Folate status in women of childbearing age in the Urban Metropolitan Region of Chile: results from the National Health Survey 2016-2017.

OBJECTIVE: To analyse serum folate levels in women of childbearing age in the Metropolitan Region (MR) of Chile. DESIGN: Cross-sectional design as part of the 2016-2017 National Health Survey (Encuesta Nacional de Salud, ENS 2016-2017), using a household-based multistage stratified random sample. Serum folate levels measured by electrochemiluminescence immunoassay in fasting venous blood samples were classified as deficient (<4·4 ng/ml), normal (4·4-20 ng/ml) or supraphysiological (>20 ng/ml). SETTING: The MR of Chile. PARTICIPANTS: Women of reproductive age (15-49 years, n 222) from the MR participated in the ENS 2016-2017. RESULTS: The mean, median and range of serum folate were 14·2 (se 0·4), 13·9 and 2·1-32·2 ng/ml, respectively. Folate deficiency was detected in 0·9 % of women, while 7·0 % had supraphysiological levels of the vitamin. No significant effects of age, educational level, marital status, parity, smoking status or nutritional status on serum folate levels were detected by univariate or multivariate analyses. Intake of folic acid supplements showed a significant association with serum folate levels, but only 1·2 % of women used supplements. CONCLUSIONS: Folate deficiency in women of reproductive age living in the MR of Chile is almost inexistent according to the ENS 2016-2017, suggesting that the current population-wide mandatory folic acid fortification of flour is an effective and equitable measure to prevent folate deficiency. These results support the option of maintaining current folic acid fortification in Chile, particularly based on the low adherence to supplementation regimes evidenced in other populations.

Red Wine Grape Pomace Attenuates Atherosclerosis and Myocardial Damage and Increases Survival in Association with Improved Plasma Antioxidant Activity in a Murine Model of Lethal Ischemic Heart Disease.

A healthy dietary pattern and high quality nutrient intake reduce atherosclerotic cardiovascular disease risk. Red wine grape pomace (RWGP)-a rich natural source of dietary fiber and antioxidants-appears to be a potential functional food ingredient. The impact of a dietary supplementation with RWGP flour was evaluated in atherogenic diet-fed SR-B1 KO/ApoER61h/h mice, a model of lethal ischemic heart disease. SR-B1 KO/ApoER61h/h mice were fed with atherogenic (high fat, cholesterol, and cholic acid, HFC) diet supplemented with: (a) 20% chow (HFC-Control), (b) 20% RWGP flour (HFC-RWGP), or (c) 10% chow/10% oat fiber (HFC-Fiber); and survival time was evaluated. In addition, SR-B1 KO/ApoER61h/h mice were fed for 7 or 14 days with HFC-Control or HFC-RWGP diets and plasma lipid levels, inflammation, oxidative damage, and antioxidant activity were measured. Atherosclerosis and myocardial damage were assessed by histology and magnetic resonance imaging, respectively. Supplementation with RWGP reduced premature death, changed TNF-α and IL-10 levels, and increased plasma antioxidant activity. Moreover, decreased atheromatous aortic and brachiocephalic plaque sizes and attenuated myocardial infarction and dysfunction were also observed. These results suggest that RWGP flour intake may be used as a non-pharmacological therapeutic approach, contributing to decreased progression of atherosclerosis, reduced coronary heart disease, and improved cardiovascular outcomes.

Gugulipid causes hypercholesterolemia leading to endothelial dysfunction, increased atherosclerosis, and premature death by ischemic heart disease in male mice.

For proper cholesterol metabolism, normal expression and function of scavenger receptor class B type I (SR-BI), a high-density lipoprotein (HDL) receptor, is required. Among the factors that regulate overall cholesterol homeostasis and HDL metabolism, the nuclear farnesoid X receptor plays an important role. Guggulsterone, a bioactive compound present in the natural product gugulipid, is an antagonist of this receptor. This natural product is widely used globally as a natural lipid-lowering agent, although its anti-atherogenic cardiovascular benefit in animal models or humans is unknown. The aim of this study was to determine the effects of gugulipid on cholesterol homeostasis and development of mild and severe atherosclerosis in male mice. For this purpose, we evaluated the impact of gugulipid treatment on liver histology, plasma lipoprotein cholesterol, endothelial function, and development of atherosclerosis and/or ischemic heart disease in wild-type mice; apolipoprotein E knockout mice, a model of atherosclerosis without ischemic complications; and SR-B1 knockout and atherogenic-diet-fed apolipoprotein E hypomorphic (SR-BI KO/ApoER61h/h) mice, a model of lethal ischemic heart disease due to severe atherosclerosis. Gugulipid administration was associated with histological abnormalities in liver, increased alanine aminotransferase levels, lower hepatic SR-BI content, hypercholesterolemia due to increased HDL cholesterol levels, endothelial dysfunction, enhanced atherosclerosis, and accelerated death in animals with severe ischemic heart disease. In conclusion, our data show important adverse effects of gugulipid intake on HDL metabolism and atherosclerosis in male mice, suggesting potential and unknown deleterious effects on cardiovascular health in humans. In addition, these findings reemphasize the need for rigorous preclinical and clinical studies to provide guidance on the consumption of natural products and regulation of their use in the general population.

Deficient Vitamin E Uptake During Development Impairs Neural Tube Closure in Mice Lacking Lipoprotein Receptor SR-BI.

