Search

AGPAT2 (1-acyl-sn-glycerol-3-phosphate-acyltransferase-2) converts lysophosphatidic acid (LPA) into phosphatidic acid (PA), and mutations of the AGPAT2 gene cause the most common form of congenital generalized lipodystrophy which leads to steatohepatitis. The underlying mechanism by which AGPAT2 deficiency leads to lipodystrophy and steatohepatitis has not been elucidated. We addressed this question using an antisense oligonucleotide (ASO) to knockdown expression of Agpat2 in the liver and white adipose tissue (WAT) of adult male Sprague-Dawley rats. Agpat2 ASO treatment induced lipodystrophy and inflammation in WAT and the liver, which was associated with increased LPA content in both tissues, whereas PA content was unchanged. We found that a controlled-release mitochondrial protonophore (CRMP) prevented LPA accumulation and inflammation in WAT whereas an ASO against glycerol-3-phosphate acyltransferase, mitochondrial ( Gpam ) prevented LPA content and inflammation in the liver in Agpat2 ASO-treated rats. In addition, we show that overnutrition, due to high sucrose feeding, resulted in increased hepatic LPA content and increased activated macrophage content which were both abrogated with Gpam ASO treatment. Taken together, these data identify LPA as a key mediator of liver and WAT inflammation and lipodystrophy due to AGPAT2 deficiency as well as liver inflammation due to overnutrition and identify LPA as a potential therapeutic target to ameliorate these conditions., Competing Interests: Competing interests statement:S.M. is an employee of Ionis Pharmaceuticals. G.I.S. consults for and has received research support from Ionis Pharmaceuticals; is a stockholder of Orsobio, which owns the Yale intellectual property for CRMP; is an inventor of the Yale patent for CRMP.

Several members of the aldehyde dehydrogenase (ALDH) family, especially ALDH1 isoenzymes, have been identified as biomarkers of cancer stem cells (CSCs), a small subpopulation of oncogenic cells with self-renewal and multipotency capability. Consistent with this contention, cell populations with high ALDH enzymatic activity exhibit greater carcinogenic potential. It has been reported that ALDH1, especially ALDH1A1, serves as a valuable biomarker for colon CSCs. However, the functional roles of ALDHs in CSCs and solid tumors of the colon tissue is not fully understood. The aim of the present study was to identify molecular signature associated with high ALDH activity in human colorectal adenocarcinoma (COLO320DM) cells by proteomics profiling. Aldefluor™ assay was performed to sort COLO320DM cells exhibiting high (ALDH high ) and low (ALDH low ) ALDH activity. Label-free quantitative proteomics analyses were conducted on these two cell populations. Proteomics profiling revealed a total of 229 differentially expressed proteins (DEPs) in ALDH high relative to ALDH low cells, of which 182 were down-regulated and 47 were up-regulated. In agreement with previous studies, ALDH1A1 appeared to be the principal ALDH isozyme contributing to the Aldefluor™ assay activity in COLO320DM cells. Ingenuity pathway analysis of the proteomic datasets indicated that DEPs were associated with mitochondrial dysfunction, sirtuin signaling, oxidative phosphorylation and nucleotide excision repair. Our proteomics study predicts that high ALDH1A1 activity may be involved in these cellular pathways to promote a metabolic switch and cellular survival of CSCs., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Alcohol consumption is a global healthcare problem with enormous social, economic, and clinical consequences. The liver sustains the earliest and the greatest degree of tissue injury due to chronic alcohol consumption and it has been estimated that alcoholic liver disease (ALD) accounts for almost 50% of all deaths from cirrhosis in the world. In this study, we used a modified Lieber-DeCarli (LD) diet to treat mice with alcohol and simulate chronic alcohol drinking. Using an untargeted metabolomics approach, our aim was to identify the various metabolites and pathways that are altered in the early stages of ALD. Histopathology showed minimal changes in the liver after 6 weeks of alcohol consumption. However, untargeted metabolomics analyses identified 304 metabolic features that were either up- or down-regulated in the livers of ethanol-consuming mice. Pathway analysis revealed significant alcohol-induced alterations, the most significant of which was in the FXR/RXR activation pathway. Targeted metabolomics focusing on bile acid biosynthesis showed elevated taurine-conjugated cholic acid compounds in ethanol-consuming mice. In summary, we showed that the changes in the liver metabolome manifest very early in the development of ALD, and when minimal changes in liver histopathology have occurred. Although alterations in biochemical pathways indicate a complex pathology in the very early stages of alcohol consumption, bile acid changes may serve as biomarkers of the early onset of ALD., (Copyright © 2022. Published by Elsevier B.V.)

