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Scholarly Works (11 results)

  • Thesis
  • Peer Reviewed
UC Davis Electronic Theses and Dissertations (2022)

Background/Objective: Nonalcoholic fatty liver disease affects 30% of the world adult population. It comprises a spectrum of liver histopathological presentation that varies from hepatocellular lipid accumulation, or nonalcoholic fatty liver (NAFL), to nonalcoholic steatohepatitis (NASH). Mechanism(s) of NAFLD onset and progression are modulated by a multilayered interaction between genetics, environmental factors, and comorbidities. In the US., the Hispanic population presents with higher rate of NAFLD and more advanced histological scores compared to other ethnicities. However, the underlying metabolic drivers of these observations are not clear. Previous metabolomic studies revealed that NAFLD is associated with several metabolic dysregulations in lipid, carbohydrate, and amino acid metabolism. Also, alterations in polyunsaturated fatty acids (PUFA) and downstream oxidized lipids as oxylipins (OXLs) and endocannabinoids (eCBs) are reported to distinguish different stages of NAFLD. However, if ethnicity-related metabolomic differences exist in NAFLD is not clear. The identification of ethnicity-related metabolomic differences will expand current knowledge with on mechanism(s) of NAFLD onset and progression. It can also identify unshared mechanism(s) that may explain the observed ethnicity-related disparity in NAFLD rate and severity. The objective of this “proof of principle” analyses is to explore metabolomic profiles of a group of Hispanic (HIS) and Caucasian (CAU) subjects with NAFLD and undergoing bariatric surgery Methods: We compared metabolomic profiles in a group of obesity NAFL-HIS and NAFL-CAU subjects to ethnicity-matched lean and healthy control subjects. We also compared the profiles of subjects with NASH to NASH-free (0-NASH) subjects. For this we profiled plasma and liver samples using untargeted, semi-quantitative metabolomic approaches for metabolites related to primary metabolism using gas chromatography/time-of-flight mass spectrometry (GC‐TOF MS); complex lipids using charged surface hybrid liquid chromatography/quadrupole time of flight mass spectrometry (CSH‐QTOF MS/MS); choline and related metabolites using hydrophilic interaction liquid chromatography/quadrupole time of flight mass spectrometer (HILIC-QTOF MS/MS). We also performed a targeted and quantitative lipidomic analysis to profile for plasma fatty acids and related downstream lipid mediators using ultra-high-performance liquid chromatography-electrospray ionization-tandem mass spectrometry (UPLC-ESI-MS/MS). Results: Findings from current analyses reveal ethnicity-related metabolic perturbation, independent of obesity. Specifically, lean, and healthy HIS subjects showed higher plasma profiles of triglycerides, acylcarnitines, free fatty acids and downstream lipid mediators. Our finding also shows that in NAFL there are ethnicity-related differences in plasma profiles of fatty acids and acylcarnitines. In specific, NAFL-CAU was characterized by higher arachidonic acid, while NAFL-HIS showed lower n-3 PUFA. With the progression to NASH, comparison within ethnicity groups indicated that the hepatic lipidomic profile in NASH-HIS was characterized higher levels of free fatty acids and lipotoxic lipids, suggesting lipotoxicity is involved in the progression of NASH. We also observed a higher hepatic triglyceride with signs of impaired mitochondrial β-oxidation. Remarkably, when comparing between ethnicity groups, the plasma OXLs and eCBs profiles discriminated ethnicities with NASH, independent of histological severity. In specific, NASH-HIS was characterized with lower arachidonic acid derived OXLs, and findings suggest a downregulated lipoxygenase(s) and upregulated soluble epoxide hydrolase(s) activities. Conclusion: The analyses presented in this dissertation highlights the existence of ethnicity-related metabolomic variations in NAFLD that could potentially modulate disease risk and severity in HIS. It also indicates that ethnicity-specific lipidomic signature can distinguish NASH, which needs to be verified in larger studies.

