Phosphatidylethanolamine N-methyltransferase Research Articles

Abstract 6379: Comparison of lipid profiles in children with solid tumors receiving chemotherapy and healthy children

Abstract Background: Lipid metabolism plays an important role in the regulation of synaptic function and plasticity.Cancer chemotherapy (chemo) can disturb the lipid profile, potentially linked with neuropsychological toxicities. This study aimed to compare the lipid profiles between children with cancer receiving chemo and healthy children. Methods: A prospective, observational study was conducted in 21 children with solid tumors enrolled from Children’s Healthcare of Atlanta (CHOA) and 16 controls. Children with solid tumors were consented precycle 2 chemo (T1) and followed at the end of chemo (T2). Controls completed one timepoint of data collection. At T1 (n=21) and T2 (n=13), fecal samples were collected for lipidomics analysis. Fecal lipids were processed using a high-resolution, untargeted lipidomics LC-MS approach. MetaboAnalyst and linear mixed models were used to analyze the data with false discovery rate correction (FDR 0.2). Results: There were no significant differences in age, sex, race, and body mass index between the groups. In total 241 lipid features were identified with high confidence and analyzed. A comparison between children with solid tumors at T2 (n=12) vs. controls (n=16) detected 8 differential lipids, which can be clustered into 4 classes, including 4 triglycerides (TG), 2 phosphatidylcholine (PC), 1 sphingomyelin (SM), and 1 lysophosphatidylcholine (LPC). Additionally, 24 differential lipids were detected for children with solid tumors from T1 to T2 (n=12), including 6 classes: 9 SM, 7 PC, 3 diacylglycerols (DG), 2 TG, 2 monoacylglycerols (MG), and 1 phosphatidylethanolamine (PE). Compared with controls, children with solid tumors at T2 had higher levels of SM (e.g., SM(d36:1)) and lower levels of TG (e.g., TG(18:0/18:0/20:0)), LPC (e.g., LPC(20:4)), and PC (e.g., PC(33:4). Compared with children at T1, those at T2 had higher levels of SM (e.g., SM(d36:1)), PE (e.g., PE(16:0p/20:4), PC (e.g., PC(30:0), and PC(36:1)), and lower levels of TG (e.g., TG(18:0/18:0/20:0) and TG(18:0/16:0/18:0)), LPC (e.g., LPC(20:4)), DG (e.g., DG(18:0/16:0) and DG(18:0/18:0)), and MG (e.g., MG(18:0)). Conclusion: Children with cancer chemo had alterations in fecal lipid profiles impacting the metabolism of PC, SM, TG, PE, MG, and DG. The increased PC and the dysregulation of LPC in comparison with controls suggest the decreased activity of phospholipase A2 (PLA2), primarily reported in inflammatory disease. Altered activity of lysophospholipid acyltransferase (LPCAT), diacylglyceride acyltransferase (DGAT), and phosphatidylethanolamine methyl transferase (PEMT) could also explain changes in PC and glycerolipids. Combined, these suggest dysregulation of the Kennedy or Lands pathways towards PC synthesis. Future work is needed to study whether the disturbed lipid metabolisms were associated with treatment toxicities and outcomes such as cognitive dysfunction based on literature. Citation Format: Jinbing Bai, Madelyn Houser, Melissa Martin, Christie Powell, Kristal Maner-Smith, Kathryn S. Sutton, Thomas Olson, Deborah W. Bruner. Comparison of lipid profiles in children with solid tumors receiving chemotherapy and healthy children [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6379.

Association between PEMT rs7946 and blood pressure levels in Chinese adolescents.

This study was to explore blood pressure levels in Chinese adolescents with different genotypes of phosphatidylethanolamine N-methyltransferase (PEMT) gene (PEMT) rs7946, as well as effects of dietary intake on blood pressure levels with different genders and different genotypes of PEMT rs7946. PEMT rs7946 genotypes were identified by PCR-restriction fragment length polymorphism and verified by DNA sequencing. Blood pressure was measured using a standard mercury sphygmomanometer. Dietary intakes were analyzed based on a 3-day diet diary, and dietary components were calculated using computer software. A total of 721 high school students (314 males and 407 females) at the age of 16.86 ± 0.59 years were included. The A allele carriers of PEMT rs7946 had increased levels of SBP, DBP, mean arterial pressure (MAP) and pulse pressure (PP) than the GG homozygotes in the female subjects. There were significant interactions between PEMT rs7946 and gender on SBP and MAP levels, regardless of whether an unadjusted or adjusted model was used. When dietary intake was taken into account, fat intake was positively associated with SBP and PP in the male GG homozygotes, while protein intake was positively associated with PP in the female A allele carriers of PEMT rs7946. This study suggests that PEMT rs7946 is significantly associated with blood pressure levels in human being. There might be interactions among PEMT rs7946, gender, and dietary intake on blood pressure levels in the adolescent population.

