Metabolic Patterning on a Chip: Towards in vitro Liver Zonation of Primary Rat and Human Hepatocytes

An important number of healthy and diseased tissues shows spatial variations in their metabolic capacities across the tissue. The liver is a prime example of such heterogeneity where the gradual changes in various metabolic activities across the liver sinusoid is termed as “zonation” of the liver. Here, we introduce the Metabolic Patterning on a Chip (MPOC) platform capable of dynamically creating metabolic patterns across the length of a microchamber of liver tissue via actively enforced gradients of various metabolic modulators such as hormones and inducers. Using this platform, we were able to create continuous liver tissues of both rat and human origin with gradually changing metabolic activities. The gradients we have created in nitrogen, carbohydrate and xenobiotic metabolisms recapitulated an in vivo like zonation and zonal toxic response. Beyond its application in recapitulation of liver zonation in vitro as we demonstrate here, the MPOC platform can be used and expanded for a variety of purposes including better understanding of heterogeneity in many different tissues during developmental and adult stages

Gut Microbial Metabolism and Nonalcoholic Fatty Liver Disease.

The gut microbiome, the multispecies community of microbes that exists in the gastrointestinal tract, encodes several orders of magnitude more functional genes than the human genome. It also plays a pivotal role in human health, in part due to metabolism of environmental, dietary, and host-derived substrates, which produce bioactive metabolites. Perturbations to the composition and associated metabolic output of the gut microbiome have been associated with a number of chronic liver diseases, including nonalcoholic fatty liver disease (NAFLD). Here, we review the rapidly evolving suite of next-generation techniques used for studying gut microbiome composition, functional gene content, and bioactive products and discuss relationships with the pathogenesis of NAFLD

Ketone Body Metabolism Preserves Hepatic Function during Adaptation to Birth and in Overnutrition

Mammalian ketone body metabolism partially oxidizes hepatic acyl-chains to ketone body intermediates, which can serve as alternative fuels in extrahapetic tissues during carbohydrate restricted states. Ketone body production (ketogenesis) occurs primarily in liver, due to hepatocyte-specific expression of the fate committing ketogenic enzyme, mitochondrial 3-hydroxymethylglutaryl-CoA synthase (HMGCS2). In contrast, the fate committing enzyme of ketone body oxidation, mitochondrial Succinyl-CoA:3-oxoacid CoA Transferase (SCOT), is expressed ubiquitously, except in liver. Here I demonstrate novel roles for ketone body metabolism during a classically ketogenic period, the transition to birth, and in a classically `non-ketogenic\u27 state, overnutrition, using novel genetic mouse models, high-resolution measures of dynamic metabolism using 13C-labeled substrates, and systems physiology approaches. I show that germline SCOT-knockout (KO) mice cannot oxidize ketone bodies in any tissue. These mice developed lethal hyperketonemia and hypoglycemia within the first 48 hr of life and died in a manner that phenocopied human sudden infant death syndrome. Nonetheless, my studies of tissue-specific SCOT-KO mice revealed that ketone body oxidation is dispensable during the transition to birth and during starvation in the adult when individually eliminated in neurons, cardiomyocytes, or skeletal myocytes, which comprise the three largest consumers of ketone bodies. Surprisingly, the inability to dispose of ketone bodies in germline SCOT-KO mice drove derangements of carbohydrate and fatty acid metabolism, oxidized redox potential, and inhibited ketogenesis in liver. Moreover, I show that adult-onset loss of HMGCS2 ablated the liver\u27s capacity to effectively convert fat into ketone bodies, and thus induced ketogenesis insufficiency. Ketogenesis insufficient mice exhibited increased hepatic gluconeogenesis from pyruvate and mild hyperglycemia in the fed state. High-fat diet feeding of ketogenesis insufficient mice caused extensive hepatocyte injury and inflammation that was associated with decreased glycemia due to fatty acid-induced sequestration of free coenzyme A that caused secondary derangements of hepatic tricarboxylic acid (TCA) cycle intermediate concentrations and impaired gluconeogenesis. Together, my studies have revealed a critical and novel role for ketone body metabolism in preservation of the dynamic intermediary metabolic network in liver during the adaptation to birth and in overnutrition

Relationships between Hematopoiesis and Hepatogenesis in the Midtrimester Fetal Liver Characterized by Dynamic Transcriptomic and Proteomic Profiles

In fetal hematopoietic organs, the switch from hematopoiesis is hypothesized to be a critical time point for organogenesis, but it is not yet evidenced. The transient coexistence of hematopoiesis will be useful to understand the development of fetal liver (FL) around this time and its relationship to hematopoiesis. Here, the temporal and the comparative transcriptomic and proteomic profiles were observed during the critical time points corresponding to the initiation (E11.5), peak (E14.5), recession (E15.5), and disappearance (3 ddp) of mouse FL hematopoiesis. We found that E11.5-E14.5 corresponds to a FL hematopoietic expansion phase with distinct molecular features, including the expression of new transcription factors, many of which are novel KRAB (Kruppel-associated box)-containing zinc finger proteins. This time period is also characterized by extensive depression of some liver functions, especially catabolism/utilization, immune and defense, classical complement cascades, and intrinsic blood coagulation. Instead, the other liver functions increased, such as xenobiotic and sterol metabolism, synthesis of carbohydrate and glycan, the alternate and lectin complement cascades and extrinsic blood coagulation, and etc. Strikingly, all of the liver functions were significantly increased at E14.5-E15.5 and thereafter, and the depression of the key pathways attributes to build the hematopoietic microenvironment. These findings signal hematopoiesis emigration is the key to open the door of liver maturation

Potential therapeutic use of the ketogenic diet in autism spectrum disorders.

The ketogenic diet (KGD) has been recognized as an effective treatment for individuals with glucose transporter 1 (GLUT1) and pyruvate dehydrogenase (PDH) deficiencies as well as with epilepsy. More recently, its use has been advocated in a number of neurological disorders prompting a newfound interest in its possible therapeutic use in autism spectrum disorders (ASD). One study and one case report indicated that children with ASD treated with a KGD showed decreased seizure frequencies and exhibited behavioral improvements (i.e., improved learning abilities and social skills). The KGD could benefit individuals with ASD affected with epileptic episodes as well as those with either PDH or mild respiratory chain (RC) complex deficiencies. Given that the mechanism of action of the KGD is not fully understood, caution should be exercised in ASD cases lacking a careful biochemical and metabolic characterization to avoid deleterious side effects or refractory outcomes

Functional Consequences of Metabolic Zonation in Murine Livers: Insights for an Old Story

Background and Aims: Zone-dependent differences in expression of metabolic enzymes along the portocentral axis of the acinus are a long-known feature of liver metabolism. A prominent example is the preferential localization of the enzyme, glutamine synthetase, in pericentral hepatocytes, where it converts potentially toxic ammonia to the valuable amino acid, glutamine. However, with the exception of a few key regulatory enzymes, a comprehensive and quantitative assessment of zonal differences in the abundance of metabolic enzymes and, much more important, an estimation of the associated functional differences between portal and central hepatocytes is missing thus far. Approach and Results: We addressed this problem by establishing a method for the separation of periportal and pericentral hepatocytes that yields sufficiently pure fractions of both cell populations. Quantitative shotgun proteomics identified hundreds of differentially expressed enzymes in the two cell populations. We used zone-specific proteomics data for scaling of the maximal activities to generate portal and central instantiations of a comprehensive kinetic model of central hepatic metabolism (Hepatokin1). Conclusions: The model simulations revealed significant portal-to-central differences in almost all metabolic pathways involving carbohydrates, fatty acids, amino acids, and detoxification