Background: Non-alcoholic fatty liver disease (NAFLD) is associated with inefficient macro- and micronutrient metabolism, and alteration of circulating phospholipid compositions defines the signature of NAFLD. This current study aimed to assess the pattern of serum phospholipids in the spectrum of NAFLD, and its related comorbidities and genetic modifications. Methods: 97 patients with diagnosed NAFLD were recruited at a single center during 2013–2016. Based on histological and transient elastography assessment, 69 patients were divided into non-alcoholic steatohepatitis (NASH) and non-alcoholic fatty liver (NAFL) subgroups. 28 patients served as healthy controls. Serum phospholipids were determined by liquid-chromatography mass spectrometry (LC-MS/MS). Results: The total content of phosphatidylcholine (PC) and sphingomyelin in the serum was significantly increased in NAFL and NASH patients, compared to healthy controls. In addition, serum lysophospatidylethanolamine levels were significantly decreased in NAFL and NASH individuals. Circulating PC species, containing linoleic and α-linolenic acids, were markedly increased in NAFLD patients with hypertension, compared to NAFLD patients without hypertension. The pattern of phospholipids did not differ between NAFLD patients with diabetes and those without diabetes. However, NAFLD patients with hyperglycemia (blood glucose level (BGL) >100 mg/dL) exhibited significantly a higher amount of monounsaturated phosphatidylethanolamine than those with low blood glucose levels. In addition, NAFLD patients with proven GG-genotype of PNPLA3, who were at higher risk for the development of progressive disease with fibrosis, showed lower levels of circulating plasmalogens, especially 16:0, compared to those with CC- and CG-allele. Conclusions: Our extended lipidomic study presents a unique metabolic profile of circulating phospholipids associated with the presence of metabolic risk factors or the genetic background of NAFLD patients.

A decrease of hepatocellular phosphatidylcholine (PC) is associated with hepatic injury, e.g., in nonalcoholic steatohepatitis (NASH). Therefore, we evaluated the hepatoprotective effect of a PC-precursor lipid specifically targeted to the liver. We synthesized the bile acid-phospholipid conjugate ursodeoxycholyl lysophosphatidylethanolamide (UDCA-LPE), which was designed to target PC to hepatocytes by way of bile-acid transport systems. We synthesized a fluorescently labeled analogue UDCA-6-[(7-nitro-2-1,3-benzoxadiazol-4-yl)amino]hexanoyl PE (UDCA-NBDPE) for uptake and metabolism studies. Unexpectedly, the majority of UDCA-NBDPE was still intact and not hydrolyzed efficiently in HepG2 cells. For targeting in vivo, NBD fluorescence from UDCA-NBDPE-injected mice was recovered in the liver the most, whereas injection of NBDPE alone resulted in an even distribution in liver, kidneys, and intestine. Cytoprotection by UDCA-LPE was tested in starvation and tumor necrosis factor alpha (TNF-α) apoptosis models using HepG2 cells. Only the intact UDCA-LPE was able to persistently stimulate growth after 36 to 120-hour starvation, and significantly inhibited TNF-α-induced apoptosis. In both models, LPC, LPE, UDCA, or UDCA added with LPE exhibited weak to no cytoprotection. UDCA-LPE stabilized mitochondrial membranes by lowering mitochondrial membrane potential. Western blot analyses of phosphorylated Akt and glycogen synthase kinase-3 (GSK-3)α/β revealed that UDCA-LPE activated phosphatidyl inositol 3-kinase (PI3K)/Akt signaling pathways. The PI3K inhibitor LY294002 or Akt small interfering (si)RNA consistently inhibited the proproliferative effects of UDCA-LPE during starvation. The TNF-α death-receptor extrinsic pathway involves caspase 8 activation, which is inhibited by cellular FLICE-inhibitory protein (cFLIP); thus, cFLIP siRNA was employed in our studies. cFLIP siRNA was able to reverse the cytoprotective effects of UDCA-LPE during TNF-α-induced apoptosis, and UDCA-LPE concomitantly upregulated protein expression of cFLIP_L. Conclusion: UDCA-LPE, which targeted the liver in vivo, elicited potent biological activities in vitro by stimulating hepatocyte growth and by inhibiting TNF-α-induced apoptosis. Thus, UDCA-LPE may be suitable for evaluation of treatment efficacy in NASH. (Hepatology 2009.)

Fatty acid (FA) metabolism is a series of processes that provide structural substances, signalling molecules and energy. Ample evidence has shown that FA uptake is mediated by plasma membrane transporters including FA transport proteins (FATPs), caveolin-1, fatty-acid translocase (FAT)/CD36, and fatty-acid binding proteins. Unlike other FA transporters, the functions of FATPs have been controversial because they contain both motifs of FA transport and fatty acyl-CoA synthetase (ACS). The widely distributed FATP4 is not a direct FA transporter but plays a predominant function as an ACS. FATP4 deficiency causes ichthyosis premature syndrome in mice and humans associated with suppression of polar lipids but an increase in neutral lipids including triglycerides (TGs). Such a shift has been extensively characterized in enterocyte-, hepatocyte-, and adipocyte-specific Fatp4-deficient mice. The mutants under obese and non-obese fatty livers induced by different diets persistently show an increase in blood non-esterified free fatty acids and glycerol indicating the lipolysis of TGs. This review also focuses on FATP4 role on regulatory networks and factors that modulate FATP4 expression in metabolic tissues including intestine, liver, muscle, and adipose tissues. Metabolic disorders especially regarding blood lipids by FATP4 deficiency in different cell types are herein discussed. Our results may be applicable to not only patients with FATP4 mutations but also represent a model of dysregulated lipid homeostasis, thus providing mechanistic insights into obesity and development of fatty liver disease.