title: Investigation into the effects of sepsis-induced hyperglycaemia on glucose-modulating hormones receptors in hepatocyte and monocyte cell lines
Creators: name: Mahmood Abdullah  Daabo, Hamid
Publisher: name: University of Liverpool Repository
Year: 2023

description: Hyperglycaemia secondary to sepsis is a common feature in critically ill patients and represents one of the leading causes of death in intensive care. Sepsis-induced hyperglycaemia is a complex condition involving altered insulin sensitivity and beta cell function. However little is known about the potential roles in sepsis-induced hyperglycaemia of other major modulators of glucose homeostasis the incretins, glucose-dependent insulinotropic polypeptide (GIP) and glucagon-like peptide-I (GLP-I). The overall aim of this study was to investigate the role of receptors for incretins and other glucose-regulating hormones in sepsis-induced hyperglycaemia. In order to achieve these aims two in vitro models was established, one of inflammation employing LPS in U937 monocytes and one of glucose metabolism employing the HUH7 hepatocyte cell line. The specific hypotheses to be tested were: Hyperglycaemia in sepsis results from the effects of endotoxin on glucose-modulating hormone receptors, in addition to insulin resistance The effects of sepsis on glucose-modulating hormone receptors are augmented by high glucose concentration Incretins are able to modulate cytokine secretion Effects of sepsis on glucose-modulating hormone receptors are altered by incretins The hepatocyte cells HUH7 and leukocyte cells U937 were used in this study as representative cell lines. Different sets of experiments were performed by culturing these cells with lipopolysaccharide (LPS) to induce an acute proinflammatory state. Analyses of receptor expression mainly by transcriptional analysis with some confirmation by immunocytochemistry and Western blot were performed. The major findings were that transcripts for receptors for GIP, GLP-I, GLP-2, insulin and adiponectin (RI &amp;R2) were detected in HUH7 cells. Expression of GIPR protein was also confirmed. Transcripts for receptors GIP, insulin and adiponectin (RI &amp;R2) were also detected in U937, but much less abundantly than for HUH7. Incubation of HUH7 and U937 with LPS for 24 hr resulted in a dose dependent decrease in expression of receptors for GIP, GLP-I, GLP-2, insulin and adiponectin (RI &amp;R2) in both cell lines. This inhibition was accompanied by an induction of cytokine IL-I β and IL6 but there was no effect on morphology and viability of the cells. IV Incubating HUH7 and U937 cells with a range of glucose concentrations alone for 24 hrs induced a dose dependent down-regulation in the expression of receptors for GIP, GLP-l, insulin and adiponectin (Rl&amp;R2) in HUH7 cells and GIP and adiponectin R2 in U937 cells. Adding LPS to the culture media-potentiated these effects markedly. Incubation of HUH7 and U937 with LPS and GLP-l showed that GLP-l decreased the LPS- induced IL-lβ, IL-6 up-regulations, and reversed the LPS effects on the INSR expression, which had been inhibited by LPS in both cell lines. Similarly, incubation of these cells with LPS and GIP partially suppressed LPS-induced IL-lβ up-regulation. Exendin-4 suppressed the LPS-induced GLP-IR, GLP-2R down-regulation in HUH7 cells, and GIPR, INSR in U937 cells and blocked the LPS-induced IL-6 up-regulation in the both cell lines, Taken collectively, the data presented in this thesis indicate that hepatocytes and monocytes express glucose-modulating hormone receptors. Furthermore, exposure of these cells to sepsis significantly inhibits the 'expression of these receptors whilst inducing cytokine expression. Suppressed expression of the glucose-modulating hormone receptors in sepsis- induced hyperglycaemia may provide a possible explanation for the impairment of the incretin effects and insulin resistance in critically ill patients or in diabetic individuals. This research suggests that prevention of the down-regulation of glucose-modulating hormone receptors in sepsis could result in improvement in the action of insulin and incretins and thereby improve glucose control during hyperglycaemia or sepsis-induced hyperglycaemia. Furthermore, this thesis presents evidence to suggest that incretins may protect against pathological inflammatory consequences during sepsis-induced hyperglycaemia.