Fatty Acid Elongation In Mitochondria Research Articles

Metabolome Profiling and Pathway Analysis in Metabolically Healthy and Unhealthy Obesity among Chinese Adolescents Aged 11-18 Years.

The underlying mechanisms of the development of unhealthy metabolic phenotypes in obese children and adolescents remain unclear. We aimed to screen the metabolomes of individuals with the unhealthy obesity phenotype and identify the potential metabolic pathways that could regulate various metabolic profiles of obesity in Chinese adolescents. A total of 127 adolescents aged 11-18 years old from China were investigated using a cross-sectional study. The participants were classified as having metabolically healthy obesity (MHO) or metabolically unhealthy obesity (MUO) based on the presence/absence of metabolic abnormalities defined by metabolic syndrome (MetS) and body mass index (BMI). Serum-based metabolomic profiling using gas chromatography-mass spectrometry (GC-MS) was undertaken on 67 MHO and 60 MUO individuals. ROC analyses showed that palmitic acid, stearic acid, and phosphate could predict MUO, and that glycolic acid, alanine, 3-hydroxypropionic acid, and 2-hydroxypentanoic acid could predict MHO (all p < 0.05) from selected samples. Five metabolites predicted MUO, 12 metabolites predicted MHO in boys, and only two metabolites predicted MUO in girls. Moreover, several metabolic pathways may be relevant in distinguishing the MHO and MUO groups, including the fatty acid biosynthesis, fatty acid elongation in mitochondria, propanoate metabolism, glyoxylate and dicarboxylate metabolism, and fatty acid metabolism pathways. Similar results were observed for boys except for phenylalanine, tyrosine and tryptophan biosynthesis, which had a high impact [0.098]. The identified metabolites and pathways could be efficacious for investigating the underlying mechanisms of the development of different metabolic phenotypes in obese Chinese adolescents.

Serum metabolic profile analysis of syndrome of Qi-yin deficiency and Yang deficiency in diabetic coronary heart disease by UPLC-Q/TOF MS

AimsQi-yin deficiency (QYD) and Yang-deficiency (YD) syndrome are two common syndromes with contrasting clinical manifestations of low fever and cold phobia. Different syndrome phenotypes must have distinct biological underpinnings in vivo. Based on serum metabolomics, this study aims to reveal the biological characteristics of the two diabetic coronary heart disease syndromes mentioned above. MethodsA total of 33 diabetic coronary heart disease (DCHD) patients were included in the study, with 13 cases of QYD, 11 cases of YD syndrome, and 9 cases of Pinghe constitution (PH). Based on UPLC-Q/TOF MS, metabolomics was used to detect and analyze serum samples from the three groups mentioned above. To find differentiating metabolites, PLS-DA (Partial least squares discrimination analysis) and database searching were used. Meanwhile, the metabolic pathways of various metabolites were investigated. ResultsThere were 17 different metabolites between QYD syndrome and YD syndrome of DCHD. Cyclic GMP, LysoPE (20:3), proline, isoleucine, N-lactoyl-tyrosine, 3-oxohexadecanoyl-CoA, and oxalosuccinic acid were lower in the (YD) group than those in the PH group, while galactitol, N-lactoyl-tyrosine, and oxalosuccinic acid were higher in the QYD group than in the PH group. The 17 differential metabolites mentioned above were linked to a variety of pathways, including amino acid metabolism, purine metabolism, fatty acid elongation in mitochondria, the tricarboxylic acid cycle, lipid metabolism, energy metabolism, etc. ConclusionDifferent DCHD syndromes, such as liver-kidney Yin deficiency and YD syndrome, have different biological bases for the same disease. To some extent, the findings of this study provide a scientific foundation for different approaches to treating the same disease in Traditional Chinese Medicine. It is also important for guiding the clinical use of herbal medicine.

Remodelin, a N-acetyltransferase 10 (NAT10) inhibitor, alters mitochondrial lipid metabolism in cancer cells.

Remodelin is a small molecule inhibitor of N-acetyltransferase 10 (NAT10), reported to reverse the effect of cancer conditions such as epithelial to mesenchymal transition, hypoxia, and drug resistance. We analysed RNA seq data of siNAT10 and found many metabolic pathways were altered, this made us perform unbiased metabolic analysis. Here we performed untargeted metabolomics in Remodelin treated cancer cells using high-performance liquid chromatography-tandem mass spectrometry. Statistical analysis revealed a total number of 138 of which 52 metabolites were significantly modified in Remodelin treated cells. Among the most significantly altered metabolites, we identified metabolites related with mitochondrial fatty acid elongation (MFAE) and mitochondrial beta-oxidation such as lauroyl-CoA, cholesterol, triglycerides, (S)-3-hydroxyhexadecanoyl-CoA, and NAD+ . Furthermore, assessment showed alteration in expression of Enoyl-CoA hydratase, short chain 1, mitochondrial (ECHS1), and Mitochondrial trans-2-enoyl-CoA reductase (MECR) genes, associated with MFAE pathway. We also found statistically significant decrease in total cholesterol and triglycerides in Remodelin treated cancer cells. Overall, our results showed that Remodelin alters mitochondrial fatty acid metabolism and lipid accumulation in cancer cells. Finally, we validated these results in NAT10 knockdown cancer cells and found that NAT10 reduction results in alteration in gene expression associated with mitochondrial fatty acid metabolism, clearly suggesting the possible role of NAT10 in maintaining mitochondrial fatty acid metabolism.

