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Sterol 27-hydroxylase (CYP27A1) is a key enzyme in bile acids (BAs) biosynthesis and a regulator of cholesterol metabolism. Cyp27a1/Apolipoprotein E double knockout (DKO) mice fed with western diet (WD) are protected from atherosclerosis via up-regulation of hepatic Cyp7a1 and Cyp3a11. Since feeding BAs ameliorates metabolic changes in Cyp27a1 KO mice, we tested BAs feeding on the development of atherosclerosis in DKO mice. DKO mice were fed for 8 weeks with WD containing 0.1% cholic acid (CA) (WD-CA) or chenodeoxycholic acid (CDCA) (WD-CDCA). Atherosclerotic lesions, plasma lipoprotein composition and functionality, hepatic lipid content, BAs amount and composition, expression of genes involved in lipid metabolism and BA signaling in liver and intestine as well as intestinal cholesterol absorption were assessed. Hepatic Cyp7a1 and Cyp3a11 expression were reduced by 60% after feeding with both WD-CA and WD-CDCA. After feeding with WD-CA we observed a 40-fold increase in the abundance of atherosclerotic lesions in the aortic valve, doubling of the levels of plasma total and low density lipoprotein cholesterol and halving of the level of high density lipoprotein cholesterol. Furthermore, in these mice plasma cholesterol efflux capacity decreased by 30%, hepatic BA content increased 10-fold, intestinal cholesterol absorption increased 6-fold. No such changes were observed in mice fed with WD-CDCA. Despite similar reduction on Cyp7a1 and Cyp3a11 hepatic expression, CA and CDCA have a drastically different impact on development of atherosclerosis, plasma and hepatic lipids, BAs composition and intestinal absorption. Reduced cholesterol absorption contributes largely to athero-protection in DKO mice.

Pre-eclampsia is a pregnancy-specific condition, leading to increased cardiovascular risk in later life. A decrease in cholesterol efflux capacity is linked to cardiovascular disease. We hypothesised that, in pre-eclampsia there would be a disruption of maternal/fetal plasma to efflux cholesterol as well as differences in the concentrations of both placental sterol 27-hydroxylase (CYP27A1) and apoA1-binding-protein (AIBP). Total, HDL- and ABCA1-mediated cholesterol efflux were performed with maternal and fetal plasma from women with pre-eclampsia and normotensive controls (both n=17). ApoA1/apoE were quantified by chemiluminescence; 27-hydroxycholesterol (27-OHC) by GC-MS. Immunohistochemistry was used to determine placental expression/localisation of CYP27A1, AIBP, apoA1, apoE and SRB1. Maternal and fetal total and HDL-mediated cholesterol efflux capacities were increased in pre-eclampsia (by 10-20%), but ABCA1-mediated efflux was decreased (by 20-35%; P

Sterol 27-hydroxylase (CYP27A1) is involved in bile acid synthesis and cholesterol homoeostasis. Cyp27a1(-/-)/Apolipoprotein E(-/-) double knockout mice (DKO) fed a western diet failed to develop atherosclerosis. Caveolin-1 (CAV-1), the main component of caveolae, is associated with lipid homoeostasis and has regulatory roles in vascular diseases. We hypothesized that liver CAV-1 would contribute to the athero-protective mechanism in DKO mice. Cyp27a1(+/+)/ApoE(-/-) (ApoE KO), Cyp27a1(+/-)/ApoE(-/-) (het), and DKO mice were fed a western diet for 2 months. Atherosclerotic plaque and CAV-1 protein were quantified in aortas. Hepatic Cav-1 mRNA was assessed using qPCR, CAV-1 protein by immunohistochemistry and western blotting. Total hepatic and plasma cholesterol was measured using chemiluminescence. Cholesterol efflux was performed in RAW264.7 cells, using mice plasma as acceptor. CAV-1 protein expression in aortas was increased in endothelial cells of DKO mice and negatively correlated with plaque surface (P < 0.05). In the liver, both CAV-1 protein and mRNA expression doubled in DKO, compared to ApoE KO and het mice (P < 0.001 for both) and was negatively correlated with total hepatic cholesterol (P < 0.05). Plasma from DKO, ApoE KO and het mice had the same efflux capacity. In the absence of CYP27A1, CAV-1 overexpression might have an additional athero-protective role by partly overcoming the defect in CYP27A1-mediated cholesterol efflux.

