Role of hemoglobin and transferrin in multi-wall carbon nanotube-induced mesothelial injury and carcinogenesis

Our plain-English summary of Role of hemoglobin and transferrin in multi-wall carbon nanotube-induced mesothelial injury and carcinogenesis, by Y. Wang, Y. Okazaki, L. Shi, et al (Cancer Science, 2016).

The study

• Authors: Y. Wang, Y. Okazaki, L. Shi, H. Kohda, M. Tanaka, K. Taki, T. Nishioka, T. Hirayama, H. Nagasawa, Y. Yamashita, S. Toyokuni
• Journal: Cancer Science
• Published: 2016;107(3):250-7
• Cited by: 32 articles, as counted by NIH iCite
• Read it: PubMed · Publisher · Free full text

Abstract

Multi-wall carbon nanotubes (MWCNT) are a form of flexible fibrous nanomaterial with high electrical and thermal conductivity. However, 50-nm MWCNT in diameter causes malignant mesothelioma (MM) in rodents and, thus, the International Agency of Research on Cancer has designated them as a possible human carcinogen. Little is known about the molecular mechanism through which MWCNT causes MM. To elucidate the carcinogenic mechanisms of MWCNT in mesothelial cells, we used a variety of lysates to comprehensively identify proteins specifically adsorbed on pristine MWCNT of different diameters (50 nm, NT50; 100 nm, NT100; 150 nm, NT150; and 15 nm/tangled, NTtngl) using mass spectrometry. We identified >400 proteins, which included hemoglobin, histone, transferrin and various proteins associated with oxidative stress, among which we selected hemoglobin and transferrin for coating MWCNT to further evaluate cytotoxicity, wound healing, intracellular catalytic ferrous iron and oxidative stress in rat peritoneal mesothelial cells (RPMC). Cytotoxicity to RPMC was observed with pristine NT50 but not with NTtngl. Coating NT50 with hemoglobin or transferrin significantly aggravated cytotoxicity to RPMC, with an increase in cellular catalytic ferrous iron and DNA damage also observed. Knockdown of transferrin receptor with ferristatin II decreased not only NT50 uptake but also cellular catalytic ferrous iron. Our results suggest that adsorption of hemoglobin and transferrin on the surface of NT50 play a role in causing mesothelial iron overload, contributing to oxidative damage and possibly subsequent carcinogenesis in mesothelial cells. Uptake of NT50 at least partially depends on transferrin receptor 1. Modifications of NT50 surface may decrease this human risk.

© 2015 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.

In plain English

This laboratory study examined multi-wall carbon nanotubes (MWCNT), a man-made fibrous material; crocidolite asbestos served only as a comparison in protein-binding tests. The authors mixed four MWCNT types (about 50, 100 and 150 nm wide, plus a tangled 15 nm type) with rat organ extracts and identified more than 400 bound proteins, including hemoglobin and transferrin. They then coated 50 nm nanotubes (NT50) with these proteins and tested them on cultured rat peritoneal mesothelial cells. Compared with untreated cells, dead cells rose approximately 1.4-fold with uncoated NT50, 1.8-fold with hemoglobin-coated and 1.9-fold with transferrin-coated NT50. Pristine and hemoglobin- or transferrin-coated tangled nanotubes showed no cytotoxicity to these cells. Coated NT50 raised catalytic ferrous iron inside cells more than uncoated NT50 and significantly increased DNA damage measures; uncoated NT50 showed no statistically significant increase. Ferristatin II, which lowered transferrin receptor levels, reduced uptake of transferrin-coated NT50 and catalytic ferrous iron in those cells. The authors concluded that their results suggest NT50's binding of proteins, especially hemoglobin and transferrin, is a major mechanism in mesothelial damage followed by carcinogenesis; the experiments measured cell injury, not cancer.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: Laboratory (cell culture and protein-binding) experiment using rat organ extracts and rat mesothelial cells
• Who was studied: Lung, heart, liver and spleen extracts from eight 24-week-old male or female Fischer-344 rats, and rat peritoneal mesothelial cells (RPMC) grown in culture; Nagoya, Japan
• Exposure: Four multi-wall carbon nanotubes from Showa Denko: NT50, NT100, NT150 (50, 100 and 150 nm diameter) and NTtngl (15 nm, tangled). NT50 and NTtngl were tested either uncoated or coated with hemoglobin, holo-transferrin or lung lysate, at 10 micrograms per square centimetre. Crocidolite asbestos (UICC) was the positive control in the protein-binding assay.
• Main results:
  • In-solution digestion identified 321 proteins bound to NT50, 131 to NT100, 231 to NT150 and 287 to NTtngl; more than 400 proteins were identified in total.
  • Apart from histones, NT50 and NTtngl adsorbed significantly larger amounts of the proteins checked than NT100, NT150 or crocidolite. NT50 bound more transferrin than NTtngl.
  • Compared with untreated control cells, uncoated NT50 produced approximately 1.4-fold dead cells, hemoglobin-coated NT50 approximately 1.8-fold, transferrin-coated NT50 approximately 1.9-fold and lung-lysate-coated NT50 approximately 2.3-fold. Pristine, hemoglobin-coated and transferrin-coated NTtngl showed no cytotoxicity to RPMC.
  • In the wound-healing assay, uncoated and transferrin-coated NT50 decreased cell proliferation by approximately 10%. Hemoglobin-coated and lung-lysate-coated NT50 caused 24% and 31% decreases.
  • Hemoglobin-coated or transferrin-coated NT50 increased catalytic ferrous iron inside the cells more than uncoated NT50 did. The NTtngl preparations did not affect it.
  • In the comet assay, uncoated NT50 caused no statistically significant increase in tail length or tail moment. Coating with hemoglobin, transferrin or lung lysate significantly increased these measures.
  • Transferrin-coated NT50 led to approximately 20% more nanotube uptake than uncoated NT50. Ferristatin II, which reduced transferrin receptor 1 levels, significantly decreased uptake of transferrin-coated NT50 and lowered catalytic ferrous iron in those cells.
• The authors' conclusion: The authors concluded that their results suggest NT50's binding of proteins, especially hemoglobin and transferrin, is a major mechanism in mesothelial damage followed by carcinogenesis. They suggested that chemically modifying carbon nanotubes to avoid this binding might decrease human risk.
• Limitations the authors state:
  • The authors stated that further studies are needed to identify and quantify the precise DNA lesions.
  • The authors said the large change in cellular iron after a modest drop in uptake may relate to iron storage and export, which needs further investigation.
  • The authors noted that many more proteins that bind to MWCNT have not yet been evaluated.

Topics

The National Library of Medicine indexes this study under subjects our site covers: Mesothelioma.

Medical terms in this study, explained in our Lexicon: Carcinogenesis, Mesothelioma, Carcinogenic, Mesothelial Cells, Oxidative Stress.

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• Dose-dependent mesothelioma induction by intraperitoneal administration of multi-wall carbon nanotubes in p53 heterozygous mice — A. Takagi, A. Hirose, et al. (2012)
• Intraperitoneal administration of tangled multiwalled carbon nanotubes of 15 nm in diameter does not induce mesothelial carcinogenesis in rats — H. Nagai, Y. Okazaki, et al. (2013)
• Osteopontin mRNA expression by rat mesothelial cells exposed to multi-walled carbon nanotubes as a potential biomarker of chronic neoplastic transformation in vitro — S. Sridharan, A. Taylor-Just, et al. (2021)
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