Hehua Lei

High-Fat Diet Induces Dynamic Metabolic Alterations in Multiple Biological Matrices of Rats

Obesity is a condition resulting from the interactions of individual biology and environmental factors causing multiple complications. To understand the systems metabolic changes associated with the obesity development and progression, we systematically analyzed the dynamic metabonomic changes induced by a high-fat diet (HFD) in multiple biological matrices of rats using NMR and GC-FID/MS techniques. Clinical chemistry and histopathological data were obtained as complementary information. We found that HFD intakes caused systematic metabolic changes in blood plasma, liver, and urine samples involving multiple metabolic pathways including glycolysis, TCA cycle, and gut microbiota functions together with the metabolisms of fatty acids, amino acids, choline, B-vitamins, purines, and pyrimidines. The HFD-induced metabolic variations were detectable in rat urine a week after HFD intake and showed clear dependence on the intake duration. B-vitamins and gut microbiota played important roles in the obesity development and progression together with changes in TCA cycle intermediates (citrate, α-ketoglutarate, succinate, and fumarate). 83-day HFD intakes caused significant metabolic alterations in rat liver highlighted with the enhancements in lipogenesis, lipid accumulation and lipid oxidation, suppression of glycolysis, up-regulation of gluconeogenesis and glycogenesis together with altered metabolisms of choline, amino acids and nucleotides. HFD intakes reduced the PUFA-to-MUFA ratio in both plasma and liver, indicating the HFD-induced oxidative stress. These findings provided essential biochemistry information about the dynamic metabolic responses to the development and progression of HFD-induced obesity. This study also demonstrated the combined metabonomic analysis of multiple biological matrices as a powerful approach for understanding the molecular basis of pathogenesis and disease progression.

Dynamic Metabolic Response of Mice to Acute Mequindox Exposure

Mequindox is used as a veterinary antibiotic drug. As part of systematic investigations into mequindox as a veterinary medicine and its subsequent applications in food safety, we conducted the investigation to assess the metabolic response of mice to mequindox using metabonomics, which combines NMR metabolic profiles of biofluids or tissues and pattern recognition data analysis. In this study, we delivered a single dose of mequindox to mice with dosage levels of 15, 75, and 350 mg/kg body weight and collected urine samples over a 7 day period, as well as plasma and liver tissues at 7 days postdose. Principal components analysis (PCA) and orthogonal projection to latent structure discriminant analysis (O-PLS-DA) were performed on (1)H NMR spectra of biofluids and liver, showing that low dose levels of mequindox exposure had no adverse effects, consistent with histological observations of the liver. High and moderate levels of mequindox exposure caused suppression of glycolysis and stimulation of fatty acid oxidation accompanied with increased levels of oxidative stress. Our metabonomic analyses also showed disruption of amino acid metabolism, consistent with liver damage observed from histopathological examinations. Furthermore, mequindox perturbed gut microbial activity manifested in the altered excretion of urinary trimethylamine (TMA), trimethylamine-N-oxide (TMAO), hippurate, phenylacetylglycine (PAG), and phenylacetate. The putative gut microbial function may also contribute to the assembly and secretion of very-low-density lipoproteins from the liver to the plasma. Our work provides important insights on the metabolic responses of mequindox.

The metabolic responses to hepatitis B virus infection shed new light on pathogenesis and targets for treatment

Chronic infection caused by the hepatitis B virus (HBV), is strongly associated with hepatitis, fatty liver and hepatocellular carcinoma. To investigate the underlying mechanisms, we characterize the metabolic features of host cells infected with the virus using systems biological approach. The results show that HBV replication induces systematic metabolic alterations in host cells. HBV infection up-regulates the biosynthesis of hexosamine and phosphatidylcholine by activating glutamine-fructose-6-phosphate amidotransferase 1 (GFAT1) and choline kinase alpha (CHKA) respectively, which were reported for the first time for HBV infection. Importantly suppressing hexosamine biosynthesis and phosphatidylcholine biosynthesis can inhibit HBV replication and expression. In addition, HBV induces oxidative stress and stimulates central carbon metabolism and nucleotide synthesis. Our results also indicate that HBV associated hepatocellular carcinoma could be attributed to GFAT1 activated hexosamine biosynthesis and CHKA activated phosphatidylcholine biosynthesis. This study provides further insights into the pathogenesis of HBV-induced diseases, and sheds new light on drug target for treating HBV infection.

