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Key Event Title
Key Event Components
Key Event Overview
AOPs Including This Key Event
|AOP Name||Role of event in AOP||Point of Contact||Author Status||OECD Status|
|Frustrated phagocytosis-induced lung cancer||MolecularInitiatingEvent||Carole Seidel (send email)||Under development: Not open for comment. Do not cite||Under Development|
|Frustrated phagocytosis leads to malignant mesothelioma||MolecularInitiatingEvent||Nureddin Mansour (send email)||Under development: Not open for comment. Do not cite|
Key Event Description
Phagocytosis is the first line of defence of the organism against foreign matter and therefore is essential for the maintenance of the homeostasis . This process, mainly performed by macrophages, is dividing in two steps, first after recognition and internalization of the foreign matter, the phagosome is formed, and second, this structure is mature in a degradative compartment .
In the lung tissue, macrophages located in the alveolar space are involved in the clearance of foreign matter inhaled. After phagocytosis, cells migrate out of the alveolar space via the mucociliary escalator or the lymphatic system.
High aspect ratio nanoparticles (HARN) are particles with a ratio length – diameter ≥ 3  . Their fibre-shaped, similar to asbestos, is causing concern about their toxicity . HARN include nanotubes, nanorods, nanowires and nanofibers in which carbon nanotubes (CNTs) are the most known and studied. CNT could enter in cells and interact with mitotic spindles as well as nuclei . Macrophages try to phagocytose these particles, however the phagocytosis if incomplete leading to a frustrated phagocytosis. Consequently, the foreign matter is retained in the body because it cannot be cleared by macrophages .
How It Is Measured or Detected
Using in vitro cell models, such as macrophages or epithelial cells, the analysis of interaction between HARN and cells could be performed in time-lapse microscopy  or backscatter electron microscopy .
The frustrated phagocytosis could be measure by different type of microscopy analysis allowing a direct measurement. For examples, time-lapse video microscopy , light microscopy , scanning electron microscopy [6, 9], bright-field microscopy  and backscatter electron microscopy  are used in the literature.
The analysis of phagocytic receptor expression such as MARCO, MSR-1, CD36, TLR4 is an indirect measurement .
The study of the capacity of macrophages to complete phagocytosis process could be performed in vitro using different type of macrophage cell lines (THP-1, NR8383, RAW267) and analysis by microscopy or gene expression or in vivo after exposure of rodents to different type of high aspect ratio nanoparticles and analysis of the remaining quantity of material in the body or the ability of macrophages to phagocyte foreign matter.
Domain of Applicability
The frustrated phagocytosis of high aspect ratio particle can occur in mammals, male or female, and is generally measured in adults.
Evidence for Perturbation by Stressor
Overview for Molecular Initiating Event
High aspect ratio nanoparticles are not completely engulfed by macrophages due to their long shape (> 10µm) [8, 10].
Some studies analysed the effect of the length of nanoparticles (NPs) on the capacity of macrophages to phagocytose them.
The study of Sweeney et al 2015 demonstrated that treatment of primary human alveolar macrophages by multi-walled carbon nanotubes (MWCNT) similar in term of diameter, specific surface area and purity but differ by their length differentially alter phagocytosis . Indeed, treatment by the longer CNT (median length 19.3 µm) induced frustrated phagocytosis and receptor expression (MARCO) as well as decreased phagocytic ability and migratory capacity in a more extend manner than the shorter CNT (median length 1.1 µm) . Another study analysed the effect of particle morphology on the ability of human monocytic cell line THP-1 to engulf carbon nanotubes . Cells were treated for 24h by two longs CNTs (men length 13 and 36 µm, dimeter 84.89 and 165.02 nm), two tangled CNTs (length 1-5 and 5-20 µm, dimeter 14.84 and 10.40 nm) and one short CNT (length 1-2 µm, diameter 25.7 nm). The authors observed by light microscopy that only the two long CNTs are protruding from the cells. A study on THP-1 cells treated for 4h was conducted with silver nanowires that possess similar diameter but different length (average length: 3, 5, 10, 14 and 28 µm) . The authors observed by bright-field microscopy that the shorter NPs (3 and 5) were fully enclosed by macrophages, while the longer NPs (14 and 28) caused frustrated phagocytosis. In addition, injection of NPs in mouse pleural cavity followed by pleural lavage demonstrated that the shorter (3 and 5) were fully phagocytosed whereas the longer (10) caused frustrated phagocytosis. The authors observed differences between in vitro and in vivo studies in term of sensitivity for the determination of the length threshold that caused frustrated phagocytosis . Finally, Padmore et al showed by time-lapse video microscopy that immortalized MH-S murine alveolar macrophages were able to internalized short glass fibres (mean length 7 µm) whereas the longer fibres were not (mean length 39.3 µm) .
All together, these studies could suggest that the threshold for frustrated phagocytosis should be around 10 µm, close to the suggestion formulated by Donaldson et al that fibres longer than 15 µm cause this process .
1. Montano F, Grinstein S, Levin R. Quantitative Phagocytosis Assays in Primary and Cultured Macrophages. 2018;1784:151-63; doi: 10.1007/978-1-4939-7837-3_15.
3. Oberdorster G, Oberdorster E, Oberdorster J. Concepts of nanoparticle dose metric and response metric. 2007;115 6:A290; doi: 10.1289/ehp.115-1892118.
4. Donaldson K, Poland CA. Inhaled nanoparticles and lung cancer - what we can learn from conventional particle toxicology. 2012;142:w13547; doi: 10.4414/smw.2012.13547.
5. Sargent LM, Shvedova AA, Hubbs AF, Salisbury JL, Benkovic SA, Kashon ML, et al. Induction of aneuploidy by single-walled carbon nanotubes. 2009;50 8:708-17; doi: 10.1002/em.20529.
6. Sweeney S, Grandolfo D, Ruenraroengsak P, Tetley TD. Functional consequences for primary human alveolar macrophages following treatment with long, but not short, multiwalled carbon nanotubes. International journal of nanomedicine. 2015;10:3115-29; doi: 10.2147/IJN.S77867.
7. Padmore T, Stark C, Turkevich LA, Champion JA. Quantitative analysis of the role of fiber length on phagocytosis and inflammatory response by alveolar macrophages. Biochimica et biophysica acta. 2017;1861 2:58-67; doi: 10.1016/j.bbagen.2016.09.031.
8. Schinwald A, Donaldson K. Use of back-scatter electron signals to visualise cell/nanowires interactions in vitro and in vivo; frustrated phagocytosis of long fibres in macrophages and compartmentalisation in mesothelial cells in vivo. Particle and fibre toxicology. 2012;9:34; doi: 10.1186/1743-8977-9-34.
9. Murphy FA, Schinwald A, Poland CA, Donaldson K. The mechanism of pleural inflammation by long carbon nanotubes: interaction of long fibres with macrophages stimulates them to amplify pro-inflammatory responses in mesothelial cells. Particle and fibre toxicology. 2012;9:8; doi: 10.1186/1743-8977-9-8.
10. Donaldson K, Murphy FA, Duffin R, Poland CA. Asbestos, carbon nanotubes and the pleural mesothelium: a review of the hypothesis regarding the role of long fibre retention in the parietal pleura, inflammation and mesothelioma. 2010;7:5; doi: 10.1186/1743-8977-7-5.