Internalization of titanium dioxide nanoparticles by glial cells is given at short times and is mainly mediated by actin reorganization-dependent endocytosis

Many nanoparticles (NPs) have toxic effects on multiple cell lines. This toxicity is assumed to be related to their accumulation within cells. However, the process of internalization of NPs has not yet been fully characterized. In this study, the cellular uptake, accumulation, and localization of titanium dioxide nanoparticles (TiO₂ NPs) in rat (C6) and human (U373) glial cells were analyzed using time-lapse microscopy (TLM) and transmission electron microscopy (TEM). Cytochalasin D (Cyt-D) was used to evaluate whether the internalization process depends of actin reorganization. To determine whether the NP uptake is mediated by phagocytosis or macropinocytosis, nitroblue tetrazolium (NBT) reduction was measured and the 5-(N-ethyl-N-isopropyl)-amiloride was used. Expression of proteins involved with endocytosis and exocytosis such as caveolin-1 (Cav-1) and cysteine string proteins (CSPs) was also determined using flow cytometry.TiO₂ NPs were taken up by both cell types, were bound to cellular membranes and were internalized at very short times after exposure (C6, 30 min; U373, 2 h). During the uptake process, the formation of pseudopodia and intracellular vesicles was observed, indicating that this process was mediated by endocytosis. No specific localization of TiO₂ NPs into particular organelles was found: in contrast, they were primarily localized into large vesicles in the cytoplasm. Internalization of TiO₂ NPs was strongly inhibited by Cyt-D in both cells and by amiloride in U373 cells; besides, the observed endocytosis was not associated with NBT reduction in either cell type, indicating that macropinocytosis is the main process of internalization in U373 cells. In addition, increases in the expression of Cav-1 protein and CSPs were observed.In conclusion, glial cells are able to internalize TiO₂ NPs by a constitutive endocytic mechanism which may be associated with their strong cytotoxic effect in these cells; therefore, TiO₂ NPs internalization and their accumulation in brain cells could be dangerous to human health.     

Potentilla anserine L. polysaccharide protects against cadmium-induced neurotoxicity

Cadmium is a toxic metal that can damage the brain and other organs. This study aimed to explore the protective effects of Potentilla anserine L. polysaccharide (PAP) against CdCl₂-induced neurotoxicity in N2a and SH-SY5Y cells and in the cerebral cortex of BALB/c mice. In addition, we aimed to identify the potential mechanisms underlying these protective effects. Relative to CdCl₂ treatment alone, pretreatment with PAP prevented the reduction in cell viability evoked by CdCl₂, decreased rates of apoptosis, promoted calcium homeostasis, decreased ROS accumulation, increased mitochondrial membrane potential, inhibited cytochrome C and AIF release, and prevented the cleavage of caspase-3 and PARP. In addition, PAP significantly decreased the CdCl₂-induced phosphorylation of CaMKII, Akt, and mTOR. In conclusion, PAP represents a potential therapeutic agent for the treatment of Cd-induced neurotoxicity, functioning in part via attenuating the activation of the mitochondrial apoptosis pathway and the Ca²⁺-CaMKII-dependent Akt/mTOR pathway.     

1H nuclear magnetic resonance-based metabolic profiling of cerebrospinal fluid to identify metabolic features and markers for tuberculosis meningitis

BackgroundTuberculosis meningitis (TBM) is the most severe form of tuberculosis, and currently lacks efficient diagnostic approaches. Metabolomics has the potential to differentiate patients with TBM from those with other forms of meningitis and meningitis-negative individuals. However, no systemic metabolomics research has compared the cerebrospinal fluid (CSF) of these patients.Methods¹H nuclear magnetic resonance (NMR) was used for CSF metabolic profiling. Principal component analysis and orthogonal signal correction-partial least squares-discriminant analysis (OPLS-DA) were used to screen for important variables. The Human Metabolome Database was used to identify metabolites, and MetaboAnalyst 4.0 was used for pathway analysis and over-representation analysis.ResultsOPLS-DA modeling could distinguish TBM from other forms of meningitis, and several significantly changed metabolites were identified. Additionally, 23, 6, and 21 metabolites were able to differentiate TBM from viral meningitis, bacterial meningitis, and meningitis-negative groups, respectively. Pathway analysis indicated that these metabolites were mainly involved in carbohydrate and amino acid metabolism, and over-representation analysis indicated that some of these pathways were over-represented.ConclusionsThe metabolites identified have the potential to serve as biomarkers for TBM diagnosis, and carbohydrate and amino acid metabolism are perturbed in the CSF of patents with TBM. Metabolomics is a valuable approach for screening TBM biomarkers. With further investigation, the metabolites identified in this study could aid in TBM diagnosis.     

