Exploiting three-dimensional human hepatic constructs to investigate the impact of rs174537 on fatty acid metabolism.

L Madison Kirk,Carl D Langefeld,Charles E Mccall,Elaheh Rahbar,John S Parks,Alexander C Bashore,Thomas J Depalma,Antonio C Renaldo,Kelli N Simms,Charlotte Mae K Waits,Aleksander Skardal,Nathaniel Hauser,Susan Sergeant,Chia-Chi Chuang Key,Allison K Meyers,Beverly Dosso,Catherine Mounier

doi:10.1371/journal.pone.0262173

Abstract

The Modern Western Diet has been associated with the rise in metabolic and inflammatory diseases, including obesity, diabetes, and cardiovascular disease. This has been attributed, in part, to the increase in dietary omega-6 polyunsaturated fatty acid (PUFA) consumption, specifically linoleic acid (LA), arachidonic acid (ARA), and their subsequent metabolism to pro-inflammatory metabolites which may be driving human disease. Conversion of dietary LA to ARA is regulated by genetic variants near and within the fatty acid desaturase (FADS) haplotype block, most notably single nucleotide polymorphism rs174537 is strongly associated with FADS1 activity and expression. This variant and others within high linkage disequilibrium may potentially explain the diversity in both diet and inflammatory mediators that drive chronic inflammatory disease in human populations. Mechanistic exploration into this phenomenon using human hepatocytes is limited by current two-dimensional culture models that poorly replicate in vivo functionality. Therefore, we aimed to develop and characterize a three-dimensional hepatic construct for the study of human PUFA metabolism. Primary human hepatocytes cultured in 3D hydrogels were characterized for their capacity to represent basic lipid processing functions, including lipid esterification, de novo lipogenesis, and cholesterol efflux. They were then exposed to control and LA-enriched media and reproducibly displayed allele-specific metabolic activity of FADS1, based on genotype at rs174537. Hepatocytes derived from individuals homozygous with the minor allele at rs174537 (i.e., TT) displayed the slowest metabolic conversion of LA to ARA and significantly reduced FADS1 and FADS2 expression. These results support the feasibility of using 3D human hepatic cultures for the study of human PUFA and lipid metabolism and relevant gene-diet interactions, thereby enabling future nutrition targets in humans.

Highlights

The Modern Western Diet (MWD) has been associated with the rising prevalence of metabolic disorders, including diabetes, obesity, cardiovascular, and chronic inflammatory diseases [1,2,3]
primary human hepatocytes (PHH) were cultured in a custom hydrogel, and were viable for up to one week in culture (Fig 1), and maintained their rounded phenotype for longer than those cultured in 2D (S1 Fig)
Given that polyunsaturated fatty acid (PUFA) metabolism in humans is dependent on fatty acid desaturase (FADS) related genetic variants, we aimed to explore the ability of the 3D human culture platform to replicate individual variability in PUFA metabolic conversion capacity

Summary

Introduction

The Modern Western Diet (MWD) has been associated with the rising prevalence of metabolic disorders, including diabetes, obesity, cardiovascular, and chronic inflammatory diseases [1,2,3]. Recent reports indicate that current consumption of LA is ~13–16 g per day, translating to about 6–8% of the US daily energy intake [7]. These extremely high LA levels are concerning as they have been linked to a variety of inflammatory diseases and cancer. In a recent prospective dietary supplementation study, the metabolic conversion of LA to ARA in humans was shown to be strongly associated with genotype at rs174537, a single nucleotide polymorphism (SNP) located downstream of FADS1 on chromosome 11 [12]. The mechanistic study of these effects within the liver remains difficult due to the lack of reliable human in vitro models

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Conclusion