Vanillin modulates activities linked to dysmetabolism in psoas muscle of diabetic rats

Skeletal muscles are important in glucose metabolism and are affected in type 2 diabetes (T2D) and its complications. This study investigated the effect of vanillin on redox imbalance, cholinergic and purinergic dysfunction, and glucose-lipid dysmetabolism in muscles of rats with T2D. Male albino rats (Sprague–Dawley strain) were fed 10% fructose ad libitum for 2 weeks before intraperitoneally injecting them with 40 mg/kg streptozotocin to induce T2D. Low (150 mg/kg bodyweight (BW)) and high (300 mg/kg BW) doses of vanillin were orally administered to diabetic rats. Untreated diabetic rats and normal rats made up the diabetic control (DC) and normal control (NC) groups, respectively. The standard antidiabetic drug was metformin. The rats were humanely put to sleep after 5 weeks of treatment and their psoas muscles were harvested. There was suppression in the levels of glutathione, activities of SOD, catalase, ENTPDase, 5′Nucleotidase and glycogen levels on T2D induction. This was accompanied by concomitantly elevated levels of malondialdehyde, serum creatine kinase-MB, nitric oxide, acetylcholinesterase, ATPase, amylase, lipase, glucose-6-phosphatase (G6Pase), fructose-1,6-biphophastase (FBPase) and glycogen phosphorylase activities. T2D induction further resulted in the inactivation of fatty acid biosynthesis, glycerolipid metabolism, fatty acid elongation in mitochondria and fatty acid metabolism pathways. There were close to normal and significant reversals in these activities and levels, with concomitant reactivation of the deactivated pathways following treatment with vanillin, which compared favorably with the standard drug (metformin). Vanillin also significantly increased muscle glucose uptake ex vivo. The results suggest the therapeutic effect of vanillin against muscle dysmetabolism in T2D as portrayed by its ability to mitigate redox imbalance, inflammation, cholinergic and purinergic dysfunctions, while modulating glucose-lipid metabolic switch and maintaining muscle histology.

L-leucine stimulation of glucose uptake and utilization involves modulation of glucose - lipid metabolic switch and improved bioenergetic homeostasis in isolated rat psoas muscle ex vivo.

The antidiabetic effect of l-leucine has been attributed to its modulatory effect on glucose uptake and lipid metabolism in muscles. However, there is a dearth on its effect on glucose metabolism in muscles. Thus, the present study investigated the effect of l-leucine - stimulated glucose uptake on glucose metabolism, dysregulated lipid metabolic pathways, redox and bioenergetic homeostasis, and proteolysis in isolated psoas muscle from Sprague Dawley male rats. Isolated psoas muscles were incubated with l-leucine (30-240μg/mL) in the presence of 11.1 mMol glucose at 37 ˚C for 2h. Muscles incubated in only glucose served as the control, while muscles not incubated in l-leucine and/or glucose served as the normal control. Metformin (6.04mM) was used as the standard antidiabetic drug. Incubation with l-leucine caused a significant increase in muscle glucose uptake, with an elevation of glutathione levels, superoxide dismutase, catalase, E-NTPDase and 5'nucleotidase activities. It also led to the depletion of malondialdehyde and nitric oxide levels, ATPase, chymotrypsin, acetylcholinesterase, glycogen phosphorylase, glucose-6-phosphatase, fructose-1,6-bisphosphatase and lipase activities. There was an alteration in lipid metabolites, with concomitant activation of glycerolipid metabolism, fatty acid metabolism, and fatty acid elongation in mitochondria in the glucose-incubated muscle (negative control). Incubation with l-leucine reversed these alterations, and concomitantly deactivated the pathways. These results indicate that l-leucine-enhanced muscle glucose uptake involves improved redox and bioenergetic homeostasis, with concomitant suppressed proteolytic, glycogenolytic and gluconeogenetic activities, while modulating glucose - lipid metabolic switch.