Molecular phenotypes behind patient stratification based on TP53 and HPV status and potential for treatment personalization with first single-cell RNA sequencing atlas of penile cancer.

8 Background: TP53 mutations and Human papillomavirus (HPV) have been suggested to impact penile squamous cell carcinoma (pSCC) prognosis and may predict treatment outcomes. Insights in differences in tumor micro-environment at single cell resolution may benefit personalized therapeutic development and are currently unavailable. Methods: We performed single cell RNA sequencing and targeted next generation sequencing (for TP53 loss-of-function mutations; p53LOF) on primary HPV-associated and HPV-independent (HPV+/-) pSCC lesions or healthy control samples of the coronal sulcus skin. Cell clusters were annotated based on known marker genes. Gene Set Enrichment Analysis (GSEA) and slingshot trajectory analysis was performed using R/Seurat package. Results: Twenty-two samples were analyzed: 4 tumors harbored p53LOF mutations (1 HPV+ p53LOF, 3 HPV- p53LOF), 6 HPV- p53WT, 6 HPV+ p53WT and 6 healthy controls. A total of 84.394 cells was annotated and we identified 68 distinct cell subtypes in pSCC. In epithelial cells, two distinct pseudotime trajectories stemming from the basal cells were identified characterized by either signals of partial epithelial-mesenchymal transition (pEMT; TGFB, ZEB2, SNAI2) or cornification ( ELF3, KRT19, PPL). Whilst the pEMT trajectory was enriched in p53LOF samples, cornification was enriched in HPV+ p53WT. HPV- p53WT samples displayed increase in metabolic changes and stemness hallmarks ( SOX2, HEY1, ZEB1). GSEA confirmed these findings and showed upregulation of pEMT gene sets in p53LOF; fatty acid metabolism and xenobiotic metabolism in HPV- p53WT, and cornification in HPV+ p53WT. We observed an immune suppressive environment in p53LOF patients with higher regulatory T-cell abundancy, and this correlated with abundancy in the epithelial pEMT clusters. Pseudo-bulk analysis on all T-cell subsets demonstrated a higher expression of inhibitory immune checkpoints ( PD-1, CTLA4, TIM3 and LAG3) in p53LOF and HPV- p53WT while HPV+ p53WT patients displayed higher abundancy of immunomodulatory and cytotoxic natural killer (NK) cells. Conclusions: We established the first comprehensive pSCC transcriptomic atlas at single cell resolution and illustrate differences in molecular phenotypes of pSCC according to TP53 mutation- and HPV status. p53LOF tumors are characterized by pEMT, which is a known driver of invasiveness, therapy resistance and metastasis. HPV- p53WT display hallmarks of stemness, which is related to and may explain chemotherapy-resistance often observed in these patients. p53LOF and HPV- p53WT display immune-evasion and exhaustion, while HPV+ p53WT are characterized by abundant cytotoxic innate-like lymphocytes. These observations provide deeper insights into cancer biology that will help advancing pSCC diagnosis and therapy.

A partial epithelial-mesenchymal transition signature for highly aggressive colorectal cancer cells that survive under nutrient restriction.

Partial epithelial-mesenchymal transition (p-EMT) has recently been identified as a hybrid state consisting of cells with both epithelial and mesenchymal characteristics and is associated with the migration, metastasis, and chemoresistance of cancer cells. Here, we describe the induction of p-EMT in starved colorectal cancer (CRC) cells and identify a p-EMT gene signature that can predict prognosis. Functional characterisation of starvation-induced p-EMT in HCT116, DLD1, and HT29 cells showed changes in proliferation, morphology, and drug sensitivity, supported by in vivo studies using the chorioallantoic membrane model. An EMT-specific quantitative polymerase chain reaction (qPCR) array was used to screen for deregulated genes, leading to the establishment of an in silico gene signature that was correlated with poor disease-free survival in CRC patients along with the CRC consensus molecular subtype CMS4. Among the significantly deregulated p-EMT genes, a triple-gene signature consisting of SERPINE1, SOX10, and epidermal growth factor receptor (EGFR) was identified. Starvation-induced p-EMT was characterised by increased migratory potential and chemoresistance, as well as E-cadherin processing and internalisation. Both gene signature and E-cadherin alterations could be reversed by the proteasomal inhibitor MG132. Spatially resolving EGFR expression with high-resolution immunofluorescence imaging identified a proliferation stop in starved CRC cells caused by EGFR internalisation. In conclusion, we have gained insight into a previously undiscovered EMT mechanism that may become relevant when tumour cells are under nutrient stress, as seen in early stages of metastasis. Targeting this process of tumour cell dissemination might help to prevent EMT and overcome drug resistance. © 2024 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

