Combining a lung microfluidic chip exposure model with transcriptomic analysis to evaluate the inflammation in BEAS-2B cells exposed to cigarette smoke

Abstract

BackgroundTypically, in vitro studies on the exposure of complex gaseous substances are performed in multi-well plate experiments by trapping and redissolving them, which could introduce potential bias into the results due to the use of inadequate trapping methods. Therefore, a more effective method is to expose complex gaseous substances in gaseous form online, such as using microfluidic chips in experiments. To address these challenges, we introduce a methodology that integrates a self-designed bionic-lung chip with transcriptome analysis to assess the impact of cigarette smoke (CS) exposure on changes in BEAS-2B cells cultured on-chip. ResultsAfter the microfluidic chip underwent online gas exposure, total RNA was extracted via in situ cell lysis, and RNA-Seq transcriptome analysis was conducted. And the RNA-Seq findings revealed the significant involvement of the MAPK signaling pathway associated with the inflammatory response in the cellular effects induced by CS exposure. Moreover, the validation of inflammatory response-related biomarkers through in situ fluorescence corroborated the outcomes of the transcriptome analysis. Besides, the experiment involving the inhibition of inflammation by DEX on the microfluidic chip provided additional confirmation of the previous experimental findings. SignificantIn this study, we present an analytical strategy that combines microfluidic-based CS in situ exposure method with RNA-Seq technology. This strategy offers an experimental scheme for in situ exposure to complex gases, transcriptome analysis, and in situ fluorescence detection. Through the integration of the comprehensiveness of transcriptome analysis with the chip's direct and intuitive in situ fluorescence detection with the stability and reliability of RT-PCR and Western blot experiments, we have successfully addressed the challenges associated with in vitro risk assessment for online exposure to complex gaseous substances.