Hepatic accumulation of intestinal cholesterol is decreased and fecal cholesterol excretion is increased in mice fed a high-fat diet supplemented with milk phospholipids

<p>Abstract</p> <p>Background</p> <p>Milk phospholipids (PLs) reduce liver lipid levels when given as a dietary supplement to mice fed a high-fat diet. We have speculated that this might be due to reduced intestinal cholesterol uptake.</p> <p>Methods</p> <p>Mice were given a high-fat diet for 3 or 5 weeks that had no added PL or that were supplemented with 1.2% by wt PL from cow's milk. Two milk PL preparations were investigated: a) a PL-rich dairy milk extract (PLRDME), and b) a commercially-available milk PL concentrate (PC-700). Intestinal cholesterol uptake was assessed by measuring fecal and hepatic radioactivity after intragastric administration of [¹⁴C]cholesterol and [³H]sitostanol. Fecal and hepatic lipids were measured enzymatically and by ESI-MS/MS.</p> <p>Results</p> <p>Both PL preparations led to significant decreases in total liver cholesterol and triglyceride (-20% to -60%, <it>P </it>< 0.05). Hepatic accumulation of intragastrically-administered [¹⁴C]cholesterol was significantly less (-30% to -60%, <it>P </it>< 0.05) and fecal excretion of [¹⁴C]cholesterol and unlabeled cholesterol was significantly higher in PL-supplemented mice (+15% to +30%, <it>P </it>< 0.05). Liver cholesterol and triglyceride levels were positively correlated with hepatic accumulation of intragastrically-administered [¹⁴C]cholesterol (<it>P </it>< 0.001) and negatively correlated with fecal excretion of [¹⁴C]cholesterol (<it>P </it>< 0.05). Increased PL and ceramide levels in the diet of mice supplemented with milk PL were associated with significantly higher levels of fecal PL and ceramide excretion, but reduced levels of hepatic PL and ceramide, specifically, phosphatidylcholine (-21%, <it>P </it>< 0.05) and monohexosylceramide (-33%, <it>P </it>< 0.01).</p> <p>Conclusion</p> <p>These results indicate that milk PL extracts reduce hepatic accumulation of intestinal cholesterol and increase fecal cholesterol excretion when given to mice fed a high-fat diet.</p

Role of ABCG1 and ABCA1 in regulation of neuronal cholesterol efflux to apolipoprotein E discs and suppression of amyloid-β peptide generation

Maintenance of an adequate supply of cholesterol is important for neuronal function, whereas excess cholesterol promotes amyloid precursor protein (APP) cleavage generating toxic amyloid-β (Aβ) peptides. To gain insights into the pathways that regulate neuronal cholesterol level, we investigated the potential for reconstituted apolipoprotein E (apoE) discs, resembling nascent lipoprotein complexes in the central nervous system, to stimulate neuronal [3H]cholesterol efflux. ApoE discs potently accelerated cholesterol efflux from primary human neurons and cell lines. The process was saturable (17.5 μg of apoE/ml) and was not influenced by APOE genotype. High performance liquid chromatography analysis of cholesterol and cholesterol metabolites effluxed from neurons indicated that <25% of the released cholesterol was modified to polar products (e.g. 24-hydroxycholesterol) that diffuse from neuronal membranes. Thus, most cholesterol (∼75%) appeared to be effluxed from neurons in a native state via a transporter pathway. ATP-binding cassette transporters ABCA1, ABCA2, and ABCG1 were detected in neurons and neuroblastoma cell lines and expression of these cDNAs revealed that ABCA1 and ABCG1 stimulated cholesterol efflux to apoE discs. In addition, ABCA1 and ABCG1 expression in Chinese hamster ovary cells that stably express human APP significantly reduced Aβ generation, whereas ABCA2 did not modulate either cholesterol efflux or Aβ generation. These data indicate that ABCA1 and ABCG1 play a significant role in the regulation of neuronal cholesterol efflux to apoE discs and in suppression of APP processing to generate Aβ peptides

Dietary Phospholipids and Intestinal Cholesterol Absorption

Experiments carried out with cultured cells and in experimental animals have consistently shown that phospholipids (PLs) can inhibit intestinal cholesterol absorption. Limited evidence from clinical studies suggests that dietary PL supplementation has a similar effect in man. A number of biological mechanisms have been proposed in order to explain how PL in the gut lumen is able to affect cholesterol uptake by the gut mucosa. Further research is however required to establish whether the ability of PLs to inhibit cholesterol absorption is of therapeutic benefit

Role of ABCG1 and ABCA1 in regulation of neuronal cholesterol efflux to apolipoprotein E discs and suppression of amyloid-β peptide generation

Maintenance of an adequate supply of cholesterol is important for neuronal function, whereas excess cholesterol promotes amyloid precursor protein (APP) cleavage generating toxic amyloid-β (Aβ) peptides. To gain insights into the pathways that regulate neuronal cholesterol level, we investigated the potential for reconstituted apolipoprotein E (apoE) discs, resembling nascent lipoprotein complexes in the central nervous system, to stimulate neuronal [3H]cholesterol efflux. ApoE discs potently accelerated cholesterol efflux from primary human neurons and cell lines. The process was saturable (17.5 μg of apoE/ml) and was not influenced by APOE genotype. High performance liquid chromatography analysis of cholesterol and cholesterol metabolites effluxed from neurons indicated that \u3c25% of the released cholesterol was modified to polar products (e.g. 24-hydroxycholesterol) that diffuse from neuronal membranes. Thus, most cholesterol (∼75%) appeared to be effluxed from neurons in a native state via a transporter pathway. ATP-binding cassette transporters ABCA1, ABCA2, and ABCG1 were detected in neurons and neuroblastoma cell lines and expression of these cDNAs revealed that ABCA1 and ABCG1 stimulated cholesterol efflux to apoE discs. In addition, ABCA1 and ABCG1 expression in Chinese hamster ovary cells that stably express human APP significantly reduced Aβ generation, whereas ABCA2 did not modulate either cholesterol efflux or Aβ generation. These data indicate that ABCA1 and ABCG1 play a significant role in the regulation of neuronal cholesterol efflux to apoE discs and in suppression of APP processing to generate Aβ peptides. © 2007 by The American Society for Biochemistry and Molecular Biology, Inc

