Abstract
Purpose :
Liver X receptors are key regulators of cholesterol homeostasis, as well as important modulators of inflammation. Synthetic LXR agonists exert anti-inflammatory effect, while endogenous oxysterol LXR agonists exert pro-inflammatory effect. We studied the paradoxical effects of LXR agonists on inflammation and explored the underlying mechanisms with transcriptome and proteome analysis.
Methods :
Human RPE cells were incubated with different concentrations of an oxysterol LXR agonists 7-KC (7-Ketocholesterol) for 36 hours. CCK-8 assay and Annexin-V FITC assay were used to detect the cell viability and apoptosis. EdU was used to detect cell proliferation. RPE cells were pretreated with 7-KC and TO90 for 12 hours, followed by stimulation with 5 μg/mL of LPS for 24 hours. The mRNA levels of IL6, IL-8, LXRα/β and ABCA1 and ABCG1 were measured. Illumina Hiseq 2500 platform and Q Exactive mass spectrometer were applied to characterize the transcriptome and proteome profiles.
Results :
Incubation with high concentration of 7-KC markedly decreased the cell viability and increased cell apoptosis. There was no cell proliferation when stimulated with various concentrations of 7-KC for 36 hours. Thus, 7.5 μM of 7-KC was used as the optimum concentration. Both 7-KC and TO90 activated LXR target genes ABCA1 and ABCG1, while they exhibited opposite effects on LPS-induced inflammatory response. 7-KC increased the mRNA levels of IL-6, IL-8, LXRα, LXRβ and ABCA1, ABCG1 in a dose dependent manner, while TO90 suppressed the inflammation. Transcriptome profile indicated that 7-KC active the toll-like receptor and MAPK signal pathways, while TO90 suppressed those pathways. Combined analysis of transcriptome and proteome revealed that 7-KC not only regulated lipid metabolism, but also increased the expression of inflammation signal pathways, including toll-like receptor, MAPK, mTOR, HIF-1, and VEGF signal pathwas. Whereas, TO90 did not directly affect those signal pathways.
Conclusions :
Both LXR agonists actived their receptors, whereas 7-KC exaggerated the LPS-induced inflammation and TO90 inhibited the inflammatory response. The paradox effects may be associated with the different regulation mechanisms to the pro-inflammatory elements. In LPS induced inflammation, 7-KC up-regulated the inflammation related signal pathways, whereas TO90 did not directly regulate those pathways.
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.