The Mediating Effect of the Choline-to-Betaine Ratio on the Association Between PEMT rs7946 and Digestive System Cancer: A Nested Case–Control Study in a Chinese Population

BackgroundThe enzyme phosphatidylethanolamine N-methyltransferase (PEMT) is responsible for synthesizing phosphatidylcholine by methylating phosphatidylethanolamine. We hypothesized that a polymorphism of the PEMT gene, rs7946, is involved in carcinogenesis. ObjectivesWe aimed to investigate the relationship between PEMT rs7946 and digestive system cancer and examine possible effect modifiers and mediators. MethodsWe conducted a nested, case–control study within the China H-type Hypertension Registry Study, including 751 cases and 1:1 matched controls. To assess the association of PEMT rs7946 and digestive system cancer, we estimated odds ratios with 95% confidence intervals (CIs) using conditional logistic regression. We used the bootstrap test to examine the potential mediating effects of related metabolites. ResultsOur results revealed that wild-type homozygous CC genotype carriers of PEMT rs7946 had a significantly increased risk [odds ratio (OR): 1.31; 95% CI: 1.04, 1.66; P = 0.023] compared with the TT/CT combined genotypes. The effect was found to be more pronounced in individuals with a lower choline-to-betaine ratio (<0.412, P-interaction = 0.021). Furthermore, the mediation analysis indicated that the choline-to-betaine ratio played a significant role in mediating 13.55% of the association between PEMT rs7946 and digestive system cancer (P = 0.018). ConclusionsOur study suggested that PEMT rs7946 may affect risk of digestive system cancer through direct and indirect pathways, and the choline-to-betaine ratio may partially mediate the indirect effect.This trial was registered at Chinese Clinical Trial Registry as ChiCTR1800017274.

Sex-Specific Perturbation of Systemic Lipidomic Profile in Newborn Lambs Impacted by Prenatal Testosterone Excess.

Gestational hyperandrogenism adversely impacts offspring health. Using an ovine model, we found that prenatal testosterone (T) excess adversely affects growth and cardiometabolic outcomes in female offspring and produces sex-specific effects on fetal myocardium. Since lipids are essential to cardiometabolic function, we hypothesized that prenatal T excess leads to sex-specific disruptions in lipid metabolism at birth. Shotgun lipidomics was performed on the plasma samples collected 48 hours after birth from female (F) and male (M) lambs of control (C) and (T) sheep (CF = 4, TF = 7, CM = 5, TM = 10) and data were analyzed by univariate analysis, multivariate dimensionality reduction modeling followed by functional enrichment, and pathway analyses. Biosynthesis of phosphatidylserine was the major pathway responsible for sex differences in controls. Unsupervised and supervised models showed separation between C and T in both sexes with glycerophospholipids and glycerolipids classes being responsible for the sex differences between C and T. T excess increased cholesterol in females while decreasing phosphatidylcholine levels in male lambs. Specifically, T excess: 1) suppressed the phosphatidylethanolamine N-methyltransferase (PEMT) phosphatidylcholine synthesis pathway overall and in TM lambs as opposed to suppression of carnitine levels overall and TF lambs; and 2) activated biosynthesis of ether-linked (O-)phosphatidylethanolamine and O-phosphatidylcholine from O-diacylglycerol overall and in TF lambs. Higher cholesterol levels could underlie adverse cardiometabolic outcomes in TF lambs, whereas suppressed PEMT pathway in TM lambs could lead to endoplasmic reticulum stress and defective lipid transport. These novel findings point to sex-specific effects of prenatal T excess on lipid metabolism in newborn lambs, a precocial ovine model of translational relevance.

Multi‐omics approach reveals disturbances in brain phosphatidylcholine metabolism in Alzheimer’s Disease