Alcoholic fatty liver disease (AFLD) is characterized by lipid accumulation and inflammation and can progress to cirrhosis and cancer in the liver. AFLD diagnosis currently relies on histological analysis of liver biopsies. Early detection permits interventions that would prevent progression to cirrhosis or later stages of the disease. Herein, we have conducted the first comprehensive time-course study of lipids using novel state-of-the art lipidomics methods in plasma and liver in the early stages of a mouse model of AFLD, i.e., Lieber-DeCarli diet model. In ethanol-treated mice, changes in liver tissue included up-regulation of triglycerides (TGs) and oxidized TGs and down-regulation of phosphatidylcholine, lysophosphatidylcholine, and 20-22-carbon-containing lipid-mediator precursors. An increase in oxidized TGs preceded histological signs of early AFLD, i.e., steatosis, with these changes observed in both the liver and plasma. The major lipid classes dysregulated by ethanol play important roles in hepatic inflammation, steatosis, and oxidative damage. Conclusion: Alcohol consumption alters the liver lipidome before overt histological markers of early AFLD. This introduces the exciting possibility that specific lipids may serve as earlier biomarkers of AFLD than those currently being used., (© 2021 The Authors. Hepatology Communications published by Wiley Periodicals, Inc., on behalf of the American Association for the Study of Liver Diseases.)

Purpose: The purpose of this study was to elucidate the role and molecular consequences of impaired glutathione (GSH) biosynthesis on eye development., Methods: GSH biosynthesis was impaired in surface ectoderm-derived ocular tissues by crossing Gclc f/f mice with hemizygous Le-Cre transgenic mice to produce Gclc f/f /Le-Cre Tg/- (KO) mice. Control mice included Gclc f/f and Gclc wt/wt /Le-Cre Tg/- mice (CRE). Eyes from all mice (at various stages of eye development) were subjected to histological, immunohistochemical, Western blot, RT-qPCR, RNA-seq, and subsequent Gene Ontology, Ingenuity Pathway Analysis and TRANSFAC analyses. PAX6 transactivation activity was studied using a luciferase reporter assay in HEK293T cells depleted of GSH using buthionine sulfoximine (BSO)., Results: Deletion of Gclc diminished GSH levels, increased reactive oxygen species (ROS), and caused an overt microphthalmia phenotype characterized by malformation of the cornea, iris, lens, and retina that is distinct from and much more profound than the one observed in CRE mice. In addition, only the lenses of KO mice displayed reduced crystallin (α, β), PITX3 and Foxe3 expression. RNA-seq analyses at postnatal day 1 revealed 1552 differentially expressed genes (DEGs) in the lenses of KO mice relative to those from Gclc f/f mice, with Crystallin and lens fiber cell identity genes being downregulated while lens epithelial cell identity and immune response genes were upregulated. Bioinformatic analysis of the DEGs implicated PAX6 as a key upstream regulator. PAX6 transactivation activity was impaired in BSO-treated HEK293T cells., Conclusions: These data suggest that impaired ocular GSH biosynthesis may disrupt eye development and PAX6 function., (Copyright © 2021 Elsevier Inc. All rights reserved.)

1,4-Dioxane (1,4-DX) is an environmental contaminant found in drinking water throughout the United States. Although it is a suspected liver carcinogen, there is no federal or state maximum contaminant level for 1,4-DX in drinking water. Very little is known about the mechanisms by which this chemical elicits liver carcinogenicity. In the present study, female BDF-1 mice were exposed to 1,4-DX (0, 50, 500, and 5,000mg/L) in their drinking water for 1 or 4 weeks, to explore the toxic effects. Histopathological studies and a multi-omics approach (transcriptomics and metabolomics) were performed to investigate potential mechanisms of toxicity. Immunohistochemical analysis of the liver revealed increased H2AXγ-positive hepatocytes (a marker of DNA double-strand breaks), and an expansion of precholangiocytes (reflecting both DNA damage and repair mechanisms) after exposure. Liver transcriptomics revealed 1,4-DX-induced perturbations in signaling pathways predicted to impact the oxidative stress response, detoxification, and DNA damage. Liver, kidney, feces, and urine metabolomic profiling revealed no effect of 1,4-DX exposure, and bile acid quantification in liver and feces similarly showed no effect of exposure. We speculate that the results may be reflective of DNA damage being counterbalanced by the repair response, with the net result being a null overall effect on the systemic biochemistry of the exposed mice. Our results show a novel approach for the investigation of environmental chemicals that do not elicit cell death but have activated the repair systems in response to 1,4-DX exposure., (© The Author(s) 2021. Published by Oxford University Press on behalf of the Society of Toxicology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