SR-BI is the main receptor for high density lipoproteins (HDL) and mediates the bidirectional transport of lipids, such as cholesterol and vitamin E, between these particles and cells. During early development, SR-BI is expressed in extraembryonic tissue, specifically in trophoblast giant cells in the parietal yolk sac. We previously showed that approximately 50% of SR-BI-/- embryos fail to close the anterior neural tube and develop exencephaly, a perinatal lethal condition. Here, we evaluated the role of SR-BI in embryonic vitamin E uptake during murine neural tube closure. Our results showed that SR-BI-/- embryos had a very low vitamin E content in comparison to SR-BI+/+ embryos. Whereas SR-BI-/- embryos with closed neural tubes (nSR-BI-/-) had high levels of reactive oxygen species (ROS), intermediate ROS levels between SR-BI+/+ and nSR-BI-/- embryos were detected in SR-BI-/- with NTD (NTD SR-BI-/-). Reduced expression of Pax3, Alx1 and Alx3 genes was found in NTD SR-BI-/- embryos. Maternal α-tocopherol dietary supplementation prevented NTD almost completely (from 54% to 2%, p < 0.001) in SR-BI-/- embryos and normalized ROS and gene expression levels. In sum, our results suggest the involvement of SR-BI in the maternal provision of embryonic vitamin E to the mouse embryo during neural tube closure.

Early onset intrauterine growth restriction in a mouse model of gestational hypercholesterolemia and atherosclerosis.

The susceptibility to develop atherosclerosis is increased by intrauterine growth restriction and prenatal exposure to maternal hypercholesterolemia. Here, we studied whether mouse gestational hypercholesterolemia and atherosclerosis affected fetal development and growth at different stages of gestation. Female LDLR KO mice fed a proatherogenic, high cholesterol (HC) diet for 3 weeks before conception and during pregnancy exhibited a significant increase in non-HDL cholesterol and developed atherosclerosis. At embryonic days 12.5 (E12.5), E15.5, and E18.5, maternal gestational hypercholesterolemia and atherosclerosis were associated to a 22-24% reduction in male and female fetal weight without alterations in fetal number/litter or morphology nor placental weight or structure. Feeding the HC diet exclusively at the periconceptional period did not alter fetal growth, suggesting that maternal hypercholesterolemia affected fetal weight only after implantation. Vitamin E supplementation (1,000 UI of α-tocopherol/kg) of HC-fed females did not change the mean weight of E18.5 fetuses but reduced the percentage of fetuses exhibiting body weights below the 10th percentile of weight (HC: 90% vs. HC/VitE: 68%). In conclusion, our results showed that maternal gestational hypercholesterolemia and atherosclerosis in mice were associated to early onset fetal growth restriction and that dietary vitamin E supplementation had a beneficial impact on this condition.

Participation of cysteine-rich secretory proteins (CRISP) in mammalian sperm-egg interaction.

Mammalian fertilization is a complex multi-step process mediated by different molecules present on both gametes. CRISP1 (cysteine-rich secretory protein 1) is an epididymal protein thought to participate in gamete fusion through its binding to egg-complementary sites. Structure-function studies using recombinant fragments of CRISP1 as well as synthetic peptides reveal that its egg-binding ability resides in a 12 amino acid region corresponding to an evolutionary conserved motif of the CRISP family, named Signature 2 (S2). Further experiments analyzing both the ability of other CRISP proteins to bind to the rat egg and the amino acid sequence of their S2 regions show that the amino acid sequence of the S2 is needed for CRISP1 to interact with the egg. CRISP1 appears to be involved in the first step of sperm binding to the zona pellucida, identifying a novel role for this protein in fertilization. The observation that sperm testicular CRISP2 is also able to bind to the egg surface suggests a role for this protein in gamete fusion. Subsequent experiments confirmed the participation of CRISP2 in this step of fertilization and revealed that CRISP1 and CRISP2 interact with common egg surface binding sites. Together, these results suggest a functional cooperation between CRISP1 and CRISP2 to ensure the success of fertilization. These observations contribute to a better understanding of the molecular mechanisms underlying mammalian fertilization.

Expression and structure-function analysis of de, a sperm cysteine-rich secretory protein that mediates gamete fusion.

Rat sperm epididymal glycoprotein DE belongs to the cysteine-rich secretory protein (CRISP) family and participates in sperm-egg fusion through its binding to complementary sites on the egg surface. To investigate the molecular mechanisms underlying the role of DE in gamete fusion, in the present work we expressed DE in a prokaryotic system, and examined the relevance of carbohydrates and disulfide bonds for the biological activity of the protein. Immunofluorescence and sperm-egg fusion assays carried out in the presence of recombinant DE (recDE) revealed that this protein exhibits the ability to bind to the DE-egg binding sites and to inhibit gamete fusion, as does native DE (nDE). Comparison of the proteins indicated, however, that the inhibitory ability of recDE was significantly lower than that of nDE. This difference would not be due to the lack of carbohydrates in the bacterially expressed protein because enzymatically deglycosylated nDE was as able as the untreated protein to inhibit gamete fusion. To examine whether disulfide bridges are involved in DE activity, the presence of sulfhydryls in nDE and recDE was evaluated by the biotin-maleimide technique. Results indicated that, unlike nDE, in which all cysteines are involved in disulfide bonds, recDE contains free thiol groups. Subsequent experiments showed that reduction of nDE with dithiothreitol significantly decreased the ability of the protein to inhibit gamete fusion. Together, these results indicate that whereas carbohydrates do not have a role in DE-mediated gamete fusion, disulfide bridges are required for full biological activity of the protein. To our knowledge, this is the first study reporting the relevance of structural components for the function of a CRISP member.