Intestinal DGAT1 deficiency reduces postprandial triglyceride and retinyl ester excursions by inhibiting chylomicron secretion and delaying gastric emptying

Acyl CoA:diacylglycerol acyltransferase (DGAT) 1 catalyzes the final step of triglyceride (TG) synthesis. We show that acute administration of a DGAT1 inhibitor (DGAT1i) by oral gavage or genetic deletion of intestinal Dgat1 (intestine-Dgat1^−/−) markedly reduced postprandial plasma TG and retinyl ester excursions by inhibiting chylomicron secretion in mice. Loss of DGAT1 activity did not affect the efficiency of retinol esterification, but it did reduce TG and retinoid accumulation in the small intestine. In contrast, inhibition of microsomal triglyceride transfer protein (MTP) reduced chylomicron secretion after oral fat/retinol loads, but with accumulation of dietary TG and retinoids in the small intestine. Lack of intestinal accumulation of TG and retinoids in DGAT1i-treated or intestine-Dgat1^−/− mice resulted, in part, from delayed gastric emptying associated with increased plasma levels of glucagon-like peptide (GLP)-1. However, neither bypassing the stomach through duodenal oil injection nor inhibiting the receptor for GLP-1 normalized postprandial TG or retinyl esters excursions in the absence of DGAT1 activity. In summary, intestinal DGAT1 inhibition or deficiency acutely delayed gastric emptying and inhibited chylomicron secretion; however, the latter occurred when gastric emptying was normal or when lipid was administered directly into the small intestine. Long-term hepatic retinoid metabolism was not impacted by DGAT1 inhibition.     

Intestine-specific expression of acyl CoA:diacylglycerol acyltransferase 1 reverses resistance to diet-induced hepatic steatosis and obesity in Dgat1−/− mice

Mice deficient in acyl-CoA:diacylglycerol acyltransferase 1 (DGAT1), a key enzyme in triacylglycerol (TG) biosynthesis, are resistant to high-fat (HF) diet-induced hepatic steatosis and obesity. DGAT1-deficient (Dgat1^−/−) mice have no defect in quantitative absorption of dietary fat; however, they have abnormally high levels of TG stored in the cytoplasm of enterocytes, and they have a reduced postprandial triglyceridemic response. We generated mice expressing DGAT1 only in the intestine (Dgat1^IntONLY) to determine whether this phenotype contributes to resistance to HF diet-induced hepatic steatosis and obesity in Dgat1^−/− mice. Despite lacking DGAT1 in liver and adipose tissue, we found that Dgat1^IntONLY mice are not resistant to HF diet-induced hepatic steatosis or obesity. The results presented demonstrate that intestinal DGAT1 stimulates dietary fat secretion out of enterocytes and that altering this cellular function alters the fate of dietary fat in specific tissues.     

DGAT1 Expression Increases Heart Triglyceride Content but Ameliorates Lipotoxicity

Intracellular lipid accumulation in the heart is associated with cardiomyopathy, yet the precise role of triglyceride (TG) remains unclear. With exercise, wild type hearts develop physiologic hypertrophy. This was associated with greater TG stores and a marked induction of the TG-synthesizing enzyme diacylglycerol (DAG) acyltransferase 1 (DGAT1). Transgenic overexpression of DGAT1 in the heart using the cardiomyocyte- specific α-myosin heavy chain (MHC) promoter led to approximately a doubling of DGAT activity and TG content and reductions of ∼35% in cardiac ceramide, 26% in DAG, and 20% in free fatty acid levels. Cardiac function assessed by echocardiography and cardiac catheterization was unaffected. These mice were then crossed with animals expressing long-chain acyl-CoA synthetase via the MHC promoter (MHC-ACS), which develop lipotoxic cardiomyopathy. MHC-DGAT1XMHC-ACS double transgenic male mice had improved heart function; fractional shortening increased by 74%, and diastolic function improved compared with MHC-ACS mice. The improvement of heart function correlated with a reduction in cardiac DAG and ceramide and reduced cardiomyocyte apoptosis but increased fatty acid oxidation. In addition, the survival of the mice was improved. Our study indicates that TG is not likely to be a toxic lipid species directly, but rather it is a feature of physiologic hypertrophy and may serve a cytoprotective role in lipid overload states. Moreover, induction of DGAT1 could be beneficial in the setting of excess heart accumulation of toxic lipids.     