Sterol 27-hydroxylase (CYP27A1) catalyzes the hydroxylation of cholesterol to 27-hydroxycholesterol (27-OHC) and regulates cholesterol homeostasis. In Cyp27a1/ Apolipoprotein E (ApoE) double knockout (KO) mice fed with Western diet (WD), the atherosclerotic phenotype found in ApoE KO mice was reversed. As protective mechanism, up-regulation of Cyp3a11 and Cyp7a1 was proposed. Cyp27a1 heterozygote/ApoE KO (het) mice, with reduced Cyp27a1 expression and normal levels of Cyp7a1 and Cyp3a11, developed more severe lesions than ApoE KO mice. To analyze the contribution of Cyp3a11 to the protection of atherosclerosis development, Cyp3a11 was induced by Rifampicin (RIF) in ApoE KO and het mice. Males were fed with WD and treated daily with RIF (10 mg/kg ip) or vehicle for 4 weeks. Atherosclerosis was quantified in the aortic valve. Plasma lipids and 27-hydroxycholesterol (27-OHC), expression of cytochromes P450 and genes involved in cholesterol transport and bile acids (BAs) signaling in liver and intestine, and intestinal cholesterol absorption were analyzed. RIF increased expression of hepatic but not intestinal Cyp3a11 4-fold in both genotypes. In ApoE KO mice treated with RIF, we found a 2-fold decrease in plasma cholesterol, and a 2-fold increase in high-density lipoprotein/low-density lipoprotein ratio and CY27A1 activity. Intestinal cholesterol absorption remained unchanged and atherosclerotic lesions decreased approximately 3-fold. In het mice, RIF had no effect on plasma lipids composition, CYP27A1 activity, and atherosclerotic plaque development, despite a reduction in cholesterol absorption. In conclusion, the antiatherogenic effect of Cyp3a11 induction by RIF was also dependent on Cyp27a1 expression.

Objectives: We have previously demonstrated an association between gestational diabetes mellitus and dysregulated placental iron transport and regulation. In this context we aimed to investigate the effect of antioxidant treatment to counterbalance the pathophysiological effects of elevated glucose and lipid levels on placental iron homeostasis. Methods: In newly established BeWo-derived cell models, we assessed the time-dependent adaption of trophoblast cells to normoglycemic, hyperglycemic and hyperglycemic combined with hyperlipidemic conditions. During 5 weeks we monitored iron uptake, expression patterns of iron transporters and regulators and cellular responses involving autophagy and oxidative stress pathways. Autophagy markers were analyzed by immunoblotting and oxidative stress by TBARS, total GSH and protein carbonylation assays. In parallel, selenium (Se) supplementation was applied to assess and counteract oxidative stress levels induced by hyperglycemia/ hyperlipidemia. Using qRT-PCR we analyzed 7 genes involved in placental iron homeostasis. The gene expression results were complemented by protein data applying a newly developed mass spectrometry-based method. Results: Elevated glucose levels in BeWo cells altered cellular morphology, affected mRNA and protein expression of iron transporters and regulators, and also reduced iron uptake confirming an impact of hyperglycemia on placental iron transport function. Our results further indicated that increased oxidative stress and changes in the autophagy pathway were involved in altering iron homeostasis under hyperglycemic conditions. Increased antioxidative potential through Se-supplementation was able to completely rescue the adverse effects of hyperglycemia/hyperlipidemia on iron homeostasis. Conclusion: Our data indicate that changes in expression, localization and function of placental iron transporters are responsible for reduced placental iron transport under hyperglycemic conditions. These adaptations could be part of a protective mechanism preventing oxidative damage for both the fetus and the placenta caused by highly oxidative iron. The successful rescue by Se supplementation suggests that antioxidant treatment could protect pregnant women with increased risk to develop GDM by balancing placental oxidative stress levels.

Introduction Pre-eclampsia is a pregnancy-specific condition affecting 2–7% of women, causing perinatal and maternal morbidity/mortality and predisposes both mother and fetus to increased risks of adult cardiovascular disease. A disturbance in the flow of cholesterol is linked to atherosclerosis and cardiovascular disease. Objective We measured cholesterol efflux capacity, apoA1, apoE and oxysterols, in maternal and fetal plasma and placental sterol 27-hydroxylase (CYP27A1) and apo-A1-binding-protein (AIBP) expression in normotensive control and pre-eclamptic pregnancies. Methods Cholesterol efflux assays were performed in RAW264.7 cells with maternal and matched fetal plasma of normotensive control or pre-eclamptic pregnancies (n = 17 in both). ApoA1, apoE were measured by ELISAs. Oxysterols were also measured by gas-chromatography–mass-spectrometry. Placental mRNA and protein expression/localisation of CYP27A1 and AIBP were measured in the same women by qPCR and immunohistochemistry. Results In pre-eclampsia, maternal and fetal cholesterol efflux capacity and fetal apoE were all increased ( P 0.05 for all), yet fetal cholesterol efflux capacity and apoE were lower in the small-for-gestational-age (SGA) subgroup (n = 6; P 0.05 for both). Fetal 27-hydroxycholesterol (27-OHC) concentrations were lower in babies born to pre-eclamptic mothers ( P 0.05 ) with no other observed differences in oxysterols. Fetal apparent CYP27A1 activities (calculated by ratios of 27-OHC/(27-OHC + total serum cholesterol) were also lower in the pre-eclampsia group ( P 0.05 ). Protein expression was localised around fetal vessels; CYP27A1 and AIBP proteins increased in pre-eclampsia (P = 0.04), but no differences were found in gene expression. Conclusions Increased maternal and fetal cholesterol efflux capacity in pre-eclampsia, which is a key step in reverse cholesterol transport, coupled with increased placental CYP27A1, could be a rescue mechanism to remove cholesterol from cells thus trying to reduce lipid peroxidation. However, increased placental AIBP suggests disturbed lipid homeostasis, which could disrupt placental angiogenesis, contributing to the endothelial dysfunction so characteristic of pre-eclampsia.