Dose-dependent effects of extracted microcystins on embryonic development, larval growth and histopathological changes of southern catfish (Silurus meridionalis)☆

A laboratory toxic experiment was conducted to examine dose-dependent effects of extracted microcystins (MCs) on embryonic development, larval growth and histopathological changes of southern catfish (Silurus meridionalis). Fertilized eggs were incubated in solutions with four concentrations of MCs (0, 1, 10, 100 microg MC-L Req l(-1)). Higher MCs retarded egg development (2-10h delays) and larval growth, reduced hatching rate (up to 45%), and caused high malformation rate (up to 15%) and hepatocytes damage (characterized by disorganization of cell structure and a loss of adherence between hepatocytes, cellular degeneration with vacuolar hepatocytes and marginal nuclei, even hepatocellular necrosis). A 10 microg MC-L Req l(-1) is close to a high concentration in natural cyanobacterial blooms, suggesting a possible existence of such toxic effects in eutrophic waters.

Distribution of toxins in various tissues of crucian carp intraperitoneally injected with hepatotoxic microcystins

An acute toxicity experiment was conducted to examine the distribution and depuration of microcystins (MCs) in crucian carp (Carassius aurutus) tissues. Fish were injected intraperitoneally with extracted MCs at a dose of 200 microg MC-LR (where L=leucine and R=arginine) equivalent/kg body weight. Microcystin concentrations in various tissues and aquaria water were analyzed at 1, 3, 12, 24, and 48 h postinjection using liquid chromatography coupled with mass spectrometry. Microcystins were detected mainly in blood (3.99% of injected dose at 1 h), liver (1.60% at 1 h), gonad (1.49% at 3 h), and kidney (0.14% at 48 h). Other tissues, such as the heart, gill, gallbladder, intestine, spleen, brain, and muscle, contained less than 0.1% of the injected MCs. The highest concentration of MCs was found in blood (526-3,753 ng/g dry wt), followed by liver (103-1,656 ng/g dry wt) and kidney (279-1,592 ng/g dry wt). No MC-LR was detectable in intestine, spleen, kidney, brain, and muscle, whereas MC-RR was found in all examined fish tissues, which might result from organ specificity of different MCs. Clearance of MC-RR in brain tissue was slow. In kidney, the MC-RR content was negatively correlated with that in blood, suggesting that blood was important in the transportation of MC-RR to kidney for excretion.

Metabolic Influence of Acute Cyadox Exposure on Kunming Mice

Cyadox is an antibiotic drug and has the potential to be used as a feedstuff additive in promoting the growth of animals. However, the toxicity of cyadox should be fully assessed before application, and this has prompted the current investigation on the metabolic responses of mice to cyadox exposure, using a metabonomic technique. Three groups of Kunming mice were respectively given a single dose of cyadox at three different concentrations (100, 650, and 4000 mg/kg body weight) via gavage. We present here the metabolic alterations of urine, plasma, liver, and renal medulla extracts induced by cyadox exposure. The metabolic alterations induced by cyadox exposure are dose-dependent, and metabolic recovery is achieved only for low and moderate levels of cyadox exposure during the experimental period. Cyadox exposure resulted in a disturbance of gut microbiota, which is manifested in depleted levels of urinary hippurate, trimethylamine-N-oxide (TMAO), dimethylamine (DMA), and trimethylamine (TMA). In addition, mice exposed to cyadox at high levels caused accumulations of amino acids and depletions of nucleotides in the liver. Furthermore, marked elevations of nucleotides and a range of organic osmolytes, such as myo-inositol, choline, and glycerophosphocholine (GPC), and decreased levels of amino acids are observed in the renal medulla of cyadox-exposed mice. These results suggest that cyadox exposure causes inhibition of amino acid metabolism in the liver and disturbance of gut microbiota community, influencing osmolytic homeostasis and nucleic acids synthesis in both the liver and the kidney. Our work provides a comprehensive view of the toxicological effects of cyadox, which is important in animal and human food safety.

Renal accumulation and effects of intraperitoneal injection of extracted microcystins in omnivorous crucian carp (Carassius auratus)

An acute toxicological experiment was designed to characterize the sequence of renal ultrastructural changes with accumulated MCs in crucian carp injected intraperitoneally (i.p.) with extracted microcystins (mainly MC-RR and -LR) at two doses, 50 and 200 μg MC-LReq. kg⁻¹ body weight. Quantitative and qualitative determinations of MCs in the kidney were conducted by HPLC and LC-MS, respectively. MC-RR content in kidney of crucian carp showed a time dose-dependent increase within 48 h post-injection, followed by a sharp decline afterward, while no MC-LR in kidney was detectable throughout the experiment. Ultrastructural changes in the kidney of crucian carp progressed with increasing accumulated MCs and exposure times within 48 h post-injection, whereas renal ultrastructural recovery of crucian carp in the 50 μg MC-LReq. kg⁻¹ dose group was evident at 168 h post-injection. Our ultrastructural observation suggests that the membranous structure is the main action site of MCs in the kidney, among which mitochondria damage in the tubules is clearly an early, and presumably a critically important effect of MCs. The increases in blood urea nitrogen (BUN) and creatinine (CR) in both dose groups further revealed severe impairment occurred in the kidney of crucian carp.

