Chenyang Lu

Modulation of the Gut Microbiota by Krill Oil in Mice Fed a High-Sugar High-Fat Diet

Multiple lines of evidence suggest that the gut microbiota plays vital roles in metabolic diseases such as hyperlipidemia. Previous studies have confirmed that krill oil can alleviate hyperlipidemia, but the underlying mechanism remains unclear. To discern whether krill oil changes the structure of the gut microbiota during the hyperlipidemia treatment, 72 mice were acclimatized with a standard chow diet for 2 weeks and then randomly allocated to receive a standard chow diet (control group, n = 12) or a high-sugar-high-fat (HSHF) diet supplemented with a low (100 μg/g·d, HSHF+LD group, n = 12), moderate (200 μg/g·d, HSHF+MD group, n = 12) or high dosage of krill oil (600 μg/g·d, HSHF+HD group, n = 12), simvastatin (HSHF+S group, n = 12) or saline (HSHF group, n = 12) continuously for 12 weeks. The resulting weight gains were attenuated, the liver index and the low-density lipoprotein, total cholesterol and triglyceride concentrations showed a stepwise reduction in the treated groups compared with those of the control group. A dose-dependent modulation of the gut microbiota was observed after treatment with krill oil. Low- and moderate- doses of krill oil increased the similarity between the composition of the HSHF diet-induced gut microbiota and that of the control, whereas the mice fed the high-dose exhibited a unique gut microbiota structure that was different from that of the control and HSHF groups. Sixty-five key operational taxonomic units (OTUs) that responded to the krill oil treatment were identified using redundancy analysis, of which 26 OTUs were increased and 39 OTUs were decreased compared with those of the HSHF group. In conclusion, the results obtained in this study suggest that the structural alterations in the gut microbiota induced by krill oil treatment were dose-dependent and associated with the alleviation of hyperlipidemia. Additionally, the high-dose krill oil treatment showed combined effects on the alleviation of hyperlipidemia and obesity.

Modulation of the gut microbiota by the mixture of fish oil and krill oil in high-fat diet-induced obesity mice

Previous studies confirmed that dietary supplements of fish oil and krill oil can alleviate obesity in mice, but the underlying mechanism remains unclear. This study aims to discern whether oil treatment change the structure of the gut microbiota during the obesity alleviation. The ICR mice received high-fat diet (HFD) continuously for 12 weeks after two weeks of acclimatization with a standard chow diet, and the mice fed with a standard chow diet were used as the control. In the groups that received HFD with oil supplementation, the weight gains were attenuated and the liver index, total cholesterol, triglyceride and low-density lipoprotein cholesterol were reduced stepwise compared with the HFD group, and the overall structure of the gut microbiota, which was modulated in the HFD group, was shifted toward the structure found in the control group. Moreover, eighty-two altered operational taxonomic units responsive to oil treatment were identified and nineteen of them differing in one or more parameters associated with obesity. In conclusion, this study confirmed the effect of oil treatment on obesity alleviation, as well as on the microbiota structure alterations. We proposed that further researches are needed to elucidate the causal relationship between obesity alleviation and gut microbiota modulation.

Dietary Apostichopus japonicus Alleviates Diabetes Symptoms and Modulates Genes Expression in Kidney Tissues of db/db Mice

The effects of Apostichopus japonicus enzymatic hydrolysate on the regulation of dyslipidemia, pathoglycemia, and transcription changes in kidney tissues of db/db mice were evaluated. In this study, the symptoms of diabetes in db/db mice were alleviated after 10 weeks of treatments with low (db/db + LD group) and high dose (db/db + HD group) of Apostichopus japonicus enzymatic hydrolysate, and the high dose treatment showed a better antidiabetic effect. Compared with the db/db group, the fasting blood glucose levels (36.84 ± 7.82 vs 25.18 ± 6.84 mmol/L, P < 0.01), the urine glucose levels (45.44 ± 3.93 vs 22.66 ± 5.58 mmol/L, P < 0.01), and the serum insulin sensitivity index (-4.65 ± 0.43 vs -4.74 ± 0.75, P > 0.05) in the db/db + HD group were decreased, whereas the fasting plasma insulin (3.12 ± 1.08 vs 5.54 ± 1.82 μg/L, P < 0.01) and the serum insulin resistance index (5.01 ± 2.02 vs 5.96 ± 2.49, P < 0.05) were increased. Subsequently, the kidney transcription profiles were measured in the db/db group and db/db + HD group via microarray, and the results show that Apostichopus japonicus hydrolysate induced differential expression of 77 genes. Among these genes, the down-regulation of genes ntrK1 and ptpN5 played vital roles, as this effect induced the further down-regulation of neurotrophin tyrosine kinase, protein tyrosine phosphatase, and other transcription factors, which are involved in the classical mitogen-activated protein kinases (MAPK) and p38MAPK signaling pathways. The inhibited MAPK and p38MAPK signaling pathways are involved in glycometabolism and the control of lipid metabolism, and they regulate the occurrence and development of diabetic nephropathy.