The etiology of colorectal cancer (CRC) is complex. Approximately, 10% of individuals with CRC have predisposing germline mutations that lead to familial cancer syndromes, whereas most CRC patients have sporadic cancer resulting from a combination of environmental and genetic risk factors. It has become increasingly clear that chronic alcohol consumption is associated with the development of sporadic CRC; however, the exact mechanisms by which alcohol contributes to colorectal carcinogenesis are largely unknown. Several proposed mechanisms from studies in CRC models suggest that alcohol metabolites and/or enzymes associated with alcohol metabolism alter cellular redox balance, cause DNA damage, and epigenetic dysregulation. In addition, alcohol metabolites can cause a dysbiotic colorectal microbiome and intestinal permeability, resulting in bacterial translocation, inflammation, and immunosuppression. All of these effects can increase the risk of developing CRC. This review aims to outline some of the most significant and recent findings on the mechanisms of alcohol in colorectal carcinogenesis. We examine the effect of alcohol on the generation of reactive oxygen species, the development of genotoxic stress, modulation of one-carbon metabolism, disruption of the microbiome, and immunosuppression.

Background: Colorectal cancer (CRC) is a leading cause of cancer mortality worldwide. Mutations in the adenomatous polyposis coli (APC) gene are pivotal in colorectal tumorigenesis. Recently, we demonstrated that aldehyde dehydrogenase 1B1 (ALDH1B1) knockdown dramatically reduced colon tumor growth in a mouse xenograft model. The purpose of the present preliminary study is to examine the effect of loss of ALDH1B1 in CRC development in an inducible colon-specific Apc mouse model., Methods: Apc W/F Cdx2 ERT2-Cre mice develop uni-allelic inactivation of Apc specifically in colon epithelial cells following tamoxifen treatment. Aldh1b1 -/- KO mice were crossed with Apc W/F Cdx2 ERT2-Cre mice. Six-month-old male Apc W/F Cdx2 ERT2-Cre /Aldh1b1 -/- , and Apc W/F Cdx2 ERT2-Cre /Aldh1b1 +/+ mice were treated with tamoxifen (50 mg/kg, i.p.) for three consecutive days. Apc W/F /Aldh1b1 -/- and Apc W/F /Aldh1b1 +/+ mice were treated with corn oil (i.e., tamoxifen vehicle control) for three consecutive days. Eighteen days later, mice were sacrificed and their colons examined microscopically, macroscopically and histologically for the presence of adenoma., Results: All Apc W/F Cdx2 ERT2-Cre /Aldh1b1 +/+ and Apc W/F Cdx2 ERT2-Cre /Aldh1b1 -/- mice treated with tamoxifen developed colorectal adenoma. The Apc W/F Cdx2 ERT2-Cre /Aldh1b1 -/- mice showed a significant decrease in the total volume of all ileal and colonic adenomas, and decreased incidence of large colonic adenoma compared to Apc W/F Cdx2 ERT2-Cre /Aldh1b1 +/+ mice. Immunohistochemical analysis of p53 and β-catenin showed a trend toward decreased expression score in colonic adenomas of Apc W/F Cdx2 ERT2-Cre /Aldh1b1 -/- mice., Conclusion: The present preliminary study suggests that deletion of ALDH1B1 may protect against the full development of colorectal cancer. Further mechanistic studies are required to elucidate how ALDH1B1 contributes for colorectal cancer., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Processing of fish in aquaculture generates considerable amounts of by-products that remain underused and/or unexploited. We evaluated the nutritive content of fish by-products (head, gills, intestines, trimmings, bones, and skin) from meagre and gilthead sea bream fish species reared in Greece in order to estimate their nutritional value for future development of high added-value products. The proximate composition of the fish samples (total protein, total lipid, ash, moisture, and macro-element content) was determined using the Association of Official Analytical Chemists (AOAC) and International Organization for Standardization (ISO) official methods. The content of fatty acids was determined using capillary gas chromatography, and the protein profile was estimated employing scientific orbitrap mass spectrophotometer methodology. The nutrient composition of fish by-products presented fluctuations among the different by-products. Skin was the most significant protein source, trimmings and bones were high in calcium, and the head, intestines, and bones were a good source of lipids. The most abundant lipid acids found in by-products were oleic, palmitic, linoleic, and eicosenoic acids, whereas the most abundant proteins were adenosine triphosphate (ATP) synthase subunit epsilon, mitochondrial nicotinamide adenine dinucleotide (NADH) dehydrogenase, and mitochondrial cytochrome b-c1 complex subunit 8. These data suggest that by-products constitute valuable sources of nutrients and could therefore be exploited in accordance with the principles of a circular economy.

