Perturbation of lipid metabolism in 3T3-L1 at different stages of preadipocyte differentiation and new insights into the association between changed metabolites and adipogenesis promoted by TBBPA or TBBPS.

Tetrabromobisphenol A (TBBPA) and tetrabromobisphenol S (TBBPS) are widely distributed brominated flame retardants. While TBBPA has been demonstrated to stimulate adipogenesis, TBBPS is also under suspicion for potentially inducing comparable effects. In this study, we conducted a non-targeted metabolomics to examine the metabolic changes in 3T3-L1 cells exposed to an environmentally relevant dose of TBBPA or TBBPS. Our findings revealed that 0.1 µM of both TBBPA and TBBPS promoted the adipogenesis of 3T3-L1 preadipocytes. Multivariate analysis showed significant increases in glycerophospholipids, sphingolipids, and steroids relative levels in 3T3-L1 cells exposed to TBBPA or TBBPS at the final stage of preadipocyte differentiation. Metabolites set composed of glycerophospholipids was found to be highly effective predictors of adipogenesis in 3T3-L1 cells exposed to TBBPA or TBBPS (revealed from the receiver operating characteristic curve with an area under curve > 0.90). The results from metabolite set enrichment analysis suggested both TBBPA and TBBPS exposures significantly perturbed steroid biosynthesis in adipocytes. Moreover, TBBPS additionally disrupted the sphingolipid metabolism in the adipocytes. Our study presents new insights into the obesogenic effects of TBBPS and provides valuable information about the metabolites associated with adipogenesis induced by TBBPA or TBBPS.

GPM6A expression is suppressed in hepatocellular carcinoma through miRNA-96 production.

GPM6A is a glycoprotein in endothelial cells, and its biological function in the development of hepatocellular carcinoma (HCC) is unknown. Through Affymetrix gene expression microarray and bioinformatic analysis, very low GPM6A expression was found in HCC tissue. The present study aims to explore the function and regulatory mechanism of GPM6A in HCC development and progression. Levels of GPM6A expression in HCC specimens from different disorders and various hepatoma cell lines were determined, and its role on cell proliferation was evaluated in hepatoma cells stably overexpressing GPM6A. Modulation of a specific microRNA (miRNA) on its expression and function was evaluated with miRNA mimetic transfection. Herein, it is reported that much lower GPM6A levels were found in HCC tissues than pericancerous liver tissues and correlated to a poor prognosis. GPM6A overexpression inhibited cell proliferation, suppressed colony formation, migration and invasion in two hepatoma cell types. Available evidence does not support that genetic and epigenetic dysregulation contributes significantly to GPM6A inactivation in HCC. Additional findings demonstrated that miR-96-5p acted directly on the 3'-UTR of the GPM6A gene and significantly decreased its mRNA and protein levels. MiR-96-5p transfection promoted proliferation, migration and invasion of SMMC-7721 and MHCC-97H hepatoma cells; whereas the function of oncogenic microRNA-96 was significantly inhibited in GPM6A-overexpressed hepatoma cells. In conclusion, GPM6A expression in HCC is commonly suppressed regardless its base disease types, and its low expression in HCC tissues is most likely attributed to upregulated miR-96-5p. GPM6A may function as a valuable biomarker for HCC progression and prognosis.

Significance of biotransformation and excretion on the enantioselective bioaccumulation of hexabromocyclododecane (HBCDD) in laying hens and developing chicken embryos.

Although (-)-α-hexabromocyclododecane (HBCDD) and (+)-γ-HBCDD are preferentially enriched in chickens, the key factors contributing to their selective bioaccumulation in hens and their potential biotransformation in developing chicken embryos remain unclear. Herein, in vivo and in ovo exposure experiments using hens and fertilized eggs were conducted to investigate the absorption, excretion, and biotransformation of HBCDDs in chickens. γ-HBCDD (76%) exhibited a higher absorption efficiency than α- (22%) and ß- (69%) HBCDDs. However, α-HBCDD was dominant in hen tissues, although γ-HBCDD accounted for >75% in the spiked feed. Moreover, chicken embryos biotransformed approximately 9.5% and 11.7% of absorbed α- and γ-HBCDDs, respectively, implying that diastereomer-selective elimination causes the predominance of α-HBCDD in hens. The concentration and enantiomer fraction (EF) of α-HBCDD in laid eggs were significantly positively correlated, suggesting enantioselective elimination. The EFs of α- and γ-HBCDDs varied between feces from the exposure and depuration periods, indicating the preferred excretion of (+)-α- and (-)-γ-HBCDDs. Furthermore, the enantioselective biotransformation of (-)-γ-HBCDD was confirmed in developing chicken embryos. These results show that excretion and biotransformation contribute to the diastereomer- and enantiomer-selective bioaccumulation of HBCDDs in chickens; The results may improve our understanding of the environmental fate and ecological risks of HBCDDs in biota.

