Sphingosine-1-phosphate receptor 2: a novel bile acid receptor and regulator of hepatic lipid metabolism?

Dysregulation of hepatic lipid metabolism is a major contributing factor for the development of hepatic steatosis and pathogenesis of non-alcoholic fatty liver disease (NAFLD), diabetes and obesity. Bile acids have lipid-lowering effects but the underlying mechanisms are very complex and not fully understood [review in ¹]. It has been known for a long time that bile acid pool size is inversely correlated to serum triglyceride levels in human patients. Bile acid sequestrants reduce bile acid pool size but increase serum triglycerides. Conversely, bile acid treatment in hypertriglyceridemia and gallstone patients decreased serum triglycerides and VLDL production. The discovery of farnesoid X receptor (FXR) as a bile acid-activated receptor has generated a plethora of research to study the roles of bile acids in the regulation of hepatic lipid metabolism in recent years [review in ²].

FXR is activated by primary bile acids, chenodeoxycholic acid and cholic acid, to inhibit bile acid synthesis by a feedback mechanism (Fig 1). In the liver, FXR induces the small heterodimer partner (SHP), which inhibits cholesterol 7α-hydroxylase (CYP7A1) gene transcription and bile acid synthesis. In the intestine, FXR induces fibroblast growth factor 15 (FGF15, or a human orthologue FGF19), which is secreted into blood circulation to hepatocytes to activate the FGF receptor 4 (FGFR4) that activates the mitogen activated protein kinase (MAPK)/extracellular receptor kinase 1/2 (ERK1/2) pathway to inhibit CYP7A1. CA and synthetic FXR agonists have been shown to reduce serum triglycerides in diabetic and obese mice. Consistently, FXR^-/- mice have increased serum triglycerides and pro-atherogenic lipoprotein profiles.

Fig 1. Complex mechanisms of bile acid signaling in regulation of lipid metabolism in hepatocytes.

Bile acids are the end products of cholesterol catabolism. Cholesterol 7α-hydroxylase (CYP7A1) is the first and rate-limiting enzyme in the bile acid synthetic pathway. Primary bile acids, cholic acid (CA) and chenodeoxycholic acid (CDCA) are endogenous ligands of FXR. In the liver, FXR induces SHP, which inhibits Cyp7A1 gene transcription. In the intestine, FXR induces fibroblast growth factors 1 (FGF15), which is secreted to blood circulation to activate a hepatic FGF receptor 4 (FGFR4) to activate mainly the MAPK/ERK1/2 pathway to inhibit CYP7A1 and sterol 12α-hydroxylase (CYP8B1) gene transcription. FXR is known to stimulate energy metabolism by inducing ApoCII and inhibiting ApoCIII to activate lipoprotein lipase (LPL), which hydrolyzes triglycerides carried by VLDL and chylomicrons in peripheral tissues. FXR also induces peroxisome proliferator-activated receptor α (PPARα) to stimulate fatty acid oxidation and clearance. FXR regulates lipoprotein metabolism by inducing VLDL receptor (VLDLR) and phospholipid transfer protein (PLTP), and inhibiting ApoB1 and microsomal triglyceride transport protein (MTTP) to inhibit VLDL secretion. FXR also has anti-inflammatory function by inhibiting NFκB-mediated inflammatory cytokine production in hepatocytes and enterocytes. FXR-independent pathways also are involved in regulation of bile acid synthesis and liver metabolism. A secondary bile acid-activated G protein-coupled receptor (GPCR), TGR5, is expressed in hepatic endothelium cells and Kupffer cells, but not hepatocytes. TGR5 may regulate lipid metabolism by indirect mechanisms. In brown adipose tissues, TGR5/cAMP signaling may stimulate energy metabolism by inducing a type 2 deiodinase, which activates T₄ to T₃. In the intestine, TGR5 signaling protects barrier function and inflammation in the intestine, and may indirectly protect hepatocyte against inflammation. Conjugated bile acids, taurocholic acid (TCA) selectively activates a sphingosine-1-phosphate receptor 2 (S1PR2) in hepatocytes. Extracellular signal activates sphingosine kinase 1(SphK1), which is translocated to plasma membrane to phosphorylate sphingosine to S1P. S1P activates membrane S1PRs by paracrine/autocrine mechanism. Growth factors (epidermal growth factor, FGFR4, etc.) and phorbol esters activate protein kinase C (PKC), which activates ERK1/2 of the mitogen-activated kinase (MAPK) pathway to phosphorylate SphK2. SphK2 is translocated to the nucleus to bind to histone deacetylase 1/2 (HDAC1/2). S1P signaling activates ERK1/2, and AKT in insulin receptor signaling pathway. Nuclear SphK2 generated S1P has been shown to bind to chromatin and inhibits HDAC1/2. Inhibition of HDAC1/2 results in activation of histone acetylases, which stimulate expression of steroid response element binding protein-1c (SREBP-1c), which induces all genes in lipogenesis (fatty acid synthase, acetyl-CoA carboxylase, etc.), and CYP7A1 in bile acid synthesis. On the other hand, the FXR/SHP pathway inhibits SREBP-1c and CYP7A1 gene transcription by recruiting HDAC1/2, histone methytransferase G9a, and histone lysine-specific demethylase LSD-1 to inhibit lipogenesis and bile acid synthesis. These positive and negative regulations modulate hepatic lipid homeostasis.