AbstractBackgroundLipid dysregulation is implicated in Alzheimer’s disease (AD) pathology, given its involvement in amyloid β processing, myelination, and cell membrane structure. Studies have demonstrated increased phosphatidylcholine (PC) hydrolysis in the brains of AD individuals, but the connections between AD and PC metabolism are not fully understood.MethodTo investigate the alterations in PC metabolism in AD, we analyzed publicly available transcriptomic, metabolomic, and lipidomic data from the ROSMAP study, which included dorsolateral prefrontal cortex (DLPFC) tissue and serum samples. DLPFC and serum samples were classified based on clinical consensus diagnosis of cognitive status at the time of death and the last valid clinical diagnosis, respectively. Concentration differences were determined with a linear model using the Limma R package.ResultAccording to the lipidomic results, the concentration of total PC‐containing glycerophospholipid was significantly decreased in AD DLPFC samples (P &lt; 0.05). Specifically, diacyl‐PC and acyl‐alkyl‐PC, the two most abundant subclasses, were significantly decreased in the brains of AD patients (P &lt; 0.05). The concentration of the PC breakdown product, glycerophosphocholine (GPC), was increased in AD patient brains (P‐value &lt; 0.05). However, GPC phosphodiesterase 1 (GPCPD1), the enzyme that further hydrolyzes GPC to free choline, was downregulated and choline abundance was reduced in AD DLPFC samples (P &lt; 0.05). The expression of phosphatidylethanolamine N‐methyltransferase (PEMT), which converts PE to PC, was reduced as was the abundance of PEMT products (PC 18:2/20:4 and PC 18:0/20:4, P &lt; 0.05). These alterations in the brain were not observed in the serum.ConclusionThese results indicate that PC metabolism is disturbed in the DLPFC of AD patients, with evidence of increased PC breakdown and reduced GPC hydrolysis as well as reduced conversion of PE to PC in AD brains. However, these changes in the brain are not reflected in serum. Further studies are needed to better understand changes in phospholipid metabolism in the brains of AD patients.

The Role of Phosphatidylethanolamine N-Methyltransferase (PEMT) and Its Waist-Hip-Ratio-Associated Locus rs4646404 in Obesity-Related Metabolic Traits and Liver Disease

In previous genome-wide association studies (GWAS), genetic loci associated with obesity and impaired fat distribution (FD) have been identified. In the present study, we elucidated the role of the PEMT gene, including the waist–hip-ratio-associated single nucleotide polymorphism rs4646404, and its influence on obesity-related metabolic traits. DNA from 2926 metabolically well-characterized subjects was used for genotyping. PEMT expression was analyzed in paired visceral (vis) and subcutaneous (sc) adipose tissue (AT) from a subset of 574 individuals. Additionally, PEMT expression was examined in vis, sc AT and liver tissue in a separate cohort of 64 patients with morbid obesity and liver disease. An in vitro Pemt knockdown was conducted in murine epididymal and inguinal adipocytes. Our findings highlight tissue-specific variations in PEMT mRNA expression across the three studied tissues. Specifically, vis PEMT mRNA levels correlated significantly with T2D and were implicated in the progression of non-alcoholic steatohepatitis (NASH), in contrast to liver tissue, where no significant associations were found. Moreover, sc PEMT expression showed significant correlations with several anthropometric- and metabolic-related parameters. The rs4646404 was associated with vis AT PEMT expression and also with diabetes-related traits. Our in vitro experiments supported the influence of PEMT on adipogenesis, emphasizing its role in AT biology. In summary, our data suggest that PEMT plays a role in regulating FD and has implications in metabolic diseases.

Role of circadian clock system in the mitochondrial trans-sulfuration pathway and tissue remodeling.

Previous studies from our laboratory revealed that the gaseous molecule hydrogen sulfide (H2S), a metabolic product of epigenetics, involves trans-sulfuration pathway for ensuring metabolism and clearance of homocysteine (Hcy) from body, thereby mitigating the skeletal muscle's pathological remodeling. Although the master circadian clock regulator that is known as brain and muscle aryl hydrocarbon receptor nuclear translocator like protein 1 (i.e., BMAL 1) is associated with S-adenosylhomocysteine hydrolase (SAHH) and Hcy metabolism but how trans-sulfuration pathway is influenced by the circadian clock remains unexplored. We hypothesize that alterations in the functioning of circadian clock during sleep and wake cycle affect skeletal muscle's biology. To test this hypothesis, we measured serum matrix metalloproteinase (MMP) activities using gelatin gels for analyzing the MMP-2 and MMP-9. Further, employing casein gels, we also studied MMP-13 that is known to be influenced by the growth arrest and DNA damage-45 (GADD45) protein during sleep and wake cycle. The wild type and cystathionine β synthase-deficient (CBS-/+) mice strains were treated with H2S and subjected to measurement of trans-sulfuration factors from skeletal muscle tissues. The results suggested highly robust activation of MMPs in the wake mice versus sleep mice, which appears somewhat akin to the "1-carbon metabolic dysregulation", which takes place during remodeling of extracellular matrix during muscular dystrophy. Interestingly, the levels of trans-sulfuration factors such as CBS, cystathionine γ lyase (CSE), methyl tetrahydrofolate reductase (MTHFR), phosphatidylethanolamine N-methyltransferase (PEMT), and Hcy-protein bound paraoxonase 1 (PON1) were attenuated in CBS-/+ mice. However, treatment with H2S mitigated the attenuation of the trans-sulfuration pathway. In addition, levels of mitochondrial peroxisome proliferator-activated receptor-gamma coactivator 1-α (PGC 1-α) and mitofusin-2 (MFN-2) were significantly improved by H2S intervention. Our findings suggest participation of the circadian clock in trans-sulfuration pathway that affects skeletal muscle remodeling and mitochondrial regeneration.

