Angiopoietin-like proteins 4 and 3

Angiopoietin-like proteins 3 and 4 are two members of the angiopoietin-like family of proteins (Angptl). Unique within this family, Angptl3 and 4 inhibit lipoprotein metabolism via their ability to inhibit the activity of lipoprotein lipase.

Angptl4 exhibits a widespread distribution of tissue expression, while Angptl3 is exclusively expressed in the liver. Treatments with various ligands of nuclear receptors reveal that Angptl3 is a target gene of liver X receptor, while Angptl4 expression is activated by ligands of all peroxisome proliferator-activated receptors. Expression of Angptl4 in adipose tissue and liver is induced by fasting, while Angptl3 expression is not appreciably affected by nutritional status. The differential regulation of Angptl3 and Angptl4 by sites of expression, nutritional status, and ligands of nuclear receptors may confer unique roles of each in lipoprotein metabolism. In vivo studies revealed that thyroid hormone down-regulates ANGPTL3 but not ANGPTL4 gene expression in hypothyroid rats.

Angiopoietin-like protein 4

Angptl4, (also called peroxisome proliferator-activated—γ angiopoietin-related protein [PGAR] and fasting-induced adipose factor [FIAF]) is a downstream target of peroxisome proliferator-activated receptors (PPARs), including PPARα (important in fatty acid oxidation) and PPARγ (involved in modulating insulin sensitivity and adipocyte differentiation). Angptl4 increases circulating triglycerides by inhibition of lipoprotein lipase. Being downstream of PPARγ, Angptl4 has beneficial effects on insulin sensitivity. Angptl4 expression is most abundant in liver and adipose tissue and is induced by fasting.

Angiopoietin-like protein 4 structure/fun­ction relations

Angptl4 contains an amino-terminal coiled-coil domain and a carboxyl-terminal fibrinogen-like domain. Angptl4 exists as variable-sized oligomers that contain intermolecular disulfide bonds and undergoes regulated proteolytic processing upon secretion.

Oligomerized Angptl4 undergoes proteolytic processing to release its carboxyl fibrinogen-like domain, which circulates as a monomer. Angptl4‘s N-terminal coiled-coil domain mediates its oligomerization, which by itself is sufficient to form higher order oligomeric structures. Adenovirus-mediated overexpression of Angptl4 in 293 cells shows that conversion of full-length, oligomerized Angptl4 is mediated by a cell-associated protease activity induced by serum. Cysteine residues at positions 76 and 80 of Angptl4, conserved among mouse, rat, and human, are required to form higher order structures. Loss of oligomerization results in decreased stability of the N-terminal coiled-coil domain of Angptl4 as well as decreased ability to increase plasma TG levels, suggesting that intermolecular disulfide bond formation plays important roles in determining the magnitude of the hyperlipidemic effect of Angptl4. Inappropriate oligomerization of Angptl4 could be associated with disorders of lipid metabolism in vivo.

Remarkably, in mice the full-length form of Angptl4 was physically associated with HDL, whereas truncated Angptl4 was associated with low density lipoprotein. In humans, both full-length and truncated Angptl4 were associated with HDL. The composite data suggest that via physical association with plasma lipoproteins, Angptl4 acts as a powerful signal from fat and other tissues to prevent fat storage and stimulate fat mobilization. These data indicate that disturbances in Angptl4 signaling might be involved in dyslipidemia.

Angiopoietin-like protein 4 in transgenic mice

In mice, Angptl4 overexpression caused a 50% reduction in adipose tissue weight, partly by stimulating fatty acid oxidation and uncoupling in fat. In addition, Angptl4 overexpression increased plasma levels of triglycerides, free fatty acids, glycerol, total cholesterol, and high density lipoprotein (HDL)-cholesterol. Functional tests indicated that Angptl4 overexpression severely impaired plasma triglyceride clearance but had no effect on very low density lipoprotein production. The effects of Angptl4 overexpression were amplified by a high fat diet, resulting in markedly elevated plasma and liver triglycerides, plasma free fatty acids, and plasma glycerol levels, and impaired glucose tolerance in Angptl transgenic mice fed a high fat diet. Angptl4-deficient mice displayed hypotriglyceridemia and increased post heparin plasma lipoproteid lipase activity, with greater effects in the fasted compared with the fed state.

Angiopoietin-like protein 4 as a blood-borne hormone

ANGPTL4 is a blood-borne hormone directly involved in regulating glucose homeostasis, lipid metabolism, and insulin sensitivity. Adenovirus-mediated expression of ANGPTL4 potently decreased blood glucose and improved glucose tolerance, whereas it induced hyperlipidemia, fatty liver, and hepatomegaly in C57 mice. In db/db diabetic mice, ANGPTL4 treatment reduced hyperglycemia to a normal level, and markedly alleviated glucose intolerance and hyperinsulinemia. Ex vivo studies on primary rat hepatocytes revealed that ANGPTL4 significantly decreased hepatic glucose production and enhanced insulin-mediated inhibition of gluconeogenesis.

Serum levels of ANGPTL4 in human subjects inversely correlated with plasma glucose concentrations and HOMA IR, the homeostasis model assessment of insulin resistance. In patients with type 2 diabetes, serum levels of ANGPTL4 were significantly lower than those in healthy subjects, suggesting that the decreased ANGPTL4 could be a causative factor of this disease. These results collectively indicate that ANGPTL4 exerts distinct effects on glucose and lipid metabolism, and that its beneficial effect on glucose homeostasis might be useful for the treatment of diabetes.

