LPL is the rate limiting enzyme

LPL is the rate-limiting enzyme in hydrolysis of TG in TG-rich lipoproteins, and LPL activity is inversely correlated with plasma TG levels (Shimizugawa et al., 2002). Support for the second mechanism linking ABCA1 and plasma TG rests on the observations that TD subjects exhibit decreased LPL activity (Alaupovic et al., 1991; Clifton-Bligh et al., 1972; Greten et al., 1974; Wang et al., 1987) and delayed clearance of postprandial lipid (Kolovou et al., 2003). Post-heparin LPL activity is also reduced in HSKO compared to WT mice, resulting in delayed clearance of postprandial TG (Chung et al., 2010b). The observations in TD patients and HSKO mice regarding LPL activity raise the question as to how the loss of ABCA1 in liver leads to a decrease in LPL activity. Here, we identified ANGPTL3 as a potential mediating factor between ABCA1 and TG levels. TD HLCs displayed elevated ANGPTL3 expression and a~10 fold increase in ANGPTL3 protein secretion. More importantly, male TD patients, including the two recruited to the current study, exhibited significantly increased plasma ANGPTL3 (Fig. 4F) and TG levels (M. Cuchel, unpublished observation). ANGPTL3 is a hepatokine that inhibits LPL activity. In agreement with its known TG regulatory effect, circulating ANGPTL3 positively correlates with TG levels in a larger cohort of TD patients and matched controls (M. Cuchel, unpublished observation). Inactivation of ANGPTL3 in mice has been recently shown to reduce VLDL-TG secretion (Tikka et al., 2014; Wang et al., 2015), which is thought to be a result of reduced free fatty pgds inhibitor supply to the liver. Therefore, the increased ANGPTL3 expression in TD hepatocytes may result in efficient secretion of ANGPTL3, confer enhanced LPL inhibition and delay TG clearance in vivo, contributing to hypertriglyceridemia in TD patients. Overall, our data are compatible with a model of increased secretion, reduced lipolysis and delayed clearance of TG as the underlying mechanisms of hypertriglyceridemia in TD patients. The increased ANGPTL3 expression and secretion with ABCA1 deficiency is intriguing. ANGPTL3 is a target gene of liver X receptors (LXR) (Tikka and Jauhiainen, 2016). HSKO mice fed with chow diet do not have altered liver lipids or evidence of LXR activation, but have phenotypes of early insulin resistance, including defective PI3 kinase activation (Chung et al., 2010b). Insulin signaling has been implicated as a negative regulator of ANGPTL3 mRNA in HepG2 cells, immortalized human hepatocytes and in vivo in human livers (Nidhina Haridas et al., 2015; Shimamura et al., 2004). The possibility of attenuated hepatic insulin signaling under conditions of ABCA1 deficiency in ANGPTL3 regulation will need to be explored in future studies. Regardless of the mechanism, ANGPTL3 as a potential mediator of altered TG metabolism in TD is an exciting finding that also has significant clinical implications. The global transcriptional profile of ABCA1-deficient human hepatocytes has never been examined. In this study, we performed microarray analysis with TD and control HLCs. As exemplified by ANGPTL3, we gained insights into transcriptional changes in human hepatic ABCA1 deficiency. Of note, some genes of immune response pathways were differentially expressed, including HLA-A and HLA-C, members of the human leukocyte antigen family. The HLA gene complex encodes the human proteins of the major histocompatibility complex (MHC), which is important for presenting processed peptide antigens (Choo, 2007; Neefjes et al., 2011). It is of interest to know whether these changes represent another link between ABCA1 and immune responses, considering the increasingly appreciated relationship between ABCA1 and inflammation, lipid-regulatory effects dependent or not (Bi et al., 2015; Bochem et al., 2015; Tall and Yvan-Charvet, 2015). The HLCs system enables future explorations using unbiased strategies to enhance our understanding of the functions of human hepatic ABCA1.