  • 1 supplemental ZIP
Cover page: Plasma and Liver Metabolomic Profiling in Hispanic and Caucasian Subjects with Nonalcoholic Fatty Liver Disease
  • Article
  • Peer Reviewed
UC Davis Previously Published Works (2020)
Copper accumulation and deficiency are reciprocally connected to lipid metabolism. In Wilson disease (WD), which is caused by a genetic loss of function of the copper-transporting P-type ATPase beta, copper accumulates mainly in the liver and lipid metabolism is dysregulated. The underlying mechanisms linking copper and lipid metabolism in WD are not clear. Copper may impair metabolic machinery by direct binding to protein and lipid structures or by generating reactive oxygen species with consequent damage to cellular organelles vital to energy metabolism. In the liver, copper overload results in mitochondrial impairment, down-regulation of lipid metabolism, and the development of steatosis with an etiology not fully elucidated. Little is known regarding the effect of copper overload on extrahepatic energy homeostasis. This review aims to discuss alterations in hepatic energy metabolism associated with WD, highlights potential mechanisms involved in the development of hepatic and systemic dysregulation of lipid metabolism, and reviews current knowledge on the effects of copper overload on extrahepatic energy metabolism.
    Cover page: Lipid and energy metabolism in Wilson disease
    • Article
    • Peer Reviewed
    UC Davis Previously Published Works (2024)
    Hepatic inflammation is commonly identified in Wilson disease (WD), a genetic disease of hepatic and brain copper accumulation. Copper accumulation is associated with increased oxidative stress and reactive oxygen species generation which may result in non-enzymatic oxidation of membrane-bound polyunsaturated fatty acids (PUFA). PUFA can be oxidized enzymatically via lipoxygenases (LOX), cyclooxygenases (COX), and cytochrome P450 monooxygenases (CYP). Products of PUFA oxidation are collectively known as oxylipins (OXL) and are bioactive lipids that modulate hepatic inflammation. We examined hepatic OXL profiles at early stages of WD in two mouse models, the toxic milk mouse from The Jackson Laboratory (tx-j) and the Atp7b knockout on a C57Bl/6 background (Atp7b-/-B6). Targeted lipidomic analysis performed by ultra-high-performance liquid chromatography-electrospray ionization-tandem mass spectrometry showed that in both tx-j and Atp7b-/-B6 mice, hepatic OXL profiles were altered with higher thromboxane and prostaglandins levels. The levels of oxidative stress marker, 9-HETE were increased more markedly in tx-j mice. However, both genotypes showed upregulated transcript levels of many genes related to oxidative stress and inflammation. Both genotypes showed higher prostaglandins, thromboxin along with higher PUFA-derived alcohols, diols, and ketones with altered epoxides; the expression of Alox5 was upregulated and many CYP-related genes were dysregulated. Pathway analyses show dysregulation in arachidonic acid and linoleic acid metabolism characterizes mice with WD. Our findings indicate alterations in hepatic PUFA metabolism in early-stage WD and suggest the upregulation of both, non-enzymatic ROS-dependent and enzymatic PUFA oxidation, which could have implications for hepatic manifestations in WD and represent potential targets for future therapies.
      Cover page: Hepatic oxylipin profiles in mouse models of Wilson disease: New insights into early hepatic manifestations.
      • Article
      • Peer Reviewed
      UC Davis Previously Published Works (2019)
      Wilson disease (WD) is a genetic copper overload condition characterized by hepatic and neuropsychiatric symptoms with a not well-understood pathogenesis. Dysregulated methionine cycle is reported in animal models of WD, though not verified in humans. Choline is essential for lipid and methionine metabolism. Defects in neurotransmitters as acetylcholine, and biogenic amines are reported in WD; however, less is known about their circulating precursors. We aimed to study choline, methionine, aromatic amino acids, and phospholipids in serum of WD subjects. Hydrophilic interaction chromatography-quadrupole time-of-flight mass spectrometry was employed to profile serum of WD subjects categorized as hepatic, neurologic, and pre-clinical. Hepatic transcript levels of genes related to choline and methionine metabolism were verified in the Jackson Laboratory toxic milk mouse model of WD (tx-j). Compared to healthy subjects, choline, methionine, ornithine, proline, phenylalanine, tyrosine, and histidine were significantly elevated in WD, with marked alterations in phosphatidylcholines and reductions in sphingosine-1-phosphate, sphingomyelins, and acylcarnitines. In tx-j mice, choline, methionine, and phosphatidylcholine were similarly dysregulated. Elevated choline is a hallmark dysregulation in WD interconnected with alterations in methionine and phospholipid metabolism, which are relevant to hepatic steatosis. The elevated phenylalanine, tyrosine, and histidine carry implications for neurologic manifestations and are worth further investigation.
        Cover page: Dysregulated Choline, Methionine, and Aromatic Amino Acid Metabolism in Patients with Wilson Disease: Exploratory Metabolomic Profiling and Implications for Hepatic and Neurologic Phenotypes.
        • Article
        • Peer Reviewed
        UC Davis Previously Published Works (2021)
        Nonalcoholic fatty liver disease (NAFLD) is a progressive condition that includes steatosis (NAFL) and nonalcoholic steatohepatitis (NASH). In the U.S., Hispanics (HIS) are afflicted with NAFLD at a higher rate and severity compared to other ethnicities. To date, the mechanisms underlying this disparity have not been elucidated. In this pilot study, we compared untargeted plasma metabolomic profiles for primary metabolism, complex lipids, choline and related compounds between a group of HIS (n =7) and White Caucasian (CAU, n =8) subjects with obesity and biopsy-characterized NAFL to ethnicity-matched lean healthy controls (n =14 HIS and 8 CAU). We also compared liver and plasma metabolomic profiles in a group of HIS and CAU subjects with obesity and NASH of comparable NAFLD Activity Scores, to BMI-matched NASH-free subjects in both ethnicities. Results highlight signs of metabolic dysregulation observed in HIS, independent of obesity, including higher plasma triglycerides, acylcarnitines, and free fatty acids. With NASH progression, there were ethnicity-related differences in the hepatic profile, including higher free fatty acids and lysophospholipids seen in HIS, suggesting lipotoxicity is involved in the progression of NASH. We also observed greater hepatic triglyceride content, higher plasma triglyceride concentrations and lower hepatic phospholipids with signs of impaired hepatic mitochondrial β-oxidation. These findings provide preliminary evidence indicating ethnicity-related variations that could potentially modulate the risk for progression of NALD to NASH.

