A Human Gut-Liver Platform with Microbiome Interactions for In Vitro Toxicology

ABSTRACT While rodents have provided valuable information on key aspects of the gut-liver axis, there are fundamental species-specific differences in the gut-liver axis, such as the composition of the microbiome, differences in liver xenobiotic metabolism, and progression of alcoholic and metabolic fatty liver disease. Therefore, human- relevant in vitro models now play important roles in complementing in vivo animal studies for basic science and for drug screening. While considerable progress has been made in the development of human models of intestine and the liver using physiologically relevant primary cells, very little has been done in engineering an integrated human gut-liver model using primary cells. We have utilized droplet microfluidics to fabricate highly monodisperse extracellular matrix (ECM)-based 3D liver microtissues containing primary human hepatocytes and liver sinusoidal endothelial cells that are rapidly generated and functionally outperform self-assembled hepatic spheroids and hepatocytes within bulk gels for 4+ weeks; furthermore, the microtissues can be further augmented with human hepatic stellate cells and Kupffer cells. We have also separately developed in vitro human intestine models using primary cells, which can be engineered to maintain gradients of O2 across the basal and luminal surfaces to recapitulate the stem cell and differentiated cell compartments of the crypt; can produce mucus; and are amenable to the incorporation of microbiota in the luminal compartment. Here, we will engineer in vitro human gut-liver models of increasing complexities, which will be used to elucidate how intestine, liver, and microbiota interact to modulate reciprocal functions. In aim 1, we will elucidate reciprocal functional changes in a human gut-liver model containing primary human colonocytes and liver cells, while in aim 2 we will introduce mucus and microbiota in the intestinal model and elucidate reciprocal crosstalk upon co-culture with human liver microtissues. This proposal will yield a first-of-its-kind scalable human gut-liver model containing primary cells for in vitro applications, such as compound screening and disease modeling.