DGAT1 deficiency decreases PPAR expression and does not lead to lipotoxicity in cardiac and skeletal muscle

Diacylglycerol (DAG) acyl transferase 1 (Dgat1) knockout ((-/-)) mice are resistant to high-fat-induced obesity and insulin resistance, but the reasons are unclear. Dgat1(-/-) mice had reduced mRNA levels of all three Ppar genes and genes involved in fatty acid oxidation in the myocardium of Dgat1(-/-) mice. Although DGAT1 converts DAG to triglyceride (TG), tissue levels of DAG were not increased in Dgat1(-/-) mice. Hearts of chow-diet Dgat1(-/-) mice were larger than those of wild-type (WT) mice, but cardiac function was normal. Skeletal muscles from Dgat1(-/-) mice were also larger. Muscle hypertrophy factors phospho-AKT and phospho-mTOR were increased in Dgat1(-/-) cardiac and skeletal muscle. In contrast to muscle, liver from Dgat1(-/-) mice had no reduction in mRNA levels of genes mediating fatty acid oxidation. Glucose uptake was increased in cardiac and skeletal muscle in Dgat1(-/-) mice. Treatment with an inhibitor specific for DGAT1 led to similarly striking reductions in mRNA levels of genes mediating fatty acid oxidation in cardiac and skeletal muscle. These changes were reproduced in cultured myocytes with the DGAT1 inhibitor, which also blocked the increase in mRNA levels of Ppar genes and their targets induced by palmitic acid. Thus, loss of DGAT1 activity in muscles decreases mRNA levels of genes involved in lipid uptake and oxidation.     

Integrin α1-null Mice Exhibit Improved Fatty Liver When Fed a High Fat Diet Despite Severe Hepatic Insulin Resistance

Hepatic insulin resistance is associated with increased collagen. Integrin α1β1 is a collagen-binding receptor expressed on hepatocytes. Here, we show that expression of the α1 subunit is increased in hepatocytes isolated from high fat (HF)-fed mice. To determine whether the integrin α1 subunit protects against impairments in hepatic glucose metabolism, we analyzed glucose tolerance and insulin sensitivity in HF-fed integrin α1-null (itga1^−/−) and wild-type (itga1^+/+) littermates. Using the insulin clamp, we found that insulin-stimulated hepatic glucose production was suppressed by ∼50% in HF-fed itga1^+/+ mice. In contrast, it was not suppressed in HF-fed itga1^−/− mice, indicating severe hepatic insulin resistance. This was associated with decreased hepatic insulin signaling in HF-fed itga1^−/− mice. Interestingly, hepatic triglyceride and diglyceride contents were normalized to chow-fed levels in HF-fed itga1^−/− mice. This indicates that hepatic steatosis is dissociated from insulin resistance in HF-fed itga1^−/− mice. The decrease in hepatic lipid accumulation in HF-fed itga1^−/− mice was associated with altered free fatty acid metabolism. These studies establish a role for integrin signaling in facilitating hepatic insulin action while promoting lipid accumulation in mice challenged with a HF diet.     

Endogenous levels of 1-O-acylceramides increase upon acidic ceramidase deficiency and decrease due to loss of Dgat1 in a tissue-dependent manner

Except for epidermis and liver, little is known about endogenous expression of 1-O-acylceramides (1-OACs) in mammalian tissue. Therefore, we screened several organs (brain, lung, liver, spleen, lymph nodes, heart, kidney, thymus, small intestine, and colon) from mice for the presence of 1-OACs by LC-MS². In most organs, low levels of about 0.25–1.3 pmol 1-OACs/mg wet weight were recorded. Higher levels were detected in liver, small and large intestines, with about 4–13 pmol 1-OACs/mg wet weight. 1-OACs were esterified mainly with palmitic, stearic, or oleic acids. Esterification with saturated very long-chain fatty acids, as in epidermis, was not observed. Western-type diet induced 3-fold increased 1-OAC levels in mice livers while ceramides were unaltered. In a mouse model of Farber disease with a decrease of acid ceramidase activity, we observed a strong, up to 50-fold increase of 1-OACs in lung, thymus, and spleen. In contrast, 1-OAC levels were reduced 0.54-fold in liver. Only in lung 1-OAC levels correlated to changes in ceramide levels - indicating tissue-specific mechanisms of regulation. Glucosylceramide synthase deficiency in liver did not cause changes in 1-OAC or ceramide levels, whereas increased ceramide levels in glucosylceramide synthase-deficient small intestine caused an increase in 1-OAC levels. Deficiency of Dgat1 in mice resulted in a reduction of 1-OACs to 30% in colon, but not in small intestine and liver, going along with constant free ceramides levels. From these data, we conclude that Dgat1 as well as lysosomal lipid metabolism contribute in vivo to homeostatic 1-OAC levels in an organ-specific manner.     

