Inhibiting Cholesterol Absorption During Lactation Programs Future Intestinal Absorption of Cholesterol in Adult Mice.

In nematodes, the intestine senses and integrates early life dietary cues that lead to lifelong epigenetic adaptations to a perceived nutritional environment-it is not clear whether this process occurs in mammals. We aimed to establish a mouse model of reduced dietary cholesterol availability from maternal milk and investigate the consequences of decreased milk cholesterol availability, early in life, on the metabolism of cholesterol in adult mice. We blocked intestinal absorption of cholesterol in milk fed to newborn mice by supplementing the food of dams (for 3 weeks between birth and weaning) with ezetimibe, which is secreted into milk. Ezetimibe interacts with the intestinal cholesterol absorption transporter NPC1l1 to block cholesterol uptake into enterocytes. Characterization of these offspring at 24 weeks of age showed a 27% decrease in cholesterol absorption (P < .001) and reduced levels of Npc1l1 messenger RNA and protein, but not other cholesterol transporters, in the proximal small intestine. We observed increased histone H3K9me3 methylation at positions -423 to -607 of the proximal Npc1l1 promoter in small intestine tissues from 24-week-old offspring fed ezetimibe during lactation, compared with controls. These findings show that the early postnatal mammalian intestine functions as an environmental sensor of nutritional conditions, responding to conditions such as low cholesterol levels by epigenetic modifications of genes. Further studies are needed to determine how decreased sterol absorption for a defined period might activate epigenetic regulators; the findings of our study might have implications for human infant nutrition and understanding and preventing cardiometabolic disease.

ABCA8 Regulates Cholesterol Efflux and High-Density Lipoprotein Cholesterol Levels.

OBJECTIVE: High-density lipoproteins (HDL) are considered to protect against atherosclerosis in part by facilitating the removal of cholesterol from peripheral tissues. However, factors regulating lipid efflux are incompletely understood. We previously identified a variant in adenosine triphosphate-binding cassette transporter A8 (ABCA8) in an individual with low HDL cholesterol (HDLc). Here, we investigate the role of ABCA8 in cholesterol efflux and in regulating HDLc levels. APPROACH AND RESULTS: We sequenced ABCA8 in individuals with low and high HDLc and identified, exclusively in low HDLc probands, 3 predicted deleterious heterozygous ABCA8 mutations (p.Pro609Arg [P609R], IVS17-2 A>G and p.Thr741Stop [T741X]). HDLc levels were lower in heterozygous mutation carriers compared with first-degree family controls (0.86±0.34 versus 1.17±0.26 mmol/L; P=0.005). HDLc levels were significantly decreased by 29% (P=0.01) in Abca8b-/- mice on a high-cholesterol diet compared with wild-type mice, whereas hepatic overexpression of human ABCA8 in mice resulted in significant increases in plasma HDLc and the first steps of macrophage-to-feces reverse cholesterol transport. Overexpression of wild-type but not mutant ABCA8 resulted in a significant increase (1.8-fold; P=0.01) of cholesterol efflux to apolipoprotein AI in vitro. ABCA8 colocalizes and interacts with adenosine triphosphate-binding cassette transporter A1 and further potentiates adenosine triphosphate-binding cassette transporter A1-mediated cholesterol efflux. CONCLUSIONS: ABCA8 facilitates cholesterol efflux and modulates HDLc levels in humans and mice.

Group IIA Secretory Phospholipase A2 Predicts Graft Failure and Mortality in Renal Transplant Recipients by Mediating Decreased Kidney Function.

The acute phase protein group IIA secretory phospholipase A2 (sPLA2-IIA) has intrinsic proatherosclerotic properties. The present prospective cohort study investigated whether plasma sPLA2-IIA associates with graft failure, cardiovascular, and all-cause mortality in renal transplant recipients (RTRs), patients with accelerated atherosclerosis formation both systemically and within the graft. In 511 RTRs from a single academic center with stable graft function >1 year, baseline plasma sPLA2-IIA was determined by ELISA. Primary end points were death-censored graft failure and mortality (median follow-up, 7.0 years). Baseline sPLA2-IIA was higher in RTRs than in healthy controls (median 384 ng/dL (range 86-6951) vs. 185 ng/dL (range 104-271), p < 0.001). Kaplan-Meier analysis demonstrated increased risk for graft failure (p = 0.002), as well as cardiovascular (p < 0.001) and all-cause mortality (p < 0.001), with increasing sPLA2-IIA quartiles. Cox regression showed strong associations of sPLA2-IIA with increased risks of graft failure (hazard ratio (HR) = 1.42 (1.11-1.83), p = 0.006), as well as cardiovascular (HR = 1.48 (1.18-1.85), p = 0.001) and all-cause mortality (HR = 1.39 (1.17-1.64), p < 0.001), dependent on parameters of kidney function. Renal function during follow-up declined faster in RTRs with higher baseline sPLA2-IIA levels. In RTRs, sPLA2-IIA is a significant predictive biomarker for chronic graft failure, as well as overall and cardiovascular disease mortality dependent on kidney function. This dependency is conceivably explained by sPLA2-IIA impacting negatively on kidney function.

Gestational oxidative stress protects against adult obesity and insulin resistance.

