Hise AG et al. (2003) J Endotoxin Res "Immunopathogenesis of Onchocerca volvulus keratitis (river blindness): a novel role for TLR4 and endosymbiotic Wolbachia bacteria."

Pearlman E, Hise AG, Gillette-Ferguson I

[J Endotoxin Res, 2003]

Infection with the parasitic nematode Onchocerca volvulus is associated with inflammation of the skin and cornea that can lead to blindness. Corneal damage is thought to occur as a result of the host inflammatory responses to degenerating microfilariae in the eye. We have utilized a murine model of corneal inflammation (keratitis) to investigate the immune and inflammatory responses associated with river blindness. Soluble extracts of O. volvulus, a filarial species that contains the endosymbiont bacteria Wolbachia or Acanthocheilonema viteae (a nematode not naturally infected with the bacteria) were injected into mouse corneas. Inflammatory responses and corneal changes were measured. We demonstrated a major role for endosymbiont Wolbachia bacteria and Toll-like receptor 4 (TLR4) in the pathogenesis of ocular onchocerciasis.

Hise AG et al. (2007) J Immunol "Innate immune responses to endosymbiotic Wolbachia bacteria in Brugia malayi and Onchocerca volvulus are dependent on TLR2, TLR6, MyD88, and Mal, but not TLR4, TRIF, or TRAM."

Taylor MJ, Fitzgerald KA, Cho E, Golenbock DT, Hise AG, McGarry HF, Gillette-Ferguson I, Daehnel K, Pearlman E, Kazura JW

[J Immunol, 2007]

The discovery that endosymbiotic Wolbachia bacteria play an important role in the pathophysiology of diseases caused by filarial nematodes, including lymphatic filariasis and onchocerciasis (river blindness) has transformed our approach to these disabling diseases. Because these parasites infect hundreds of millions of individuals worldwide, understanding host factors involved in the pathogenesis of filarial-induced diseases is paramount. However, the role of early innate responses to filarial and Wolbachia ligands in the development of filarial diseases has not been fully elucidated. To determine the role of TLRs, we used cell lines transfected with human TLRs and macrophages from TLR and adaptor molecule-deficient mice and evaluated macrophage recruitment in vivo. Extracts of Brugia malayi and Onchocerca volvulus, which contain Wolbachia, directly stimulated human embryonic kidney cells expressing TLR2, but not TLR3 or TLR4. Wolbachia containing filarial extracts stimulated cytokine production in macrophages from C57BL/6 and TLR4(-/-) mice, but not from TLR2(-/-) or TLR6(-/-) mice. Similarly, macrophages from mice deficient in adaptor molecules Toll/IL-1R domain-containing adaptor-inducing IFN-beta and Toll/IL-1R domain-containing adaptor-inducing IFN-beta-related adaptor molecule produced equivalent cytokines as wild-type cells, whereas responses were absent in macrophages from MyD88(-/-) and Toll/IL-1R domain-containing adaptor protein (TIRAP)/MyD88 adaptor-like (Mal) deficient mice. Isolated Wolbachia bacteria demonstrated similar TLR and adaptor molecule requirements. In vivo, macrophage migration to the cornea in response to filarial extracts containing Wolbachia was dependent on TLR2 but not TLR4. These results establish that the innate inflammatory pathways activated by endosymbiotic Wolbachia in B. malayi and O. volvulus filaria are dependent on TLR2-TLR6 interactions and are mediated by adaptor molecules MyD88 and TIRAP/Mal.

Starnes DL et al. (2015) Environ Pollut "Impact of sulfidation on the bioavailability and toxicity of silver nanoparticles to Caenorhabditis elegans."

