Stress during Lactation Induces Insulin Resistance Associated with an Increase in Type 1 Cannabinoid Receptors in Liver and Adipose Tissue.

Several reports have shown that stress during lactation causes long-term metabolic and hormonal disruptions. In this study, we designed experiments to evaluate the effects of stress during lactation on the abundance of Type 1 cannabinoid/endocannabinoid receptors (CB1R) in epididymal fat and liver and development of insulin resistance in adult mice. During the whole lactation, male mice pups were daily subcutaneously injected (days 1-21) with a saline solution to produce a soft nociceptive stress (NS). Mice body weight and food intake were periodically evaluated. Adult animals were subsequently subjected to an insulin tolerance test and some days later sacrificed to evaluate the amount of epididymal fat and abundance of CB1R and adipophilin in liver and epididymal adipose tissue. Lipoprotein lipase (LPL) activity and circulating levels of leptin, adiponectin, and corticosterone were also evaluated. In this model, NS during lactation significantly increased the amount of epididymal fat and induced insulin resistance in adult mice. In addition, a significantly increased abundance of CB1R and adipophilin in epididymal fat and liver was observed, together with elevated circulating levels of leptin and corticosterone. Adult NS animals also had low plasmatic adiponectin and, although nonsignificant, had a sustained trend to a greater LPL activity associated with epididymal fat. These results indicate that increased abundance of CB1R in liver and epididymal fat alters tissue functionality likely associated with development of systemic metabolic alterations such as insulin resistance in adult mice. All these pathophysiological facts are long-term consequences of nociceptive stress during lactation.

High levels of circulating folate concentrations are associated with DNA methylation of tumor suppressor and repair genes p16, MLH1, and MGMT in elderly Chileans.

BACKGROUND: Changes in DNA methylation, one of the most studied epigenetic mechanisms, are considered an initial marker for early cancer detection. We evaluated how availability of dietary factors (folates and vitamin B12) involved in one-carbon metabolism may contribute to DNA methylation changes of cancer-related genes in human subjects. METHODS: We studied, by pyrosequencing, the methylation of tumor suppressor gene p16, DNA repair genes MLH1 and MGMT, and the repetitive element LINE-1 (as a surrogate for global DNA methylation), in blood of elderly individuals (n = 249) who had been exposed to folic acid (FA) through FA-fortified wheat flour during the last 12 years. RESULTS: We found that serum folate and to a lesser extent, vitamin B12 concentrations, were significantly correlated with DNA methylation of p16, MLH1, and MGMT, but not with LINE-1. High serum folate concentrations (>45.3 nmol/L) were present in 31.1% of the participants. Although the methylated fraction of CpG sites in p16, MLH1, and MGMT was low (1.17-3.8%), high folate concentrations were significantly associated with methylation at the 3rd tertile of specific CpG sites in all genes with OR between 1.97 and 4.17. CONCLUSIONS: This study shows that a public policy, like food fortification with FA that increases circulating serum folate levels, could affect methylation levels of specific genes linked to cancer risk. Our present results deserve additional studies to clarify the real impact of high FA levels for risk of cancer in a whole population chronically exposed to a fortified food such as wheat flour. TRIAL REGISTRATION: ISRCTN 48153354 and ISRCTN 02694183.

Impact of cadmium exposure during pregnancy on hepatic glucocorticoid receptor methylation and expression in rat fetus.

Adverse fetal environment due to maternal undernutrition or exposure to environmental chemicals alters glucocorticoid (GC) metabolism increasing the risk of metabolic disorders in adulthood. In this study, we investigated the effects of maternal exposure to cadmium (Cd, 50 ppm) during pregnancy in the methylation of fetal hepatic glucocorticoid receptor promoter (GR) and the correlation with its expression and that of the DNA methyltransferases (DNMT1a and 3a). We also studied the expression of liver phosphoenolpyruvate carboxykinase (PEPCK) and acyl-CoA oxidase (AOX), two enzymes involved in the metabolism of carbohydrates and lipids respectively. The methylation of the rat GR gene exon 1(10) (GR1(10)) in nucleotides -2536 to -2361 was analyzed by pyrosequencing. Quantitative real time PCR was used to assess hepatic GR, PEPCK and AOX mRNA, and their protein levels using Western blotting analysis. Differential methylation was noted across groups at all CpG sites in the GR exon 1(10) in a sex-dependent manner. In males, CpG were more methylated than the controls (185 ± 21%, p<0.001) but only CpG sites 1,6,7 and 9 showed a significantly different extent of methylation. In addition, a lower expression of GR (mRNA and protein) was found. On the contrary, in females, CpG were less methylated than the controls (62 ± 11%, p<0.05) and overexpressed, affecting PEPCK and AOX expression, which did not change in males. The GR methylation profile correlates with DNMT3a expression which may explain epigenetic sex-dependent changes on GR1(10) promoter induced by Cd treatment. In conclusion, Cd exposure during pregnancy affects fetal liver DNMT3a resulting in sex-dependent changes in methylation and expression of GR1(10). Although these effects do not seem to be directly involved in the low birth weight and height, they may have relevant implications for long-term health.

