Pathogenesis of Otitis Media: The utilisation of fetal middle ear epithelial cells as a model for Non-typeable Haemophilus influenzae-induced acute otitis media

Otitis media (OM) is an inflammation of the middle ear (ME). 80% of children will experience a case of OM during their lifetime. The disease presents a significant health burden to both high and low-income countries. In high-income countries, OM is the leading cause for paediatric antibiotic prescription and administration of general anaesthesia and in low-income countries, lack of access to healthcare facilities leads to uncontrolled progression of disease, which commonly culminates in hearing loss, but can also lead to meningitis and death. This project aims to investigate 3 branches of the study of OM. 1) A better understanding of the ME embryonic development, will provide new insights into the ME cavity and its epithelial lining, and consequently the development of OM. To achieve this, immunohistochemistry staining for embryonic and epithelium differentiation markers of fetal ME tissue from 23CS to 17 post-conception weeks was performed. This part of the study presented significant challenges, particularly in the identification of very early embryonic tissues, however, new staining patterns were identified allowing novel insights into the early development of this part of a human embryo. 2) Finding sustainable alternatives to the current OM research tools is fundamental as currently, this is a significant limitation. A single human cell line is available (hMEEC-1), with most OM research being conducted in animal models. ME primary cells recovered from patients undergoing surgery also present limitations and few in vitro human 3D models have been established. Here, I report the recovery of ME epithelial cells from the fetal ME of early terminations of pregnancy (MEEC). MEEC present similar characteristics to hMEEC-1, while being primary cells. Furthermore, the cells naturally present extended lifespans when compared to previously published studies using primary cells recovered from paediatric patients for 3D model development. 3) OM is a disease of primarily bacterial sources, thus a strong knowledge of the interactions between otopathogens and the ME epithelium will inform decisions on future prevention and treatment methods. As NTHi emerges as a central otopathogen, replacing Pneumococcal spp., it is fundamental to understand the ability of this bacteria to inhabit the ME epithelium and the cellular responses to infection. This study reports, for the first time, the ability of NTHi to intracellularly invade ME epithelium cells. Furthermore, the response of MEEC to infection by NTHi is proinflammatory and follows a chronological similar pattern to that seen in in vivo responses, according to the literature. Thus, MEEC are representative of OM disease and may prove to be an extremely useful tool in the research of OM pathogenesis.