Excessive consumption of alcohol is a leading cause of lifestyle-induced morbidity and mortality worldwide. Although long-term alcohol abuse has been shown to be detrimental to the liver, brain and many other organs, our understanding of the exact molecular mechanisms by which this occurs is still limited. In tissues, ethanol is metabolized to acetaldehyde (mainly by alcohol dehydrogenase and cytochrome p450 2E1) and subsequently to acetic acid by aldehyde dehydrogenases. Intracellular generation of free radicals and depletion of the antioxidant glutathione (GSH) are believed to be key steps involved in the cellular pathogenic events caused by ethanol. With continued excessive alcohol consumption, further tissue damage can result from the production of cellular protein and DNA adducts caused by accumulating ethanol-derived aldehydes. Much of our understanding about the pathophysiological consequences of ethanol metabolism comes from genetically-engineered mouse models of ethanol-induced tissue injury. In this review, we provide an update on the current understanding of important mouse models in which ethanol-metabolizing and GSH-synthesizing enzymes have been manipulated to investigate alcohol-induced disease.

Diagnostic markers are needed for accidental or deliberate radiation exposure that could cause acute and chronic radiation toxicity. Biomarkers of temporal, dose-dependent, aging-attenuated and multiple radiation exposures have been previously described by others. However, the physiological origin and biochemical networks that generate these biomarkers and their association at the molecular level have yet to be explored. Hence, the discovery and identification of total-body-irradiation-induced tissue specific biomarkers remains an enormous challenge within radiation biodosimetry research. To determine the tissue level response of total-body exposure (6 Gy), metabolomics analysis was carried out on radiosensitive tissues bone marrow, ileum, liver, muscle and lung as well as serum and on urine within 12 h postirradiation. Differences in the metabolic signatures between the sham and gamma-irradiated groups were analyzed by hydrophilic interaction liquid chromatography (HILIC)-based ultra-performance liquid chromatography-electrospray ionization-quadrupole time-of-flight mass spectrometry (UPLC-ESI-QTOFMS). A panel of 67 biomarkers identified in radiosensitive tissues and biofluids (serum and urine) at a 6 Gy dose. Among the identified biomarkers, 3-methylglutarylcarnitine (3-MGC) was found to be a novel metabolite in liver, serum and urine that could potentially be an early radiation response marker. The degree of metabolic changes among different tissues showed perturbations in pathways including DNA methylation, energy, nucleic acid, amino acid, glutathione and bile acid metabolism. These results highlight metabolomics as a potential novel approach to understand functional alterations in the metabolome that could be adapted for use in the rapid assessment of radiation exposure and triage protocols in the case of nuclear incidents.

Despite widespread use as well as epidemiologic indications, there have been no investigations into the effect of St. John's wort (SJW) extract on colorectal carcinogenesis in vivo. This study reports a systematic evaluation of the impact of dietary supplementation of SJW extract on azoxymethane-induced colorectal carcinogenesis in mice. Mice were fed with either AIN-93G (control) diet or SJW extract-supplemented diet (SJW diet) prior to azoxymethane treatment. SJW diet was found to significantly improve the overall survival of azoxymethane-treated mice. Pretreatment with the SJW diet significantly reduced body weight loss as well as decrease of serum albumin and cholesterol levels associated with azoxymethane-induced colorectal tumorigenesis. SJW diet-fed mice showed a significant decrease in tumor multiplicity along with a decrease in incidence of large tumors and a trend toward decreased total tumor volume in a dose-dependent manner. A short-term study, which examined the effect of SJW prior to rectal bleeding, also showed decrease in colorectal polyps in SJW diet-fed mice. Nuclear factor kappa B (NF-κB) and extracellular signal-regulated kinase (ERK1/2) pathways were attenuated by SJW administration. SJW extract resulted in early and continuous attenuation of these pathways in the colon epithelium of SJW diet-fed mice under both short-term and long-term treatment regimens. In conclusion, this study demonstrated the chemopreventive potential of SJW extract against colorectal cancer through attenuation of proinflammatory processes., (©2015 American Association for Cancer Research.)