Cardiac tissue injury resistance during myocardial infarction at adulthood by developmental exposure to cadmium.

It has been suggested that prenatal exposure to cadmium may alter the cardiovascular function during adulthood. Using the left coronary artery ligation model of acute myocardial infarction, we studied the cardiac function of female adult offspring rats exposed to cadmium (30 ppm) during gestation. The cardiac ischemic zone in the control and cadmium-exposed groups was measured 72 h post-ligation using the TPT staining technique. Offspring from cadmium-treated dams showed a significantly smaller infarcted area compared with the control group (7.1 ± 1.5 vs. 19.6 ± 2.8%, P ≤ 0.05). We also performed echocardiographic and biochemical studies, which positively correlated with the differences observed previously. To evaluate whether the effects were associated to pre-infarct tissue damage and/or angiogenic molecules, we performed histological studies and measured the expression of vascular endothelial growth factor (VEGF), and platelet endothelial cellular adhesion molecule-1 (PECAM-1). Results revealed a higher heart vascularization in the exposed offspring that was associated with an increase in PECAM and a decrease in VEGF expression. We conclude that prenatal exposure to cadmium induces fetal adaptive responses involving changes in the expression of some cardiac angiogenic molecules resulting in long-term resistance to infarction.

The CB1 receptor antagonist SR141716A reverses adult male mice overweight and metabolic alterations induced by early stress.

Perinatal stress may cause metabolic and hormonal disruptions during adulthood. The aim of this study was to evaluate the effects of early postnatal nociceptive stimulation (NS) on body weight and other metabolic parameters during adulthood and to determine whether CB1 endocannabinoid receptors (CB1Rs) may be involved in these effects. Male mice were subjected to NS during lactation with a daily subcutaneous injection of saline solution. Subsequently, both control and NS-mice were treated from day 40 to 130, with an oral dose (1 µg/g body weight) of SR141716A, a specific CB1R antagonist/inverse agonist. Mice body weight and food intake was periodically evaluated. Adult animals were then killed to evaluate epididymal fat pads and metabolic parameters. NS did not influence food intake in adult animals, but caused significant increases in body weight, epididymal fat pads, and circulating levels of leptin, corticosterone, and triglycerides (TGs). Chronic treatment with SR141716A normalized these parameters, with the exception of corticosterone levels. This treatment also reduced plasma levels of glucose, insulin, and total cholesterol in both adult control and NS-mice. In addition, fatty acid (FA) amide hydrolase (FAAH) activity (the enzyme able to hydrolyze endocannabinoids) from liver and epididymal fat of adult NS-mice was decreased by 40-50% in comparison to activities found in same tissues of control mice. Results suggest that overactive liver and epididymal fat CB1R due to early NS may be involved in late metabolic alterations, which are sensitive to chronic treatment with SR141716A.

Specific strychnine binding sites on acrosome-associated membranes of golden hamster spermatozoa.

This study demonstrates for the first time, that membrane vesicles originated from the hamster sperm head after the occurrence of the acrosome reaction, possess specific strychnine binding sites. [3H]Strychnine binding was saturable and reversible, being displaced by unlabeled strychnine (IC(50)=26.7+/-2.3 microM). Kinetic analysis revealed one binding site with K(d)=120nM and B(max)=142fmol/10(6) spermatozoa. Glycine receptor agonists beta-alanine and taurine inhibited strychnine binding by 20-30%. Surprisingly, glycine stimulated binding by about 40-50%. Results obtained in this study strongly suggest the presence of glycine receptors-with distinctive kinetic properties on the periacrosomal plasma membrane of hamster spermatozoa. Localization of this receptor fits well with its previously proposed role in acrosomal exocytosis during mammalian fertilization.