Angiopoietin-like protein 3

Angiopoietin-like 3 (ANGPTL3) is a secreted protein that is mainly expressed in the liver and regulates lipid metabolism by inhibiting the lipolysis of triglycerid rich lipoproteins. The N-terminal coiled-coil domain-containing fragment-( 17–207) and not the C-terminal fibrinogen-like domain-containing fragment-(207–460) increased the plasma triglyceride levels in mice. The N-terminal region 17–165 was required to increase plasma triglyceride levels in mice and a substitution of basic amino acid residues in the region 61–66 of the fragment showed no increase in the plasma triglyceride levels and no inhibition of lipolysis by lipoprotein lipase. In addition, when analyzed in human plasma, cleaved fragments of ANGPTL3 were detected. By analyzing recombinant ANGPTL3 in mouse plasma, it was found that it was cleaved in the linker region between the coiled-coil domain and the fibrinogen-like domain. Furthermore, a cleavage-resistant mutant of ANGPTL3 was determined to be less active than wild-type ANGPTL3 in increasing mouse plasma triglyceride levels but not in inhibiting lipoprotein lipase activity. These findings suggest that the cleavage of ANGPTL3 is important for the activation of ANGPTL3 in vivo

Angiopoietin-like protein 3 regulation

Angiopoietin-like protein 3 and Liver X Receptor

The Angptl3 gene is a direct target of the liver X receptor (LXR). In Angptl3– deficient C57BL/6J mice, LXR ligand did not cause hypertriglyce­ridemia but accumulation of triglyceride in the liver. Induction of hepatic Angptl3 accounts for hypertriglyce­ridemia associated with the treatment of LXR ligand. Angptl3 expression and plasma ANGPTL3 levels were increased in leptin-resistant C57BL/6J(db/db) and -deficient C57BL/6J(ob/ob) mice, relative to the control. Leptin supplements decreased Angptl3 gene expression and plasma ANGPTL3 in C57BL/6J(ob/ob) mice. The changes of Angptl3 were associated with alterations of plasma TG and FFA levels.

Angiopoietin-like protein 3 and thyroid hormone

Thyroid hormone down-regulates ANGPTL3 gene expression in hypothyroid rats. ANGPTL3 is a novel TRbeta target gene and provides a new potential mechanism to explain the hypotriglyceridemic properties of TRbeta agonists in vivo.

Angiopoietin-like protein 3 and leptin

Leptin treatment directly suppressed Angptl3 gene expression in hepatocytes.

Angiopoietin-like protein 3 and insulin

Insulin treatment of hepatocytes decreased Angptl3 gene expression and protein secretion. These results suggest that elevated ANGPTL3 by leptin- or insulin-resistance is attributed to increased plasma TG and FFA concentrations in obesity. Angptl3 gene expression and plasma protein levels were also increased in insulin-deficient streptozotocin-treated mice.

Angiopoietin-like protein 3 action

Human ANGPTL3 treatment increased both plasma triglyceride and FFA. ANGPTL3 inhibited the activity of lipoprotein lipase, which accounted for the increase of plasma triglyceride. ANGPTL3 targets on adipose cells and induces lipolysis.

In mice, reduction of Angptl3 expression is protective against hyperlipidemia and atherosclerosis, even in the absence of apoE, owing to the enhanced catabolism and clearance of TG-rich lipoproteins.

Angiopoietin-like protein 3 in obese and diabetic mice

The level of ANGPTL3 mRNA was increased approximately 2.2-fold in the livers of streptozotocin (STZ) diabetic mice, and this effect was reversed by administration of insulin. Furthermore, the level of ANGPTL3 mRNA was increased more than 3.0-fold in type 2 diabetic obese mice, db/db mice, as compared with age matched lean littermates. The hepatic level of ANGPTL3 protein was also increased in these diabetic mice to an extent similar to that of ANGPTL3 mRNA. Thus, the expression of ANGPTL3 was enhanced in both insulin-deficient and -resistant diabetic states. These results strongly suggest ANGPTL3 to play an important role in hyperlipidemia in diabetes.

Angiopoietin-like protein 3 in Circulation

It was found that probands with metabolic syndrome or familiar hypercholeste­rolemia had higher Angptl3 values than healthy individuals. Angptl3 values correlated with systolic blood pressure, LDL and adipocyte fatty acid –binding protein A-FABP.

Carotid artery intima-media thickness showed a significant positive correlation with plasma ANGPTL3 and other classical risk factors such as age, blood pressure, plasma glucose and lipid levels. The positive association between ANGPTL3 and carotid artery intima-media thickness remained significant after adjustment for age, sex, smoking, body mass index, systolic blood pressure, plasma glucose, insulin resistance index, triglyceride, and high-density and low-density lipoprotein cholesterol levels. ANGPTL3 also showed a positive association with FA-IMT independent of these factors

Catalog Number	Species	Analyte	Assay	Regulatory	Format
RD191092200R	Human	Angiopoietin-like Protein 3	Sandwich ELISA, Biotin-labelled antibody	RUO	96 wells (1 kit)