During mammalian development, some methylated cytosines (5mC) in CG dinucleotides are iteratively oxidized by TET dioxygenases to 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC), and 5-carboxylcytosine (5caC). The effect of these cytosine oxidative products on the sequence-specific DNA binding of transcription factors is being actively investigated. Here, we used the electrophoretic mobility shift assay (EMSA) to examine C/EBPα and C/EBPβ homodimers binding to all 25 chemical forms of a CG dinucleotide for two DNA sequences: the canonical C/EBP 8-mer TTGC|GCAA and the chimeric C/EBP|CRE 8-mer TTGC|GTCA. 5hmC in the CG dinucleotide in the C/EBP|CRE motif 8-mer TGAC|GCAA inhibits binding of C/EBPβ but not C/EBPα. Binding was increased by 5mC, 5fC and 5caC. Circular dichroism monitored thermal denaturations for C/EBPβ bound to the C/EBP|CRE motif confirmed the EMSA. The structural differences between C/EBPα and C/EBPβ that may account for the difference in binding 5hmC in the 8-mer TGAC|GCAA are explored., (Published by Elsevier B.V.)

Three oxidative products of 5-methylcytosine (5mC) occur in mammalian genomes. We evaluated if these cytosine modifications in a CG dinucleotide altered DNA binding of four B-HLH homodimers and three heterodimers to the E-Box motif CGCAG|GTG. We examined 25 DNA probes containing all combinations of cytosine in a CG dinucleotide and none changed binding except for carboxylation of cytosine (5caC) in the strand CGCAG|GTG. 5caC enhanced binding of all examined B-HLH homodimers and heterodimers, particularly the Tcf3|Ascl1 heterodimer which increased binding ~10-fold. These results highlight a potential function of the oxidative products of 5mC, changing the DNA binding of sequence-specific transcription factors., (Published by Elsevier Inc.)

Background: JAG1 is an evolutionarily conserved ligand for Notch receptor and functions in the cell fate decisions, cell-cell interactions throughout the development of heart especially right heart development. Tetralogy of Fallot (TOF) is essentially a right sided heart disease with characteristic features of ventricular septal defect, right ventricular outflow tract obstruction, aortic dextroposition and right ventricular hypertrophy. Hence, the present study was investigated to identify mutations of JAG1 gene in an Indian cohort of patients with TOF., Methods: The clinical data and blood samples from 84 unrelated subjects with TOF were collected and evaluated in comparison with 87 healthy individuals. PCR based single strand conformation polymorphism analysis and subsequent bidirectional DNA sequencing of conformers was carried in the exon 6 of JAG1 gene., Results: The DNA sequences aligned with NCBI-BLAST led to the identification of four novel variations including one nonsense 765 C>A, two missense 814 G>T, 834 G>T; and one silent alteration 816 G>T in TOF patients. The protein structure of JAG1 predicts that these variations effect first and second epidermal growth factor like repeat and might disturb ligand-receptor binding ability. The presence of similar variations was not observed in healthy controls. The software CLUSTAL-W showed the inter species conservation of altered amino acids in missense mutations., Conclusion: Disease-associating novel JAG1 gene variations were found in TOF patients, and seem to play an important role in the causation of the disease., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Background: Researchers have determined that Indians face a higher risk of heart disease, despite the fact that nearly half of them are vegetarians and lack many of the other traditional risk factors. In the below-30 age group, coronary artery disease mortality among Indians is three-fold higher than in the whites in United Kingdom and ten-fold higher than the Chinese in Singapore. High levels of homocysteine have been widely linked to the early onset of heart diseases in other populations, although a definite proof among Indians is lacking, which needs to be investigated by way of screening for factors responsible for high homocysteine levels., Objective: To screen for genetic factors responsible for hyperhomocysteinemia and the risk for premature coronary artery disease., Materials and Methods: A total of 100 individuals with proven premature coronary artery disease and 200 age-and-sex matched controls were screened for polymorphisms in Methylenetetrahydrofolate reductase (MTHFR) (C677T) Methionine synthase (MS) genes (A2756G, C2758G), and the B12 and Folate levels were estimated., Results: Results from the mutational analysis revealed that in the study group, seven individuals had a polymorphism for the C677T allele in the MTHFR gene (one homozygous and six heterozygous) (Fischer's Exact test P > 0.046) (OR: 0.2711 95% CI 0.0774 to 0.9491). Six were heterozygous for the A2756G polymorphism in the MS gene (Fischer's Exact test P > 0.0012). None showed a polymorphism at the C2758G allele in the MS gene. Four controls showed heterozygosity for the C677T polymorphism and none for the MS gene. The B12 and Folate levels were significantly lower in the study group as compared to the controls., Conclusions: It is important to know which factors determine the total homocysteine concentrations. In the general population, the most important modifiable determinants of tHcy are folate intake and coffee consumption. Smoking and alcohol consumption are also associated with the total homocysteine concentrations, but more research is necessary to elucidate whether these relations are not originating from residual confounding due to other lifestyle factors.