Endotoxin Contamination in Nanomaterials Leads to the Misinterpretation of Immunosafety Results

Li, Y.; Fujita, Mayumi; Boraschi, Diana

doi:10.3389/fimmu.2017.00472

Cited by 75 publications

(71 citation statements)

References 39 publications

(47 reference statements)

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“…It is well‐established that materials containing endotoxins can trigger the TLR4 signaling pathway in immune cells and induce the secretion of IL‐6, IL‐1β, TNF‐α, and IL‐8; however, many groups fail to report or use inadequate methods to detect possible LPS residues, which may lead to debatable results. Owing to the broad differences reported in GBMs immunological properties it is vital to consider the possible presence of endotoxins, since due to their hydrophobic nature, endotoxins would bind strongly to hydrophobic materials such as graphene .…”

Section: Discussionmentioning

confidence: 99%

Exfoliation in Endotoxin‐Free Albumin Generates Pristine Graphene with Reduced Inflammatory Properties

et al. 2018

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mechanical strength. Within the GBMs, pristine graphene (pG) presents key advantages; its virtually defect-free surface enhances its mechanical properties and the absence of oxygen groups increases its electrical and optical conductivity compared to functionalized graphene, [7] presenting opportunities to develop new biomedical devices. Moreover, since no additional chemical modifications are required, the use of pG would have a positive environmental and economic impact over functionalized graphenes, such as graphene oxide (GO). Nevertheless, pG has been less studied in a biological context, due to its intrinsic hydrophobicity, restricting dispersibility and stability in aqueous solution, unless potentially toxic stabilizers are added. Thus, the use of biocompatible dispersants is crucial.We currently lack a complete understanding of the toxic and inflammatory effects of GBMs and conflicting results have been reported. While the toxicity of other graphene-based materials (e.g., GO, rGO), has been extensively studied, [1,[8][9][10] limited work has been done on pristine graphene. [11][12][13] Studies highlight key roles for factors such as lateral size (small vs large), [10] surface functionalization (e.g., GO vs pG), [12,14] purity [15] and hydrophobicity [16] on GBM-cell interaction, thus it is vital to address the intrinsic toxicological effects of pG and elucidate its immunomodulatory properties in order to assess its specific potential as a material for biomedical applications. According Graphene-based materials are under consideration as a new generation of biomaterials for medical applications, owning to their distinctive physicochemical properties. A major obstacle to clinical translation of novel biomaterials is the presence of residual endotoxin contaminants that can trigger deleterious inflammatory responses and unrelated immunomodulatory effects. To overcome this challenge, a liquid exfoliation methodology to prepare endotoxin-free bovine serum albumin (BSA)-exfoliated pristine graphene (EF-Gr) was developed and compared with conventional BSA-exfoliated graphene (C-Gr) and sodium cholate-exfoliated graphene (SC-Gr). In contrast to SC-Gr, protein-exfoliated graphene exhibits low toxicity. Moreover, BSAexfoliated EF-Gr does not trigger cytokine secretion or increase costimulatory molecule expression in dendritic cells (DCs) or macrophages; however it retains the ability to activate the NLRP3 inflammasome in DCs primed withToll-like receptor (TLR) agonists. The results suggest that some reported immunostimulatory effects of graphene preparations may have resulted from contaminants in the formulations. When injected into mice, the inflammatory response induced is comparable in magnitude to that induced by the safe and effective vaccine adjuvant alum. Overall, the study demonstrates the feasibility of producing endotoxin-free pristine graphene that facilitates assessment of the potential of pristine graphene for novel biomedical applications without the confounding effects of toxic and inflammatory conta...

