Comparative analysis of N/TERT-1 and N/TERT-2G keratinocyte responses to oxidative stress and immune challenges

Abstract

The responses of N/TERT-1 and N/TERT-2G keratinocyte cell lines to oxidative stress and immune challenges were investigated to assess their suitability for dermatological testing. The cell lines were exposed to various stimuli, including PAMPs, DAMPs, H₂O₂, and menadione, to assess cytokine production, oxidative stress markers, cell viability, apoptosis, and membrane integrity. IL-1α, IL-6, IL-8, TNF-α, and TGF-β levels significantly increased in N/TERT-1 cells following exposure to LPS, while N/TERT-2G cells remained unaffected. Both cell lines showed increased production of IL-1α, IL-1β, TNF-α, IL-6, and IL-8 in response to dsDNA and LMW and HMW Poly I:C, although TGF-β significantly decreased only in N/TERT-1 cells. In response to H₂O₂, a dose-dependent increase in cytokine levels was observed in N/TERT-2G, whereas N/TERT-1 did not exhibit a clear dose-dependent response. Markers of oxidative stress, including SOD and GSH, displayed similar patterns in both cell lines, with N/TERT-2G showing slightly higher sensitivity. Lipid peroxidation and mitochondrial membrane potential fluctuations were more pronounced in N/TERT-2G, suggesting greater oxidative stress sensitivity. The baseline GSH levels were higher in N/TERT-1 cells, which may contribute towards the enhanced resilience to oxidative stress. Despite decreased viability in MTT assays following H₂O₂ exposure, the lack of significant changes in cleaved Caspase-3 levels indicated that apoptosis was not the primary mechanism of cell death. These findings highlight the distinct characteristics of N/TERT-1 and N/TERT-2G cells, with N/TERT-1 showing higher baseline resilience to oxidative stress and N/TERT-2G displaying greater sensitivity, particularly to H₂O₂. The study underscores the importance of selecting the appropriate cell line for specific research applications in skin biology and disease modelling, considering the differences in their responses to oxidative and immune challenges.

Keywords: Hydrogen peroxide; Inflammatory markers; Membrane integrity; N/TERT-1 & N/TERT-2G keratinocytes; Oxidative stress.