A Scalable Primary Cortical Tri-Culture Model to Study the Influence for Gut Microbiome-Derived Bioactive Compounds on Neuroinflammation

University Of California At Davis, Davis CA

Investigators

Abstract

PROJECT SUMMARY Bioactive compounds produced by native or transient members of the gut microbiota are finding increasing use as therapeutics for a variety of complex human diseases, including obesity, diabetes, cardiovascular disease, and neurological disorders. Once these compounds cross the blood-brain-barrier, some can result in beneficial (e.g., anti-inflammatory, neuroprotective) or detrimental (e.g., pro-inflammatory, neurodegenerative) effects on the central nervous system (CNS). Specific cell types in the CNS are involved in this process, yet how the bioactive compounds interact with them remains poorly understood. Cell culture models offer better experimental control and scalability than animal models and avoid the limits of working with different species. However, most in vitro models suffer from low biological relevance in part due to not being able to simultaneously contain the critical cell types (i.e., neurons, astrocytes, microglia). There is a need for scalable in vitro models that can capture phenomenological outcomes of bioactive compound-CNS interactions and alterations to neural function observed in vivo and thus allow further mechanistic studies, bioactive compound discovery, and translation to human use. To address this critical need, we will employ a novel primary rat cortical cell tri-culture (primary neuron, astrocyte, microglia) model of neuroinflammation and integrated extracellular recording electrodes. In contrast to the co- culture of just neurons and astrocytes, the tri-culture model more faithfully captures neurotoxic and neuroprotective features observed in vivo. Since the tri-culture model is maintained simply by including IL-34, TGF-β, and cholesterol supplements in the conventional co-culture media, it is amenable to scale-up and screening studies in multi-well formats. In the proposed project, we will use the tri-culture model to simulate neuroinflammation that is present in many disorders that range from cancer to neurodegeneration. Microglia plays a particularly important role in neuroinflammation, where various phenotypic changes are observed, including impaired phagocytic capacity. Here, we will use a lipopolysaccharide (LPS)-induced neuroinflammation model. By introducing the bioactive molecules, before, during, or after LPS treatment, we will further mimic scenarios such as the normal presence of gut microbiota bioactive molecules (e.g., prevention) vs. their post- symptom introduction (e.g., therapeutic). Across two aims, we will reveal the effects of bioactive compounds on neuroinflammatory responses via morphological, and proteomic read-outs, as well as on cellular function by evaluating microglial phagocytic capacity and neuronal electrophysiological activity. We expect that this project will create a scalable in vitro tool to study the influence of gut microbiome-derived bioactive compounds on neuroinflammation. This tool can then be used broadly for mechanistic studies and for discovering new bioactive molecules that inform regular diet or prescribed as therapeutics for improving and maintaining neurological health.