The nasal microbiome, nasal transcriptome, and pet sensitization

Petallergies are common in children with asthma. Microbiota and host responses may mediate allergen sensitization. We sought to uncover host-microbe relationships in petallergen sensitization via joint examination of the nasal microbiome and nasal transcriptome. We collected nasal samples from 132 children with asthma for parallel 16S rRNA and RNA sequencing. Specific IgE levels for cat and dog dander were measured. Analyses of the nasal microbiome, nasal transcriptome, and their correlations were performed with respect to pet sensitization status. Among the 132 children, 91 (68.9%) were cat sensitized and 96 (72.7%) were dog sensitized. Cat sensitization was associated with lower nasal microbial diversity by Shannon index (P= .021) and differential nasal bacterial composition by weighted UniFrac distance (permutational multivariate ANOVA P= .035). Corynebacterium sp and Staphylococcus epidermidis were significantly less abundant, and the metabolic process "fatty acid elongation in mitochondria" was lower in pet-sensitized versus unsensitized children. Correlation networks revealed that the nasal expression levels of 47 genes representing inflammatory processes were negatively correlated with the relative abundances of Corynebacterium sp and S epidermidis. Thus, these species were directly associated not only with the absence of pet sensitization but also with the underexpression of host gene expression of inflammatory processes that contribute to allergen sensitization. Causal mediation analyses revealed that the associations between these nasal species and pet sensitization were mediated by nasal gene expression. Higher abundances of nasal Corynebacterium sp and S epidermidis are associated with absence of pet sensitization and correlate with lower expression of inflammatory genes.

Buddleja saligna Willd (Loganiaceae) Inhibits Angiotensin‐Converting Enzyme Activity in Oxidative Cardiopathy with Concomitant Modulation of Nucleotide Hydrolyzing Enzymatic Activities and Dysregulated Lipid Metabolic Pathways

Buddleja saligna Willd (Loganiaceae), mostly indigenous to South Africa is traditionally used in the treatment cardio‐dysfunctional related ailments amongst other diseases. The cardio‐protective effect of B. saligna was investigated in ferric‐induced oxidative cardiopathy. Hearts harvested from healthy male SD rats were incubated with 0.1 mM FeSO4 to induce oxidative damage and co‐incubated with B. saligna extract. Reaction mixtures without the extract served as negative control, while tissues without the extract or standard antioxidant (gallic acid) and pro‐oxidant served as the normal control. The tissues were analyzed for levels of glutathione, malondialdehyde, and nitric oxide as well as cholinergic, angiotensin‐converting enzyme (ACE), lipase, and purinergic enzymes activities, lipid profiles, fatty acid metabolic pathways and metabolites. Induction of oxidative damage significantly (p&lt;0.05) depleted the levels of GSH, SOD, catalase, and ENTPDase activities, while concomitantly elevating the levels of MDA, NO, ACE, acetylcholinesterase, lipase and ATPase activities. These levels and activities were significantly reversed on treatment with B. saligna. Treatment with B. saligna also led to depletion of cardiac cholesterol and LDL‐c levels, while elevating triglyceride and HDL‐c level. It also depleted oxidative‐induced lipid metabolites with concomitant generation of thirteen other metabolites. B. saligna also inactivated oxidative‐induced pathways for beta oxidation of very long chain fatty acids, glycerolipid metabolism, and fatty acid elongation in mitochondria. These results suggest that B. saligna protects against ferric‐induced oxidative cardiopathy by mitigating oxidative stress, while concomitantly inhibiting ACE, acetylcholinesterase and lipase activities, and modulating lipid spectrum and dysregulated metabolic pathways.Support or Funding InformationThis work was supported by research funding from the National Research Foundation; and the Department of Science and Innovation (DSI) through the IKS‐Based Technology Innovations, Pretoria, South Africa

Buddleja saligna Willd (Loganiaceae) inhibits angiotensin-converting enzyme activity in oxidative cardiopathy with concomitant modulation of nucleotide hydrolyzing enzymatic activities and dysregulated lipid metabolic pathways.

Buddleja saligna Willd (Loganiaceae), mostly indigenous to South Africa is traditionally used in the treatment cardio-dysfunctional related ailments amongst other diseases. The cardio-protective effect of B. saligna was investigated in ferric-induced oxidative cardiopathy. Hearts harvested from healthy male SD rats were incubated with 0.1 mM FeSO4 to induce oxidative damage and co-incubated with B. saligna extract. Reaction mixtures without the extract served as negative control, while tissues without the extract or standard antioxidant (gallic acid) and pro-oxidant served as the normal control. The tissues were analyzed for levels of glutathione, malondialdehyde, and nitric oxide as well as cholinergic, angiotensin-converting enzyme (ACE), lipase, and purinergic enzymes activities, lipid profiles, fatty acid metabolic pathways and metabolites. Induction of oxidative damage significantly (p < 0.05) depleted the levels of GSH, SOD, catalase, and ENTPDase activities, while concomitantly elevating the levels of MDA, NO, ACE, acetylcholinesterase, lipase and ATPase activities. These levels and activities were significantly reversed on treatment with B. saligna. Treatment with B. saligna also led to depletion of cardiac cholesterol and LDL-c levels, while elevating triglyceride and HDL-c level. It also depleted oxidative-induced lipid metabolites with concomitant generation of thirteen other metabolites. B. saligna also inactivated oxidative-induced pathways for beta oxidation of very long chain fatty acids, glycerolipid metabolism, and fatty acid elongation in mitochondria. These results suggest that B. saligna protects against ferric-induced oxidative cardiopathy by mitigating oxidative stress, while concomitantly inhibiting ACE, acetylcholinesterase and lipase activities, and modulating lipid spectrum and dysregulated metabolic pathways.