ITRAQ-based proteomics identifies proteins associated with betaine accumulation in Lycium barbarum L

In order to better understand the mechanism of betaine accumulation in Lycium barbarum L. (LBL), we used iTRAQ (Isotope relative and absolute quantitative labeling) proteomics to screen and identify differentially abundant proteins (DAPs) at five stages (S1-young fruit stage, S2-green fruit stage, S3-early yellowing stage, S4-late yellowing stage, S5-ripening stage). A total of 1799 DAPs and 171 betaine-related DAPs were identified, and phosphatidylethanolamine N-methyltransferase (NMT), choline monooxygenase (CMO), and betaine aldehyde dehydrogenase (BADH) were found to be the key enzymes related to betaine metabolism. These proteins are mainly involved in carbohydrates, amino acids and their derivatives, fatty acids, carboxylic acids, photosynthesis and photoprotection, isoquinoline alkaloid biosynthesis, peroxisomes, and glycine, serine, and threonine metabolism. Three of the key enzymes were also up- and down-regulated to different degrees at the mRNA level. The study provide new insights into the of mechanism of betaine accumulation in LBL. SignificanceBetaine, a class of naturally occurring, water-soluble alkaloids, has been found to be widespread in animals, higher plants, and microbes. In addition to being an osmotic agent, betaine has biological functions such as hepatoprotection, neuroprotection, and antioxidant activity. Betaine metabolism (synthesis and catabolism) is complexly regulated by developmental and environmental signals throughout the life cycle of plant fruit maturation. As a betaine-accumulating plant, little has been reported about the regulatory mechanisms of betaine metabolism during the growth and development of Lycium barbarum L. (LBL) fruit. Therefore, this study used iTRAQ quantitative proteomics technology to investigate the abundance changes of betaine-related proteins in LBL fruit, screen and analyze the differential abundance proteins related to betaine metabolism, and provide theoretical references for the in-depth study of the mechanism of betaine metabolism in LBL fruit.

FRI103 Developmental Programming: Sex-specific Differences In Lipid Metabolism Of Newborn Lambs Exposed To Prenatal Testosterone Excess

Abstract Disclosure: N. Saadat: None. J.N. Ciarelli: None. B. Pallas: None. V. Padmanabhan: None. A.K. Vyas: None. Gestational hyperandrogenism such as that seen in Polycystic Ovary Syndrome (PCOS), leads to adverse offspring health outcomes in both humans and animals. Prenatal T excess (T) in a sheep model of PCOS phenotype induces peripheral insulin resistance, dyslipidemia, adipocyte defects, liver steatosis, pathological cardiac remodeling and hypertension in the female offspring. Whether these adverse cardiometabolic outcomes evidenced in the female offspring also extends to male offspring needs to be addressed. We hypothesized that prenatal T excess will induce sex-specific disruptions in lipid metabolism of newborn lambs thereby contributing to the aberrant cardiometabolic phenotype. Shotgun Lipidomics was performed on the plasma samples collected at 48 hours after birth from prenatal T treated (100mg T-propionate in corn oil from days 30 to 90 of gestation intramuscularly; Term: 147days) and control (C, vehicle only) lambs (N=26, Female C=4, T=7 and Male C=5, T=10). The data was analyzed by multivariate dimensionality reduction modeling followed by functional enrichment and pathway analyses. Unsupervised 3D principal component analysis and supervised models showed separation between C and T groupings in female and male lambs. Most lipids responsible for this separation were from glycerophospholipids and glycerolipids classes in both sexes. Prenatal T excess led to an increase in cholesterol levels only in female lambs as opposed to a decrease in levels of carnitine species (CAR) CAR 16:1 and CAR 14:1 and phosphatidylcholine species (PC) PC 39:4, PC 39:5 and PC 38:5 in male lambs. To investigate the impact of prenatal T excess on lipid metabolism pathways and active reactions, 738 lipid species were mapped, of which 147 were found to be active in reactions. The active reactions involving conversion of ether linked diglycerides (DG) to ether linked phosphatidylethanolamines (PE, O-DG→O-PE, z-score 3.367, p &amp;lt;0.05) and ether linked PC (O-DG →O-PC, z-score 2.299, p &amp;lt; 0.05) were increased in female lambs. In contrast, the conversion of PE to PC to phosphatidic acids (PA) was suppressed (PE→PC→PA, z-score 2.133, p &amp;lt;0.05) in prenatal T-treated male lambs pointing towards the suppression of Phosphatidylethanolamine N-methyltransferase (PEMT) phosphatidylcholine synthesis pathway. Higher cholesterol levels in T female lambs could contribute toward the adverse cardiometabolic phenotype evidenced earlier in this model, whereas suppressed PEMT pathway could lead to endoplasmic reticulum stress, defects in membrane permeability and lipid transport that may lead to liver steatosis in T male lambs. These novel findings of sex-specific effects of gestational exposure to T excess on lipid metabolism of newborn lamb, a precocial model of translational relevance, may have sex-specific functional outcomes in cardiometabolic systems. Presentation: Friday, June 16, 2023