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Section: Discussionmentioning

confidence: 99%

Exfoliation in Endotoxin‐Free Albumin Generates Pristine Graphene with Reduced Inflammatory Properties

et al. 2018

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“…In this context, the presence of an ubiquitous contaminant such as bacterial endotoxin (lipopolysaccharide, LPS) should be thoroughly assessed in newly developed engineered NP preparations, since LPS is a robust, persistent, ubiquitous, potent immune/inflammatory activator, and toxic molecule, which is difficult to detect in NPs and could be responsible for the majority of inflammatory/toxic effects generally attributed to engineered NPs Li, Fujita, and Boraschi 2017).…”

Section: Introductionmentioning

confidence: 99%

Bacterial endotoxin (lipopolysaccharide) binds to the surface of gold nanoparticles, interferes with biocorona formation and induces human monocyte inflammatory activation

Shi

Radauer-Preiml

et al. 2017

Nanotoxicology

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Nanoparticles (NPs) are easily contaminated by bacterial endotoxin (lipopolysaccharide [LPS]). The presence of LPS can be responsible for many immune/inflammatory effects attributed to NPs. In this study, we examined the effects of LPS adsorption on the NP surface on the formation of a biocorona in biological fluids and on the subsequent inflammation-inducing activity of NPs. Different gold (Au) NPs with sizes ranging from 10 to 80 nm and with different surface functionalization (sodium citrate, lipoic acid, and branched polyethyleneimine (BPEI), or polyethylene glycol (PEG)) were exposed to E. coli LPS under different conditions. The binding capacity of LPS to the surface of AuNPs was dose-and time-dependent. LPS attached to sodium citrate and lipoic acid coatings, but did not adhere to BPEI-or PEG-coated NPs. By computational simulation, the binding of LPS to AuNPs seems to follow the Langmuir absorption isotherm. The presence of LPS on AuNP surface interfered and caused a decrease in the formation of the expected biomolecular corona upon incubation in human plasma. LPS-coated AuNPs, but not the LPS-free NPs, induced significant inflammatory responses in vitro. Notably, while free LPS did also induce an anti-inflammatory response, LPS bound to NPs appeared unable to do so. In conclusion, the unintentional adsorption of LPS onto the NP surface can affect the biocorona formation and the inflammatory properties of NPs. Thus, for an accurate interpretation of NP interactions with cells, it is extremely important to be able to distinguish the intrinsic NP biological effects from those caused by biologically active contaminants such as endotoxin. ARTICLE HISTORY

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“…Endotoxin or lipopolysaccharide (LPS) is a molecule found in the outer membrane of Gram-negative bacteria. [9] It is nearly ubiquitous, due also to the bacterial flora that every human carries. LPS can easily persist in the absence of these bacteria and due to its high thermostability, it is resistant towards most standard sterilization methods applied in biological laboratories.…”

Section: Characteristics Of Endotoxinmentioning

confidence: 99%

When Would Immunologists Consider a Nanomaterial to be Safe? Recommendations for Planning Studies on Nanosafety

Himly

Geppert

Hofer

et al. 2020

Small

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The immune system is professional in recognizing and responding to non-self, including nanomaterials. Immune responses by professional and nonprofessional immune cells are thus nearly inevitable upon exposure of cells and organisms to such materials. The state of research into taking the immune system into account in nanosafety studies is reviewed and three aspects in which further improvements are desirable are identified: 1) Due to technical limitations, more stringent testing for endotoxin contamination should be made. 2) Since under overdose conditions immunity shows unphysiological responses, all doses used should be justified by being equivalent to tissue-delivered doses. 3) When markers of acute inflammation or cell stress are observed, functional assays are necessary to distinguish between homeostatic fluctuation and genuine defensive or tolerogenic responses. Since immune activation can also indicate that the immune system considers a stimulus to be harmless and induces tolerance, activation markers by themselves do not necessarily imply a danger to the body. Guidelines such as these are necessary to approach the point where specific nanomaterials are classified as safe based on reliable testing strategies.

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Endotoxin Contamination in Nanomaterials Leads to the Misinterpretation of Immunosafety Results

Cited by 75 publications

References 39 publications

Exfoliation in Endotoxin‐Free Albumin Generates Pristine Graphene with Reduced Inflammatory Properties

Exfoliation in Endotoxin‐Free Albumin Generates Pristine Graphene with Reduced Inflammatory Properties

Bacterial endotoxin (lipopolysaccharide) binds to the surface of gold nanoparticles, interferes with biocorona formation and induces human monocyte inflammatory activation

When Would Immunologists Consider a Nanomaterial to be Safe? Recommendations for Planning Studies on Nanosafety

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