Exposure to the antimicrobial chemical triclosan disrupts keratinocyte function and skin integrity in a model of reconstructed human epidermis
Updated June 8, 2023
February 2023
NIOSH Dataset RD-1054-2023-0
Introduction
Triclosan is an antimicrobial chemical incorporated into products that are applied to the skin of healthcare workers. Exposure to triclosan has previously been shown to be immunomodulatory and associated with allergic disease. Additionally, we have shown that exposure to triclosan dermally activates the NLRP3 inflammasome and disrupts the skin barrier integrity in mice. The skin is the largest organ in the body and plays an important role as a physical barrier and regulator of the immune system. Alterations in the barrier and immune regulatory functions of the skin have been demonstrated to increase the risk of sensitization and development of allergic disease. In this study, the impact of triclosan exposure on the skin barrier and keratinocyte function was investigated using a model of reconstructed human epidermis. The apical surface of reconstructed human epidermis was exposed to triclosan once for 6, 24, or 48 hours or daily for 5 consecutive days.
- Single Exposure Permeability Epidermal Thickness 2022-07-13 [XLS – 3 KB]
- Single Exposure LDH 2022-07-13 [XLS – 2 KB]
- Single Exposure Gene Expression 2022-07-13 [XLS – 40 KB]
- Single Exposure Cytokine Release 2022-07-13 [XLS – 9 KB]
- Repeated Exposure Permeability 2022-07-13 [XLS – 748 B]
- Repeated Exposure LDH 2022-07-13 [XLS – 3 KB]
- Repeated Exposure Gene Expression 2022-07-13 [XLS – 8 KB]
- Repeated Exposure Cytokine Release 2022-07-13 [XLS – 17 KB]
- Data Dictionary [PDF – 89 KB]
- Methods Collection [PDF – 139 KB]
Data Collection Methods
- Tissues
- EpiDerm (MatTek) tissues were incubated in 0.9 mL or 5 mL media/well
- Triclosan Exposures
- Tissues were exposed on the apical side to acetone or triclosan (0.05-0.2%) once for 6, 24, or 48 hours or once/day for 5 days
- Tissues were incubated at 37 °C with 5% CO2 during the exposure
- LDH Release Assay
- The LDH-Cytotoxicity Colorimetric Assay Kit II (BioVision) was used
- Absorbance (450 nm) was measured using a microplate spectrophotometer in duplicate for each sample
- Gene Expression Analysis
- Tissues were washed with 100 µL DPBS immediately prior to tissue collection
- Tissues were disrupted and homogenized in 700 µL QIAzol using a steel bead and TissueLyser II or with a pellet pestle homogenizer
- Total RNA was isolated from lysates using the miRNAeasy kit (Qiagen)
- RNA purity and yield were determined on a NanoDrop Spectrophotometer
- Reverse transcription was performed using a High-Capacity cDNA Reverse Transcription Kit (Applied Biosystems)
- TaqMan Fast Universal PCR Master Mix (Applied Biosystems), cDNA, and gene-specific primers (TaqMan Gene Expression Assays) were combined and real-time quantitative PCR was performed
- Genes evaluated include FLG, FLG2, IVL, LOR, TJP1, OCLN, KRT10, KRT14, CDH1, TSLP, S100A8, IL-1A, IL-1B, TNF, CXCL1, CXCL2, CXCL8
- Cytokine Release
- Evaluated with a custom human premixed multi-analyte kit (LXSAHM; R&D Systems) and analyzed on a MagPix (Luminex)
- Analytes measured: EGF, IL-1A, IL-6, IL-18, IL-33, S100A8, TNF-A, VEGF, GM-CSF, IL-1B, IL-8, IL-31, IL-36B, TGF-A, TSLP
- Permeability Assay
- Tissues were washed with 100 µL DPBS immediately prior to assay
- Tissues were exposed on the apical side to Lucifer Yellow CH dilithium salt (Sigma) and incubated at 37 °C with 5% CO2 for 2 hours
- Media was collected and fluorescence intensity was measured in triplicate
- Read on a microplate reader (excitation 428 nm, emission 536 nm)
- Histology
- Formalin-fixed paraffin-embedded tissues (5 µm) were mounted on slides and stained with hematoxylin and eosin
- Slides were brightfield imaged and epidermal thickness was measured
Publications Based on Dataset
Baur R, Kashon M, Lukomska E, Weatherly LM, Shane HL, Anderson SE. Exposure to the anti-microbial chemical triclosan disrupts keratinocyte function and skin integrity in a model of reconstructed human epidermis. J Immunotoxicol. 2023 Dec;20(1):1-11. doi: 10.1080/1547691X.2022.2148781.
Acknowledgements
This project was supported by the National Institute for Occupational Safety and Health (NIOSH). When a publication makes use of this dataset, acknowledgement of the development of the dataset should be attributed to the NIOSH Health Effects Laboratory Division.
We would also like to recognize the work of Rachel Baur, Michael Kashon, Ewa Lukomska, Lisa M. Weatherly, Hillary L. Shane, and Stacey E. Anderson.
Contact
NIOSH/Health Effects Laboratory Division
Allergy and Clinical Immunology Branch
(304) 285-6024