Mapping the response of human fibroblast growth factor 21 (FGF21) promoter to serum availability and lipoic acid in HepG2 hepatoma cells.

The hormone-like polypeptide, fibroblast growth factor 21 (FGF21), is a major modulator of lipid and glucose metabolism and an exploratory treatment strategy for obesity related metabolic disorders. The costs of recombinant FGF21 and mode of delivery by injection are important constraints to its wide therapeutic use. The stimulation of endogenous FGF21 production through diet is being explored as an alternative approach. To that end, we examined the mechanism(s) by which serum manipulation and lipoic acid (a dietary activator of FGF21) induce FGF21 in human hepatocellular carcinoma HepG2 cells. Serum withdrawal markedly induced FGF21 mRNA levels (88 fold) and FGF21 secreted in the media (19 fold). Lipoic acid induced FGF21 mRNA 7 fold above DMSO-treated control cells and FGF21 secretion 3 fold. These effects were several-fold greater than those of PPARα agonist, Wy14643, which failed to induce FGF21 above and beyond the induction seen with serum withdrawal. The use of transcription inhibitor, actinomycin D, revealed that de novo mRNA synthesis drives FGF21 secretion in response to serum starvation. Four previously unrecognized loci in FGF21 promoter were nucleosome depleted and enriched in acetylated histone H3 revealing their role as transcriptional enhancers and putative transcription factor binding sites. FGF21 did not accumulate to a significant degree in induced HepG2 cells, which secreted FGF21 time dependently in media. We conclude that lipoic acid cell signaling connects with the transcriptional upregulation of FGF21 and it may prove to be a safe and affordable means to stimulate FGF21 production.

α-Lipoic acid as a triglyceride-lowering nutraceutical.

Considering the current obesity epidemic in the United States (>100 million adults are overweight or obese), the prevalence of hypertriglyceridemia is likely to grow beyond present statistics of ∼30% of the population. Conventional therapies for managing hypertriglyceridemia include lifestyle modifications such as diet and exercise, pharmacological approaches, and nutritional supplements. It is critically important to identify new strategies that would be safe and effective in lowering hypertriglyceridemia. α-Lipoic acid (LA) is a naturally occurring enzyme cofactor found in the human body in small quantities. A growing body of evidence indicates a role of LA in ameliorating metabolic dysfunction and lipid anomalies primarily in animals. Limited human studies suggest LA is most efficacious in situations where blood triglycerides are markedly elevated. LA is commercially available as dietary supplements and is clinically shown to be safe and effective against diabetic polyneuropathies. LA is described as a potent biological antioxidant, a detoxification agent, and a diabetes medicine. Given its strong safety record, LA may be a useful nutraceutical, either alone or in combination with other lipid-lowering strategies, when treating severe hypertriglyceridemia and diabetic dyslipidemia. This review examines the current evidence regarding the use of LA as a means of normalizing blood triglycerides. Also presented are the leading mechanisms of action of LA on triglyceride metabolism.

Characterization of genome-wide transcriptional changes in liver and adipose tissues of ZDF (fa/fa) rats fed R-α-lipoic acid by next-generation sequencing.

We report on the characterization of lipogenic tissue transcriptional networks that support physiological responses of obese rats to a lipid-lowering bioactive food compound, R-α-lipoic acid (LA). Nine-week-old male Zucker diabetic fatty (fa/fa) rats were fed a chow diet supplemented with 3 g LA per kg diet or pair fed for 2 wk. At the end of the trial, high-quality RNA was extracted from the liver and epididymal fat and subjected to transcriptome analysis by RNA-Seq technology. Results showed a substantially higher number of differentially expressed genes [DEG, false discovery rate adjusted P ≤ 0.05 and absolute log2 (fold change) ≥ 1] in the liver (110 genes) vs. epididymal fat (10 genes). Most epididymal fat DEG were also differentially expressed in liver and shared directionality of change. Gene Ontology (GO) analysis of these transcripts revealed significant enrichment of GO categories related to immune response, stress response, lipid metabolism, and carboxylic acid metabolic processes. Of interest, interferon-related genes involved in defense against microorganisms and innate immune response were induced by LA. Lipid metabolism-related transcript changes observed in LA-fed animals included downregulation of lipogenic genes (Pnpla3, Pnpla5, Elovl6, Acly, Gpam, and Aacs) and concomitant upregulation of short-, medium-, and long-chain fatty acid metabolic processes (Acot1, Acot2, Acsf2, and Crat). Transcriptional changes were accompanied by the lowering of abdominal adiposity and blood triacylglycerol levels. We conclude that LA dietary supplementation induces prominent gene expression changes in liver in support of significant improvement of whole-body lipid status.

Reversal of obesity-induced hypertriglyceridemia by (R)-α-lipoic acid in ZDF (fa/fa) rats.

