Mixture-Dependent Toxicity of E-Cigarette Constituents: Cytotoxicity, Inflammation, and DNA-Repair-Associated Protein Responses in Bronchial Epithelial Cells.
- 1School of Health Sciences, Purdue University, West Lafayette, IN 47907, USA.
Abstract
Electronic cigarette (e-cigarette) aerosols contain complex mixtures of flavoring chemicals, synthetic cooling agents, and metals derived from device components. The biological effects of these constituents in combination remain incompletely characterized. We evaluated selected flavoring chemicals and nickel oxide (NiO), individually and in combination, in BEAS-2B human bronchial epithelial cells. Cells were exposed for 24 h to vanillin, WS23, ethyl vanillin, or p-anisaldehyde over an experimental concentration range of 10-1000 µM. Vanillin-WS23 and ethyl vanillin-p-anisaldehyde mixtures were evaluated at a fixed total concentration of 1000 µM over multiple component ratios. Separate experiments evaluated 100 µM ethyl vanillin, 50 µg/cm² NiO, and their combination. Vanillin, WS23, ethyl vanillin, and p-anisaldehyde reduced viability-associated metabolic activity and cellular ATP in a concentration-dependent manner. The lowest concentrations associated with significant reductions in the MTT endpoint were 200 µM for vanillin, 50 µM for WS23, 300 µM for ethyl vanillin, and 200 µM for p-anisaldehyde. Chou-Talalay analysis identified synergistic interactions between vanillin and WS23 (combination index [CI] < 1) and antagonistic interactions between ethyl vanillin and p-anisaldehyde (CI > 1). Ethyl vanillin and NiO produced no statistically significant decrease in viability but increased inflammatory cytokines in conditioned medium. Both IL-6 and IL-8 levels were significantly elevated with NiO and the combined ethyl vanillin-NiO exposure. Total H2AX abundance did not differ significantly among groups at 24 h. NEIL2 abundance showed increasing trends following NiO alone (p = 0.0518) and combined ethyl vanillin-NiO exposure (p = 0.0760). PNKP increased significantly after ethyl vanillin (p = 0.0261) and combined ethyl vanillin-NiO (p = 0.0041) treatments. These findings show mixture-dependent biological responses that differed from the effects observed with individual constituents. Flavoring chemical mixtures produced either synergistic or antagonistic effects, and ethyl vanillin-NiO exposure produced inflammatory and DNA-repair-associated protein responses under conditions that did not significantly reduce cell viability. By demonstrating non-additive flavoring chemical interactions together with inflammatory and DNA-repair-associated protein responses to chemical-metal exposure, this study evaluated e-cigarette toxicity beyond single-constituent hazard characterization. The NEIL2 and PNKP findings support altered base-excision repair (BER) pathways but do not establish oxidative DNA lesions or increased DNA repair activity.
Related
- MAGI3 Acts as an Intestinal Epithelial Gatekeeper in Inflammation‐Associated Colorectal Cancer by Suppressing the STAT3‐CCL2 Axis and TAM‐Mediated Immunosuppression
- The Critical Clock: Time-Dependent Inflammation, Catabolism and Immunosuppression Syndrome Is Associated with ICU Mortality
- Mechanism Analysis of Chronic Inflammation in Polycystic Ovary Syndrome and Targeted Therapeutic Strategies Based on Bioactive Constituents of Traditional Chinese Medicine.
- Assessing the Skin Sensitizing Potential of Constituents and Leachables from Wearable Devices Using GARDskin and Associated Applications
- PARG inhibition elicits cytotoxicity by sequestering XRCC1 and other PAR-binding proteins into PAR-dependent nuclear condensates
BibTeX
@article{kabir2026mixturedependent,
title = {{Mixture-Dependent Toxicity of E-Cigarette Constituents: Cytotoxicity, Inflammation, and DNA-Repair-Associated Protein Responses in Bronchial Epithelial Cells.}},
author = {Ehsanul Kabir and Joshua Furst and Thivanka Muthumalage},
journal = {Toxics},
year = {2026},
month = oct,
doi = {10.3390/toxics14100910},
url = {https://europepmc.org/article/MED/42931697},
note = {CC BY},
}
RIS
TY - JOUR
TI - Mixture-Dependent Toxicity of E-Cigarette Constituents: Cytotoxicity, Inflammation, and DNA-Repair-Associated Protein Responses in Bronchial Epithelial Cells.
AU - Ehsanul Kabir
AU - Joshua Furst
AU - Thivanka Muthumalage
PY - 2026
DA - 2026/10/15
JO - Toxics
DO - 10.3390/toxics14100910
UR - https://europepmc.org/article/MED/42931697
AB - Electronic cigarette (e-cigarette) aerosols contain complex mixtures of flavoring chemicals, synthetic cooling agents, and metals derived from device components. The biological effects of these constituents in combination remain incompletely characterized. We evaluated selected flavoring chemicals and nickel oxide (NiO), individually and in combination, in BEAS-2B human bronchial epithelial cells. Cells were exposed for 24 h to vanillin, WS23, ethyl vanillin, or p-anisaldehyde over an experimental concentration range of 10-1000 µM. Vanillin-WS23 and ethyl vanillin-p-anisaldehyde mixtures were evaluated at a fixed total concentration of 1000 µM over multiple component ratios. Separate experiments evaluated 100 µM ethyl vanillin, 50 µg/cm² NiO, and their combination. Vanillin, WS23, ethyl vanillin, and p-anisaldehyde reduced viability-associated metabolic activity and cellular ATP in a concentration-dependent manner. The lowest concentrations associated with significant reductions in the MTT endpoint were 200 µM for vanillin, 50 µM for WS23, 300 µM for ethyl vanillin, and 200 µM for p-anisaldehyde. Chou-Talalay analysis identified synergistic interactions between vanillin and WS23 (combination index [CI] < 1) and antagonistic interactions between ethyl vanillin and p-anisaldehyde (CI > 1). Ethyl vanillin and NiO produced no statistically significant decrease in viability but increased inflammatory cytokines in conditioned medium. Both IL-6 and IL-8 levels were significantly elevated with NiO and the combined ethyl vanillin-NiO exposure. Total H2AX abundance did not differ significantly among groups at 24 h. NEIL2 abundance showed increasing trends following NiO alone (p = 0.0518) and combined ethyl vanillin-NiO exposure (p = 0.0760). PNKP increased significantly after ethyl vanillin (p = 0.0261) and combined ethyl vanillin-NiO (p = 0.0041) treatments. These findings show mixture-dependent biological responses that differed from the effects observed with individual constituents. Flavoring chemical mixtures produced either synergistic or antagonistic effects, and ethyl vanillin-NiO exposure produced inflammatory and DNA-repair-associated protein responses under conditions that did not significantly reduce cell viability. By demonstrating non-additive flavoring chemical interactions together with inflammatory and DNA-repair-associated protein responses to chemical-metal exposure, this study evaluated e-cigarette toxicity beyond single-constituent hazard characterization. The NEIL2 and PNKP findings support altered base-excision repair (BER) pathways but do not establish oxidative DNA lesions or increased DNA repair activity.
SN - 2305-6304
ER -