Oxysterols serve as endogenous ligands of liver X receptors

Oxysterols serve as endogenous ligands of liver X receptors (LXRα (NR1H3)/LXRβ (NR1H2)) (Janowski et al., 1996). In addition to their established role in cholesterol metabolism (Janowski et al., 1999), LXRs also suppress inflammatory gene expression, thus controlling immune response and inflammation (Huang et al., 2011; Ogawa et al., 2004; Pascual et al., 2005). Importantly, LXR null mice, even in the absence of hyperglycemia, present with pathological changes seen in the diabetic retina, dramatically reduced CAC function and increased levels of activated macrophages (Hazra et al., 2012). Another key metabolic regulator, Sirtuin 1 (SIRT1), was recently shown to exert its beneficial effect through LXR-dependent mechanisms (Li et al., 2007). SIRT1 is the mammalian ortholog of yeast Sir2 gene that has been shown to play a role in aging, apoptosis, neural protection, glucose metabolism and neural development (Miranda et al., 2015; Purushotham et al., 2009; Wang et al., 2011). SIRT1 interacts with LXR to promote its deacetylation at a single conserved lysine (K432 in LXRα and K433 in LXRβ) adjacent to the ligand-regulated activation domain, leading to LXR activation (Li et al., 2007). Via LXR activation, SIRT1 promotes insulin secretion, reduces glucose tolerance and decreases body weight (Zabolotny and Kim, 2007; Purushotham et al., 2009; Wang et al., 2011). Importantly, SIRT1 is critical to vascular health (Kitada et al., 2016). High glucose adversely influences CAC function by reducing SIRT1, while increasing SIRT1 in CACs of diabetic origin corrects their dysfunction (Balestrieri et al., 2008, 2013; Yuen et al., 2012). Animal and in vitro studies have demonstrated the importance of SIRT1 regulation of inflammation (Zabolotny and Kim, 2007; Howitz et al., 2003; Liang et al., 2009). Activation of SIRT1, followed by LXR deacetylation resulted in inhibition NFκB activity induced by high-fat diet, and sustained repression of inflammatory fatty acid amide hydrolase (Zeng et al., 2013). By contrast, SIRT1-KO mice on a high-fat diet were prone to liver inflammation (Purushotham et al., 2009). In primary macrophages, deficiency of SIRT1 decreased the induction of the LXR target gene, ABCA1, impairing cholesterol export. Activation of SIRT1 enhanced LXRα, activity and ABCA1 and ABCG1 expression (Li et al., 2007). SIRT1-LXRα axis is also significantly reduced in U937 cells during foam cell formation (Zeng et al., 2013). Finally, decreased SIRT1 levels have recently been implicated in the pathogenesis of microvascular complications, specifically DR (Kowluru et al., 2014). The interplay of SIRT1 with LXR in the pathogenesis of DR has not been previously investigated and represents the focus of this study.
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Results
Discussion Activation of the LXR signaling pathway has dual beneficial consequences. First, LXR signaling has been shown to directly cause downstream activation of the RCT pathway via ABCA1 and ABCG1 activation. These membrane-associated protein transporters are the first step in the RCT pathway and are major regulators of cellular cholesterol by acting as cholesterol efflux pumps that efflux cholesterol to lipid poor apolipoproteins (ApoA1 and ApoE) (Demina et al., 2016). The RCT pathway is actively used by the retina to transport cholesterol back to the liver in a multi-step process (Fliesler, 2015). Genetic manipulation studies have demonstrated that downregulation of ABCA1 leads to pathologic vessel thickening and hardening (Aiello et al., 2003). Loss of ABCA1 and ABCG1 also leads to accumulation of inflammatory macrophage foam cells in mice myocardium, lung, liver spleen and thymus (Yvan-Charvet et al., 2007). Additionally, individuals with ABCA1 mutations are diagnosed with Tangier disease, a disease that is characterized by a severe reduction in HDL. Activated LXR has also been shown to bind to and stabilize NCoR Complex, thus inhibiting signaling through NFκB response element on inflammatory gene promoters resulting in reduced inflammation (Huang et al., 2011).