Dietary sphingomyelin lowers hepatic lipid levels and inhibits intestinal cholesterol absorption in high-fat-fed mice.

Controlling intestinal lipid absorption is an important strategy for maintaining lipid homeostasis. Accumulation of lipids in the liver is a major risk factor for metabolic syndrome and nonalcoholic fatty liver disease. It is well-known that sphingomyelin (SM) can inhibit intestinal cholesterol absorption. It is, however, unclear if dietary SM also lowers liver lipid levels. In the present study (i) the effect of pure dietary egg SM on hepatic lipid metabolism and intestinal cholesterol absorption was measured with [(14)C]cholesterol and [(3)H]sitostanol in male C57BL/6 mice fed a high-fat (HF) diet with or without 0.6% wt/wt SM for 18 days; and (ii) hepatic lipid levels and gene expression were determined in mice given a HF diet with or without egg SM (0.3, 0.6 or 1.2% wt/wt) for 4 weeks. Mice supplemented with SM (0.6% wt/wt) had significantly increased fecal lipid and cholesterol output and reduced hepatic [(14)C]cholesterol levels after 18 days. Relative to HF-fed mice, SM-supplemented HF-fed mice had significantly lower intestinal cholesterol absorption (-30%). Liver weight was significantly lower in the 1.2% wt/wt SM-supplemented mice (-18%). Total liver lipid (mg/organ) was significantly reduced in the SM-supplemented mice (-33% and -40% in 0.6% wt/wt and 1.2% wt/wt SM, respectively), as were triglyceride and cholesterol levels. The reduction in liver triglycerides was due to inactivation of the LXR-SREBP-1c pathway. In conclusion, dietary egg SM has pronounced hepatic lipid-lowering properties in mice maintained on an obesogenic diet

Suppression of ABCE1-Mediated mRNA Translation Limits N-MYC-Driven Cancer Progression

The ability of the N-MYC transcription factor to drive cancer progression is well-demonstrated in neuroblastoma, the most common extracranial pediatric solid tumor, where MYCN-amplification heralds a poor prognosis with only 11% of patients surviving past 5 years. However, decades of attempts of direct inhibition of N-MYC or its paralogues has led to the conclusion that this protein is 'undruggable'. Therefore, targeting pathways upregulated by N-MYC signaling presents an alternative therapeutic approach. Here we show that MYCN-amplified neuroblastomas are characterized by elevated rates of protein synthesis and that high expression of ABCE1, a translation factor directly upregulated by N-MYC, is itself a strong predictor of poor clinical outcome. Despite the potent ability of N-MYC in heightening protein synthesis and malignant characteristics in cancer cells, suppression of ABCE1 alone selectively negated this effect, returning the rate of translation to baseline levels and significantly reducing the growth, motility, and invasiveness of MYCN-amplified neuroblastoma cells and patient-derived xenograft tumors in vivo. The growth of non-malignant cells or MYCN-non-amplified neuroblastoma cells remained unaffected by reduced ABCE1, supporting a therapeutic window associated with targeting ABCE1. Neuroblastoma cells with c-MYC overexpression also required ABCE1 to maintain cell proliferation and translation. Taken together, ABCE1-mediated translation constitutes a critical process in the progression of N-MYC-driven and c-MYC-driven cancers that warrants investigations into methods of its therapeutic inhibitionThis work was supported by the National Health and Medical Research Council (APP1016699 and APP1132608 to M. Haber and M.D. Norris), Cancer Institute NSW (10/TPG/1-03 and 14/TPG/1-13 to M. Haber and M.D. Norris), Tour de Cure (RG162423 to M.J. Henderson), the Australian Postgraduate Award (to J. Gao), and Children's Cancer Institute and Cancer Therapeutics CRC PhD Top-up Scholarship (to J. Gao)

Body weight, liver weight and food intake of mice fed a high-fat diet with or without SM.

<p>Male C57BL/6 mice were fed a high-fat (HF) diet without or with SM (HFSM) (0.3%. 0.6% or 1.2% wt/wt) and sacrificed after 4 weeks. Body and liver weights were recorded. Liver lipids were quantified as described in “<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0055949#s2" target="_blank">Methods</a>”. Values represent means±SEM. Significant differences between HF and HFSM groups were determined by one-way ANOVA:</p>*<p><i>P</i><0.05;</p>***<p><i>P</i><0.001.</p

Cholesterol, triglyceride, sphingomyelin and ceramide levels in livers of mice fed a high-fat diet supplemented with SM.

<p>Male mice were fed a high-fat diet (HF) without or with 0.6% wt/wt SM (HFSM) for 4 weeks. Liver lipids were quantified as described in “<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0055949#s2" target="_blank">Methods</a>”. Values represent means±SEM. Percentage difference between groups is given in italics. Significant differences between the HF and HFSM groups were determined by Student's t-test:</p>†<p><i>P_corrected</i><0.05.</p