Controlling elevated blood triacylglycerol translates into substantial health benefits. The present study aimed to evaluate the triacylglycerol-lowering properties of (R)-α-lipoic acid (LA) once circulating triacylglycerol levels have become elevated, and identify the molecular targets of LA. Nine-week old male ZDF (fa/fa) rats were fed a chow diet supplemented with 3g LA per kg diet or pair fed for two weeks (8 rats per treatment). We determined changes in blood triacylglycerol, insulin, non-esterified fatty acids, and ketone bodies concentrations. We analyzed the expression of genes and proteins involved in fatty acid and triacylglycerol metabolism in liver, epididymal fat, and skeletal muscle. Feeding LA to ZDF rats (a) corrected severe hypertriglyceridemia, (b) lowered abdominal fat mass, (c) raised circulating fibroblast growth factor-21 and Fgf21 liver gene expression, (d) repressed lipogenic gene expression of ATP-citrate synthase (Acly), acetyl-coA carboxylase 1 (Acaca), fatty acid synthase (Fasn), sn-glycerol-3-phosphate acyltransferase 1 (Gpam), adiponutrin (Pnpla3) in the liver and adipose tissue, (e) decreased hepatic protein levels of ACC1/2, FASN and 5'-AMP-activated protein kinase catalytic subunit α (AMPKα), (f) did not change phospho-AMPKα/AMPKα and phospho-ACC/ACC ratios, (g) stimulated liver gene expression of PPARα target genes carnitine O-palmitoyltransferase 1ß (Cpt1b) and acyl-CoA thioesterase 1 (Acot1) but not carnitine O-palmitoyltransferase 1α (Cpt1a). This is evidence that short-term LA feeding to obese rats reverses severe hypertriglyceridemia. FGF21 may mediate the beneficial metabolic effects of LA.

[Comparative study of SNP diversity and calculation of the effective size of population in chicken].

A region (200 kb) of Contig. 060226.1 on GGA1 was selected to study the average diversity of Red Jungle Fowl (RJF), Taihe Silk chicken (TS), and White Recessive Rock (WRR), and estimate the effective population size (Ne) of chicken. The mean heterozygosities of RJF, TS and WRR were 0.28533+/-0.034747, 0.32926+/-0.039191 and 0.30168+/-0.040382, respectively. Significant test indicted that there is not significant difference between them (P=0.2368>0.05). The initial chicken effective population size was estimated to be about 20 000-150 000. Chicken has undergone serious population bottleneck effect during the first stage of domestication. However, this bottleneck effect did not result in a substantial loss of diversity as revealed by SNP. The possible explanations for similar SNP diversity between domesticated chicken and Red Jungle Fowl might due to many factors, such as a big Ne in earlier phase of domestication, population expending in breed differentiation, abroad crossing between breeds (especially crossing with RJF), together with high recombination rate in chicken genome.

Tumour oxygen dynamics measured simultaneously by near-infrared spectroscopy and 19F magnetic resonance imaging in rats.

Simultaneous near-infrared spectroscopy (NIRS) and magnetic resonance imaging (MRI) were used to investigate the correlation between tumour vascular oxygenation and tissue oxygen tension dynamics in rat breast 13762NF tumours with respect to hyperoxic gas breathing. NIRS directly detected global variations in the oxygenated haemoglobin concentration (Delta[HbO(2)]) within tumours and oxygen tension (pO(2)) maps were achieved using (19)F MRI of the reporter molecule hexafluorobenzene. Multiple correlations were examined between rates and magnitudes of vascular (Delta[HbO(2)]) and tissue (pO(2)) responses. Significant correlations were found between response to oxygen and carbogen breathing using either modality. Comparison of results for the two methods showed a correlation between the vascular perfusion rate ratio and the mean pO(2) values (R(2) > 0.7). The initial rates of increase of Delta[HbO(2)] and the slope of dynamic pO(2) response, d(pO(2))/dt, of well-oxygenated voxels in response to hyperoxic challenge were also correlated. These results demonstrate the feasibility of simultaneous measurements using NIRS and MRI. As expected, the rate of pO(2) response to oxygen is primarily dependent upon the well perfused rather than poorly perfused vasculature.

Effect of photothermal therapy on breast tumor vascular contents: noninvasive monitoring by near-infrared spectroscopy.