NLRP3 deficiency did not attenuate NASH development under high fat calorie diet plus high fructose and glucose in drinking water.

NOD-like receptor protein 3 (NLRP3) promotes the inflammatory response during progression of nonalcoholic fatty liver (NAFL) to nonalcoholic steatohepatitis (NASH). This study aimed to further delineate the role of NLRP3 in NASH development by abolishing its expression in mice. A high-fat and calorie diet plus high fructose and glucose in drinking water (HFCD-HF/G) was used to establish NASH in both wild-type (WT) and NLRP3 knock-out (KO) mice. Hepatocellular injury, hepatic steatosis and fibrosis, as well as inflammatory response and insulin resistance in the liver and epidydimal white adipose tissue (eWAT) were determined. Elevated body weight, liver weight and serum alanine transaminase level, increased hepatic triglyceride accumulation and collagen deposition, and worsened systemic insulin resistance were observed in Nlrp3-/- mice compared to WT mice under HFCD-HF/G feeding. Upregulated hepatic transcription of tumor necrosis factor-α (TNF-α) and monocyte chemotactic protein-1 (MCP-1), and enhanced infiltration of inducible nitric oxide synthase-positive (iNOS+) M1 macrophages were also documented in HFCD-HF/G-fed Nlrp3-/- mice in comparison to HFCD-HF/G-fed WT mice. Moreover, transcription of TNF-α and MCP-1 and infiltration of iNOS+ M1 macrophages were increased in the liver of Nlrp3-/- mice under control diet. NLRP3 deficiency did not attenuate, but instead aggravated NASH development under HFCD-HF/G feeding. The worsened extent of NASH might be attributed to enhanced hepatic MCP-1 expression and M1 macrophage infiltration in Nlrp3-/- mice. Our study points to additional caution when NLRP3 blockade is considered as a therapeutic strategy in the treatment of human NASH.

Serum lipocalin-2 is a potential biomarker for the clinical diagnosis of nonalcoholic steatohepatitis.

BACKGROUND/AIMS: Nonalcoholic steatohepatitis (NASH) is a progressive form of nonalcoholic fatty liver disease (NAFLD) characterized by hepatic steatosis, inflammation, hepatocellular injury, and fibrosis. We aimed to investigate the usefulness of a key biomarker, lipocalin-2 (LCN2), for the detection of NASH progression. METHODS: A mouse NASH model was established using a high-fat diet and a high-sugar drinking water. Gene expression profile of the NASH model was analyzed using RNA sequencing. Moreover, 360 NAFLD patients (steatosis, 83; NASH, 277), 40 healthy individuals, and 87 patients with alcoholic fatty liver disease were recruited. RESULTS: Inflammatory infiltration, focal necrosis in the leaflets, steatosis, and fibrosis were documented in the mouse liver. In total, 504 genes were differentially expressed in the livers of NASH mice, and showed significant functional enrichment in the inflammation-related category. Upregulated liver LCN2 was found to be significantly interactive with various interleukins and toll-like receptors. Serum LCN2 levels were significantly increased in NAFLD patients. Serum LCN2 levels were correlated with steatosis, intralobular inflammation, semiquantitative fibrosis score, and nonalcoholic fatty liver disease activity score. The area under the curve of serum LCN2 was 0.987 with a specificity of 100% and a sensitivity of 93.5% for NASH diagnosis, and 0.977 with almost the same specificity and sensitivity for steatosis. CONCLUSION: LCN2 might be involved in the transition from NAFL to NASH by mediating inflammation. Serum LCN2 levels might be a novel biomarker for the diagnosis of NASH.

Effect of laying sequence and selection of maternal tissues in assessment of maternal transfer of organohalogenated contaminants during chicken egg formation: A pilot study.

