HEPSYS-I MICROARRAY EXPRESSION PROFILING OF AN IN VITRO 'NUTRIENT EXCESS' MODEL OF NAFLD IN HUMAN HEPATOCYTES

Leonard Nelson, Philipp Treskes, Donald Dunbar, Jonathan R. Manning, K. Ann Lockman, Steven D Morley, Peter Hayes, John Plevris

Research output: Contribution to conferencePosterpeer-review

Abstract

Non-alcoholic fatty liver disease (NAFLD) is the most common liver disease worldwide, characterized by excessive triglyceride accumulation (steatosis) and oxidative stress. We have developed an in vitro
‘nutrient overload' NAFLD model in human hepatic C3A cells, to recapitulate the sequence of events
proposed to occur in NAFLD.
Objectives: To develop a systems biology approach (HepSys) combining identification of distinct patterns
of perturbed hepatic metabolism, with downstream integration of proteomics and metabolomics data sets.
Methods: C3A cells were loaded with a 'nutrient cocktail' (72h) of Lactate, Pyruvate, Octanoate and
Ammonia (LPON) to promote the dysregulated metabolic NAFLD phenotype. Microarray RNA expression
was measured using Illumina® Whole Human Genome BeadChip H12 Microarray. Data was normalized,
validated and analysed to examine differences between Control (C3A untreated) and LPON-treated
groups.
Results: NAFLD phenotype was confirmed using morphological [BODIPY] and biochemical parameters
[MitoSOX]. Microarray data was collated onto the HepSys Microarray Portal with capacity for Proteomics
data integration. 12 distinct biochemical pathways were identified [Control v LPON] with significantly
upregulated differential expression patterns [Filters/ Statistics: >2x fold Deltaexpression; P<0.05, Rank
Products pfp; and linked to KEGG Metabolic Pathway]; including: Metabolic pathways [30 genes; eg
FASN, involved in fatty acid metabolism, ACAT2 (synthesizes Acetoacetyl CoA, essential for cholesterol
synthesis; PCK2 key mitochondrial enzyme in gluconeogenesis; Steroid biosynthesis [5 genes: MSMO1,
TM7SF2 FDFT1, DHCR7, EBP (EBP is involved in cholesterol biosynthesis/ lipoprotein internalisation];
Oxidative phosphorylation [6 genes total: eg NADH dehydrogenases: NDUFA3, NDUFB10, NDUFB7;
Cytochrome c reductase: CYC1; ATP6V0E2 (H+-ATPase) and ATP5D (catalyzes ATP synthesis). Global
changes in expression levels were visualized using Heatmap [138 log2 expression values, samples
clustered via complete linkage].
Conclusions: Our results show NAFLD-related morphological/ biochemical changes are associated with
clear differential gene expression patterns, which implicate several distinct pathways in the dysregulated
metabolism of NAFLD. Future validatory studies of gene expression signatures, including qRT-PCR
analyses, and correlation/ integration with cognate Proteomic data sets may uncover hitherto unknown
pathways involved in NAFLD pathogenesis.
Original languageEnglish
Publication statusPublished - Feb 2013
EventlEASL Monothematic Conference: Systems Biology of the Liver: Systems Biology and Clinics Face-à-Face - Luxemburg, Luxembourg
Duration: 21 Feb 201323 Feb 2013

Conference

ConferencelEASL Monothematic Conference: Systems Biology of the Liver: Systems Biology and Clinics Face-à-Face
CountryLuxembourg
CityLuxemburg
Period21/02/1323/02/13

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