The adipose tissue and liver in human non-alcoholic fatty liver disease: Characterization on a single-cell level

Non-alcoholic fatty liver disease (NAFLD) is the most common cause of chronic liver disease, that encompasses a disease spectrum ranging from simple steatosis, non-alcoholic steatohepatitis (NASH), cirrhosis, to the development of hepatocellular carcinoma (HCC). Patients with simple steatosis (an accumulation of lipids in hepatocytes) are largely asymptomatic and only a fraction of patients will develop the more severe NASH, or NASH with cirrhosis. We were able to describe the various hepatic cell subtypes and their dysfunctions exacerbating NAFLD utilizing single-nucleus RNA sequencing. Here we observed that the hepatocyte lipid metabolism was modulated, likely promoting lipotoxicity and cell death in NASH. However, also macrophages are affected during NAFLD. Macrophages play a central role in initiation and maintenance of inflammation, as well as fibrogenesis through the activation of hepatic stellate cells. To exert these diverse functions they are naturally very versatile cells that can adapt their activation status according to cues from their microenvironment (e.g. lipotoxic molecules, cytokines, danger signals, necrotic cell debris). We observed that lipid-associated macrophages (LAMs) were elevated in NASH livers, while Kupffer cells (KCs) were decreased. KCs are important for the homeostasis in the liver, and a loss of KCs could aggravate NAFLD progression. Interestingly, during NASH these LAMs not only were functionally involved in lipid clearance but also inflammatory pathways, events that may exacerbate NASH. Furthermore, it has been proposed that the adipose tissue is a major contributor to the development of NAFLD. Hereby, adipose tissue inflammation is linked to the development of NASH, and surgical adipose tissue removal or adipose tissue macrophage (ATM) ablation ameliorated NASH formation. Therefore we utilized single-cell RNA sequencing, to demonstrate that the visceral adipose tissue comprises of multiple macrophage subtypes that contribute to the adipose tissue homeostasis. However, we show that the depletion and dysfunction of a metabolically active ATM and resident vasculature-associated ATM, was associated with a breach of adipose tissue vascular integrity during NAFLD. This could result in extravasation of lipotoxic and inflammatory molecules to the liver, thereby driving NAFLD progression. The findings presented highlight that NAFLD is a systemic disease, and while the liver is severely dysfunctional during NAFLD, also the adipose tissue needs to be considered.