FXR signaling may regulate hepatic lipid metabolism by several mechanisms. It has been reported that activation of FXR by bile acids or FXR agonists inhibits steroid response element binding protein-1c (SREBP-1c)-mediated hepatic lipogenesis. It was suggested that the FXR/SHP pathway might inhibit expression of the genes in lipogenesis and VLDL metabolism (Fig 1). FXR agonists stimulate triglyceride clearance. Transgenic overexpression of FGF19 has been shown to increase metabolic rate in mice, but the mechanism is not clear. More importantly, recent studies showed that bile acids and FXR agonists had an anti-inflammatory function through the inhibition of NFκB-mediated inflammatory cytokine production in hepatocytes and enterocytes ^{3, 4}. However, how FXR signaling inhibits NF-κB activity is not clear. Expression of the SREBP-1c and its target genes is not reduced in FXR^-/- mice, suggesting that FXR-independent mechanisms may also be involved in the triglyceride-lowering effect of bile acids.

Secondary bile acids (lithocholic acid and deoxycholic acid) and their taurine conjugates are activators of Gαs protein-coupled receptor, TGR5 (aka Gpbar1). In the digestive tract, TGR5 is expressed in the gallbladder epithelium and in the intestine. In the liver, TGR5 is expressed in Kupffer cells and sinusoidal endothelial cells, as well as in cholangiocytes, but not in hepatocytes. Activation of TGR5 signaling stimulates energy metabolism in brown adipose tissue, protects the intestinal barrier function and alleviates inflammation in animal models of inflammatory bowel diseases. However, TGR5^-/- mice have no obvious liver metabolic phenotype. Activation of TGR5 has been shown to protect liver from inflammation but the underlying mechanism in not clear. TGR5 agonists increase insulin secretion from β cells and glycogen like peptide-1 secretion from enteroendocrine cells, and may contribute to the amelioration of liver inflammation and insulin resistance.

Another GPCR, sphingosine-1-phosphate receptor 2 (S1PR2) has recently been reported as a conjugated bile acid-activated receptor ⁵. Taurocholic acid (TCA) is the most abundant bile acid in humans and mice and is an efficacious activator of S1PR2 in hepatocytes. S1P activates a S1P family of Gαi-coupled receptors (S1PR1 to 5). S1P kinases (SphK1 and SphK2) phosphorylate sphingosine to S1P. SphK1 is located in the cytosol, whereas SphK2 is localized in the nucleus. In hepatocytes, S1PR2 signaling activates ERK1/2 in the MAPK signaling pathways and AKT in insulin signaling (Fig 1). Sphingolipid metabolites have diverse functions in endothelial cells, including immune response, lymphatic trafficking, and vascular integrity ⁶. In endothelial cells, S1P/S1PRs mediated signaling activates NF-κB in pro-inflammatory responses. The role of S1PR2/SphK2/S1P on hepatic lipid metabolism has not been studied.

