ABSTRACT
We investigated whether: (1) liver X receptor (LXR)-driven induction of high-density lipoprotein cholesterol (HDL-C) and other LXR-mediated effects on cholesterol metabolism depend on intestinal cholesterol absorption; and (2) combined treatment with the LXR agonist GW3965 and the cholesterol absorption inhibitor ezetimibe results in synergistic effects on cholesterol metabolism that could be beneficial for treatment of atherosclerosis. Mice were fed 0.2 % cholesterol and treated with GW3965+ezetimibe, GW3965 or ezetimibe. GW3965+ezetimibe treatment elevated serum HDL-C and Apolipoprotein (Apo) AI, effectively reduced the intestinal cholesterol absorption and increased the excretion of faecal neutral sterols. No changes in intestinal ATP-binding cassette (ABC) A1 or ABCG5 protein expression were observed, despite increased mRNA expression, while hepatic ABCA1 was slightly reduced. The combined treatment caused a pronounced down-regulation of intestinal Niemann-Pick C1-like 1 (NPC1L1) and reduced hepatic and intestinal cholesterol levels. GW3965 did not affect the intestinal cholesterol absorption, but increased serum HDL-C and ApoAI levels. GW3965 also increased Apoa1 mRNA levels in primary mouse hepatocytes and HEPA1-6 cells. Ezetimibe reduced the intestinal cholesterol absorption, ABCA1 and ABCG5, but did not affect the serum HDL-C or ApoAI levels. Thus, the LXR-driven induction of HDL-C and ApoAI was independent of the intestinal cholesterol absorption and increased expression of intestinal or hepatic ABCA1 was not required. Inhibited influx of cholesterol via NPC1L1 and/or low levels of intracellular cholesterol prevented post-transcriptional expression of intestinal ABCA1 and ABCG5, despite increased mRNA levels. Combined LXR activation and blocked intestinal cholesterol absorption induced effective faecal elimination of cholesterol.
Subject(s)
ATP Binding Cassette Transporter 1/metabolism , Cholesterol, HDL/metabolism , Cholesterol/metabolism , Orphan Nuclear Receptors/metabolism , ATP Binding Cassette Transporter 1/genetics , ATP Binding Cassette Transporter, Subfamily G, Member 5 , ATP-Binding Cassette Transporters/genetics , ATP-Binding Cassette Transporters/metabolism , Animals , Anticholesteremic Agents/pharmacology , Apolipoprotein A-I/blood , Apolipoprotein A-I/metabolism , Azetidines/pharmacology , Benzoates/pharmacology , Benzylamines/pharmacology , Blotting, Western , Cell Line, Tumor , Cells, Cultured , Cholesterol/blood , Cholesterol, HDL/blood , Ezetimibe , Feces/chemistry , Gene Expression/drug effects , Hepatocytes/drug effects , Hepatocytes/metabolism , Intestinal Absorption/drug effects , Intestinal Mucosa/metabolism , Intestines/drug effects , Lipoproteins/genetics , Lipoproteins/metabolism , Liver/drug effects , Liver/metabolism , Liver X Receptors , Male , Membrane Transport Proteins/genetics , Membrane Transport Proteins/metabolism , Mice , Mice, Inbred C57BL , Orphan Nuclear Receptors/agonists , Reverse Transcriptase Polymerase Chain ReactionABSTRACT
Endocannabinoids have been implicated in cancer development and cause heterogenous effects in tumor cells, by inducing apoptosis, reducing migration, causing anti-angiogenic activity and alterations in the cell cycle resulting in growth arrest. Recently, several novel amides of fatty acids that are structurally related to endocannabinoids have been isolated from mammalian sources, although the functions of these fatty amides are not well studied. One group of these novel fatty acid amides are the N-acyl taurines (fatty acids conjugated to the amino acid taurine). This study examined if N-acyl taurines, specifically N-arachidonoyl taurine and N-oleoyl taurine could function in a similar way to endocannabinoids and result in cell cycle alterations or growth arrest in the human prostate adenocarcinoma cell line PC-3. PC-3 cells were treated with various concentrations of N-arachidonoyl taurine and N-oleoyl taurine and cell proliferation and viability was measured using resazurin and colony formation assays. Effects of N-acyl taurines on the cell cycle was measured using FACS analysis. Treatment with N-arachidonoyl taurine and N-oleoyl taurine resulted in a significant reduction in proliferation of PC-3 cells, even at concentrations as low as 1 µM. Treatment with N-oleoyl taurine resulted in an increased number of cells in the subG1 population, suggesting apoptosis, and a lower number of cells in S-phase of the cell cycle. In summary, our results show that novel biologically active lipids, the N-acyl taurines, result in reduced proliferation in PC-3 cells.
Subject(s)
Adenocarcinoma/drug therapy , Arachidonic Acids/pharmacology , Prostate/drug effects , Prostatic Neoplasms/drug therapy , Taurine/analogs & derivatives , Adenocarcinoma/pathology , Apoptosis/drug effects , Arachidonic Acids/chemistry , Cannabinoid Receptor Modulators/chemistry , Cannabinoid Receptor Modulators/pharmacology , Cell Count , Cell Cycle/drug effects , Cell Line, Tumor , Cell Proliferation/drug effects , Cell Survival/drug effects , Dose-Response Relationship, Drug , Flow Cytometry , Humans , Male , Oxazines , Prostate/pathology , Prostatic Neoplasms/pathology , Taurine/chemistry , Taurine/pharmacology , XanthenesABSTRACT
To investigate the effects of abolished cholic acid (CA) synthesis in the ApoE knockout model [apolipoprotein E (apoE) KO],a double-knockout (DKO) mouse model was created by crossbreeding Cyp8b1 knockout mice (Cyp8b1 KO), unable to synthesize the primary bile acid CA, with apoE KO mice. After 5 months of cholesterol feeding, the development of atherosclerotic plaques in the proximal aorta was 50% less in the DKO mice compared with the apoE KO mice. This effect was associated with reduced intestinal cholesterol absorption, decreased levels of apoB-containing lipoproteins in the plasma, enhanced bile acid synthesis, reduced hepatic cholesteryl esters, and decreased hepatic activity of ACAT2. The upregulation of Cyp7a1 in DKO mice seemed primarily caused by reduced expression of the intestinal peptide FGF15. Treatment of DKO mice with the farnesoid X receptor (FXR) agonist GW4064 did not alter the intestinal cholesterol absorption, suggesting that the action of CA in this process is confined mainly to formation of intraluminal micelles and less to its ability to activate the nuclear receptor FXR. Inhibition of CA synthesis may offer a therapeutic strategy for the treatment of hyperlipidemic conditions that lead to atherosclerosis.