Spatial,temporal and host factors structure the Ceratomyxa shasta (Myxozoa) population in the Klamath River basin

The myxozoan parasite Ceratomyxa shasta is a virulent pathogen of salmonid fish in the Klamath River, Oregon/California, USA. We previously defined four principal genotypes of the parasite (O, I, II, III) based on a trinucleotide repeat (ATC)_0–3 in Internal Transcribed Spacer region 1 sequences. Genotypes occur in sympatry and show marked host preference: I in Chinook salmon (Oncorhynchus tschawytscha) and II in non-native rainbow trout (O. mykiss). In the present study, we sequenced the parasite from river water samples collected in May, June and September at three localities below, above and between the Klamath's five dams. We also sampled adult and juvenile coho salmon (O. kisutch), steelhead trout (O. mykiss, anadromous form) and native redband rainbow trout (O. mykiss, freshwater form) and additional Chinook salmon and non-native rainbow trout. We found that the C. shasta population was highly structured spatially, temporally and with respect to fish host species. Genotype O was present in water throughout the basin but detected almost exclusively in steelhead and native rainbow trout. Genotype I was in water only below the dams and detected only in Chinook salmon. Genotype II was detected in coho salmon below the dams, and in non-native rainbow trout exposed both above and below the dams. The same genotypes were detected in adult and juvenile fish of the same species. These findings have major implications for the design of effective surveillance and control programs for this economically and ecologically important fish parasite.     

Genetic diversity of the Leptospiral immunoglobulin-like (Lig) genes in pathogenic Leptospira spp.

Recent serologic, immunoprotection, and pathogenesis studies identified the Lig proteins as key virulence determinants in interactions of leptospiral pathogens with the mammalian host. We examined the sequence variation and recombination patterns of ligA, ligB, and ligC among 10 pathogenic strains from five Leptospira species. All strains were found to have intact ligB genes and genetic drift accounting for most of the ligB genetic diversity observed. The ligA gene was found exclusively in L. interrogans and L. kirschneri strains, and was created from ligB by a two-step partial gene duplication process. The aminoterminal domain of LigB and the LigA paralog were essentially identical (98.5 ± 0.8% mean identity) in strains with both genes. Like ligB, ligC gene variation also followed phylogenetic patterns, suggesting an early gene duplication event. However, ligC is a pseudogene in several strains, suggesting that LigC is not essential for virulence. Two ligB genes and one ligC gene had mosaic compositions and evidence for recombination events between related Leptospira species was also found for some ligA genes. In conclusion, the results presented here indicate that Lig diversity has important ramifications for the selection of Lig polypeptides for use in diagnosis and as vaccine candidates. This sequence information will aid the identification of highly conserved regions within the Lig proteins and improve upon the performance characteristics of the Lig proteins in diagnostic assays and in subunit vaccine formulations with the potential to confer heterologous protection.     

Polyphenolic compounds from propolis modulate immune responses and increase host resistance to tumour cells

Propolis contains a variety of polyphenolic compounds. We investigated the effect of a water-soluble derivatives of propolis (WSDP) and polyphenolic compounds, components of propolis, on growth of Ehrlich ascites tumour (EAT) in mice. Tumour in peritoneal cavity was produced by 2×10⁶ EAT cells. WSDP and polyphenolic compounds (caffeic acid-CA, caffeic acid phenethyl ester-CAPE and quercetin-QU) were given to mice perorally (po). It was found that the volume of ascitic fluid induced by EAT cells and total number of cells present in the peritoneal cavity was markedly reduced in EAT-bearing mice treated with test components and the survival time of treated mice was prolonged. Inhibition of EAT growth was due to their effect on the immune system of mice. When innate and acquired immune responses were evaluated, a dose-related increase of cytotoxic T-cell, NK and B cells activity was observed in test components-treated mice. Furthermore, exposure of animals to test components increased functional activity of macrophages to produce factors regulating the function of B-, T-, and NK- cells respectively. In conclusion, these findings imply that the antitumour activity of WSDP and polyphenolic compounds of propolis enhanced host resistance in the EAT tumour model, increasing the activities of macrophages, cytotoxic T cells, B cells and NK cells.