In humans, sterol 27-hydroxylase (CYP27A1) deficiency leads to cholesterol deposition in tendons and vasculature. Thus, in addition to its role in bile acid synthesis, where it converts cholesterol to 27-hydroxycholesterol (27-OHC), CYP27A1 may also be atheroprotective. Cyp27A1-deficient (Cyp27A1−/−) mice were crossed with apolipoprotein E (apoE)-deficient mice. Cyp27A1+/+/apoE−/− [ApoE-knockout (KO)], Cyp27A1+/−/apoE−/− heterozygous (het), and Cyp27A1−/−/apoE−/− [double-knockout (DKO)] mice were challenged with a Western diet (WD) for 3 and 6 mo. ApoE-KO mice fed a chow diet or a WD were used as the control. The severity of atherosclerosis in DKO mice was reduced 10-fold. Compared with the control, the DKO mice had no 27-OHC, total plasma cholesterol and low-density lipoprotein and very low density lipoprotein (LDL/VLDL) concentrations were reduced 2-fold, and HDL was elevated 2-fold. Expression of hepatic CYP7A1, CYP3A, and CYP8B1 were 5- to 10-fold higher. 3-Hydroxy-3-methyl-glutaryl-CoA reductase (HMGR) activity increased 4-fold. Fecal cholesterol was increased. In contrast, het mice fed a WD developed accelerated atherosclerosis and severe skin lesions, possibly because of reduced reverse cholesterol transport due to diminished 27-OHC production. CYP27A1 activity is involved in the control of cholesterol homeostasis and development of atherosclerosis with a distinct gene dose-dependent effect.—Zurkinden, L., Solcà, C., Vögeli, I. A., Vogt, B., Ackermann, D., Erickson, S. K., Frey, F. J., Sviridov, D., Escher, G. Effect of Cyp27A1 gene dosage on atherosclerosis development in ApoE-knockout mice.

Introduction Pre-eclampsia (PE), increases the risks of later cardiovascular disease possibly through endothelial dysfunction. Atherosclerosis has been associated with low cholesterol efflux from peripheral macrophages to HDL particles. We hypothesise that there is a link to atherosclerosis via impaired cholesterol flow and thus future cardiovascular risk. Objective We measured cholesterol efflux capacity, apoA1 and apoE, in maternal plasma at 6-8 weeks postpartum from women who had undergone a normotensive pregnancy (NC; n =23) or had experienced PE ( n =67). Methods Cholesterol efflux was performed in RAW264.7 cells with maternal plasma of NC or PE pregnancies as acceptor. ApoA1 and apoE were measured by ELISA. Results Maternal cholesterol efflux capacity was reduced in PE compared to NC (% cholesterol efflux; median [IQR]; NC: 114.7 [107, 127.9]; PE: 108.8 [95.7, 121.6]; P =0.02). ApoE concentrations were also reduced in these women (median [IQR]; NC: 7.02 [5.85, 8.10]; PE: 5.39 [3.94, 7.37]g/L; P =0.004). However, no differences were observed in apoA1 concentrations ( P >0.05). When the PE data were analysed by onset of disease (early-onset before 34 weeks and late-onset after 34 weeks), the late-onset PE group had significantly lower cholesterol efflux compared to the early-onset PE group ( P =0.03). Conclusions Decreased maternal cholesterol efflux capacity persists 6–8 weeks following PE. A decrease in the key step in reverse cholesterol transport, coupled with reduced apoE concentrations, suggest a disruption in cholesterol homeostasis increasing the probability of future cardiovascular disease. The further reduction in the late-onset PE group implies that these women may have had pre-existing metabolic disease/endothelial dysfunction that only manifests following the stress test of pregnancy. Further work is required to validate these initial findings.