The identification of tyrosine-nitrated human serum albumin in airborne particulate matter from Japan

https://doi.org/10.1016/j.apr.2021.03.013Get rights and content

Highlights

  • Human serum albumin was identified in atmospheric particulate matter in Japan.

  • Tyrosine nitration was detected in human serum albumin of PM.

  • The amount of albumin contained in PM2.5 varies by region.

Abstract

Atomospheric particulate matter (PM) has been accepted as a risk to human health worldwide. PM contains a variety of components, but it is not clear which components are harmful to human health. For example, there have been many reports on metals, but there have been only a few studies on proteins. Based on an analysis of serum antibodies in a mouse model of PM-induced asthma reported previously, we have been conducting research focusing on the proteins contained in PM. In the present study, we found for the first time that human serum albumin (HSA) is bound to PM. HSA was detected in total suspended particulate matter (TSP) and PM2.5, from various regions in Japan, during all seasons. The presence of HSA in PM was confirmed by proteomics analysis, amino acid analysis, immunoblotting and BD140, which emits fluorescence when bound to a specific HSA site. Moreover, we discovered that TSP-binding HSA was tyrosine-nitrated, one of a chemical modification via environmental atmospheric pollutants using high performance liquid chromatography-electrochemical detection. It was concluded that although the origin of HSA in PM was unclear, HSA is present in the airborne PM of Japan, partially modified for tyrosine nitration.

Introduction

It has been suggested that daily exposure to atomospheric particulate matter (PM2.5, PM10) may cause premature mortality, cardiovascular mortality, and respiratory mortality (Bruewer et al., 2005; Liu et al., 2019). Although the complexity of how PMs enter the human body through the respiratory system is becoming clearer, but the molecular mechanism of their toxicity is not fully understood (Horie et al., 2012). Generally, the factors contributing to the health effects and toxicity of PM are thought to be related to the contaminants contained in the particles.

PM samples in urban areas have been chemically characterized for inorganic ions, total carbon (elemental or black carbon), elements (mineral dust and trace elements from anthropogenic sources), polycyclic aromatic hydrocarbons (PAHs), and biological products (endotoxin) (Brook et al., 2010). In the atmosphere, cellular debris and various proteins form bio-aerosol particles, or PMs, and react with organic or inorganic materials (Jaenicke et al., 2005). The bio-aerosol particles in atmospheric PM include viruses, pollen, fungal spores, bacteria, and debris from vertebrates (e.g., humans), and other biota (e.g., plants and insects) (Castillo et al., 2012). Among these bio-aerosol particles, various proteins such as the debris from keratinocytes have been identified (Clark et al., 1973; Fox et al., 2008). In fact, proteins may account for up to ~5% or 0.5–2% of urban PM (Franze et al., 2005; Abe et al., 2006), and their considerably high protein content in PM has been found to have physicochemical effects on atmospheric particles. It has previously been demonstrated that the protein in PM proteins may contribute to allergic airway inflammation in mice (Ogino et al., 2014a, 2014b, 2018). Additionally, PM2.5 easily induces allergic airway inflammation in the presence of HSA, a non-allergen, illustrating another of its action mechanisms other than as an adjuvant (Nagaoka et al., 2019). Chemically modified tyrosine-nitrated proteins in atmospheric PM have also been measured, and the results suggest that protein reactions with environmental pollutants such as ozone and NOx (Ito et al., 2018), and the generation of 3-nitrotyrosine (3-NT) in the tyrosine residues of proteins is associated with allergies (Gruijthuijsen et al., 2006).

This study was initiated by the unexpected detection of an antibody against PM2.5-bound protein in experimental asthmatic mice (Ogino et al., 2014b). We identified a PM protein using immunoglobulin G (IgG) serum from experimentally asthmatic mice, and evaluated the biochemical characteristics of the identified proteins in PM.

Section snippets

Sources of airborne PM

The total suspended particles (TSP) were collected as previously reported (Ogino et al., 2014a). These samples were collected from seven locations in Japan (Table S1). PM2.5 was collected from low-volume air samplers (Okayama City and Fukue Island of Nagasaki Prefecture) (Ogino et al., 2014b), or cyclone system (Okayama City) (Ogino et al., 2017). TSP and PM2.5, on the filters and cyclone particles of PM2.5, were handled by gloved hands and then frozen at −30 °C. For the TSP and PM2.5, filters

Immunoreactive proteins against PM2.5 in mice serum with PM2.5-induced allergic airway inflammation

A PM2.5 filter was randomly selected for each month from 2010 to 2011, and the precipitate fraction of PM2.5 following elution was analyzed for western blots using the serum from mice for antiserum, which demonstrated acute airway inflammation (Ogino et al., 2014b). The PM content (30 μg) was electrophoresed by SDS-PAGE, and the immunoblot is shown in Fig. 1A. There was a single immunoreactive band near the molecular weight of 50 kDa.

Proteomic analysis and protein sequence analysis

Immunoprecipitation was performed with TSP proteins and

Discussion and conclusion

There have been a few reports on the presence of proteins in atmospheric PM and aerosols (Clark et al., 1973; Castillo et al., 2012), and plant and bacterial proteins have also reported as bio-aerosols (Liu F et al., 2016). As for human sources, skin fragments and human K10 epithelial keratin have been detected in atmospheric PM (Fox et al., 2008), but there was no previous evidence for the presence of HSA in atmospheric PM.

We were convinced of the presence of candidate HSA by proteomic and

Authors' contributions

Noriyoshi Ogino; Investigation, Writing – original draft. Keiki Ogino; Conceptualization, Data curation, Formal analysis, Funding acquisition, Project administration, Writing – review & editing. Masamitsu Eitoku; Supervision. Narufumi Suganuma; Project administration. Kenjiro Nagaoka; Methodology. Yasuhiro Kuramitsu; Methodology.

Declaration of competing interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Acknowledgements

We thank Emeritus Prof. Kazuyuki Nakamura of Yamaguchi University for his help with the proteomic analysis. We thank Prof. Hiroyuki Nakamura of Kanazawa University, Emeritus Prof. Hiroyuki Katsunuma of Tokyo Medical University, Specially Appointed Prof. Nakaji of Hirosaki University, and Assoc. Prof. Yasushi Obase of Nagasaki University for their help with the TSP/PM2.5 sampling in Kanazawa City, Tokyo, Nagasaki City, and Fukue Island. We thank Dr. Masayuki Kubo of the US Environmental

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