The Role of Mesenchymal Cells in the Regeneration of Salivary Gland Tissue

Abstract

Primary Sjogren’s Syndrome (PSS) patients will suffer a variable degree of damage to their salivary glands, caused by lesions of immune infiltrates into the tissue surrounding duct structures. Some patients present with extensive lesions that destroy any residual secretory acinar tissue, leading to hyposalivation. This damage, and reduced salivation, can lead to deleterious effects on the oral health, such as increased risk of caries and periodontitis, and will impact the general quality of life of the patients and can increase their risk of developing other, more serious conditions. Previously, there has been limited knowledge and characterisation of how different cell populations are impacted by PSS. Therefore, this project developed clear characterisation of labial salivary gland tissue from patients presenting with PSS compared to non-PSS patient samples. It was shown that regions that were not directly associated to lesions present with the same phenotypical profile as observed in non-PSS patients, indicating a very limited focal impact of the disorder. These regions stained positively for E-cadherin and Vimentin in patterns that were comparable to non-PSS patient samples. Simultaneously, the directly affected lesion regions presented with complete absence of any healthy salivary gland cell markers. Peri-lesion tissue presented with subtle changes to marker expression; here we could identify changes to mesenchymal cell markers such as PDGFRb and CD29 between the PSS and non-PSS tissue. The established immunofluorescent markers could further be used in later experimentations to identify each cell population, particularly when developing cell cultures and organoid models. 23 In order to investigate the impact of PSS on the ability of cells to proliferate and perform a rudimentary regenerative function, the group developed cell cultures that were representative of epithelial and mesenchymal cell types and recombined them to form organoid models. These models were first developed using mouse derived cell cultures before being adapted for use with human derived cells. Using these organoid models, it was possible to demonstrate that mesenchymal cells isolated from PSS positive patients had a reduced ability to direct the regenerative capacity of the epithelial spheroid structures, resulting in smaller, less complex, and less functionally advanced organoids than those developed using non-PSS derived cells. During this study, standard techniques were used such as quantitative real-time PCR of relative mRNA expression levels, immunofluorescent staining, and laser capture microdissection (LCM), whilst a novel organoid culture technique was optimised. Since these models have been established, it is thought that they could be adapted to investigate other disease or environmental impact on the salivary gland tissue, with some preliminary studies attempting to investigate the impact of x-ray irradiation exposure on the tissue. Furthermore, the technique could possibly be extrapolated to other tissue types to aid the understanding of the effect of other autoimmune conditions on their target tissues. In conclusion, this project shows mesenchymal cells appear to play an important and necessary role within tissue regeneration models. There is a significant change to cell fate determination pathways in PSS positive patient biopsies, and these mechanistic changes now need to be studied further within the constraints of the organoid model that has been developed.