Metabonomic Analysis Reveals Efficient Ameliorating Effects of Acupoint Stimulations on the Menopause-caused Alterations in Mammalian Metabolism

Acupoint stimulations are effective in ameliorating symptoms of menopause which is an unavoidable ageing consequence for women. To understand the mechanistic aspects of such treatments, we systematically analyzed the effects of acupoint laser-irradiation and catgut-embedding on the ovariectomy-induced rat metabolic changes using NMR and GC-FID/MS methods. Results showed that ovariectomization (OVX) caused comprehensive metabolic changes in lipid peroxidation, glycolysis, TCA cycle, choline and amino acid metabolisms. Both acupoint laser-irradiation and catgut-embedding ameliorated the OVX-caused metabonomic changes more effectively than hormone replacement therapy (HRT) with nilestriol. Such effects of acupoint stimulations were highlighted in alleviating lipid peroxidation, restoring glucose homeostasis and partial reversion of the OVX-altered amino acid metabolism. These findings provided new insights into the menopause effects on mammalian biochemistry and beneficial effects of acupoint stimulations in comparison with HRT, demonstrating metabonomics as a powerful approach for potential applications in disease prognosis and developments of effective therapies.

Systems Responses of Rats to Mequindox Revealed by Metabolic and Transcriptomic Profiling

Mequindox is used as an antibiotic drug in livestock; however, its toxicity remains largely unclear. Previously, we investigated metabolic responses of mice to mequindox exposure. In order to evaluate dependences of animal species in response to mequindox insult, we present the metabolic consequences of mequindox exposure in a rat model, by employing the combination of metabonomics and transcriptomics. Metabolic profiling of urine revealed that metabolic recovery is achieved for rats exposed to a low or moderate dose of mequindox, whereas high levels of mequindox exposure trigger liver dysfunction, causing no such recovery. We found that mequindox exposure causes suppression of the tricarboxylic acid cycle and stimulation of glycolysis, which is in contrast to a mouse model previously investigated. In addition, mequindox dosage induces promotion of β-oxidation of fatty acids, which was confirmed by elevated expressions of acox1, hsd17b2, and cpt1a in liver. Furthermore, altered levels of N-methylnicotinate, 1-methylnicotinamide, and glutathione disulfide highlighted the promotion of vitamin B3 antioxidative cycle in rats exposed to mequindox. Moreover, mequindox exposure altered levels of gut microbiotal related co-metabolites, suggesting a perturbation of the gut microflora of the host. Our work provides a comprehensive view of the toxicological effects of mequindox, which is important in the usage of mequindox in animal and human food safety.

Combined NMR and GC-MS Analyses Revealed Dynamic Metabolic Changes Associated with the Carrageenan-Induced Rat Pleurisy

Inflammation is closely associated with pathogenesis of various metabolic disorders, cardiovascular diseases, and cancers. To understand the systems responses to localized inflammation, we analyzed the dynamic metabolic changes in rat plasma and urine associated with the carrageenan-induced self-limiting pleurisy using NMR spectroscopy in conjunction with multivariate data analysis. Fatty acids in plasma were also analyzed using GC-FID/MS with the data from clinical chemistry and histopathology as complementary information. We found that in the acute phase of inflammation rats with pleurisy had significantly lower levels in serum albumin, fatty acids, and lipoproteins but higher globulin level and larger quantity of pleural exudate than controls. The carrageenan-induced inflammation was accompanied by significant metabolic alterations involving TCA cycle, glycolysis, biosyntheses of acute phase proteins, and metabolisms of amino acids, fatty acids, ketone bodies, and choline in acute phase. The resolution process of pleurisy was heterogeneous, and two subgroups were observed for the inflammatory rats at day-6 post treatment with different metabolic features together with the quantity of pleural exudate and weights of thymus and spleen. The metabolic differences between these subgroups were reflected in the levels of albumin and acute-phase proteins, the degree of returning to normality for multiple metabolic pathways including glycolysis, TCA cycle, gut microbiota functions, and metabolisms of lipids, choline and vitamin B3. These findings provided some essential details for the dynamic metabolic changes associated with the carrageenan-induced self-limiting inflammation and demonstrated the combined NMR and GC-FID/MS analysis as a powerful approach for understanding biochemical aspects of inflammation.