        Highlights

        Hispanic subjects show signs of metabolic dysregulations independent of obesity. In nonalcoholic fatty liver, plasma metabolomic profiles are differentially altered between ethnicities. In nonalcoholic steatohepatitis, the hepatic lipidomic profiles are differentially altered between ethnicities. In Hispanics, nonalcoholic steatohepatitis shows higher triglycerides and lipotoxic lipids with signs of impaired hepatic mitochondrial β-oxidation. Our results indicate evidence of ethnicity-related variations in the pathogenesis of nonalcoholic fatty liver disease.

        Graphical Abstract

          • Article
          • Peer Reviewed
          UC Davis Previously Published Works (2022)
          Non-alcoholic fatty liver disease (NAFLD) is a common liver pathology that includes steatosis, or non-alcoholic fatty liver (NAFL), and non-alcoholic steatohepatitis (NASH). Without a clear pathophysiological mechanism, it affects Hispanics disproportionately compared to other ethnicities. Polyunsaturated fatty acids (PUFAs) and inflammatory lipid mediators including oxylipin (OXL) and endocannabinoid (eCB) are altered in NAFLD and thought to contribute to its pathogenesis. However, the existence of ethnicity-related differences is not clear. We employed targeted lipidomic profiling for plasma PUFAs, non-esterified OXLs and eCBs in White Hispanics (HIS, n = 10) and Caucasians (CAU, n = 8) with biopsy-confirmed NAFL, compared with healthy control subjects (HC; n = 14 HIS; n = 8 CAU). NAFLD was associated with diminished long chain PUFA in HIS, independent of histological severity. Differences in plasma OXLs and eCBs characterized ethnicities in NASH, with lower arachidonic acid derived OXLs observed in HIS. The secondary analysis comparing ethnicities within NASH (n = 12 HIS; n = 17 CAU), confirms these ethnicity-related differences and suggests lower lipoxygenase(s) and higher soluble epoxide hydrolase(s) activities in HIS compared to CAU. While causes are not clear, these lipidomic differences might be with implications for NAFLD severity and are worth further investigation. We provide preliminary data indicating ethnicity-specific lipidomic signature characterizes NASH which requires further validation.
            Cover page: Plasma Oxylipin Profile Discriminates Ethnicities in Subjects with Non-Alcoholic Steatohepatitis: An Exploratory Analysis.
            • Article
            • Peer Reviewed
            UC Davis Previously Published Works (2021)

