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Efficient and simple genetic engineering of enteroids using mouse isolated crypts for investigating intestinal
Shuya Ohira1, Yuki Yokoi2, Tokiyoshi Ayabe2
1Innate Immunity Laboratory, Graduate School of Life Science, Hokkaido University, Kita-21, Nishi-11, Kita-ku, Sapporo, Hokkaido, 001-0021, Japan.
Biochemical and Biophysical Research Communications
|November 19, 2022
Summary
Researchers developed a faster, more efficient method to create transgenic enteroids for studying intestinal epithelial cells. This new technique involves gene transfer into isolated crypts, improving high-throughput investigation of gut function.
Area of Science:
- Gastroenterology
- Cell Biology
- Biotechnology
Background:
- Intestinal epithelial cells (IECs) are crucial for gut function, separating the lumen from tissues.
- Enteroids, 3D cultures of IECs, are used for functional analysis, with transgenic models aiding molecular studies.
- Conventional transgenic enteroid generation is inefficient and time-consuming due to single stem cell transfection.
Purpose of the Study:
- To develop a more efficient and rapid method for generating transgenic enteroids.
- To enable high-throughput investigation of intestinal epithelial cell functions.
- To facilitate a deeper understanding of molecular mechanisms in intestinal biology.
Main Methods:
- Gene transfer into isolated small intestinal crypts using lentiviral vectors without single-cell dissociation.
- Culturing transfected crypts to generate transgenic enteroids.
- Assessing enteroid function, including drug transport and innate immunity.
Main Results:
- Transgenic enteroids were generated efficiently and quickly (within 7 days) from transfected crypts.
- A high yield of 56% was achieved for transgenic enteroid generation.
- The generated enteroids maintained key intestinal functions, such as drug transport and innate immunity.
Conclusions:
- Gene transfer into isolated crypts offers an efficient and rapid approach for transgenic enteroid production.
- This method overcomes the limitations of conventional techniques, enabling faster research.
- The improved method supports high-throughput studies, advancing the understanding of intestinal epithelial cell biology and function.