A metabonomic analysis on the response of Enterobacter cloacae from coastal outfall for land-based pollutant under phoxim stress

Enterobacter cloacae is an opportunistic pathogen widely distributed in human and animal intestinal systems. The secretion of extended-spectrum β-lactamases (ESBLs) and cephalosporinase (AmpC) endows E. cloacae with strong drug resistance. In a previous study by our group, protein expression of E. cloacae under phoxim stress was measured by two-dimensional electrophoresis. Here, nuclear magnetic resonance was used to detect differences in E. cloacae metabonomics when under phoxim stress. We determined that there are 29 types of metabolites that differ between phoxim stress and normal culture conditions. Among these, 6 types of metabolites were upregulated in the phoxim stress group, and 23 types of metabolites were inhibited. Though enrichment analysis, seven pathways were identified by different expression levels of metabolites, which were involved in DNA and RNA synthesis, DNA damage repair, antioxidation and functions of the cell membrane and cell wall. The mechanism underlying how phoxim affects E. cloacae was determined by studying the results of both two-dimensional electrophoresis in our prior work and the analysis of E. cloacae metabonomic changes under phoxim stress.

Sex-Based Differences in Gut Microbiota Composition in Response to Tuna Oil and Algae Oil Supplementation in a D-galactose-Induced Aging Mouse Model

Our previous work indicated that a mixture of tuna oil and algae oil treatment in male mice effectively relieved D-galactose (D-gal)-induced aging and resulted in gut microbiota alterations, and that the best anti-aging effects were observed for a tuna oil to algae oil ratio of 1:2. However, the possibility of a sex-based difference in the anti-aging effect of the tuna oil and algae oil mixture or gut microbiota variation, has rarely been investigated. In this study, the anti-aging effect of an oil mixture (1:2) in male and female mice was measured, and oil treatment improved the learning and cognition of mice that were damaged by D-gal, increased the activities of anti-oxidative enzymes, and decreased the level of MDA, which acted as a hallmark of oxidative damage to lipids. Male mice showed better anti-aging effects than female mice with a specific oil mixture ratio, and the clinical drug donepezil showed a similar or better effect on aging alleviation than oil treatments in both sexes. On the other hand, the same oil treatment led to different gut microbiota composition alterations in male and female mice. Redundancy analysis (RDA) identified 31 and 30 key operational taxonomic units (OTUs) in the male and female mice, respectively, and only three of these OTUs overlapped. Moreover, the abundance of Lactobacillus and several probiotic-like butyric acid producers was higher in male mice than in female mice, whereas the abundance of some inflammation-related genera, such as Clostridium XlVa, was lower in male mice. In conclusion, this study indicated the sex-based differences related to the anti-aging effects of tuna oil and algae oil treatment are accompanied by sex-based differences in gut microbiota modulation.

Proteomics and 1 H NMR-based metabolomics analysis of pathogenic Vibrio vulnificus aquacultures isolated from sewage drains

Vibrio bacteria live in both marine and freshwater habitats and are associated with aquatic animals. Vibrio vulnificus is a pathogenic bacterium that infects people and livestock. It is usually found in offshore waters or within fish and shellfish. This study presents a comparative proteomic analysis of the outer membrane protein (OMP) changes in V. vulnificus proteins after stimulation with sewage from sewage drains. Using two-dimensional electrophoresis followed by MALDI-TOF MS/MS, 32 protein spots with significant differences in abundance were identified and characterized. These identified proteins were found to be involved in various functional categories, including catalysis, transport, membrane proteins progresses, receptor activity, energy metabolism, cytokine activity, and protein metabolism. The mRNA expression levels of 12 differential proteins were further assessed by qRT-PCR. Seven genes including carboxypeptidase, hemoglobin receptor, succinate dehydrogenase iron-sulfur subunit, ATP synthase subunit alpha, thioredoxin, succinyl-CoA synthetase subunit, and alanine dehydrogenase were downregulated upon stimulation, whereas the protein expression levels HupA receptor, type I secretion outer membrane protein, glutamine synthetase, superoxide dismutase, OmpU, and VuuA were upregulated. 1H NMR spectra showed 18 dysregulated metabolites from V. vulnificus after the sewage stimulation and the pathogenicity was enhanced after that.

