HPLC separation of cobalamin analogues (CorA).

<p>Corrinoids were enzymatically released from immunoprecipitated HC derived from a pool of serum from mothers (A) and cord serum (B), exposed to KCN and separated by reversed-phase HPLC at a gradient of 5–30% MeCN in 0.05 mol/L phosphoric acid pH 5 from 4 to 24 min. Corrinoids in the post column fractions were quantified with the corrinoid-assay (……) and the Cbl-assay (----). The elution pattern for CorA was calculated as the difference between the two (<b>——</b>). Three distinct CorA peaks were identified in serum from both mother and newborn eluting at 13.5, 14.5 and 16.5 min. Retention times for CNCbl (15 min) (1) and CN₂Cbi (16.5 min) (2) are indicated.</p

HC-CorA in cord blood in relation to gestational age of the newborns.

<p>The concentration of CorA in cord serum correlated to the gestational age (r² = 0.19, p = 0.0004) of the newborns (n = 63).</p

Cobalamin analogues (CorA) and cobalamins (Cbl) on haptocorrin (HC) in maternal and cord sera from 69 mother-baby pairs.

<p>(A) Results for mother and newborn showed a weak correlation for HC-CorA (p = 0.01), and (B) a strong correlation for HC-Cbl (p<0.0001). (C) The absolute and relative (HC-CorA/HC-corrinoids)) amounts (mean and SEM) of HC-CorA was higher in serum from the newborns (390±20 pmol/L, (0.62±0.016)) than in serum from the mothers (170±6 pmol/L, (0.53±0.014)), p<0.0001 (paired or unpaired t-test) for both parameters.</p

Measurements of cobalamin (HC-Cbl) and cobalamin analogues (HC-CorA) attached to serum HC.

<p>Measurements of cobalamin (HC-Cbl) and cobalamin analogues (HC-CorA) attached to serum HC.</p

Main characteristics of the mothers and the newborn babies.

<p>Main characteristics of the mothers and the newborn babies.</p

Mean Subjective visual vertical measure.

<p>Mean values of the SVV for the five different conditions (OKN+SVV at 80°/s toward the left and the right, at 120°/s toward the left and the right, and the condition No OKN+SVV) for adults and children. </p

Postural parameters for the different conditions.

<p>Mean of Surface of CoP (A), of length of CoP in the medio-lateral axis (B), of standard deviation of medio-lateral body sway (C) and of mean speed of CoP (D) for the four different conditions (OKN+SVV, No OKN+SVV, DARK+FIX and DARK+EC) for adults and children. Verticals bars indicate the standard error.</p

Hepatic Methionine Homeostasis Is Conserved in C57BL/6N Mice on High-Fat Diet Despite Major Changes in Hepatic One-Carbon Metabolism

<div><p>Obesity is an underlying risk factor in the development of cardiovascular disease, dyslipidemia and non-alcoholic fatty liver disease (NAFLD). Increased hepatic lipid accumulation is a hallmark in the progression of NAFLD and impairments in liver phosphatidylcholine (PC) metabolism may be central to the pathogenesis. Hepatic PC biosynthesis, which is linked to the one-carbon (C1) metabolism by phosphatidylethanolamine N-methyltransferase, is known to be important for hepatic lipid export by VLDL particles. Here, we assessed the influence of a high-fat (HF) diet and NAFLD status in mice on hepatic methyl-group expenditure and C1-metabolism by analyzing changes in gene expression, protein levels, metabolite concentrations, and nuclear epigenetic processes. In livers from HF diet induced obese mice a significant downregulation of cystathionine β-synthase (CBS) and an increased betaine-homocysteine methyltransferase (BHMT) expression were observed. Experiments <i>in vitro</i>, using hepatoma cells stimulated with peroxisome proliferator activated receptor alpha (PPARα) agonist WY14,643, revealed a significantly reduced Cbs mRNA expression. Moreover, metabolite measurements identified decreased hepatic cystathionine and L-α-amino-n-butyrate concentrations as part of the transsulfuration pathway and reduced hepatic betaine concentrations, but no metabolite changes in the methionine cycle in HF diet fed mice compared to controls. Furthermore, we detected diminished hepatic gene expression of <i>de novo</i> DNA methyltransferase 3b but no effects on hepatic global genomic DNA methylation or hepatic DNA methylation in the Cbs promoter region upon HF diet. Our data suggest that HF diet induces a PPARα-mediated downregulation of key enzymes in the hepatic transsulfuration pathway and upregulates BHMT expression in mice to accommodate to enhanced dietary fat processing while preserving the essential amino acid methionine.</p> </div

Influence of HF diet on hepatic Dnmt gene expression and global DNA methylation.

<p>(<b>A–C</b>) Quantification of Dnmt gene expression after 12 weeks of feeding (n = 5–6). Control mice showed stronger expression for Dnmt1 (Ct = 25.13±0.06) and <i>de novo</i> Dnmt3a (Ct = 28.10±0.15) than for <i>de novo</i> Dnmt3b (Ct = 30.55±0.31). Upon HF feeding, Dnmt1 mRNA expression was unaltered (Ct = 25.03±0.11), but gene expression of Dnmt3a (Ct = 28.28±0.18) and Dnmt3b (Ct = 31.37±0.14) decreased, respectively. (<b>D</b>) Analysis of hepatic global DNA methylation of control and HF animals (n = 6). DNA methylation was calculated from the (<i>Hpa</i>II/<i>Msp</i>I) ratio, whereby a ratio of 1 indicates 0% methylation and a ratio approaching 0 corresponds to 100% DNA methylation at the investigated sites. Data are presented as mean ± SEM. Open and grey bars represent control and HF animals, respectively. Asterisk indicates statistical significance (p<0.05).</p

Impact of HF diet on selected hepatic metabolite concentrations after 12 weeks of feeding.

<p>Data are presented as box and whisker plot. (<b>A</b>) Selected data for taurine, L-glutamine, L-α-amino-n-butyrate, L-citrulline, L-ornithine, hydroxyproline and L-methionine (n = 9–11). (<b>B</b>) Analysis of S-adenosyl-methionine, S-adenosyl-homocysteine, L-homocysteine, cystathionine (n = 5–6) and choline, betaine and dimethylglycine (n = 7–9). (<b>C</b>) Selected ratios between measured hepatic metabolite concentrations. Open and grey bars represent control and HF mice, respectively. Asterisk indicates statistical significance (p<0.05).</p