Two Carotenoid Oxygenases Contribute to Mammalian Provitamin A Metabolism

Mammalian genomes encode two provitamin A-converting enzymes as follows: the β-carotene-15,15′-oxygenase (BCO1) and the β-carotene-9′,10′-oxygenase (BCO2). Symmetric cleavage by BCO1 yields retinoids (β-15′-apocarotenoids, C₂₀), whereas eccentric cleavage by BCO2 produces long-chain (>C₂₀) apocarotenoids. Here, we used genetic and biochemical approaches to clarify the contribution of these enzymes to provitamin A metabolism. We subjected wild type, Bco1^−/−, Bco2^−/−, and Bco1^−/−Bco2^−/− double knock-out mice to a controlled diet providing β-carotene as the sole source for apocarotenoid production. This study revealed that BCO1 is critical for retinoid homeostasis. Genetic disruption of BCO1 resulted in β-carotene accumulation and vitamin A deficiency accompanied by a BCO2-dependent production of minor amounts of β-apo-10′-carotenol (APO10ol). We found that APO10ol can be esterified and transported by the same proteins as vitamin A but with a lower affinity and slower reaction kinetics. In wild type mice, APO10ol was converted to retinoids by BCO1. We also show that a stepwise cleavage by BCO2 and BCO1 with APO10ol as an intermediate could provide a mechanism to tailor asymmetric carotenoids such as β-cryptoxanthin for vitamin A production. In conclusion, our study provides evidence that mammals employ both carotenoid oxygenases to synthesize retinoids from provitamin A carotenoids.     

5-Aminoimidazole-4-carboxamide-1-β-d-ribofuranoside (AICAR) Effect on Glucose Production,but Not Energy Metabolism,Is Independent of Hepatic AMPK in Vivo

Metabolic stress, as well as several antidiabetic agents, increases hepatic nucleotide monophosphate (NMP) levels, activates AMP-activated protein kinase (AMPK), and suppresses glucose production. We tested the necessity of hepatic AMPK for the in vivo effects of an acute elevation in NMP on metabolism. 5-Aminoimidazole-4-carboxamide 1-β-d-ribofuranoside (AICAR; 8 mg·kg⁻¹·min⁻¹)-euglycemic clamps were performed to elicit an increase in NMP in wild type (α1α2^lox/lox) and liver-specific AMPK knock-out mice (α1α2^lox/lox + Albcre) in the presence of fixed glucose. Glucose kinetics were equivalent in 5-h fasted α1α2^lox/lox and α1α2^lox/lox + Albcre mice. AMPK was not required for AICAR-mediated suppression of glucose production and increased glucose disappearance. These results demonstrate that AMPK is unnecessary for normal 5-h fasting glucose kinetics and AICAR-mediated inhibition of glucose production. Moreover, plasma fatty acids and triglycerides also decreased independently of hepatic AMPK during AICAR administration. Although the glucoregulatory effects of AICAR were shown to be independent of AMPK, these studies provide in vivo support for the AMPK energy sensor paradigm. AICAR reduced hepatic energy charge by ∼20% in α1α2^lox/lox, which was exacerbated by ∼2-fold in α1α2^lox/lox + Albcre. This corresponded to a ∼6-fold rise in AMP/ATP in α1α2^lox/lox + Albcre. Consistent with the effects on adenine nucleotides, maximal mitochondrial respiration was ∼30% lower in α1α2^lox/lox + Albcre than α1α2^lox/lox livers. Mitochondrial oxidative phosphorylation efficiency was reduced by 25%. In summary, these results demonstrate that the NMP capacity to inhibit glucose production in vivo is independent of liver AMPK. In contrast, AMPK promotes mitochondrial function and protects against a more precipitous fall in ATP during AICAR administration.     