Collin BE, Oostveen EK, Ma R, Bertsch PM, Starnes CP, Lowry GV, Tsyusko OV, Starnes DL, Unrine JM

[Environ Pollut, 2015]

Sulfidation is a major transformation product for manufactured silver nanoparticles (Ag-MNPs) in the wastewater treatment process.We studied the dissolution, uptake, and toxicity of Ag-MNP and sulfidized Ag-MNPs (sAg-MNPs) to a model soil organism, Caenorhabditis elegans. Our results show that reproduction was the most sensitive endpoint tested for both Ag-MNPs and sAg-MNPs. We also demonstrate that sulfidation not only decreases solubility of Ag-MNP, but also reduces the bioavailability of intact sAg-MNP. The relative contribution of released Ag(+) compared to intact particles to toxicity was concentration dependent. At lower total Ag concentration, a greater proportion of the toxicity could be explained by dissolved Ag, whereas at higher total Ag concentration, the toxicity appeared to be dominated by particle specific effects.

Starnes, Daniel L et al. (2013) International Worm Meeting "Toxicogenomic responses of Caenorhabditis elegans to silver nanomaterials."

Unrine, J., Tsyusko, O., Collin, B., Starnes, Daniel L, Bertsch, P., Oostveen, E., Smith, J., Starnes, C.

[International Worm Meeting, 2013]

There are over 2000 consumer products containing manufactured nanomaterials (MNMs) available today. One of the MNMs of major concern is silver nanoparticles (AgNPs). During wastewater treatment, AgNPs can undergo transformations resulting in partially or fully sulfidized AgNPs. Our objective is to understand the bioavailability and toxicity of polyvinylpyrrolidone (PVP) coated AgNPs (Ag-PVP) and fully sulfidized AgNPs (Ag-S) to a model organism Caenorhabditis elegans using a toxicogenomic approach. Since AgNP toxicity can also be determined by dissolution and release of Ag+, to differentiate between particle and ion-specific toxicity we included AgNO3 as an additional treatment. Our results showed that for AgNPs, Ag-PVP are more toxic to C. elegans than Ag-S due in part to a greater bioavailability and uptake as evidenced by synchrotron-based x-ray microscopy. Evidence also suggests that the observed toxicity is partially particle specific for both Ag-PVP and Ag-S because nematodes exposed to particle free supernatants showed very low mortality. Among endpoints screened, reproduction was the most sensitive and was used for the microarray study. The transcriptomic data indicate that each treatment produced a distinct genomic response. Ag+ had the largest number of differentially expressed genes (312) followed by Ag-S (223), and lastly by Ag-PVP (136). Of the total number of significant differentially expressed genes only 3% were shared among all of the treatments and 86% were uniquely expressed for the respective treatments. The genomic data support the hypothesis that the observed responses are partially particle specific, since both Ag-PVP and Ag-S have a unique set of genes that are not shared with the Ag+ treatment. In addition, our results demonstrate that C. elegans transcriptomic responses to the Ag-S are distinct from those to Ag-PVP resulting in their different toxicities.

Yang X et al. (2014) Environ Sci Technol "Silver nanoparticle behavior, uptake, and toxicity in Caenorhabditis elegans: effects of natural organic matter."

Wiesner MR, Meyer J, Badireddy AR, Yang X, Hinton DE, Dykstra M, Jiang C, Hsu-Kim H

[Environ Sci Technol, 2014]

Significant progress has been made in understanding the toxicity of silver nanoparticles (Ag NPs) under carefully controlled laboratory conditions. Natural organic matter (NOM) is omnipresent in complex environmental systems, where it may alter the behavior of nanoparticles in these systems. We exposed the nematode Caenorhabditis elegans to Ag NP suspensions with or without one of two kinds of NOM, Suwannee River and Pony Lake fulvic acids (SRFA and PLFA, respectively). PLFA rescued toxicity more effectively than SRFA. Measurement of total tissue silver content indicated that PLFA reduced total organismal (including digestive tract) uptake of ionic silver, but not of citrate-coated Ag NPs (CIT-Ag NPs). The majority of the CIT-Ag NP uptake was in the digestive tract. Limited tissue uptake was detected by hyperspectral microscopy but not by transmission electron microscopy. Co-exposure to PLFA resulted in the formation of NOM-Ag NP composites (both in medium and in nematodes) and rescued AgNO3- and CIT-Ag NP-induced cellular damage, potentially by decreasing intracellular uptake of CIT-Ag NPs.