Many studies have reported maternal transfer of organic contaminants in oviparous species, with inconsonant results. Egg-laying sequence and selected maternal tissues may impact on assessment of potential maternal transfer of contaminants. Here, this hypothesis was verified by exposing chickens (Gallus domesticus) to polychlorinated biphenyls (PCBs), polybrominated diphenyl ethers (PBDEs), and dechlorane plus (DPs). Concentrations in eggs laid during exposure exhibited periodic fluctuations (conforming to egg-laying cycles) and a decreasing trend during depuration. Fluctuation patterns of DPs and BDE209 differed from those of other compounds. The PBDE congener profiles in eggs were dominated by BDE209 during exposure and by BDE100 and 153 during depuration. The abundance of PCB congener (CB138) which is recalcitrant to metabolism increased with laying sequence. Maternal transfer potential was negatively correlated (P = 0.0014, R2 = 0.7874) to the log KOW of chemicals (log KOW >7) when the muscle, heart, lung, or stomach was used. No correlations were found when the liver, fat, kidneys, or intestine was used (log KOW >7), although DPs and BDE209 showed the highest maternal transfer potential. Different fluctuation patterns of DPs and BDE209 in eggs and increased abundance of BDE209 in eggs laid in the initial egg-laying period imply that the liver, fat, kidney, or intestinal tissues could be more appropriate in assessing maternal transfer of the target analytes.

Comparative study of dechlorane plus (DP) in adult chickens and developing embryos: Stereo-selective bioaccumulation of DP in chickens.

Although isomer-specific bioaccumulation of dechlorane plus (DP) has been addressed in many studies, it remains unclear which factors determine this process and whether biotransformation of DP occurs in organisms. Comparative experiments were conducted in both in vivo and in ovo incubation using hens and eggs to identify the dominant factors determining the bioaccumulation of DP. Hens and fertilized eggs were exposed to DP isomers (syn- and anti-DP) by feeding and spiking, respectively, to investigate absorption, elimination, and metabolism. No significant differences were found between absorption efficiencies of DP isomers in the adult hens. Following first-order kinetics, anti-DP exhibited a slightly longer half-life than syn-DP as well as an elevated anti-DP fraction in laid eggs, thereby suggesting selective enrichment of anti-DP in adult hens. However, chicken embryos metabolized approximately 12% and 28% of the absorbed syn- and anti-DP, respectively, thereby verifying that anti-DP was preferably metabolized. This result indicated that stereo-selective excretion of syn-DP, rather than preferred metabolism of anti-DP, played a more prominent role in isomer-specific bioaccumulation of DP in chickens. Further studies on metabolites of DP are crucial to understanding the fate of DP in organisms.

Iron Porphyrin Catalyzed Insertion Reaction of N-Tosylhydrazone-Derived Carbenes into X-H (X = Si, Sn, Ge) Bonds.

An efficient Fe(TPP)Cl catalyzed insertion reaction of in situ generated benzylic carbenes from N-tosylhydrazones into X-H (X = Si, Sn, Ge) was developed. Silanes bearing tertiary, secondary, and primary (3°, 2°, and 1°) Si-H bonds all reacted well to afford insertion products in moderate to high yields (up to 97%), and the reaction time could be significantly shortened to 1 h under microwave irradiation. A programmable stepwise double insertion strategy was developed for the synthesis of unsymmetrical tetrasubstituted silanes.

In ovo uptake, metabolism, and tissue-specific distribution of chiral PCBs and PBDEs in developing chicken embryos.

Fertilized chicken eggs were injected with environmental doses of 4 chiral polychlorinated biphenyls (PCBs) and 8 polybrominated biphenyl ethers (PBDEs) to investigate their uptake, metabolism in the embryo, and distribution in the neonate chicken. PCB95 uptake was the most efficient (80%) whereas BDE209 was the least (56%). Embryos metabolized approximately 52% of the PCBs absorbed. Though some degree of metabolism in the first 18 days, most of the PCBs and PBDEs was metabolized in the last three days, when BDE85, 99, 153, and 209 decrease by 11-37%. Enantioselective metabolism of the (+) enantiomers of PCB95, 149, and 132 and the (-) enantiomer of PCB91 was observed. The enantioselective reactivity was higher with the two penta-PCBs than the two tetra-PCBs. Liver, exhibited high affinity for high lipophilic chemicals, enrich all chemicals that was deflected in other tissues except for some special chemicals in a given tissues. Lipid composition, time of organ formation, and metabolism contribute to the distribution of chemicals in the neonate chicken. The result of this study will improve our understanding on the fate and potential adverse effects of PCBs and PBDEs in the neonate chicken.

Tracing the biotransformation of polychlorinated biphenyls (PCBs) in common carp (Cryprinus carpio): Enantiomeric fraction and compound-specific stable carbon isotope analyses.

