Fe₂O₃‑Promoted Intermolecular Chlorotrifluoromethylthiolation of Alkenes

A simple, convenient method for intermolecular chlorotrifluoromethylthiolation of alkenes by using a low-cost and more abundant iron catalyst has been developed. This protocol provides a straightforward way to synthesize a variety of useful SCF₃-containing chlorides from a wide range of alkenes, including electron-deficient, aromatic, and unactivated alkenes. Mechanistic studies indicate that this is a free radical transformation, and the stronger electrophilic trifluoromethylthiolating reagent CF₃SCl was generated in situ under the employed conditions. The synthetic applications of this approach were also explored by a variety of synthetically useful transformations

Betaine Supplementation in Maternal Diet Modulates the Epigenetic Regulation of Hepatic Gluconeogenic Genes in Neonatal Piglets

<div><p>In this study, gestational sows were fed control or betaine-supplemented diets (3 g/kg) throughout the pregnancy, and the newborn piglets were used to elucidate whether maternal dietary betaine affected offspring hepatic gluconeogenic genes through epigenetic mechanisms. Neonatal piglets born to betaine-supplemented sows had significantly higher serum and hepatic betaine contents, together with significantly greater expression of methionine metabolic enzymes in the liver. Interestingly, significantly higher serum concentrations of lactic acid and glucogenic amino acids, including serine, glutamate, methionine and histidine, were detected in the piglets born to betaine-supplemented sows, which were coincident with higher hepatic glycogen content and PEPCK1 enzyme activity, as well as greater protein expression of gluconeogenic enzymes, pyruvate carboxylase (PC), cytoplasmic phosphoenolpyruvate carboxykinase (PEPCK1), mitochondrional phosphoenolpyruvate carboxykinase (PEPCK2) and fructose-1, 6-bisphosphatase (FBP1). Moreover, maternal betaine significantly changed the methylation status of both CpGs and histones on the promoter of gluconeogenic genes. The lower <i>PEPCK1</i> mRNA was associated with DNA hypermethylation and more enriched repression histone mark H3K27me3, while the up-regulated <i>PEPCK2</i> and <i>FBP1</i> mRNA was associated with DNA hypomethylation and more enriched activation histone mark H3K4me3. Furthermore, the expression of two miRNAs predicted to target <i>PC</i> and 6 miRNAs predicted to target <i>PEPCK1</i> was dramatically suppressed in the liver of piglets born to betaine-supplemented sows. Our results provide the first evidence that maternal betaine supplementation affects hepatic gluconeogenic genes expression in newborn piglets through enhanced hepatic methionine metabolism and epigenetic regulations, which involve DNA and histone methylations, and possibly miRNAs-mediated post-transcriptional mechanism.</p></div

p53 Cooperates with Sp1 to Regulate Breed-Dependent Expression of Glucocorticoid Receptor in the Liver of Preweaning Piglets

<div><p>Previous studies indicate that Chinese indigenous pig breeds demonstrate distinct pattern of glucocorticoid receptor (GR) expression, which is associated with their unique growth and metabolic phenotypes. Here we sought to unravel the transcriptional mechanisms underlying the breed-specific hepatic GR expression in preweaning Chinese Erhualian (EHL) and Western Large White (LW) piglets. Total GR mRNA and the predominant GR mRNA variant 1–9/10 were expressed significantly higher in EHL compared with LW piglets (<i>P</i><0.01), which was associated with more enriched histone H3 acetylation on 1–9/10 promoter (<i>P</i><0.05). Nuclear content of transcription factor specificity protein 1 (Sp1) was significantly lower in EHL piglets, yet its binding to GR 1–9/10 promoter was significantly higher in EHL piglets, as revealed by chromatin immunoprecipitation assays. Although p53 binding to GR promoter 1–9/10 did not differ between breeds, expression of p53 mRNA and protein, as well as its binding to Sp1, were significantly higher in EHL piglets. Moreover, p53 activator doxorubicin significantly enhanced GR 1–9/10 promoter activity in HepG2 cells at 100 nM, which was associated with significantly higher protein content of p53 and GR. Sp1 inhibitor, mithramycin A, significantly inhibited (<i>P</i><0.05) the basal activity of GR promoter 1–9/10 and completely blocked doxorubicin -induced activation of GR promoter 1–9/10. These data indicate that higher hepatic GR expression in EHL piglets attributes mainly to the enhanced transcription of GR promoter 1–9/10, which is achieved from breed-specific interaction of p53 and Sp1 on porcine GR 1–9/10 promoter.</p></div

Body and liver weight, betaine concentration in serum and liver, hepatic glycogen content, biochemical metabolites, hormones and amino acids in serum of newborn piglets.

<p>Values are means ± SEM, n = 16 (8 males plus 8 females).</p><p>*Different from Control, P<0.05.</p

Expression of mtDNA-encoded genes and COX enzyme activity in liver.

<p>(A) Expression of mtDNA-encoded genes in liver of male newborn piglets. (B) Expression of mtDNA-encoded genes in liver of female newborn piglets. (C) COX enzyme activity in liver of both male and female newborn piglets. Filled bar, standard protein diet; blank bar, low-protein diet. Values are mean ± SEM, * <i>P</i><0.05 vs. SP of the same sex. Means without a common letter differ, <i>P</i><0.05.</p

MicroRNAs targeting <i>PC</i> (A) and <i>PEPCK1</i> (B) 3′UTR in the liver of newborn piglets.

<p>Values are means ± SEM, n = 16 (8 males plus 8 females). Different from control, ^*<i>P</i><0.05 and ^**<i>P</i><0.01. PC, pyruvate carboxylase; PEPCK1, phosphoenolpyruvate carboxykinase 1.</p

Expression of p53 and its associations with GR and Sp1 in the liver of preweaning piglets.

<p>(A) Expression of p53 mRNA in liver tissue, 18S was used as the reference gene; (B) Nuclear p53 protein content; Histone H1 was used as a loading control for Western Blot of p53, data were presented as the fold change relative to LW. (C) Correlation of p53 and GR mRNA levels; (D) and (E) Physical interaction of Sp1 and p53 was revealed by co-immunoprecipitation and Western blot. Lane 1 means negative control, lane 2 means LW, lane 3 mean EHL. Data were presented as the fold change relative to LW. Values are mean ± SEM, n = 4. *, <i>P</i><0.05, compared with LW piglets.</p

Effect of maternal LP diet on hepatic ATP, ADP, AMP, NADH and NAD in newborn piglets.

<p>Values are mean ± SEM, Means without a common letter differ, <i>P</i><0.05. D, diet; EC, energy charge; LP, low protein; S, sex; SP, standard protein. EC was calculated using the following formula:</p><p>EC  =  ([ATP] +1/2[ADP])/([ATP] + [ADP] + [AMP]).</p

The mtDNA copy number in liver. Values are mean ± SEM, Means without a common letter differ, <i>P</i><0.05.

<p>Filled bar, standard protein diet; blank bar, low-protein diet. mtDNA, mitochondrial DNA.</p

Hepatic mRNA abundance of genes (A), protein content and western blot bands for relevant proteins (B) involved in methionine metabolism in newborn piglets.

<p>Values are means ± SEM, n = 16 (8 males plus 8 females). Different from control, ^*<i>P</i><0.05. AHCYL1, adenosylhomocysteine hydrolase-like 1; BHMT, betaine-homocysteine methyltransferase; MAT2B, methionine adenosyltransferase II beta.</p