Studies on the oxysterol receptor LXRβ : linking cholesterol metabolism to water transport and cell proliferation

Abstract

Liver X Receptor β (LXRβ) is a nuclear receptor, belonging to the superfamily of ligand-­activated transcription factors. With its α isoform (LXRα), LXRβ shares more than 78% homology in its amino acid sequence, a common profile of oxysterol ligands and the heterodimerization partner, Retinoid X Receptor. LXRs have a crucial role in lipid metabolism, in particular in preventing cholesterol accumulation, in glucose homeostasis and in macrophage inflammatory response. The first evidence that, in spite of all the common properties, LXRα and LXRβ have distinct functions, came in 2001 with the creation of knock-­out mice for each LXR isoform. LXRα-­/-­ mice fed with 2% cholesterol diet show a severe cholesterol accumulation in the liver due to an inability to increase bile acid synthesis in response to high cholesterol intake. Surprisingly, LXRβ-­/-­ mice had the same compensatory capacity of WT mice to avoid hepatic cholesterol accumulation suggesting that LXRβ may have a completely distinct role from LXRα. Indeed in 2005, it was shown that specifically in LXRβ-­/-­ male mice cholesterol accumulates in big motor neurons of the spinal cord leading to their death and inducing a significant motor function impairment like amyotrophic lateral sclerosis (ALS). Starting from this neurological phenotype of LXRβ-­/-­ mice and comparing the characteristics of knock-­out mice for each LXR isoform, this thesis aims to define new and specific functions of the oxysterol receptor LXRβ.

Paper I of this thesis aims to investigate the neurological phenotype of LXRβ-­/-­ mice focusing in particular on the role of β-­sitosterol in the pathogenesis ALS-­Parkinson-­Dementia complex. Administration of β-­sitosterol to LXRβ-­/-­ mice creates a severe motor-­impairment and loss of dopaminergic neurons in the substantia nigra, activates microglia and decreases brain cholesterol indicating that LXRβ may have a protective role against the toxic action of β-­ sitosterol on the central nervous system.

Paper II investigates the resistance to gain weight, characteristic of LXRβ-­/-­ mice and demonstrates that they are affected by a severe pancreatic insufficiency with low serum levels of amylase, lipase, low fecal protease and abundant inflammatory infiltrates all around medium size pancreatic ducts. The water channel aquaporin-­1 (AQP-­1), responsible of transporting water into the pancreatic ductal lumen was markedly decreased in LXRβ-­/-­ mice leading to the presence of plugs inside the ducts and in turn to a pancreatic insufficiency. In the digestive system AQP-­1 is strongly expressed in the cholangiocytes of the gallbladder being, together with AQP-­8, the mediator of the absorbing-­secretory functions of this organ.

Paper III shows that the male gallbladder cholangiocytes of LXRβ-­/-­ mice express very low mRNA and protein levels of both AQP-­1 and AQP-­8 and morphologically they appear shrunk with loss of cell polarization. Treatment of WT mice with LXR-­agonist increases the expression of the two water channels in the gallbladder together with the cholesterol transporters ATP Binding Cassette G5/G8 and it is associated with cholesterol crystals in the bile.

The morphology of female LXRβ-­/-­ gallbladders was studied in paper IV: at the age of 12 months a wide range of preneoplastic lesions are detectable, from dysplasia to metaplasia and adenomas, degenerating into carcinoma in situ, when the mice become 19 months old. The pathogenesis involves a complex interplay between LXRβ, Transforming Growth Factor β (TGFβ) and estrogens. Indeed, ovariectomy of LXRβ-­/-­ mice prevents the development of preneoplastic lesions and normalizes the TGFβ signaling that is upregulated in LXRβ-­/ mice. In conclusion, this thesis describes new emerging and specific roles for LXRβ in controlling not only cholesterol homeostasis in the central nervous system but also water channels in pancreas and gallbladder as well neoplastic transformation of cholangiocytes.