Two-pore Channels (TPC2s) and Nicotinic Acid Adenine Dinucleotide Phosphate (NAADP) at Lysosomal-Sarcoplasmic Reticular Junctions Contribute to Acute and Chronic β-Adrenoceptor Signaling in the Heart

Ca²⁺-permeable type 2 two-pore channels (TPC2) are lysosomal proteins required for nicotinic acid adenine dinucleotide phosphate (NAADP)-evoked Ca²⁺ release in many diverse cell types. Here, we investigate the importance of TPC2 proteins for the physiology and pathophysiology of the heart. NAADP-AM failed to enhance Ca²⁺ responses in cardiac myocytes from Tpcn2^−/− mice, unlike myocytes from wild-type (WT) mice. Ca²⁺/calmodulin-dependent protein kinase II inhibitors suppressed actions of NAADP in myocytes. Ca²⁺ transients and contractions accompanying action potentials were increased by isoproterenol in myocytes from WT mice, but these effects of β-adrenoreceptor stimulation were reduced in myocytes from Tpcn2^−/− mice. Increases in amplitude of L-type Ca²⁺ currents evoked by isoproterenol remained unchanged in myocytes from Tpcn2^−/− mice showing no loss of β-adrenoceptors or coupling mechanisms. Whole hearts from Tpcn2^−/− mice also showed reduced inotropic effects of isoproterenol and a reduced tendency for arrhythmias following acute β-adrenoreceptor stimulation. Hearts from Tpcn2^−/− mice chronically exposed to isoproterenol showed less cardiac hypertrophy and increased threshold for arrhythmogenesis compared with WT controls. Electron microscopy showed that lysosomes form close contacts with the sarcoplasmic reticulum (separation ∼25 nm). We propose that Ca²⁺-signaling nanodomains between lysosomes and sarcoplasmic reticulum dependent on NAADP and TPC2 comprise an important element in β-adrenoreceptor signal transduction in cardiac myocytes. In summary, our observations define a role for NAADP and TPC2 at lysosomal/sarcoplasmic reticulum junctions as unexpected but major contributors in the acute actions of β-adrenergic signaling in the heart and also in stress pathways linking chronic stimulation of β-adrenoceptors to hypertrophy and associated arrhythmias.     

Mechanism of Hyperinsulinism in Short-chain 3-Hydroxyacyl-CoA Dehydrogenase Deficiency Involves Activation of Glutamate Dehydrogenase

The mechanism of insulin dysregulation in children with hyperinsulinism associated with inactivating mutations of short-chain 3-hydroxyacyl-CoA dehydrogenase (SCHAD) was examined in mice with a knock-out of the hadh gene (hadh^−/−). The hadh^−/− mice had reduced levels of plasma glucose and elevated plasma insulin levels, similar to children with SCHAD deficiency. hadh^−/− mice were hypersensitive to oral amino acid with decrease of glucose level and elevation of insulin. Hypersensitivity to oral amino acid in hadh^−/− mice can be explained by abnormal insulin responses to a physiological mixture of amino acids and increased sensitivity to leucine stimulation in isolated perifused islets. Measurement of cytosolic calcium showed normal basal levels and abnormal responses to amino acids in hadh^−/− islets. Leucine, glutamine, and alanine are responsible for amino acid hypersensitivity in islets. hadh^−/− islets have lower intracellular glutamate and aspartate levels, and this decrease can be prevented by high glucose. hadh^−/− islets also have increased [U-¹⁴C]glutamine oxidation. In contrast, hadh^−/− mice have similar glucose tolerance and insulin sensitivity compared with controls. Perifused hadh^−/− islets showed no differences from controls in response to glucose-stimulated insulin secretion, even with addition of either a medium-chain fatty acid (octanoate) or a long-chain fatty acid (palmitate). Pull-down experiments with SCHAD, anti-SCHAD, or anti-GDH antibodies showed protein-protein interactions between SCHAD and GDH. GDH enzyme kinetics of hadh^−/− islets showed an increase in GDH affinity for its substrate, α-ketoglutarate. These studies indicate that SCHAD deficiency causes hyperinsulinism by activation of GDH via loss of inhibitory regulation of GDH by SCHAD.     