Evaluation of circulating leukocyte transcriptome and its relationship with immune function and blood markers in dairy cows during the transition period

Dairy cows during the transition period are faced with important physiological changes which include a dysfunctional immune system and an increased inflammatory state. New data are necessary to understand the key factors involved in the immune system regulation. Six dairy cows were sampled during transition period to investigate the leukocyte transcriptome changes and its relationship with blood biomarkers. Blood samples were collected at - 20 ± 2, - 3 ± 1, 3, and 7 days from parturition (DFP). Leukocyte transcriptome was analyzed by deep sequencing technology (Hiseq1000 Illumina, USA). Plasma was analyzed for metabolic biomarkers. Differentially expressed genes (DEG) were used to run an enrichment analysis through the Dynamic Impact Approach (DIA). Considering - 20 DFP as references time, the main KEGG impacted pathways were activated before calving (- 3 DFP) and were connected to lipid metabolism, lipid transport in plasma, and phagosome. The greatest differences were found after parturition with 281 DEG (179 upregulated and 102 downregulated). The activated pathways were mainly related to immunity and endocrine aspects, while metabolic pathways related to lipid and amino acid metabolism were inhibited. Plasma BHBA had a substantial inhibitory impact on KEGG pathways related to DNA replication and cell cycle, while plasma IL-1β had an inhibitory impact on fatty acid elongation in mitochondria and an activated impact in several pathways related to cellular energy metabolism. Overall, this study confirmed that many changes in lipid metabolism and immune competence of the circulating leukocytes occurred in dairy cow around calving. Interestingly, BHBA and IL-1β connected with the transcriptome.

Metabolomics profiling in a mouse model reveals protective effect of Sancao granule on Con A-Induced liver injury

Sancao granule (SCG) is a traditional Chinese herb formula, which has been used for autoimmune liver disease for decades. Previous study demonstrated that there was an exactly therapeutic effect of SCG on autoimmune hepatitis (AIH) by improving liver function and alleviating the clinical symptoms. However, studies of the mechanism by which SCG alleviates Con A-induced liver injury (CILI) should be complemented. An ultraperformance liquid chromatography with quadrupole time-of-flight mass spectrometry (UHPLC-QTOF/MS)-based metabolomics approach combined with principle component analysis (PCA) and orthogonal projection to latent structures discriminate analysis (OPLS-DA) were integrated applied to obtain metabolites for clarifying mechanisms of disease. In accordance with previously study, the present study demonstrated that SCG could obviously improve the liver injury in mouse induced by Con A via downregulating serum biochemical indexes, alleviating the histological damage and inhibiting the neutrophil infiltration in liver tissues. Different expression of 9 metabolites related to 8 pathways, including fatty acid biosynthesis, arachidonic acid metabolisms, linoleic acid metabolisms, sphingolipid metabolisms, fatty acid elongation in mitochondria, glycerophospholipid metabolism, fatty acid metabolism, pyrimidine metabolism were demonstrated responsible for the efficacy of SCG in treating CILI. In sum up, SCG has been indicated favorable therapeutic effect on Con A induced liver injury. And metabolomics could be a promising approach, which provide insights into mechanisms of SCG in treating CILI.

Urinary metabolomics for discovering metabolic biomarkers of laryngeal cancer using UPLC-QTOF/MS

The incidence of laryngeal cancer (LYC) is second only to lung cancer, which is also the second most common cancer in head and neck cancer. Risk assessment metabolomics biomarkers to diagnose LYC have not been found by now. To profile the metabolites in healthy controls (HCs) and LYC patients (LYCs), urine metabonomics study was performed based on reversed phase liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (RPLC-QTOF/MS). Six urine differentially expressed metabolites (Variable Importance in Projection >1, p < 0.05) were identified by searching reference library or comparing with standard based on OPLS-DA (orthogonal partial least squares-discriminant analysis) model. d-pantothenic acid, palmitic acid, myristic acid, oleamide, sphinganine and phytosphingosine were identified as differential metabolites associated with the LYC and they might play roles in sphingolipid metabolism, fatty acid biosynthesis, fatty acid elongation in mitochondria, pantothenate and coenzyme A (CoA) biosynthesis, beta-Alanine metabolism and fatty acid metabolism. These six differential metabolites were combined to test the potentiality of diagnosis of LYC. Results revealed that the area under the curve (AUC) value, sensitivity and specificity of receiving operator characteristic (ROC) curve were 0.97, 95% and 97%, respectively, indicating that this diagnosis method could be used to distinguish LYCs from HCs with good sensitivity and specificity.

