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Blue mussel proteins rescue polystyrene nanoparticles-induced

cellular toxicity in human dermal fibroblasts

through upregulation of the Nrf2-cytoprotective pathway

Choi, Yun-Sik , Goh Eun Hye , and Oh, Yunok *
3
2
1

Convergence Research Center for Smart Healthcare, Department of Pharmacology, Kyungsung University, Busan, Korea
1

Department of Pharmaceutical Science and Technology, Kyungsung University, Busan, Korea
2

Convergence Research Center for Smart Healthcare, Kyungsung University, Busan, Korea
3

Abstract

With the increased use of plastics, the potential toxicological effects of micro- and nanoplastics on human health have become a global concern. Health and beauty products

containing microbeads for dermal applications could lead to dermal exposure. But, there is little research that has specifically looked at the toxicity of nanosplastics (NPs) to

skin penetration. In this study, we investigated the cellular toxicity and skin penetration of polystyrene-amine nanoplastics using Human Dermal Fibroblasts (HDFs). Next, we

evaluated cytoprotective effects against NPs toxicity in HDFs using blue mussel proteins-derived antioxidant (BMP). BMP showed excellent antioxidant activity in a dose-

dependent manner, assessed by DPPH assay. Immunostaining and western blot analysis revealed that BMP pre-treatment significantly prevented NPs cellular toxicity by

inhibiting of mitochondrial ROS generation and caspase-3 activation, and up-regulation of Nrf-2 expression. These results suggest that BMP treatment attenuated cytotoxicity of

NPs through Nrf2-mediated antioxidant defense mechanism. Therefore, BMP could be considered for use as an antioxidant functional health food against NPs-induced toxicity.

Introduction Results

• Membrane damage

• Oxidative stress A B

• Immune response

Skin contact

of plastic particles Skin penetration
BMP (< 1 kDa)

Cytoprotective effects of BMP on PS-NPs-induced cellular toxicity in human dermal fibroblasts

Fig. 1. Schematic illustration showing the pathway of human skin exposure to nanoplastics. Fig. 5. The effects of BMP on mitochondrial membrane potential (MMP) loss. (A) MMP disruption

identified by Mito Tracker TM Red kit under a fluorescence microscope, and (B) graphical

Results quantification showing decrease of MMP level on HDF after 24 of treatment with NPs and BMP at

A B C different concentrations. * p < 0.05 vs. control, # p < 0.05 vs NPs only.

BMP concentration (g/mL)

Fig. 2. Antioxidant abilities of blue mussel hydrolysates with various proteolytic enzymes

determined by (A) DPPH scavenging assay and (B) ORAC assay. (C) DPPH scavenging

assay of three concentrations of blue mussel hydrolysate by pepsin (BMP). a–d Different Fig. 6. Immunofluorescence staining of cleaved caspase-3 in HDF cells. BMP was treated with

letters on the bar charts indicated significant differences (p < 0.05). * p < 0.05 vs. control. various concentration (10, 25, and 50 g/mL) with 0.01% NPs in HDF cells.

A B

A B

Fig. 3. Cytoprotective effects on human dermal fibroblasts (HDF) after of treatment with (A)

NPs only and (B) NPs with BMP at different concentrations. The cells were pre-treated with

various concentration of BMP for 1 h followed by 0.0.1% NPs exposure and 24 h incubation. Fig. 7. Representative western blotting of Nrf-2, and (B) the relative expression level of cleaved

* p < 0.05 vs. control, # p < 0.05 vs NPs only. BMP was pre-treated before 1 h followed Nrf-2 protein level in NPs-induced cellular damage. Cells were pretreated with BMP for 1 h

A followed by 0.0.1% NPs exposure and 24 h incubation.

Summary

• In this study, we demonstrated the anti-atherosclerotic effect of blue mussel hydrolysates by

pepsin (BMP).

B C • The antioxidant effect by BMP was attributed to the promotion of intracellular antioxidant

defense capacity, such as the inhibition of ROS generation, MMP disruption, caspase-3 activation,

and upregulation of Nrf-2 expression in NPs-induced HDF damage.

• Our findings provide new insight of potential use of marine proteins and BMP might considered

natural antioxidant ingredients for antioxidant functional health food against NPs-induced cellular

toxicity.

Reference

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