Loss of stearoyl-CoA desaturase 1 rescues cardiac function in obese leptin-deficient mice

The heart of leptin-deficient ob/ob mice is characterized by pathologic left ventricular hypertrophy along with elevated triglyceride (TG) content, increased stearoyl-CoA desaturase (SCD) activity, and increased myocyte apoptosis. In the present study, using an ob/ob;SCD1^−/− mouse model, we tested the hypothesis that lack of SCD1 could improve steatosis and left ventricle (LV) function in leptin deficiency. We show that disruption of the SCD1 gene improves cardiac function in ob/ob mice by correcting systolic and diastolic dysfunction without affecting levels of plasma TG and FFA. The improvement is associated with reduced expression of genes involved in FA transport and lipid synthesis in the heart, as well as reduction in cardiac FFA, diacylglycerol, TG, and ceramide levels. The rate of FA β-oxidation is also significantly lower in the heart of ob/ob;SCD1^−/− mice compared with ob/ob controls. Moreover, SCD1 deficiency reduces cardiac apoptosis in ob/ob mice due to increased expression of antiapoptotic factor Bcl-2 and inhibition of inducible nitric oxide synthase and caspase-3 activities. Reduction in myocardial lipid accumulation and inhibition of apoptosis appear to be one of the main mechanisms responsible for improved LV function in ob/ob mice caused by SCD1 deficiency.     

Role of Stearoyl-CoA Desaturase-1 in Skin Integrity and Whole Body Energy Balance

The skin is the single largest organ in humans, serving as a major barrier to infection, water loss, and abrasion. The functional diversity of skin requires the synthesis of large amounts of lipids, such as triglycerides, wax esters, squalene, ceramides, free cholesterol, free fatty acids, and cholesterol and retinyl esters. Some of these lipids are used as cell membrane components, signaling molecules, and a source of energy. An important class of lipid metabolism enzymes expressed in skin is the Δ⁹-desaturases, which catalyze the synthesis in Δ⁹-monounsaturated lipids, primarily oleoyl-CoA (18:1n-9) and palmitoyl-CoA (16:1n-7), the major monounsaturated fatty acids in cutaneous lipids. Mice with a deletion of the Δ⁹-desaturase-1 isoform (SCD1) either globally (Scd1^−/−) or specifically in the skin (skin-specific Scd1-knockout; SKO) present with marked changes in cutaneous lipids and skin integrity. Interestingly, these mice also exhibit increased whole body energy expenditure, protection against diet-induced adiposity, hepatic steatosis, and glucose intolerance. The increased energy expenditure in skin-specific Scd1-knockout (SKO) mice is a surprising phenotype, as it links cutaneous lipid homeostasis with whole body energy balance. This minireview summarizes the role of skin SCD1 in regulating skin integrity and whole body energy homeostasis and offers a discussion of potential pathways that may connect these seemingly disparate phenotypes.     

The Farnesoid X Receptor Regulates Adipocyte Differentiation and Function by Promoting Peroxisome Proliferator-activated Receptor-γ and Interfering with the Wnt/β-Catenin Pathways

The bile acid receptor farnesoid X receptor (FXR) is expressed in adipose tissue, but its function remains poorly defined. Peroxisome proliferator-activated receptor-γ (PPARγ) is a master regulator of adipocyte differentiation and function. The aim of this study was to analyze the role of FXR in adipocyte function and to assess whether it modulates PPARγ action. Therefore, we tested the responsiveness of FXR-deficient mice (FXR^−/−) and cells to the PPARγ activator rosiglitazone. Our results show that genetically obese FXR^−/−/ob/ob mice displayed a resistance to rosiglitazone treatment. In vitro, rosiglitazone treatment did not induce normal adipocyte differentiation and lipid droplet formation in FXR^−/− mouse embryonic fibroblasts (MEFs) and preadipocytes. Moreover, FXR^−/− MEFs displayed both an increased lipolysis and a decreased de novo lipogenesis, resulting in reduced intracellular triglyceride content, even upon PPARγ activation. Retroviral-mediated FXR re-expression in FXR^−/− MEFs restored the induction of adipogenic marker genes during rosiglitazone-forced adipocyte differentiation. The expression of Wnt/β-catenin pathway and target genes was increased in FXR^−/− adipose tissue and MEFs. Moreover, the expression of several endogenous inhibitors of this pathway was decreased early during the adipocyte differentiation of FXR^−/− MEFs. These findings demonstrate that FXR regulates adipocyte differentiation and function by regulating two counteracting pathways of adipocyte differentiation, the PPARγ and Wnt/β-catenin pathways.     

