3D Flipwell Engineering for Developing Asynchronous Systems for Toxicologic and Immunomodulatory Therapies in Bacterial, Gut, and Immune Cells

Maria A. Beamer; Saori Furuta

doi:10.3791/68701

3D Flipwell Engineering for Developing Asynchronous Systems for Toxicologic and Immunomodulatory ...

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Cancer Research

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3D Flipwell Engineering for Developing Asynchronous Systems for Toxicologic and Immunomodulatory Therapies in Bacterial, Gut, and Immune Cells

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11:00 min

October 17, 2025

DOI: 10.3791/68701-v

Maria A. Beamer¹, Saori Furuta²

¹Division of Pediatric Rheumatology, Department of Pediatrics,University of Michigan, Ann Arbor, ²MetroHealth Medical Center,Case Western Reserve University School of Medicine, Case Comprehensive Cancer Center

Overview

This article provides a detailed protocol for engineering and utilizing a 3D co-culture system that mimics the gut mucosal environment. It highlights the assembly of co-culture insert stacks for studying interactions among gut bacteria, epithelial cells, and macrophages.

Key Study Components

Area of Science

Neuroscience
Microbiology
Immunology

Background

Existing culture systems for gut mucosal modeling are complex and costly.
This study presents a simpler and more affordable co-culture system.
The system aims to evaluate drug responses and cellular interactions.
It focuses on both aerobic and anaerobic bacterial strains.

Purpose of Study

To develop a 3D co-culture system for studying gut mucosal interactions.
To evaluate drug responses in a controlled environment.
To uncover immune-modulating effects of bacterial metabolites.

Methods Used

Assembly of co-culture insert stacks.
Co-culturing gut bacteria, epithelial cells, and macrophages.
Evaluation of drug responses and cellular interactions.
Analysis of proteinogenic drug effects on host immunity.

Main Results

The co-culture system revealed coordinated responses among gut bacteria, epithelial, and immune cells.
Proteinogenic drugs activated host immunity through bacterial metabolites.
Cross-species communication was mediated by these interactions.
The findings pave the way for new immunotherapeutic strategies.

Conclusions

The developed co-culture system is a viable alternative to complex models.
It facilitates the study of gut mucosal interactions and drug responses.
Future research can explore the therapeutic potential of bacterial metabolites.

Frequently Asked Questions

What is the main advantage of the 3D co-culture system?

It is simpler and more cost-effective compared to existing complex systems.

What types of cells are used in the co-culture?

Gut bacteria, epithelial cells, and macrophages are co-cultured.

How does the system help in drug screening?

It allows for evaluation of drug responses in a controlled gut-like environment.

What are the implications of the study's findings?

The findings may lead to new immunotherapeutic strategies based on bacterial metabolites.

Can the system be used for both aerobic and anaerobic bacteria?

Yes, the study aims to investigate both types of bacterial strains.

What is the role of bacterial metabolites in the study?

Bacterial metabolites are believed to activate host immunity and mediate interactions.

This article provides a detailed protocol with key steps to guide the 3D Flipwell engineering and utilization. We describe and show how to assemble the co-culture insert stacks and utilize them for co-culturing stratified layers of gut bacteria, gut epithelia, and macrophages to model the gut mucosal environment.

We have developed a 3D co-culture system that mimics the gut mucosal environment to study the serial crosstalk and to evaluate the drug responses which could be utilized for drug screening and for understanding mucosal cellular interactions. Different groups have developed several different culture system to model the gut mucosal environment by using very complex bioengineering techniques. But instead, we developed a co-culture system which is much easier to make using less expensive materials.

Our co-culture system revealed that a proteinogenic drug triggers coordinated responses among gut bacteria, epithelial and immune cells, activating host immunity likely with our bacterial metabolites and mediating cross-species communication. We aim to study both aerobic and anerobic bacterial strains and their metabolites to uncover immune modulating effects, paving the way for new immunotherapeutic strategies based on bacterial metabolites. To begin, open a Petri dish cover and set it aside.

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