Characterization and expression profiles of muscle transcriptome in Schizothoracine fish, Schizothorax prenanti

Schizothorax prenanti is a cold-water fish species with great economic importance in aquaculture in Western China, and the underlying mechanisms of muscle development and growth in S. prenanti remain to be elucidated. In this study, deep RNA sequencing was performed to provide an in-depth view of the transcriptome of skeletal muscle of S. prenanti with the specific objective to identify expressed genes in the skeletal muscle of S. prenanti at 30 days post-hatching (S01), 1 year (S02), and 3 years (S03). De novo assembly of high-quality reads generated 132,784 transcripts with an average length of 1282 bp and 67,596 unigenes with an average length of 1559 bp. 2445 unigenes were differentially expressed with 1483 up-regulated and 962 down-regulated in the skeletal muscle of S. prenanti at S01 and S02 stages, and 1936 unigenes were significantly impacted at S02 and S03 stages with 1153 increased and 783 decreased. GO analysis showed that the differentially expressed genes are involved in various biological processes with dominance by cell & cell part, binding & catalytic activity, and cellular process & metabolic process. KEGG enrichment suggested that there are considerable differences in the physiological processes at different stages of muscle development and growth of S. prenanti. PPAR signaling pathway, cardiac muscle contraction, fatty acid metabolism, tight junction, and focal adhesion were the top pathways enriched in comparison between S01 and S02 stages. Whereas significant enrichment of the TCA cycle, fatty acid metabolism, fatty acid elongation in mitochondria, valine, leucine and isoleucine degradation, porphyrin and chlorophyll metabolism, and propanoate metabolism pathway was found in differentially expressed genes identified between S02 and S03 stages. This study provides not only an overall insight into the global gene expression landscape in the skeletal muscle of S. prenanti, but also candidate genes or markers that can be used for further investigations of the underlying mechanisms of skeletal muscle development and growth of S. prenanti.

Multiplatform plasma fingerprinting in cancer cachexia: a pilot observational and translational study.

BackgroundCachexia is a metabolic syndrome that affects up to 50–80% of cancer patients. The pathophysiology is characterized by a variable combination of reduced food intake and abnormal metabolism, including systemic inflammation and negative protein and energy balance. Despite its high clinical significance, defined diagnostic criteria and established therapeutic strategies are lacking. The ‘omics’ technologies provide a global view of biological systems. We hypothesize that blood‐based metabolomics might identify findings in cachectic patients that could provide clues to gain knowledge on its pathophysiology, and eventually postulate new therapeutic strategies.MethodsThis is a cross‐sectional observational study in two cohorts of cancer patients, with and without cachexia. Patients were consecutively recruited from routine clinical practice of a General Oncology Department at ‘12 de Octubre’ University Hospital. Selected clinical and biochemical features were collected. Blood metabolite fingerprinting was performed using three analytical platforms, gas chromatography coupled to mass spectrometry (GC–MS), capillary electrophoresis coupled to mass spectrometry (CE–MS), and liquid chromatography coupled to mass spectrometry (LC–MS). Besides, we performed pathway‐based metabolite analyses to obtain more information on biological functions.ResultsA total of 15 subjects were included in this study, 8 cachectic and 7 non‐cachectic patients. Metabolomic analyses were able to correctly classify their samples in 80% (GC–MS), 97% (CE–MS), 96% [LC–MS (positive mode)], and 89% [LC–MS (negative mode)] of the cases. The most prominent metabolic alteration in plasma of cachectic patients was the decrease of amino acids and derivatives [especially arginine, tryptophan, indolelactic acid, and threonine, with 0.4‐fold change (FC) compared with non‐cachectic patients], along with the reduction of glycerophospholipids [mainly lysophosphatidylcholines(O‐16:0) and lysophosphatidylcholines(20:3) sn‐1, FC = 0.1] and sphingolipids [SM(d30:0), FC = 0.5]. The metabolite with the highest increase was cortisol (FC = 1.6). Such alterations suggest a role of the following metabolic pathways in the pathophysiology of cancer cachexia: arginine and proline metabolism; alanine, aspartate, and glutamate metabolism; phenylalanine metabolism; lysine degradation; aminoacyl‐tRNA biosynthesis; fatty acid elongation in mitochondria; tricarboxylic acids cycle; among others.ConclusionsThese findings suggest that plasma amino acids and lipids profiling has great potential to find the mechanisms involved in the pathogenesis of cachexia. Metabolic profiling of plasma from cancer patients show differences between cachexia and non‐cachexia in amino acids and lipids that might be related to mechanisms involved in its pathophysiology. A better understanding of these mechanisms might identify novel therapeutic approaches to palliate this unmet medical condition.