Membrane-bound sn-1,2-diacylglycerols explain the dissociation of hepatic insulin resistance from hepatic steatosis in MTTP knockout mice

Microsomal triglyceride transfer protein (MTTP) deficiency results in a syndrome of hypolipidemia and accelerated NAFLD. Animal models of decreased hepatic MTTP activity have revealed an unexplained dissociation between hepatic steatosis and hepatic insulin resistance. Here, we performed comprehensive metabolic phenotyping of liver-specific MTTP knockout (L-Mttp^−/−) mice and age-weight matched wild-type control mice. Young (10–12-week-old) L-Mttp^−/− mice exhibited hepatic steatosis and increased DAG content; however, the increase in hepatic DAG content was partitioned to the lipid droplet and was not increased in the plasma membrane. Young L-Mttp^−/− mice also manifested normal hepatic insulin sensitivity, as assessed by hyperinsulinemic-euglycemic clamps, no PKCε activation, and normal hepatic insulin signaling from the insulin receptor through AKT Ser/Thr kinase. In contrast, aged (10-month-old) L-Mttp^−/− mice exhibited glucose intolerance and hepatic insulin resistance along with an increase in hepatic plasma membrane sn-1,2-DAG content and PKCε activation. Treatment with a functionally liver-targeted mitochondrial uncoupler protected the aged L-Mttp^−/− mice against the development of hepatic steatosis, increased plasma membrane sn-1,2-DAG content, PKCε activation, and hepatic insulin resistance. Furthermore, increased hepatic insulin sensitivity in the aged controlled-release mitochondrial protonophore-treated L-Mttp^−/− mice was not associated with any reductions in hepatic ceramide content. Taken together, these data demonstrate that differences in the intracellular compartmentation of sn-1,2-DAGs in the lipid droplet versus plasma membrane explains the dissociation of NAFLD/lipid-induced hepatic insulin resistance in young L-Mttp^−/− mice as well as the development of lipid-induced hepatic insulin resistance in aged L-Mttp^−/− mice.     

Up-regulation of Cholesterol Absorption Is a Mechanism for Cholecystokinin-induced Hypercholesterolemia

Excessive absorption of intestinal cholesterol is a risk factor for atherosclerosis. This report examines the effect of cholecystokinin (CCK) on plasma cholesterol level and intestinal cholesterol absorption using the in vivo models of C57BL/6 wild-type and low density lipoprotein receptor knock-out (LDLR^−/−) mice. These data were supported by in vitro studies involving mouse primary intestinal epithelial cells and human Caco-2 cells; both express CCK receptor 1 and 2 (CCK1R and CCK2R). We found that intravenous injection of [Thr²⁸,Nle³¹]CCK increased plasma cholesterol levels and intestinal cholesterol absorption in both wild-type and LDLR^−/− mice. Treatment of mouse primary intestinal epithelial cells with [Thr²⁸,Nle³¹]CCK increased cholesterol absorption, whereas selective inhibition of CCK1R and CCK2R with antagonists attenuated CCK-induced cholesterol absorption. In Caco-2 cells, CCK enhanced CCK1R/CCK2R heterodimerization. Knockdown of both CCK1R and CCK2 or either one of them diminished CCK-induced cholesterol absorption to the same extent. CCK also increased cell surface-associated NPC1L1 (Niemann-Pick C1-like 1) transporters but did not alter their total protein expression. Inhibition or knockdown of NPC1L1 attenuated CCK-induced cholesterol absorption. CCK enhanced phosphatidylinositide 3-kinase (PI3K) and Akt phosphorylation and augmented the interaction between NPC1L1 and Rab11a (Rab-GTPase-11a), whereas knockdown of CCK receptors or inhibition of G protein βγ dimer (Gβγ) diminished CCK-induced PI3K and Akt phosphorylation. Inhibition of PI3K and Akt or knockdown of PI3K diminished CCK-induced NPC1L1-Rab11a interaction and cholesterol absorption. Knockdown of Rab11a suppressed CCK-induced NPC1L1 translocation and cholesterol absorption. These data imply that CCK enhances cholesterol absorption by activation of a pathway involving CCK1R/CCK2R, Gβγ, PI3K, Akt, Rab11a, and NPC1L.