Kleiven M et al. (2018) Environ Toxicol Chem "Characterizing NM300K Silver Nanoparticles Behavior, Uptake and Toxicity in Caenorhabditis elegans."

Gallego-Urrea JA, Brede DA, Oughton DH, Kleiven M, Coutris C, Rossbach LM

[Environ Toxicol Chem, 2018]

Using Caenorhabditis elegans as a model organism, this study addresses the potential linkage between toxicity of NM300K Ag nanoparticles (AgNPs), their particle size distribution and the presence of dissolved Ag in the test media. Of the three endpoints assessed (growth, fertility and reproduction), reproduction was the most sensitive, with 50% effect concentration (EC50) ranging from 0.26-0.84mg Ag L^-1and 0.08-0.11mg Ag L^-1for NM300K and AgNO₃, respectively. Silver uptake by C. elegans was similar for both forms of Ag, while bioaccumulation was higher in AgNO₃exposure. The observed differences in toxicity between NM300K and AgNO₃did not correlate to bioaccumulated Ag, which suggests the toxicity to be a function of the type of exposing agent (AgNPs vs AgNO₃) and their mode of action. Before addition of the food source, E. coli, size fractionation revealed that dissolved Ag comprised 13-90% and 4-8% of total Ag in the AgNO₃and NM300K treatments, respectively. No dissolved Ag was detectable in the actual test media, due to immediate Ag adsorption to bacteria. Results from the current study highlight that information on behavior and characterization of exposure conditions is essential for nanotoxicity studies. This article is protected by copyright. All rights reserved.

Yang X et al. (2012) Environ Sci Technol "Mechanism of silver nanoparticle toxicity is dependent on dissolved silver and surface coating in Caenorhabditis elegans."

Marinakos SM, Hsu-Kim H, Yang X, Liu J, Meyer J, Gondikas A, Auffan M

[Environ Sci Technol, 2012]

The rapidly increasing use of silver nanoparticles (Ag NPs) in consumer products and medical applications has raised ecological and human health concerns. A key question for addressing these concerns is whether Ag NP toxicity is mechanistically unique to nanoparticulate silver, or if it is a result of the release of silver ions. Furthermore, since Ag NPs are produced in a large variety of monomer sizes and coatings, and since their physicochemical behavior depends on the media composition, it is important to understand how these variables modulate toxicity. We found that a lower ionic strength medium resulted in greater toxicity (measured as growth inhibition) of all tested Ag NPs to Caenorhabditis elegans and that both dissolved silver and coating influenced Ag NP toxicity. We found a linear correlation between Ag NP toxicity and dissolved silver, but no correlation between size and toxicity. We used three independent and complementary approaches to investigate the mechanisms of toxicity of differentially coated and sized Ag NPs: pharmacological (rescue with trolox and N-acetylcysteine), genetic (analysis of metal-sensitive and oxidative stress-sensitive mutants), and physicochemical (including analysis of dissolution of Ag NPs). Oxidative dissolution was limited in our experimental conditions (maximally 15% in 24 h) yet was key to the toxicity of most Ag NPs, highlighting a critical role for dissolved silver complexed with thiols in the toxicity of all tested Ag NPs. Some Ag NPs (typically less soluble due to size or coating) also acted via oxidative stress, an effect specific to nanoparticulate silver. However, in no case studied here was the toxicity of a Ag NP greater than would be predicted by complete dissolution of the same mass of silver as silver ions.

Jiang X et al. (2020) J Fluoresc "A Water-Soluble Fluorescent Probe for the Selective Sensing of Ag+ and its Application in Imaging of Living Cells and Nematodes."