            Background & aims

            The pathogenesis of Wilson disease (WD) involves hepatic and brain copper accumulation resulting from pathogenic variants affecting the ATP7B gene and downstream epigenetic and metabolic mechanisms. Prior methylome investigations in human WD liver and blood and in the Jackson Laboratory (Bar Harbor, ME) C3He-Atp7btx-j/J (tx-j) WD mouse model revealed an epigenetic signature of WD, including changes in histone deacetylase (HDAC) 5. We tested the hypothesis that histone acetylation is altered with respect to copper overload and aberrant DNA methylation in WD.

            Methods

            We investigated class IIa HDAC4 and HDAC5 and H3K9/H3K27 histone acetylation in tx-j mouse livers compared with C3HeB/FeJ (C3H) control in response to 3 treatments: 60% kcal fat diet, D-penicillamine (copper chelator), and choline (methyl group donor). Experiments with copper-loaded hepatoma G2 cells were conducted to validate in vivo studies.

            Results

            In 9-week tx-j mice, HDAC5 levels increased significantly after 8 days of a 60% kcal fat diet compared with chow. In 24-week tx-j mice, HDAC4/5 levels were reduced 5- to 10-fold compared with C3H, likely through mechanisms involving HDAC phosphorylation. HDAC4/5 levels were affected by disease progression and accompanied by increased acetylation. D-penicillamine and choline partially restored HDAC4/5 and H3K9ac/H3K27ac to C3H levels. Integrated RNA and chromatin immunoprecipitation sequencing analyses revealed genes regulating energy metabolism and cellular stress/development, which, in turn, were regulated by histone acetylation in tx-j mice compared with C3H mice, with Pparα and Pparγ among the most relevant targets.

            Conclusions

            These results suggest dietary modulation of class IIa HDAC4/5, and subsequent H3K9/H3K27 acetylation/deacetylation can regulate gene expression in key metabolic pathways in the pathogenesis of WD.
              Cover page: Wilson Disease: Intersecting DNA Methylation and Histone Acetylation Regulation of Gene Expression in a Mouse Model of Hepatic Copper Accumulation
              • Article
              • Peer Reviewed
              UC Davis Previously Published Works (2020)

              Background and aims

              Wilson disease (WD) is caused by mutations in the copper transporter ATP7B, with its main pathology attributed to copper-mediated oxidative damage. The limited therapeutic effect of copper chelators and the early occurrence of mitochondrial deficits, however, undermine the prevalence of this mechanism.

              Methods

              We characterized mitochondrial DNA copy number and mutations as well as bioenergetic deficits in blood from patients with WD and in livers of tx-j mice, a mouse model of hepatic copper accumulation. In vitro experiments with hepatocytes treated with CuSO4 were conducted to validate in vivo studies.

              Results

              Here, for the first time, we characterized the bioenergetic deficits in WD as consistent with a mitochondrial DNA depletion-like syndrome. This is evidenced by enriched DNA synthesis/replication pathways in serum metabolomics and decreased mitochondrial DNA copy number in blood of WD patients as well as decreased mitochondrial DNA copy number, increased citrate synthase activity, and selective Complex IV deficit in livers of the tx-j mouse model of WD. Tx-j mice treated with the copper chelator penicillamine, methyl donor choline or both ameliorated mitochondrial DNA damage but further decreased mitochondrial DNA copy number. Experiments with copper-loaded HepG2 cells validated the concept of a direct copper-mitochondrial DNA interaction.