Pregnancy complications such as preeclampsia cause increased fetal oxidative stress and fetal growth restriction, and associate with a higher incidence of adult metabolic syndrome. However, the pathophysiological contribution of oxidative stress per se is experimentally difficult to discern and has not been investigated. This study determined, if increased intrauterine oxidative stress (IUOx) affects adiposity, glucose and cholesterol metabolism in adult Ldlr-/-xSod2+/+ offspring from crossing male Ldlr-/-xSod2+/+ mice with Ldlr-/-xSod2 +/- dams (IUOx) or Ldlr-/-xSod2 +/- males with Ldlr-/-xSod2+/+ dams (control). At 12 weeks of age mice received Western diet for an additional 12 weeks. Adult male IUOx offspring displayed lower body weight and reduced adiposity associated with improved glucose tolerance compared to controls. Reduced weight gain in IUOx was conceivably due to increased energy dissipation in white adipose tissue conveyed by higher expression of Ucp1 and an accompanying decrease in DNA methylation in the Ucp1 enhancer region. Female offspring did not show comparable phenotypes. These results demonstrate that fetal oxidative stress protects against the obesogenic effects of Western diet in adulthood by programming energy dissipation in white adipose tissue at the level of Ucp1.

Milk cholesterol concentration in mice is not affected by high cholesterol diet- or genetically-induced hypercholesterolaemia.

Breast milk cholesterol content may imply to affect short- and long-term cholesterol homeostasis in the offspring. However, mechanisms of regulating milk cholesterol concentration are only partly understood. We used different mouse models to assess the impact of high cholesterol diet (HC)- or genetically-induced hypercholesterolaemia on milk cholesterol content. At day 14 postpartum we determined milk, plasma and tissue lipids in wild type (WT), LDL receptor knockout (Ldlr-/-), and ATP-binding cassette transporter G8 knockout (Abcg8-/-) mice fed either low- or 0.5% HC diet. In chow-fed mice, plasma cholesterol was higher in Ldlr-/- dams compared to WT. HC-feeding increased plasma cholesterol in all three models compared to chow diet. Despite the up to 5-fold change in plasma cholesterol concentration, the genetic and dietary conditions did not affect milk cholesterol levels. To detect possible compensatory changes, we quantified de novo cholesterol synthesis in mammary gland and liver, which was strongly reduced in the various hypercholesterolaemic conditions. Together, these data suggest that milk cholesterol concentration in mice is not affected by conditions of maternal hypercholesterolaemia and is maintained at stable levels via ABCG8- and LDLR-independent mechanisms. The robustness of milk cholesterol levels might indicate an important physiological function of cholesterol supply to the offspring.

High-cholesterol diet does not alter gut microbiota composition in mice.

INTRODUCTION: Western diet containing both saturated fat and cholesterol impairs cardio-metabolic health partly by modulating diversity and function of the microbiota. While diet containing only high fat has comparable effects, it is unclear how diets only enriched in cholesterol impact the microbiota. Therefore, we aimed to characterize the response of host and microbiota to a high cholesterol (HC) diet in mice susceptible to cardio-metabolic disease. METHODS: LDLR knockout mice received either 1.25% HC or no cholesterol containing control diet (NC) for 12 weeks before characterizing host cholesterol metabolism and intestinal microbiota composition (next generation sequencing). RESULTS: HC diet substantially increased plasma (1.6-fold) and liver cholesterol levels (21-fold), biliary cholesterol secretion (4.5-fold) and fecal neutral sterol excretion (68-fold, each p < 0.001) but not fecal bile acid excretion. Interestingly, despite the profound changes in intestinal cholesterol homeostasis no differences in microbial composition between control and HC-fed mice were detected. In both groups the main phyla were Bacteroidetes (55%), Firmicutes (27%) and Verrucomicrobia (14%). CONCLUSION: Our results demonstrate that in mice HC diet alone does not alter the microbiota composition despite inducing substantial adaptive changes in whole body cholesterol homeostasis. The impact of Western diet on intestinal microbiota thus appears to be mediated exclusively by its high fat content.

The predictive value of the antioxidative function of HDL for cardiovascular disease and graft failure in renal transplant recipients.

BACKGROUND: Protection of low-density lipoproteins (LDL) against oxidative modification is a key anti-atherosclerotic property of high-density lipoproteins (HDL). This study evaluated the predictive value of the HDL antioxidative function for cardiovascular mortality, all-cause mortality and chronic graft failure in renal transplant recipients (RTR). METHODS: The capacity of HDL to inhibit native LDL oxidation was determined in vitro in a prospective cohort of renal transplant recipients (RTR, n = 495, median follow-up 7.0 years). RESULTS: The HDL antioxidative functionality was significantly higher in patients experiencing graft failure (57.4 ± 9.7%) than in those without (54.2 ± 11.3%; P = 0.039), while there were no differences for cardiovascular and all-cause mortality. Specifically glomerular filtration rate (P = 0.001) and C-reactive protein levels (P = 0.006) associated independently with antioxidative functionality in multivariate linear regression analyses. Cox regression analysis demonstrated a significant relationship between antioxidative functionality of HDL and graft failure in age-adjusted analyses, but significance was lost following adjustment for baseline kidney function and inflammatory load. No significant association was found between HDL antioxidative functionality and cardiovascular and all-cause mortality. CONCLUSION: This study demonstrates that the antioxidative function of HDL (i) does not predict cardiovascular or all-cause mortality in RTR, but (ii) conceivably contributes to the development of graft failure, however, not independent of baseline kidney function and inflammatory load.