthers ◽  
Y. Suzuki
Keyword(s):  
Toxoplasma Gondii ◽  
Toxoplasma Gondii Infection ◽  
The Brain
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