Hepatic levels of S-adenosylmethionine regulate the adaptive response to fasting

There has been an intense focus to uncover the molecular mechanisms by which fasting triggers the adaptive cellular responses in the major organs of the body. Here, we show that in mice, hepatic S-adenosylmethionine (SAMe)-the principal methyl donor-acts as a metabolic sensor of nutrition to fine-tune the catabolic-fasting response by modulating phosphatidylethanolamine N-methyltransferase (PEMT) activity, endoplasmic reticulum-mitochondria contacts, β-oxidation, and ATP production in the liver, together with FGF21-mediated lipolysis and thermogenesis in adipose tissues. Notably, we showthat glucagon induces the expression of the hepatic SAMe-synthesizing enzyme methionine adenosyltransferase α1 (MAT1A), which translocates to mitochondria-associated membranes. This leads to the production of this metabolite at these sites, which acts as a brake to prevent excessive β-oxidation and mitochondrial ATP synthesis and thereby endoplasmic reticulum stress and liver injury. This work provides important insights into the previously undescribed function of SAMe as a new arm of the metabolic adaptation to fasting.

Mass Spectrometry Imaging Reveals Early Metabolic Priming of Cell Lineage in Differentiating Human-Induced Pluripotent Stem Cells.

Induced pluripotent stem cells (iPSCs) hold great promise in regenerative medicine; however, few algorithms of quality control at the earliest stages of differentiation have been established. Despite lipids having known functions in cell signaling, their role in pluripotency maintenance and lineage specification is underexplored. We investigated the changes in iPSC lipid profiles during the initial loss of pluripotency over the course of spontaneous differentiation using the co-registration of confocal microscopy and matrix-assisted laser desorption/ionization (MALDI) mass spectrometry imaging. We identified phosphatidylethanolamine (PE) and phosphatidylinositol (PI) species that are highly informative of the temporal stage of differentiation and can reveal iPS cell lineage bifurcation occurring metabolically. Several PI species emerged from the machine learning analysis of MS data as the early metabolic markers of pluripotency loss, preceding changes in the pluripotency transcription factor Oct4. The manipulation of phospholipids via PI 3-kinase inhibition during differentiation manifested in the spatial reorganization of the iPS cell colony and elevated expression of NCAM-1. In addition, the continuous inhibition of phosphatidylethanolamine N-methyltransferase during differentiation resulted in the enhanced maintenance of pluripotency. Our machine learning analysis highlights the predictive power of lipidomic metrics for evaluating the early lineage specification in the initial stages of spontaneous iPSC differentiation.

Hepatic oleate regulates one-carbon metabolism during high carbohydrate feeding

Obesity is a major risk factor for type 2 diabetes, coronary heart disease, and strok. These diseases are associated with profound alterations in gene expression in metabolic tissues. Epigenetic-mediated regulation of gene expression is one mechanism through which environmental factors, such as diet, modify gene expression and disease predisposition. However, epigenetic control of gene expression in obesity and insulin resistance is not fully characterized. We discovered that liver-specific stearoyl-CoA desaturase-1 (Scd1) knockout mice (LKO) fed a high-carbohydrate low-fat diet exhibit dramatic changes in hepatic gene expression and metabolites of the folate cycle and one-carbon metabolism respectively for the synthesis of S-adenosylmethionine (SAM). LKO mice show an increased ratio of S-adenosylmethionine to S-adenosylhomocysteine, a marker for increased cellular methylation capacity. Furthermore, expression of DNA and histone methyltransferase genes is up-regulated while the mRNA and protein levels of the non-DNA methyltransferases including phosphatidylethanolamine methyltransferase (PEMT), Betaine homocysteine methyltransferase (Bhmt), and the SAM-utilizing enzymes such as glycine-N-methyltransferase (Gnmt) and guanidinoacetate methyltransferase (Gamt) are generally down-regulated. Feeding LKO mice a high carbohydrate diet supplemented with triolein, but not tristearin, and increased endogenous hepatic synthesis of oleate but not palmitoleate in Scd1 global knockout mice normalized one carbon gene expression and metabolite levels. Additionally, changes in one carbon gene expression are independent of the PGC-1α-mediated ER stress response previously reported in the LKO mice. Together, these results highlight the important role of oleate in maintaining one-carbon cycle homeostasis and point to observed changes in one-carbon metabolism as a novel mediator of the Scd1 deficiency-induced liver phenotype.