Variants in the 3' untranslated region of the ovine acetyl-coenzyme A acyltransferase 2 gene are associated with dairy traits and exhibit differential allelic expression.

The acetyl-CoA acyltransferase 2 (ACAA2) gene encodes an enzyme of the thiolase family that is involved in mitochondrial fatty acid elongation and degradation by catalyzing the last step of the respective β-oxidation pathway. The increased energy needs for gluconeogenesis and triglyceride synthesis during lactation are met primarily by increased fatty acid oxidation. Therefore, the ACAA2 enzyme plays an important role in the supply of energy and carbon substrates for lactation and may thus affect milk production traits. This study investigated the association of the ACAA2 gene with important sheep traits and the putative functional involvement of this gene in dairy traits. A single nucleotide substitution, a T to C transition located in the 3' untranslated region of the ACAA2 gene, was used in mixed model association analysis with milk yield, milk protein yield and percentage, milk fat yield and percentage, and litter size at birth. The single nucleotide polymorphism was significantly associated with total lactation production and milk protein percentage, with respective additive effects of 6.81 ± 2.95 kg and -0.05 ± 0.02%. Additionally, a significant dominance effect of 0.46 ± 0.21 kg was detected for milk fat yield. Homozygous TT and heterozygous CT animals exhibited higher milk yield compared with homozygous CC animals, whereas the latter exhibited increased milk protein percentage. Expression analysis from age-, lactation-, and parity-matched female sheep showed that mRNA expression of the ACAA2 gene from TT animals was 2.8- and 11.8-fold higher in liver and mammary gland, respectively. In addition, by developing an allelic expression imbalance assay, it was estimated that the T allele was expressed at an average of 18% more compared with the C allele in the udder of randomly selected ewes. We demonstrated for the first time that the variants in the 3' untranslated region of the ovine ACAA2 gene are differentially expressed in homozygous ewes of each allele and exhibit allelic expression imbalance within heterozygotes in a tissue-specific manner, supporting the existence of cis-regulatory DNA variation in the ovine ACAA2 gene. This is the first study reporting differential allelic imbalance expression of a candidate gene associated with milk production traits in dairy sheep.

\u03b2-Thalassemia Patients Revealed a Significant Change of Untargeted Metabolites in Comparison to Healthy Individuals

β-Thalassemia is one of the most prevalent forms of congenital blood disorders characterized by reduced hemoglobin levels with severe complications, affecting all dimensions of life. The mechanisms underlying the phenotypic heterogeneity of β-thalassemia are still poorly understood. We aimed to work over metabolite biomarkers to improve mechanistic understanding of phenotypic heterogeneity and hence better management of disorder at different levels. Untargeted serum metabolites were analyzed after protein precipitation and SPE (solid phase extraction) from 100 β-thalassemia patients and 61 healthy controls using GC-MS. 40 metabolites were identified having a significance difference between these two groups at probability of 0.05 and fold change >1.5. Out of these 40 metabolites, 17 were up-regulated while 23 were down-regulated. PCA and PLS-DA model was also created that revealed a fine separation with a sensitivity of 70% and specificity of 100% on external validation of samples. Metabolic pathway analysis revealed alteration in multiple pathways including glycolysis, pyruvate, propanoate, glycerophospholipid, galactose, fatty acid, starch and sucrose metabolism along with fatty acid elongation in mitochondria, glycerolipid, glyoxylate and dicarboxylate metabolism pointing towards the shift of metabolism in β-thalassemia patients in comparison to healthy individuals.

Lipidomics Biomarkers of Diet-Induced Hyperlipidemia and Its Treatment with Poria cocos.

Hyperlipidemia is a major cause of atherosclerotic cardiovascular disease. Poria cocos (PC) is a medicinal product widely used in Asia. This study was undertaken to define the alterations of lipid metabolites in rats fed a high-fat diet to induce hyperlipidemia and to explore efficacy and mechanism of action of PC in the treatment of diet-induced hyperlipidemia. Plasma samples were then analyzed using UPLC-HDMS. The untreated rats fed a high-fat diet exhibited significant elevation of plasma triglyceride and total and low-density lipoprotein (LDL) cholesterol concentrations. This was associated with marked changes in plasma concentrations of seven fatty acids (palmitic acid, hexadecenoic acid, hexanoylcarnitine, tetracosahexaenoic acid, cervonoyl ethanolamide, 3-hydroxytetradecanoic acid, and 5,6-DHET) and five sterols [cholesterol ester (18:2), cholesterol, hydroxytestosterone, 19-hydroxydeoxycorticosterone, and cholic acid]. These changes represented disorders of biosynthesis and metabolism of the primary bile acids, steroids, and fatty acids and mitochondrial fatty acid elongation pathways in diet-induced hyperlipidemia. Treatment with PC resulted in significant improvements of hyperlipidemia and the associated abnormalities of the lipid metabolites.