In this issue of Hepatology, Nagahashi et al. ⁷ reported a novel finding that the S1PR2/SphK2/S1P signaling pathway might play a key role in control of hepatic lipid metabolism in mice. A previous in vitro study reported that the nuclear SphK2-generated S1P bound to and inhibited histone deacetylases 1 and 2 (HDAC1/2) activity and increased histone acetylation and expression of cyclin-dependent kinase inhibitor p21 and cFos of MCF-7 and HeLa cells ⁸. The current in vivo study showed that in S1PR2^-/- and SphK2^-/- mice acetylated histone 3 and histone 4 levels were reduced and these reductions correlated to reduced mRNA expression levels of SREBP-1c and its regulated genes in lipid metabolism and bile acid synthesis genes (not CYP7A1). Furthermore, overexpression of S1PR2 induced these hepatic genes (including CYP7A1) in wild-type and S1PR2^-/- mice, but not in SphK2^-/- mice suggesting that SphK2 is necessary for mediating S1PR2 signaling in lipid metabolism. A high fat/high cholesterol diet and a lithogenic diet (containing high cholesterol and cholic acid) induced SphK2, but not SphK1 expression in wild-type mouse liver. It is important to note that SphK2^-/-mice on a chow diet had markedly elevated serum triglycerides and AST and ALT levels compared to wild type mice. High fat diet feeding increased serum triglycerides and AST and ALT levels in wild-type mice as expected, but did not further increase serum triglyceride and AST/ALT level in SphK2^-/- mice. These data may imply that TCA/S1PR2/SphK2/S1P signaling may play a key role in mediating the anti-inflammatory function of bile acids in hepatocytes. It is important to study whether or not the TCA/S1PR2/ SphK2/S1P signaling pathway modulates NF-κB-mediated pro-inflammatory cytokine production via epigenetic mechanism in hepatocytes.

Paradoxically, both SHP and CYP7A1 mRNA expression levels were significantly increased in wild-type and S1PR2^-/- mice by overexpressing S1PR2. It has been reported that SHP recruits HDAC1/2, histone lysine-specific demethylase 1 (LSD-1), histone methytransferase G9a and other chromatin modifying enzymes to inhibit CYP7A1 gene transcription ⁹. Thus, TCA may activate the SHP/HDAC/LSD1/G9a pathway to inhibit and activate the S1PR2/SphK2/S1P/HDAC1/2 pathway to induce CYP7A1 expression levels to modulate bile acid homeostasis. Histone acetylation and methylation of the chromatin in gene promoters has emerged as a key mechanism for regulation of liver metabolism.

This study uncovered a novel role of the TCA/S1PR2/SphK2/S1P pathway in control of hepatic lipid metabolism. Further detailed study is needed to: 1) identify the liver metabolic phenotypes and differentially regulated genes and pathways in S1PR2^-/- and SphK2^-/- mice; 2) confirm the role of the S1PR2/SphK2/S1P pathway in control of hepatic lipid gene expression by epigenetic mechanism; and 3) identify other metabolic pathways and mechanisms that may be involved in the pleiotropic effects of S1P and its implication in pathogenesis of NAFLD. Inhibitors targeted to HDAC and histone methytransferase activity are currently being developed as drug therapies for treatment of cancer and other human diseases, and may also have potential for treating inflammatory liver-related metabolic diseases, such as NAFLD and diabetes.

Abbreviation list

CYP7A1

cholesterol 7α-hydroxylase

ERK1/2

extracellular receptor kinase 1/2

FXR

farnesoid X receptor

FGF15

fibroblast growth factor 15

MAPK

mitogen activated protein kinase

NAFLD

non-alcoholic fatty liver disease

SHP

small heterodimer partner

S1PR

sphingosine-1-phosphate receptor

S1P

sphingosine-1-phosphate

SphK2

S1P kinase 2

SREBP-1c

steroid response element binding protein-1c

TCA

taurocholic acid

TGR5 (aka Gpbar1)

G protein bile acid receptor 1