Hepatocyte-derived DPP4 regulates portal GLP-1 bioactivity, glucose production, and its absence alters liver disease progression.

Elevated circulating dipeptidyl-peptidase 4 is a biomarker for liver disease, but its involvement in gluconeogenesis and in metabolic-associated fatty liver disease (MAFLD) progression remains unclear. Here we identified that DPP4 in hepatocytes but not Tie2+ endothelial cells regulates the local bioactivity of incretin hormones and gluconeogenesis. However, the complete absence of DPP4 (Dpp4-/-) in aged mice with metabolic syndrome accelerates liver fibrosis without altering dyslipidemia and steatosis. Analysis of transcripts from the livers of whole body Dpp4-/- displayed enrichment for inflammasome, p53, and senescence programs compared to littermate controls. High-fat high-cholesterol (HFHC)-feeding decreased Dpp4 expression in F4/80+ cells, with only minor changes in immune signaling. Moreover, in a lean mouse model of severe non-alcoholic fatty liver disease (NAFLD), phosphatidylethanolamine N-methyltransferase (Pemt -/-) mice fed with HFHC diet, we observed a 4-fold increase in circulating DPP4, disassociating its release from obesity. Lastly, we evaluated DPP4 levels in patients with hepatitis C infection with dysglycemia (HOMA-IR > 2) who underwent direct antiviral treatment (with or without ribavirin). DPP4 protein levels decreased with viral clearance, and DPP4 activity levels were reduced at longer-term follow-up in ribavirin-treated patients, although metabolic factors did not improve. These data suggest elevations in DPP4 during HCV infection are not primarily regulated by metabolic disturbances.

Dietary choline intake is necessary to prevent systems-wide organ pathology and reduce Alzheimer's disease hallmarks.

There is an urgent need to identify modifiable environmental risk factors that reduce the incidence of Alzheimer's disease (AD). The B-like vitamin choline plays key roles in body- and brain-related functions. Choline produced endogenously by the phosphatidylethanolamine N-methyltransferase protein in the liver is not sufficient for adequate physiological functions, necessitating daily dietary intake. ~90% of Americans do not reach the recommended daily intake of dietary choline. Thus, it's imperative to determine whether dietary choline deficiency increases disease outcomes. Here, we placed 3xTg-AD, a model of AD, and non-transgenic (NonTg) control mice on either a standard laboratory diet with sufficient choline (ChN; 2.0g/kg choline bitartrate) or a choline-deficient diet (Ch-; 0.0g/kg choline bitartrate) from 3 to 12 (early to late adulthood) months of age. A Ch- diet reduced blood plasma choline levels, increased weight, and impaired both motor function and glucose metabolism in NonTg mice, with 3xTg-AD mice showing greater deficits. Tissue analyses showed cardiac and liver pathology, elevated soluble and insoluble Amyloid-β and Thioflavin S structures, and tau hyperphosphorylation at various pathological epitopes in the hippocampus and cortex of 3xTg-AD Ch- mice. To gain mechanistic insight, we performed unbiased proteomics of hippocampal and blood plasma samples. Dietary choline deficiency altered hippocampal networks associated with microtubule function and postsynaptic membrane regulation. In plasma, dietary choline deficiency altered protein networks associated with insulin metabolism, mitochondrial function, inflammation, and fructose metabolic processing. Our data highlight that dietary choline intake is necessary to prevent systems-wide organ pathology and reduce hallmark AD pathologies.

Phosphatidylethanolamine N-methyltransferase: from Functions to Diseases.

Locating on endoplasmic reticulum and mitochondria associated membrane, Phosphatidylethanolamine N-methyltransferase (PEMT), catalyzes phosphatidylethanolamine methylation to phosphatidylcholine. As the only endogenous pathway for choline biosynthesis in mammals, the dysregulation of PEMT can lead to imbalance of phospholipid metabolism. Dysregulation of phospholipid metabolism in the liver or heart can lead to deposition of toxic lipid species that adversely result in dysfunction of hepatocyte/cardiomyocyte. Studies have shown that PEMT-/- mice increased susceptibility of diet-induced fatty liver and steatohepatitis. However, knockout of PEMT protects against diet-induced atherosclerosis, diet-induced obesity, and insulin resistance. Thus, novel insights to the function of PEMT in various organs should be summarized. Here, we reviewed the structural and functional properties of PEMT, highlighting its role in the pathogenesis of obesity, liver diseases, cardiovascular diseases, and other conditions.

ATP8B1 Deficiency Results in Elevated Mitochondrial Phosphatidylethanolamine Levels and Increased Mitochondrial Oxidative Phosphorylation in Human Hepatoma Cells.