Transcriptome-Wide Analysis Reveals the Role of PPARγ Controlling the Lipid Metabolism in Goat Mammary Epithelial Cells.

To explore the large-scale effect of peroxisome proliferator-activated receptor γ (PPARG) in goat mammary epithelial cells (GMEC), an oligonucleotide microarray platform was used for transcriptome profiling in cells overexpressing PPARG and incubated with or without rosiglitazone (ROSI, a PPARγ agonist). A total of 1143 differentially expressed genes (DEG) due to treatment were detected. The Dynamic Impact Approach (DIA) analysis uncovered the most impacted and induced pathways “fatty acid elongation in mitochondria,” “glycosaminoglycan biosynthesis-keratan sulfate,” and “pentose phosphate pathway.” The data highlights the central role of PPARG in milk fatty acid metabolism via controlling fatty acid elongation, biosynthesis of unsaturated fatty acid, lipid formation, and lipid secretion; furthermore, its role related to carbohydrate metabolism promotes the production of intermediates required for milk fat synthesis. Analysis of upstream regulators indicated that PPARG participates in multiple physiological processes via controlling or cross talking with other key transcription factors such as PPARD and NR1H3 (also known as liver-X-receptor-α). This transcriptome-wide analysis represents the first attempt to better understand the biological relevance of PPARG expression in ruminant mammary cells. Overall, the data underscored the importance of PPARG in mammary lipid metabolism and transcription factor control.

Acute mammary and liver transcriptome responses after an intramammary Escherichia coli lipopolysaccharide challenge in postpartal dairy cows.

The study investigated the effect of an intramammary lipopolysaccharide (LPS) challenge on the bovine mammary and liver transcriptome and its consequences on metabolic biomarkers and liver tissue composition. At 7 days of lactation, 7 cows served as controls (CTR) and 7 cows (LPS) received an intramammary Escherichia coli LPS challenge. The mammary and liver tissues for transcriptomic profiling were biopsied at 2.5 h from challenge. Liver composition was evaluated at 2.5 h and 7 days after challenge, and blood biomarkers were analyzed at 2, 3, 7 and 14 days from challenge. In mammary tissue, the LPS challenge resulted in 189 differentially expressed genes (DEG), with 20 down-regulated and 169 up-regulated. In liver tissue, there were 107 DEG in LPS compared with CTR with 42 down-regulated and 65 up-regulated. In mammary, bioinformatics analysis highlighted that LPS led to activation of NOD-like receptor signaling, Toll-like receptor signaling, RIG-I-like receptor signaling and apoptosis pathways. In liver, LPS resulted in an overall inhibition of fatty acid elongation in mitochondria and activation of the p53 signaling pathway. The LPS challenge induced changes in liver lipid composition, a systemic inflammation (rise of blood ceruloplasmin and bilirubin), and an increase in body fat mobilization. The data suggest that cells within the inflamed mammary gland respond by activating mechanisms of pathogen recognition. However, in the liver the response likely depends on mediators originating from the udder that affect liver functionality and specifically fatty acid metabolism (β-oxidation, ketogenesis, and lipoprotein synthesis).

Insilico analysis of hypothetical proteins unveils putative metabolic pathways and essential genes in Leishmania donovani.

Leishmaniasis is a parasitic disease caused by the protozoan Leishmania, which is active in two broad forms namely, Visceral Leishmaniasis (VL or Kala Azar) and Cutaneous Leishmaniasis (CL). The disease is most prevalent in the tropical regions and poses a threat to over 70 countries across the globe. About 200 million people are estimated to be at risk of developing VL in the Indian subcontinent, and this refers to around 67% of the global VL disease burden. The Indian state of Bihar alone accounts for 50% of the total VL cases. While no vaccination exists, several pentavalent antimonials and drugs like Paromomycin, Amphotericin, Miltefosine etc. are used in the treatment of Leishmaniasis. However, due to their low efficacies and the resistance developed by the bug to these medications, there is an urgent need to look into newer species specific targets. The proteome information available suggests that among the 7960 proteins in Leishmania donavani, a staggering 65% remains classified as a hypothetical uncharacterized set. In this background, we have attempted to assign probable functions to these hypothetical sequences present in this parasite, to explore their plausible roles as druggable receptors. Thus, putative functions have been defined to 105 hypothetical proteins, which exhibited a GO term correlation and PFAM domain coverage of more than 50% over the query sequence length. Of these, 27 sequences were found to be associated with a reference pathway in KEGG as well. Further, using homology approaches, four pathways viz., Ubiquinone biosynthesis, Fatty acid elongation in Mitochondria, Fatty Acid Elongation in ER and Seleno-cysteine Metabolism have been reconstructed. In addition, 7 new putative essential genes have been mined with the help of Eukaryotic Database of Essential Genes (DEG). All these information related to pathways and essential genes indeed show promise for exploiting the select molecules as potential therapeutic targets.

