Development and characterisation of air-liquid interface exposure systems for cell and tissue cultures

Majeed, Shoaib

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Majeed, Shoaib (2025) Development and characterisation of air-liquid interface exposure systems for cell and tissue cultures. Doctoral thesis, London Metropolitian University.

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Abstract

Introduction:
Monitoring the delivery of inhaled materials to the respiratory tract requires consideration of particle deposition and gas dissolution within the complex structure of the respiratory system. Until the last decade or so, methodology relied heavily on animal models or limited, submerged cell culture models that did not capture air-liquid interactions, identifying the need for more comprehensive in vitro aerosol exposure systems to examine how aerosolised substances impact respiratory tissues, cellular responses, toxicity, and overall health. Aerosols possess opposing qualities, serving both as hazards and therapeutic tools, which underscores the critical necessity for rigorous safety testing. This thesis describes the development and characterisation of in vitro aerosol and smoke exposure systems for future testing, the Vitrocell® 24/48 and the InHALES systems.

Results:
The Vitrocell® 24/48 system was developed to simulate physiological conditions at the air-liquid interface using a stagnation flow setup with controlled temperature (37°C). Comprehensive characterisation revealed excellent reproducibility with a coefficient of variation of 12.2% across inserts and 15.6% overall between experimental runs. Quartz crystal microbalance measurements confirmed linear particle deposition (R² > 0.95) across dilution ranges from 7% to 69% cigarette smoke concentration. Chemical analysis revealed a consistent delivery of key smoke constituents, with eight carbonyls and nicotine exhibiting a strong linear correlation (R² > 0.96) with smoke concentration. Biological testing using A549 and BEAS-2B cells also showed concentration-dependent toxicity. Additionally, the InHALES system is described, created to offer a more precise exposure platform in which the inhalation-exhalation cycle can be modelled and controlled. The research extends to practical applications, including the effects of cigarette smoke and aerosols on dental resin composites, where cigarette smoke caused 10-fold greater surface discolouration on tooth enamel compared to heated tobacco product aerosols after 14 days of exposure. Studies on 3D bronchial and nasal tissue cultures have revealed significant reductions in ciliary beating frequency following exposure to cigarette smoke, providing quantitative evidence of respiratory epithelial dysfunction. The research concludes with the application of the developed methodologies to address contemporary challenges, including the aerosol filtration testing of over 300 fabrics for the development of reusable masks, and the aerosol administration of chloroquine, which contributes valuable insights for public health improvements, especially during the COVID-19 pandemic.

Conclusion:
The Vitrocell and InHALES platforms discussed in this thesis also bypass ethical concerns associated with traditional animal testing and propel biomedical research by offering a more relevant and compassionate approach. While much attention has been focused on cigarette smoke and tobacco-related aerosols, these in vitro systems have broader implications in inhalation science, extending to examining the effects of wood combustion and evaluating the impact of occupational and environmental aerosols on human health through inhalation. By bridging laboratory research with real-world applications during the COVID-19 pandemic, this research demonstrates how advanced exposure technologies can rapidly pivot to address urgent public health challenges, ultimately providing a more humane, accurate, and versatile approach to understanding the complex interactions between inhaled substances and human respiratory health.

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