In silico analysis and in vivo tests of the tuna dark muscle hydrolysate anti-oxidation effect

Hydrolysate is a mixture of various peptides with specific functions. However, functional identification of hydrolysate with high throughput is still a difficult task. Furthermore, using in vivo tests via animal or cell experiments is time and labor-intensive. In this study, the peptides component of hydrolysate derived from the tuna dark muscle was measured via MALDI-TOF/TOF-MS, and the functions of the KEFT (Lys-Glu-Phe-Thr), EEASA (Glu-Glu-Ala-Ser-Ala) and RYDD (Arg-Tyr-Asp-Asp) peptides, which were found with the highest proportion, were predicted via Discovery Studio 2016 software. All three peptides were predicted to bind to the Keap1 protein with the highest fit-value and to affect the activity of Keap1, which is involved in anti-oxidation pathways. Subsequently, mice experiments showed that administration of tuna dark muscle hydrolysate increased the levels of superoxide dismutase and glutathione peroxidase in the serum and liver (P 0.05), which are consistent with the in silico analysis results using Discovery Studio 2016 software. The combination of in silico analysis and in vivo tests provided an alternative strategy for identifying hydrolysate function and provided insight into high-value utilization of protein hydrolysate.

Tuna Oil Alleviates d-Galactose Induced Aging in Mice Accompanied by Modulating Gut Microbiota and Brain Protein Expression

To discern whether tuna oil modulates the expression of brain proteins and the gut microbiota structure during aging induced by d-galactose, we generated an aging mouse model with d-galactose treatment, and the mice showed aging and memory deterioration symptoms according to physiological and biochemical indices. Treatment with different doses of tuna oil alleviated the symptoms; the high dose showed a better effect. Subsequently, brain proteomic analysis showed the differentially expressed proteins were involved in damaged synaptic system repairment and signal transduction system enhancement. In addition, tuna oil treatment restored the diversity of gut microbiota, 27 key operational taxonomic units, which were identified using a redundancy analysis and were significantly correlated with at least one physiological index and three proteins or genes. These findings suggest that the combination of proteomics and gut microbiota is an effective strategy to gain novel insights regarding the effect of tuna oil treatment on the microbiota-gut-brain axis.

Heavy metal detoxification by recombinant ferritin from Apostichopus japonicus

Ferritin is a bionanomaterial that is widely applied in magnetic resonance imaging, drug delivery systems, biocompatible fluorescence, neutron-capture therapy and electrochemical markers. Ferritin also has great potential for use in environmental detection and heavy metal removal due to its hollow cage sequestration and maintenance of iron in a nontoxic and bio-available form. In this study, the heavy metal binding activity of ferritin from Apostichopus japonicus (AjFER) was elucidated using scanning electron microscopy (SEM). It was observed that ferritin aggregation morphology changed dramatically upon exposure to five metals. The reaction systems formed ferritin-Cd, ferritin-As, ferritin-Hg, ferritin-Pb, and ferritin-Cr aggregations. The aggregations of horse spleen ferritin (HSF) and AjFER are relatively unified rules. The enrichment capacity of ferritin was further analyzed using Inductively Coupled Plasma Mass spectrometry. The contents of Cd2+, Hg2+, Cr3+, Pb2+, and As3+ enriched by recombinant AjFER were higher than that for the standard HSF in ordinary groups. Moreover, the concentration levels of enriched heavy metal ions in groups treated with phosphate were higher than those in ordinary dialysis control AjFER groups. The conformation stability of AjFER binding to different heavy metal ions was determined using Circular Dichroism (CD) spectroscopy, which revealed that the second structure of Mn+-ferritin has relative stability. The affinity of ferritin for metal ions was determined using Isothermal Titration Calorimetry (ITC), and the order of binding constant was Pb-Fer > Hg-Fer > Cd-Fer > As-Fer > Cr-Fer. The ion channel was measured using site-directed mutagenesis, which revealed that heavy metal ions primarily enter the protein cage via a three-fold channel and that the ferroxidase center may be one of the key sites for metal ion diffusion into the interior of the protein shell.

NMR-based metabolomics reveals the metabolite profiles of Vibrio parahaemolyticus under ferric iron stimulation

Vibrio parahaemolyticus is a halophilic bacterium endemic to coastal areas, and its pathogenicity has caused widespread seafood poisoning. In our previous research, the protein expression of V. parahaemolyticus in Fe3+ medium was determined using isobaric tags for relative and absolute quantitation (iTRAQ). Here, nuclear magnetic resonance (NMR) was used to detect changes in the V. parahaemolyticus metabolome. NMR spectra were obtained using methanol-water extracts of intracellular metabolites from V. parahaemolyticus under various culture conditions, and 62 metabolites were identified, including serine, arginine, alanine, ornithine, tryptophan, glutamine, malate, NAD+, NADP+, oxypurinol, xanthosine, dCTP, uracil, thymine, hypoxanthine, and betaine. Among these, 21 metabolites were up-regulated after the stimulation of the cells by ferric iron, and 9 metabolites were down-regulated. These metabolites are involved in amino acid and protein synthesis, energy metabolism, DNA and RNA synthesis and osmolality. Based on these results, we conclude that Fe3+ influences the metabolite profiles of V. parahaemolyticus.