Modulation of Kupffer cell activity : physio-pathological consequences on hepatic metabolism

Classically, the maintenance and control of liver homeostasis are assigned to the metabolic activity of parenchymal cells. However, recent evidence highlights complex and tightly regulated interactions between hepatocytes and other intra-hepatic cells. Such a viewpoint is supported by the findings that nonparenchymal sinusoidal cells have the capacity to synthesize various molecules, which can have profound influences on hepatocytes. Kupffer cells -the resident macrophages of the liver- are able to release a tremendous array of mediators upon inflammatory conditions, such as infection, and their role in innate immunity is well described in the literature. However, the impact of these Kupffer cell-derived mediators on liver homeostasis is unknown. The aim of our study was to investigate the physiological involvement of Kupffer cells in the regulation of hepatic metabolism. Therefore, it was first necessary to validate the use of a compound able to selectively deplete Kupffer cells. Since gadolinium chloride (GdCl3) has been extensively used as a Kupffer cell inhibitor to study their role under various pathological situations, we analysed several parameters assessing the presence and the activities of Kupffer cells after GdCl3 administration. We confirmed that GdCl3 injection to rats eliminated ED2-positive Kupffer cells and strongly decreased both their phagocytic and peroxidase activities. In addition, secretion of key Kupffer cell-derived mediators was assessed by using an original in vitro model allowing to maintain intact hepatic architecture and cellular heterogeneity. We demonstrated that precision-cut liver slices (PCLS) obtained from GdCl3-treated rats released lower amounts of cytokine (TNF-alpha), prostaglandin (PGE2) and reactive intermediate (nitric oxide), independently of inflammatory stimulus. We proposed to use GdCl3 prior to PCLS preparation in order to investigate the role of Kupffer cells in the control of hepatic metabolism. Among various metabolic functions of the liver, we focused, in particular, on paracetamol and lipid metabolism as example of drug and intermediary metabolism, respectively. Our results suggest that the presence of Kupffer cells in liver tissue can affect the viability of PCLS in culture and are involved in the regulation of paracetamol metabolism, in particular the glucuronidation pathway. Furthermore, inhibition of Kupffer cells leads to lipid accumulation in liver tissue, supporting a role of Kupffer cells in the regulation of intra-hepatic lipid metabolism. In fact, we have shown that PGE2 inhibited lipid synthesis in cultured hepatocytes. Since, both serum PGE2 and PGE2 released by PCLS in culture decreased after Kupffer cell inhibition by GdCl3-treatment, we conclude that Kupffer cell-derived PGE2 may be involved in the physiological regulation of hepatic lipid metabolism. Final objectives of this work were devoted to highlight the impact of the diet on Kupffer cell activity, offering possibilities to modulate Kupffer cell functions by nutrients under various physio-patholical conditions, such as inflammation. These results can be summarized as follows. (1) We failed to demonstrate involvement of Kupffer cells in fasting-refeeding nutritional transition leading to lipid metabolism alteration in the liver similar to the one developed in the obese Zucker rat. However, their role in the pathogenesis of nonalcoholic fatty liver disease remains an attractive hypothesis to investigate. (2) A supplementation of glycine, a simple amino acid, in the diet could influence lipid metabolism, namely in the liver. A direct relationship between those metabolic effects and Kupffer cell activity has not been demonstrated. Nevertheless, we have shown that the use of glycine in vitro offers the possibility to elucidate complex interactions between Kupffer cells and hepatocytes, by using PCLS in culture, and could constitute an alternative tool to inhibit in vitro Kupffer cell-derived mediators such as TNF-alpha (3) Dietary oligofructose, a fermentable and nondigestible carbohydrate that is known to promote gut bacteria growth, was able to increase production of Kupffer cell-derived mediators by PCLS. Our results indicate that improvement of Kupffer cell phagocytic activity as well as their secretory capacity might be involved in the hepatoprotection against septic challenge observed after a diet enriched with oligofructose.Thèse de doctorat en sciences pharmaceutiques (FARM 3) -- UCL, 200