ATP8B1 is a phospholipid flippase that is deficient in patients with progressive familial intrahepatic cholestasis type 1 (PFIC1). PFIC1 patients suffer from severe liver disease but also present with dyslipidemia, including low plasma cholesterol, of yet unknown etiology. Here we show that ATP8B1 knockdown in HepG2 cells leads to a strong increase in the mitochondrial oxidative phosphorylation (OXPHOS) without a change in glycolysis. The enhanced OXPHOS coincides with elevated low-density lipoprotein receptor protein and increased mitochondrial fragmentation and phosphatidylethanolamine levels. Furthermore, expression of phosphatidylethanolamine N-methyltransferase, an enzyme that catalyzes the conversion of mitochondrial-derived phosphatidylethanolamine to phosphatidylcholine, was reduced in ATP8B1 knockdown cells. We conclude that ATP8B1 deficiency results in elevated mitochondrial PE levels that stimulate mitochondrial OXPHOS. The increased OXPHOS leads to elevated LDLR levels, which provides a possible explanation for the reduced plasma cholesterol levels in PFIC1 disease.

Effects of acute intravenous lipopolysaccharide administration on the plasma lipidome and metabolome in lactating Holstein cows experiencing hyperlipidemia.

The effects of lipopolysaccharides (i.e., endotoxin; LPS) on metabolism are poorly defined in lactating dairy cattle experiencing hyperlipidemia. Our objective was to explore the effects of acute intravenous LPS administration on metabolism in late-lactation Holstein cows experiencing hyperlipidemia induced by intravenous triglyceride infusion and feed restriction. Ten non-pregnant lactating Holstein cows (273 ± 35 d in milk) were administered a single bolus of saline (3 mL of saline; n [Formula: see text] 5) or LPS (0.375 [Formula: see text]g of LPS/kg of body weight; n [Formula: see text] 5). Simultaneously, cows were intravenously infused a triglyceride emulsion and feed restricted for 16 h to induce hyperlipidemia in an attempt to model the periparturient period. Blood was sampled at routine intervals. Changes in circulating total fatty acid concentrations and inflammatory parameters were measured. Plasma samples were analyzed using untargeted lipidomics and metabolomics. Endotoxin increased circulating serum amyloid A, LPS-binding protein, and cortisol concentrations. Endotoxin administration decreased plasma lysophosphatidylcholine (LPC) concentrations and increased select plasma ceramide concentrations. These outcomes suggest modulation of the immune response and insulin action. Lipopolysaccharide decreased the ratio of phosphatidylcholine to phosphatidylethanomanine, which potentially indicate a decrease in the hepatic activation of phosphatidylethanolamine N-methyltransferase and triglyceride export. Endotoxin administration also increased plasma concentrations of pyruvic and lactic acids, and decreased plasma citric acid concentrations, which implicate the upregulation of glycolysis and downregulation of the citric acid cycle (i.e., the Warburg effect), potentially in leukocytes. Acute intravenous LPS administration decreased circulating LPC concentrations, modified ceramide and glycerophospholipid concentrations, and influenced intermediary metabolism in dairy cows experiencing hyperlipidemia.

Genetic Variants in One-Carbon Metabolism and Their Effects on DHA Biomarkers in Pregnant Women: A Post-Hoc Analysis.

The delivery of docosahexanoic acid (DHA) to the fetus is dependent on maternal one-carbon metabolism, as the latter supports the hepatic synthesis and export of a DHA-enriched phosphatidylcholine molecule via the phosphatidylethanolamine N-methyltransferase (PEMT) pathway. The following is a post-hoc analysis of a choline intervention study that sought to investigate whether common variants in one-carbon metabolizing genes associate with maternal and/or fetal blood biomarkers of DHA status. Pregnant women entering their second trimester were randomized to consume, until delivery, either 25 (n = 15) or 550 (n = 15) mg choline/d, and the effects of genetic variants in the PEMT, BHMT, MTHFD1, and MTHFR genes on DHA status were examined. Variant (vs. non-variant) maternal PEMT rs4646343 genotypes tended to have lower maternal RBC DHA (% total fatty acids) throughout gestation (6.9% vs. 7.4%; main effect, p = 0.08) and lower cord RBC DHA at delivery (7.6% vs. 8.4%; main effect, p = 0.09). Conversely, variant (vs. non-variant) maternal MTHFD1 rs2235226 genotypes exhibited higher cord RBC DHA (8.3% vs. 7.3%; main effect, p = 0.0003) and higher cord plasma DHA (55 vs. 41 μg/mL; main effect, p = 0.05). Genotype tended to interact with maternal choline intake (p < 0.1) to influence newborn DHA status for PEMT rs4646343 and PEMT rs7946. These data support the need to consider variants in one-carbon metabolic genes in studies assessing DHA status and requirements during pregnancy.