The goal of this study was to investigate the effect of photothermal laser irradiation on rat breast tumor (DMBA-4) vascular contents. An 805-nm diode laser was used in our experiment with a power density ranging from 0.32 to 1.27 W/cm2. The dynamic changes of oxygenated hemoglobin and total hemoglobin concentrations, delta[HbO2] and delta[Hb]total, in rat tumors during photothermal irradiation were noninvasively monitored by a near-infrared spectroscopy system. A multichannel thermal detection system was also used simultaneously to record temperatures at different locations within the tumors. Our experimental results showed that: (1) photoirradiation did have the ability to induce hyperthermic effects inside the rat breast tumors in a single exponential trend; (2) the significant changes (P < 0.005) of delta[HbO2] and delta[Hb]total in response to a low dosage of laser irradiation (0.32 W/cm2) have a single exponential increasing trend, similar to that seen in the tumor interior temperature; and (3) the increase in magnitude of delta[HbO2] is nearly two times greater than that of delta[Hb]total, suggesting that photoirradiation may enhance tumor vascular oxygenation. The last observation may be important to reveal the hidden mechanism of photoirradiation on tumors, leading to improvement of tumor treatment efficiency.

Three promoters regulate the transcriptional activity of the human holocarboxylase synthetase gene.

Holocarboxylase synthetase (HLCS) is the only protein biotin ligase in the human proteome. HLCS-dependent biotinylation of carboxylases plays crucial roles in macronutrient metabolism. HLCS appears to be an essential part of multiprotein complexes in the chromatin that cause gene repression and contribute toward genome stability. Consistent with these essential functions, HLCS knockdown causes strong phenotypes including shortened life span and low stress resistance in Drosophila melanogaster, and de-repression of long-terminal repeats in humans, other mammalian cell lines and Drosophila. Despite previous observations that the expression of HLCS depends on biotin status in rats and in human cell lines, little is known about the regulation of HLCS expression. The goal of this study was to identify promoters that regulate the expression of the human HLCS gene. Initially, the human HLCS locus was interrogated in silico using predictors of promoters including sequences of HLCS mRNA and expressed sequence tags, CpG islands, histone marks denoting transcriptionally poised chromatin, transcription factor binding sites and DNaseI hypersensitive regions. Our predictions revealed three putative HLCS promoters, denoted P1, P2 and P3. Promoters lacked a TATA box, which is typical for housekeeping genes. When the three promoters were cloned into a luciferase reporter plasmid, reporter gene activity was at least three times background noise in human breast, colon and kidney cell lines; activities consistently followed the pattern P1>>P3>P2. Promoter activity depended on the concentration of biotin in culture media, but the effect was moderate. We conclude that we have identified promoters in the human HLCS gene.

Noninvasive monitoring of estrogen effects against ischemic stroke in rats by near-infrared spectroscopy.

The aim of this study was to assess hemodynamic changes by near-infrared spectroscopy (NIRS) during acute focal cerebral ischemia and reperfusion. The study also has evaluated the therapeutic effects of estrogen against vascular dysfunction. Focal cerebral ischemia was induced in nine bilaterally ovariectomized rats, using an endovascular occlusion technique of the middle cerebral artery (MCA). Four out of nine rats had estrogen pretreatment before MCA occlusion (MCAO). The other five rats had MCAO with no pretreatment. The occlusion time was 60 min, followed by 40-60 min of reperfusion. Real-time monitoring of changes in hemoglobin concentrations was performed by a steady-state, two-channel, NIRS system through the period of occlusion and reperfusion. Both changes in total and oxygenated hemoglobin concentrations (D[HbT] and D[HbO(2)]) display apparent periodic fluctuations during occlusion for the rats without estrogen pretreatment, while no rhythmic fluctuation was observed in the rats with the pretreatment. This rhythmic fluctuation is a microvascular dysfunction. Fourier power spectral analysis was performed on the D[HbO(2)] profiles in both rat groups. The results show that the cumulative frequency power of D[HbO(2)] in the range of 0.0025-0.01 Hz for the rats without pretreatment is significantly higher than that with pretreatment. The study implies that the dysfunctional fluctuations disappear in the rats with estrogen pretreatment, demonstrating a new application of NIRS, i.e., to detect focal cerebral ischemia and to monitor cerebral responses to therapy against vascular dysfunction in animal models.

SNP mapping of QTL affecting growth and fatness on chicken GGA1.

An F2 chicken population was established from a crossbreeding between a Xinghua line and a White Recessive Rock line. A total of 502 F2 chickens in 17 full-sib families from six hatches was obtained, and phenotypic data of 488 individuals were available for analysis. A total of 46 SNP on GGA1 was initially selected based on the average physical distance using the dbSNP database of NCBI. After the polymorphism levels in all F0 individuals (26 individuals) and part of the F1 individuals (22 individuals) were verified, 30 informative SNP were potentially available to genotype all F2 individuals. The linkage map was constructed using Cri-Map. Interval mapping QTL analyses were carried out. QTL for body weight (BW) of 35 d and 42 d, 49 d and 70 d were identified on GGA1 at 351-353 cM and 360 cM, respectively. QTL for abdominal fat weight was on GGA1 at 205 cM, and for abdominal fat rate at 221 cM. Two novel QTL for fat thickness under skin and fat width were detected at 265 cM and 72 cM, respectively.