              Conclusions

              This study underlines the relevance of targeting the copper-mitochondrial DNA pool in the treatment of WD separate from the established copper-induced oxidative stress-mediated damage.
                • Article
                • Peer Reviewed
                UC Davis Previously Published Works (2023)

                Background

                The clinical manifestations of Wilson disease (WD) are related to copper accumulation in the liver and the brain, but little is known about other tissue involvement regarding metabolic changes in WD. In vitro studies suggested that the loss of intestinal ATP7B affects metabolic dysregulation in WD. We tested this hypothesis by evaluating the gut microbiota and lipidome in 2 mouse models of WD and by characterizing a new mouse model with a targeted deletion of Atp7b in the intestine.

                Methods

                Cecal content 16S sequencing and untargeted hepatic and plasma lipidome analyses in the Jackson Laboratory toxic-milk and the Atp7b null global knockout mouse models of WD were profiled and integrated. Intestine-specific Atp7b knockout mice (Atp7bΔIEC) were generated and characterized using targeted lipidome analysis following a high-fat diet challenge.

                Results

                Gut microbiota diversity was reduced in animal models of WD. Comparative prediction analysis revealed amino acid, carbohydrate, and lipid metabolism functions to be dysregulated in the WD gut microbial metagenome. Liver and plasma lipidomic profiles showed dysregulated triglyceride and diglyceride, phospholipid, and sphingolipid metabolism in WD models. However, Atp7bΔIEC mice did not show gut microbiome differences compared to wild type. When challenged with a high-fat diet, Atp7bΔIEC mice exhibited profound alterations to fatty acid desaturation and sphingolipid metabolism pathways as well as altered APOB48 distribution in intestinal epithelial cells.

                Conclusions

                Gut microbiome and lipidome underlie systemic metabolic manifestations in murine WD. Intestine-specific ATP7B deficiency affected both intestinal and systemic response to a high-fat challenge but not the microbiome profile, at least at early stages. WD is a systemic disease in which intestinal-specific ATP7B loss and diet influence the phenotype and the lipidome profile.
                  Cover page: The role of intestine in metabolic dysregulation in murine Wilson disease
                  • Article
                  • Peer Reviewed
                  UC Davis Previously Published Works (2021)
                  Nonalcoholic fatty liver disease (NAFLD) is a progressive condition that includes steatosis (NAFL) and nonalcoholic steatohepatitis (NASH). In the U.S., Hispanics (HIS) are afflicted with NAFLD at a higher rate and severity compared to other ethnicities. To date, the mechanisms underlying this disparity have not been elucidated. In this pilot study, we compared untargeted plasma metabolomic profiles for primary metabolism, complex lipids, choline and related compounds between a group of HIS (n = 7) and White Caucasian (CAU, n = 8) subjects with obesity and biopsy-characterized NAFL to ethnicity-matched lean healthy controls (n = 14 HIS and 8 CAU). We also compared liver and plasma metabolomic profiles in a group of HIS and CAU subjects with obesity and NASH of comparable NAFLD Activity Scores, to BMI-matched NASH-free subjects in both ethnicities. Results highlight signs of metabolic dysregulation observed in HIS, independent of obesity, including higher plasma triglycerides, acylcarnitines, and free fatty acids. With NASH progression, there were ethnicity-related differences in the hepatic profile, including higher free fatty acids and lysophospholipids seen in HIS, suggesting lipotoxicity is involved in the progression of NASH. We also observed greater hepatic triglyceride content, higher plasma triglyceride concentrations and lower hepatic phospholipids with signs of impaired hepatic mitochondrial β-oxidation. These findings provide preliminary evidence indicating ethnicity-related variations that could potentially modulate the risk for progression of NALD to NASH.
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