Metabolites of polychlorinated biphenyls (PCBs) in fish are difficult to detect in vivo due to the complexity of biometabolism. In the present study, atropisomeric fraction analysis of chiral PCB congeners and compound-specific isotopic analysis (CSIA) were applied to trace the biotransformation of PCBs in fish by exposure of common carp (Cryprinus carpio) to the commercial PCB mixture Aroclor 1242. Stereoselective elimination of the chiral PCB congeners 91, 95, and 136 was observed, indicating a stereoselective biotransformation process. The δ(13)C values of PCBs 5/8, 18, and 20/33 in fish were increased compared with those in the spiked food, while PCBs 47/48 and 49 showed significant heavy isotope depletion. These results suggested a significant biotransformation of the corresponding individual PCB congeners although the potential PCB metabolites, hydroxylated PCBs (OH-PCBs) and methylsulfone PCBs (MeSO2-PCBs), were not detected in the fish tissue samples throughout this experiment. The results of the present study demonstrated that a combination of chiral analysis and CSIA is a promising new approach for investigating the biotransformation of PCBs in biota.

Distribution of organophosphorus flame retardants in sediments from the Pearl River Delta in South China.

Twelve organophosphorus flame retardants (PFRs) were identified in the sediments and the sediment core collected from the rivers and the estuary in the Pearl River Delta, with the aim of investigating their spatial and vertical distributions. The concentrations of PFRs ranged from 8.3 to 470 ng/g dry weight with high levels of PFRs in the urban area and the e-waste recycling region. Generally, TPhP, TCPP, TEHP, TCEP, and TBEP were the dominant compounds of the PFRs, the composition of which varied across the different regions, reflecting the different sources of PFRs. In the estuary, the PFRs mainly derived from the Xijiang River and the Shunde sections. Increased concentrations of halogen-containing PFRs have been observed in the upper layers of the sediment core. Conversely, relatively high concentrations of halogen-free PFRs were observed in the lower layers of the sediment core, indicating different usage patterns or environmental behaviors between the halogen and the non-halogen PFRs in the study area.

Legacy and emerging halogenated organic pollutants in marine organisms from the Pearl River Estuary, South China.

A suite of legacy and emerging halogenated organic pollutants (HOPs) were measured in marine organisms (coastal fish and invertebrates) from the Pearl River Estuary, South China, to investigate the current contamination status after the Stockholm Convention was implemented in China. Dichlorodiphenyltrichloroethane and its metabolites (DDTs), polychlorinated biphenyls (PCBs), and polybrominated diphenyl ethers (PBDEs) were detected in all samples at concentrations of 54-1500, 16-700, and 0.56-59ng/g lipid weight, respectively. Dechlorane Plus (DP), decabromodiphenyl ethane (DBDPE), 2,3,5,6-tetrabromo-p-xylene (pTBX), and pentabromotoluene (PBT) were also found at concentrations of ND (non-detectable) to 37ng/g lipid weight. The concentrations of these investigated contaminants in the present study were at moderate levels, as compared with those reported in other regions. Significant interspecies differences were found in the levels of DDTs, PCBs, PBDEs and the alternative halogenated flame retardants (AHFRs). DDTs were the predominant HOPs in those species and represented >50% of the total HOPs, followed by PCBs, PBDEs, and AHFRs. The total estimated daily intakes (EDIs) of DDTs, PCBs, PBDEs, and AHFRs were 28, 12, 1.0, and 0.18 (ng/kg)/d, respectively, via seafood consumption. These concentrations are not expected to pose health risks to humans.

An eight year (2005-2013) temporal trend of halogenated organic pollutants in fish from the Pearl River Estuary, South China.

Dichlorodiphenyltrichloroethane and its metabolites (DDTs), hexachlorocyclohexanes (HCHs), polychlorinated biphenyls (PCBs), polybrominated diphenyl ethers (PBDEs), dechlorane plus (DP), 2,3,5,6-tetrabromo-p-xylene (pTBX) and pentabromotoluene (PBT) were measured in baby croaker (Collichthys lucidus) and mullet (Osteomugil ophuyseni) collected in 2005 and 2013 from the Pearl River Estuary. DDTs, HCHs, PCBs, and PBDEs were detected in two fish species at concentrations of 150-8100, 1.4-120, 22-560, 2.2-280 ng/g lipid wt., respectively. The levels of these chemicals were significantly lower in 2013 than in 2005. The compositions for DDTs, HCHs, and PBDEs in 2013 differed from those in 2005, indicating source changes between the two sampling periods. DP, pTBX and PBT were detected at concentrations of ND-130 ng/g lipid wt. No clear temporal trends were found for these contaminants. Overall, these results indicated the effectiveness of regulations and source controls in substantively reducing inputs of these contaminants to the Pearl River Estuary.