Deng M, Wu J, Jiang X, Li H, Yang Y, Zhou X, Qi X, Lu M, Liang S

[J Fluoresc, 2020]

In this study, an imidazole-coumarin based fluorescent probe was developed for the selective and sensitive detection of Ag⁺ in aqueous solution. Using a combination of Job plot, NMR titrations, and DFT calculations, the binding properties between Ag⁺ and the probe were deeply investigated, and the results revealed a 1:1 binding stoichiometry between the probe and Ag⁺ with a binding constant of 1.02x10⁶M^-1. The detection limit was found to be 150nM, which satisfies the requirement for the quantitative detection of Ag⁺ in real water samples. Moreover, the new probe, Ic, was successfully applied to sense Ag⁺ in HeLa and HepG2 cells as well as in C. elegans, indicating that it could be a useful tool for the environmental monitoring of Ag⁺ pollution. These results demonstrated that Ic could serve as a high-efficiency and low-cost fluorescent probe for tracking Ag⁺ in an aquatic environment and biological organisms.

Farrer T et al. (2002) Nucleic Acids Res "Analysis of the role of Caenorhabditis elegans GC-AG introns in regulated splicing."

Farrer T, Kent WJ, Roller AB, Zahler AM

[Nucleic Acids Res, 2002]

GC-AG introns represent 0.7% of total human pre-mRNA introns. To study the function of GC-AG introns in splicing regulation, 196 cDNA-confirmed GC-AG introns were identified in Caenorhabditis elegans. These represent 0.6% of the cDNA- confirmed intron data set for this organism. Eleven of these GC-AG introns are involved in alternative splicing. In a comparison of the genomic sequences of homologous genes between C.elegans and Caenorhabditis briggsae for 26 GC-AG introns, the C at the +2 position is conserved in only five of these introns. A system to experimentally test the function of GC-AG introns in alternative splicing was developed. Results from these experiments indicate that the conserved C at the +2 position of the tenth intron of the let-2 gene is essential for developmentally regulated alternative splicing. This C allows the splice donor to function as a very weak splice site that works in balance with an alternative GT splice donor. A weak GT splice donor can functionally replace the GC splice donor and allow for splicing regulation. These results indicate that while the majority of GC-AG introns appear to be constitutively spliced and have no evolutionary constraints to prevent them from being GT-AG introns, a subset of GC-AG introns is involved in alternative splicing and the C at the +2 position of these introns can have an important role in splicing regulation.

Wamucho A et al. (2019) Environ Pollut "Genomic mutations after multigenerational exposure of Caenorhabditis elegans to pristine and sulfidized silver nanoparticles."

Farman M, Kieran TJ, Glenn TC, Svendsen C, Spurgeon DJ, Unrine JM, Schultz CL, Wamucho A, Tsyusko OV

[Environ Pollut, 2019]

Our previous study showed heritable reproductive toxicity in the nematode Caenorhabditis elegans after multigenerational exposure to AgNO₃ and silver nanoparticles (Ag-NPs). The aim of this study was to determine whether such inheritable effects are correlated with induced germline mutations in C.elegans. Individual C.elegans lineages were exposed for 10 generations to equitoxic concentrations at EC₃₀ of AgNO₃, Ag-NPs, and sulfidized Ag-NPs (sAg-NPs), a predominant environmentally transformed product of pristine Ag-NPs. The mutations were detected via whole genome DNA sequencing approach by comparing F₀ and F₁₀ generations. An increase in the total number of variants, though not statistically significant, was observed for all Ag treatments and the variants were mainly contributed by single nucleotide polymorphisms (SNPs). This potentially contributed towards reproductive as well as growth toxicity shown previously after ten generations of exposure in every Ag treatment. However, despite Ag-NPs and AgNO₃ inducing stronger reproductive toxicity than sAg-NPs, exposure to sAg-NPs resulted in higher mutation accumulation with significant increase in the number of transversions. Thus our results suggest that other mechanisms of inheritance, such as epigenetics, may be at play in Ag-NP- and AgNO₃-induced multigenerational and transgenerational reproductive toxicity.