Functional Enrichment Analysis by David in Transgenic MYCN Zebrafish Model

AbstractAbstract 5114 IntroductionThe MYCN oncogene encodes a basic helix-loop-helix/leucine zipper (bHLH/LZ) transcription factor that is frequently overexpressed in hematologic malignancies neoplasms (including acute leukemia, T-cell lymphoma, and so on). MYCN acts as a poor prognostic marker to promote an aggressive phenotype. However, the mechanisms of action and pathways affected by MYCN are still largely unclear. MethodsWe induced murine MYCN gene overexpression in embryonic zebrafish through heat-shock promoter and established stable germline Tg(MYCN:HSE:EGFP) zebrafish. RNA was extracted at 3 days post fertilization from wild type (WT) and transgenic zebrafish F1 generation (TG) embryo hematopoietic cells, collected by the flow cytometer, for microarray analysis. The samples were processed and subsequently analyzed in triplicate on Zebrafish Oligo Microarrays (Agilent Technologies), containing 43, 554 sets of probe, at the Advanced Throughput Inc. The microarrays were scanned in an Agilent DNA Microarray Scanner and the images were processed using Feature Extraction software. A False Discovery Rate≤0. 05 for overall interactions effect and P<0. 001 between comparisons were used to determine differentially expressed genes (DEG). Ingenuity Pathway Analysis and DAVID performed the functional analysis of DEG. ResultsMicroarray analysis revealed 626 (342 genes up-regulated and 284 genes down-regulated) DEG that showed >2-fold change in TG comparing with that of WT. Using functional enrichment analysis by DAVID, several signaling pathways were regulated in TG samples (Table 1). MAPK signaling pathway was high activated through FGF, PDGF, BDNF and CACN high expression, promoting up-regulated of Ras and MKP, enhancing phosphorylation and leading to increase of cells proliferation. TGFβ signaling was inhibited by up-regulation of IFN Ã and Smad 6/7, which negative control of TGFβR and Smad 2/3. Further, we found that MYCN enhances the expression of skp2, via decreased p21 and increased CDK2, promoting cell cycle progression (Fig. 1). In addition, overexpression of MYCN weakened the function of mismatch repair, base excision repair, while increased apoptosis pathway mediated by p53 (up-regulated Bid gene). Meanwhile, Glycolysis/gluconeogenesis pathway was significantly up-regulated in TG fish. ConclusionsOverexpression of MYCN induced up-regulation of cell proliferation and Glycolysis/gluconeogenesis pathway (as the Warburg effect in rapidly proliferating tumors), attenuation of repair function, all of which are phenomena associated with proliferation and malignancies transformation of blood cell feature. [Display omitted] We found that MYCN down-regulates p27kip1, p57kip2 and p21cip1 through up-regulate Skp2, thus up-regulates CDK2, CycA, CycB, CycD and CycE. All above changes shortened the time taken to progress through the cell cycle. Increased MARK signaling and decreased TGFβ signaling pathways also contributed to promote cell cycle. (Red star marks the up-regulated genes).Table 1Changes in Tg(MYCN:HSE:EGFP) zebrafish hematopoietic cellssignaling pathwayregulateCount*%#P-ValueGlycolysis/Gluconeogenesis+201.20.00000037Cell cycle+423.30.00000045Tryptophan metabolism+140.80.0000086Fatty acid metabolism+130.80.0000094Cardiac muscle contraction+201.20.000038Biosynthesis of unsaturated fatty acids+90.50.000057Phenylalanine metabolism+90.50.00014Tyrosine metabolism+110.70.0004Pyruvate metabolism+110.70.001p53 signaling pathway+1310.0021Fatty acid elongation in mitochondria+60.40.0022Arginine and proline metabolism+130.80.0038Valine, leucine and isoleucine degradation+100.60.0069Alanine, aspartate and glutamate metabolism+90.50.0069Butanoate metabolism+80.50.008Mismatch repair−141.15.9E-10Homologous recombination−120.90.00000097Base excision repair−1310.0000019Spliceosome−231.80.000033Nucleotide excision repair−1310.000038Pyrimidine metabolism−191.50.000056TGFβ−81.60.00063One carbon pool by folate−60.50.0021+: up-regulated; -: down-regulated.*: genes involved in the term.#: involved genes/total genes. P-Value: the threshold of EASE Score, a modified Fisher Exact P-Value, for gene-enrichment analysis (≤0. 01). Disclosures:No relevant conflicts of interest to declare.