The role of innate immunity in obesity-induced tissue inflammation

Abstract: The prevalence of obesity is rapidly increasing globally, which causes an increase in associated diseases such as type 2 diabetes, cardiovascular disease and non-alcoholic fatty liver disease (NAFLD). In addition to the detrimental effect of obesity on several organs such as the liver and adipose tissue, obesity also has a profound effect on the immune system. The immune system consists of a complex interplay of different immune cells and other factors interacting with each other and the surrounding tissue in order to protect the body from both external (e.g., infectious) and internal (e.g., cancer) threats. As part of the innate immune system, natural killer (NK) cells play an important part in the defense against viral infections as well as malignancies. Another innate immune cell population, macrophages, are common immune cells that can perform a multitude of different functions, ranging from initiating immune responses, act as scavengers and contribute to wound healing. Recent studies have suggested that obesity, and the chronic low-grade inflammation that accompanies it, disturbs the innate immune system which could lead to impairments in its protective functions and thereby contribute to disease development. However, the complexity of the heterogeneous functions and forms of innate immunity are only partially understood. The overall aim of this thesis is to add knowledge on how innate immunity is affected by obesity in order to improve our understanding and, in the future, treatment of obesity-related diseases. In study I, the NK cell population in patients with the hepatic presentation of the metabolic syndrome nonalcoholic fatty liver disease (NAFLD) was examined. The receptor repertoire and function of circulating NK cells from patients with non-alcoholic fatty liver (NAFL), nonalcoholic steatohepatitis (NASH) and healthy controls were compared. In addition, tissue-derived NK cells derived from adipose tissue and liver were assessed. The results show that while some receptors, such as NKG2D, had increased on NK cells in NASH compared to healthy controls, the NK cell population remains mainly unaltered by NAFLD. This is of interest as NK cells are believed to possess anti-fibrotic properties, whilst they are also important mediators of innate immunity in other liver diseases, such as hepatitis C. In addition, analysis of adipose tissue derived NK cells demonstrated a population of cells with tissue residency markers such as CD49 and CD69. While further explored since then, this was at the time one of the first reports of tissue resident NK cells in adipose tissue. However, future studies are warranted to increase the knowledge of these cells and how they are related to human disease and obesity development. In study II the surface proteome of adipose tissue macrophages (ATMs) was examined to increase the knowledge of this heterogeneous population of immune cells. Adipose tissue macrophages are key players in adipose tissue inflammation. Since the composition and localization of adipose tissue depots in humans affect the risk of metabolic complications, macrophages derived from different anatomical niches such as visceral and subcutaneous adipose tissue were studied. The results include novel surface proteins associated with pro-and anti-inflammatory phenotypes of adipose tissue macrophages, as well as considerable differences between macrophages derived from subcutaneous adipose tissue (SAT) and visceral adipose tissue (VAT). Three novel surface markers (CD85a, CD48 and CD371) were identified on adipose tissue derived M1-like macrophages and CD51 and α9β1 on M2-macrophages. Finally, the presence of insulin resistance was linked to the relative increase in pro-inflammatory macrophages in both SAT and VAT, supporting an association between adipose tissue inflammation and insulin resistance. In the final project, study III, the immune proteome in serum of NASH patients was measured to find a signature that separates NASH patients from those without ongoing liver inflammation (NAFL) in order to improve diagnostics. Thus, serum was collected from NAFL-patients, NASH-patients as well as from healthy, lean controls. In total, 92 inflammatory serum proteins were measured using a proximity extension assay (PEA). The results revealed a NASH-specific inflammatory imprint with 13 serum proteins which, independently of comorbidities and fibrosis stage, differed between NAFL and NASH patients. Out of these proteins, sulfotransferase A1 (ST1A1) and stem cell factor (SCF) were found in higher levels in NASH-patients compared to NAFL and healthy controls. Pathway analysis of deregulated proteins revealed dysregulation in several pathways in NASH, including NFκβ and IL-18 signaling. Analysis of IL-18, EN-RAGE and ST1A1 in a pre-existing single cell transcription data set derived from liver cells suggested involvement of hepatic macrophages and periportal hepatocytes in NASH pathogenesis. Finally, the differentially expressed proteins could also be used to subgroup NASH patients into different clusters. Taken together, study III increases our understanding of the circulating serum proteome in NASH, suggesting a way to distinguish NASH-patients from NAFL and provides insight into possible subgroups and pathogenesis. However, these results need further validation in larger cohorts as well as further analysis both over time and regarding disease progression. In conclusion, the results presented in this thesis provide novel data on the effect obesity has on key players of innate immunity. While there is an inflammatory imprint in the serum proteome of NASH-patients, circulating NK cells retain their phenotype compared to controls. While tissue-resident NK cells previously have been identified in the liver, we provide one of the early reports of NK cells in visceral adipose tissue that display tissue residency markers. In addition, we demonstrate significant heterogeneity within the macrophage population in adipose tissue and we compare ATMs from subcutaneous and visceral adipose tissue, linking this to the development of insulin resistance. In conclusion, we have added a couple of pieces to the jigsaw puzzle that is human immunology, but still future studies are warranted to further elucidate and validate our results and to in-depth investigate their effect on human disease development.

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