Liver X receptors balance lipid stores in hepatic stellate cells through Rab18, a retinoid responsive lipid droplet protein

Abstract

Liver X receptors (LXRs) are determinants of hepatic stellate cell (HSC) activation and liver fibrosis. Freshly isolated HSCs from Lxrαβ−/− mice have increased lipid droplet (LD) size but the functional consequences of this are unknown. Our aim was to determine whether LXRs link cholesterol to retinoid storage in HSCs and how this impacts activation. Primary HSCs from Lxrαβ−/− and wild-type (WT) mice were profiled by gene array during in vitro activation. Lipid content was quantified by HPLC and mass spectroscopy. Primary HSCs were treated with nuclear receptor ligands, transfected with siRNA and plasmid constructs, and analyzed by immunocytochemistry. Lxrαβ−/− HSCs have increased cholesterol and retinyl esters (CEs & REs). The retinoid increase drives intrinsic retinoic acid receptor (RAR) signaling and activation occurs more rapidly in Lxrαβ−/− HSCs. We identify Rab18 as a novel retinoic acid responsive, lipid droplet associated protein that helps mediate stellate cell activation. Rab18 mRNA, protein, and membrane insertion increase during activation. Both Rab18 GTPase activity and isoprenylation are required for stellate cell lipid droplet loss and induction of activation markers. These phenomena are accelerated in the Lxrαβ−/− HSCs, where there is greater retinoic acid flux. Conversely, Rab18 knockdown retards lipid droplet loss in culture and blocks activation, just like the functional mutants. Rab18 is also induced with acute liver injury in vivo. Conclusion Retinoid and cholesterol metabolism are linked in stellate cells by the LD associated protein, Rab18. Retinoid overload helps explain the pro-fibrotic phenotype of Lxrαβ−/− mice and we establish a pivotal role for Rab18 GTPase activity and membrane insertion in wild-type stellate cell activation. Interference with Rab18 may have significant therapeutic benefit in ameliorating liver fibrosis.

Pathobiology of liver fibrosis: a translational success story

“…34 Moreover, cholesterol and retinoid metabolism are linked by a lipid-associated protein, Rab18, that is a retinoic acid-responsive gene. 35 …”

Hepatic stellate cells as key target in liver fibrosis

“…Perilipin 2, adipose TG lipase (ATGL), and comparative gene identification-58 (CGI-58) are lipid droplet-associated proteins in rat HSC line, HSC-T6 [302]. LXRs balance lipid stores in HSCs through Rab18, a retinoid responsive lipid droplet protein [303]. Autophagy regulates turnover of lipid droplets via ROS-dependent Rab25 activation in HSC [304].…”

Autophagy regulates turnover of lipid droplets via ROS-dependent Rab25 activation in hepatic stellate cell

“…It has been reported that the small GTPase Rab7 was highly activated under nutrient deprivation and was required to promote direct physical interactions between autophagy and LDs in live hepatocellular carcinoma cells [24]. Besides, Rab18 has also been reported to be relevant Rab-GTPase for stellate cells as it dynamically modulated both LD size and cellular activation [19]. Interference with Rab18 may therefore have significant therapeutic benefit in ameliorating liver fibrosis [19].…”

“…Previous studies [11], [15], including ours [11], have reported that Diacylglycerol acyltransferase 1 (DGAT1), Adipose triglyceride lipase (ATGL), or Hepatic lipase (LIPC) are all new target molecules, and attract extensive concern of numerous scholars. Moreover, transcriptional regulation is also required to maintain the adipogenic phenotype of HSCs [17], [18], [19]. The major transcription factors involved in cell adipocyte differentiation include PPARα [16], PPARγ [17], Nrf2 [18], LXR [19], and so on.…”

“…Moreover, transcriptional regulation is also required to maintain the adipogenic phenotype of HSCs [17], [18], [19]. The major transcription factors involved in cell adipocyte differentiation include PPARα [16], PPARγ [17], Nrf2 [18], LXR [19], and so on. However, these findings are not enough to fully elucidate the molecular mechanisms of LD disappearance.…”