Dataset Information

A flow cytometric analysis of macrophage- nanoparticle interactions in vitro: induction of altered Toll-like receptor expression.

ABSTRACT:

Background

Nanoparticles exhibit unique physiochemical characteristics that provide the basis for their utilization. The diversity of potential and actual applications compels a thorough understanding regarding the consequences of their containment within the cellular environment.

Purpose

This paper presents a flow cytometric examination of the biologic effects associated with the internalization of citrate-buffered silver (Ag) nanoparticles (NP) by the murine macrophage cell line, RAW264.7.

Materials and methods

Cells were cultured with varying concentrations of citrate-buffered Ag nanoparticle and analyzed for changes in cellular volume, fluorescence emissions, and surface receptor expression.

Results

Notable changes in side scatter (SSC) signal occurred following the phagocytosis of citrate-buffered Ag NP representative of the 10 nm, 50 nm, and 100 nm particle size by cultured RAW 264.7 cells. A characteristic associated with the internalization of all the citrated Ag NP sizes tested, was the detection of emitted infra-red and near-infrared wavelength emissions. This characteristic consistently permitted the detection of 10 nm, 50 nm, and 100 nm Ag NP particles internalized within the RAW cells by flow cytometry. A functional distinction between monocyte subsets within the RAW 264.7 cell line was noted as Ag NP are taken up by the F4/80+ subset of cells within the culture. Further, the internalization of Ag NP by the cells resulted in an increased cell surface expression of the Toll-like receptor (TLR) 3, but not TLR4.

Conclusion

Taken together, these results implicate the more mature macrophage in the ingestion of Ag NP; and an influence upon at least one of the Toll receptors present in macrophages following exposure to Ag NP. Further, our flow cytometric approach presents a potentially viable detection method for the identification of occult Ag NP material using an indicator cell line.

SUBMITTER: Njoroge JM

PROVIDER: S-EPMC6296684 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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Publications

A flow cytometric analysis of macrophage- nanoparticle interactions in vitro: induction of altered Toll-like receptor expression.

Njoroge Joyce M JM Yourick Jeffrey J JJ Principato Mary Ann MA

International journal of nanomedicine 20181212

<h4>Background</h4>Nanoparticles exhibit unique physiochemical characteristics that provide the basis for their utilization. The diversity of potential and actual applications compels a thorough understanding regarding the consequences of their containment within the cellular environment.<h4>Purpose</h4>This paper presents a flow cytometric examination of the biologic effects associated with the internalization of citrate-buffered silver (Ag) nanoparticles (NP) by the murine macrophage cell line ...[more]

PMID: 30587965

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Project description:Implanted biomaterials elicit an immune-mediated foreign body reaction (FBR) that results in the fibrous encapsulation of the implant and can critically impact the performance of some implants. Consequently, understanding the molecular mechanisms that underpin cell-materials interactions that initiate biomaterial-induced inflammation and fibrosis is critical to improving the performance of biomaterial implants negatively impacted by the FBR. Damage-associated molecular patterns (DAMPs) are endogenous mediators of inflammation that are released upon tissue injury and induce sterile inflammation via Toll-like receptors (TLRs). However, the prevalence of DAMPs within the adsorbed protein layer on material surfaces and their role mediating cell-material interactions is unclear. Previously, our group demonstrated that molecules in fibroblast lysates adsorbed to various biomaterials and induced a potent TLR2-dependent inflammatory response in macrophages at 24 h. In this study, we examined the extended response of RAW-Blue reporter macrophages on lysate or serum-adsorbed Teflon™ AF surfaces to understand the potential role of adsorbed DAMPs in macrophage-material interactions at later time points. Lysate-conditioned surfaces maintained increased nuclear factor kappa B (NF-κB) and activator protein 1 (AP-1) transcription factor activity and increased expression Regulated upon Activation, Normal T Cell Expressed and Presumably Secreted (RANTES/CCL5) at 72 h and 120 h, compared to FBS-conditioned surfaces. In contrast, monocyte chemoattractant protein 1 (MCP-1/CCL2) was only elevated at 72 h in lysate conditions. Transforming growth factor beta 1 (TGF-β1) secretion was significantly increased on lysate-conditioned surfaces, while conditioned media from macrophages on lysate-conditioned surfaces induced alpha smooth muscle actin (αSMA) expression in 3T3 fibroblasts. TLR2 neutralizing antibody treatment significantly decreased NF-κB/AP-1 activity and attenuated TGF-β1 expression at both time points, and MCP-1 and RANTES at 72 h. Finally, multinucleated cells were observed on lysate-conditioned surfaces at 72 h, indicating adsorbed DAMPs induced a fusion permissive environment for adherent macrophages. This study demonstrates that adsorbed DAMPs continue to influence macrophage-material responses beyond the initial 24-h period and maintain a pro-inflammatory and fibrotic response that models aspects of the early FBR. Furthermore, the transient inhibition of TLR2 continued to exert an effect at these later time points, suggesting TLR2 may be a target for therapeutic interventions in FBR.

Dataset Information

A flow cytometric analysis of macrophage- nanoparticle interactions in vitro: induction of altered Toll-like receptor expression.

Background

Purpose

Materials and methods

Results

Conclusion

Publications

A flow cytometric analysis of macrophage- nanoparticle interactions in vitro: induction of altered Toll-like receptor expression.

Similar Datasets

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