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Tissue Engineering

Tissue Engineering - Poster Session E

Poster L20 - Using Organoids As a Model System to Study Intestinal Epithelial Growth and Morphology

Saturday, October 26, 2024
10:00 AM – 11:00 AM EST
Location: Exhibit Hall E, F & G

Introduction:  Organoids are three-dimensional (3D) miniature versions of organs or tissues that are stem cell derived. They are able to replicate the morphology and structure of the in vitro organ. Organoids are also able to proliferate spontaneously. They are currently being used in various biomedical studies due to their capability to provide good 3D models of the organs in our body. Organoid models are currently being used for drug research, disease modeling, transplantable organs, and regenerative medicine.

In this study we used intestinal organoids (IOs) as our model system. The intestine is vital for digestive function and nutrient absorption. The intestine is lined with the epithelium and forms the crypt-villus architecture. This is vital in the body, because it acts as a protective barrier to the organ, maintains the structure of the intestinal epithelium, and maintains gut homeostasis. This system allows for nutrient and drug absorption to happen seamlessly while protecting the organ from toxins and disease. The cells on the villi shed when dying and are renewed by cells from the crypt. This allows the protective barrier to maintain structure while protecting the cells. So, in this study, we used mouse intestinal organoids as a model system to look into the growth and morphology of intestinal organoids over time.

Materials and

Methods:  1. Intestinal organoid culture
Mouse intestinal organoids are embedded in extracellular matrix (ECM) hydrogel drops and injected into the well plates for drop culture analysis.
Medium is changed every other day and between day 5-7 passaging occurs. The passaging process includes breaking down the cells and gel so that cells can be redistributed into more wells to allow for more room to grow and proliferate. Passaging is done usually 4x and if passaging does not take place cells will die due to lack of space and growth factors.

2. Image analysis
Image J is used to detect organoid size and edges. This software allows us to measure the organoids and provides a clear image of the edges due to the edge detection filter plugin. Next, the organoids go through a series of steps to stop the growth phases at certain time points to prepare for immunostaining. This targets actin filaments and then we can image the structure and cell morphology.

Results, Conclusions, and Discussions: Results and Discussions: Currently Figure 1 shows the organoid increasing in size over the course of a 9-day culture. The organoids started around 150 um in diameter at day 1, to around 400 um in diameter at day 9. This shows that the organoids are now 2.5x bigger than originally. This means organoids are developing and proliferating. The budding that is shown in Figure 2 shows the morphological development over time. These organoids are designed for their nutrient absorption and this crypt villus structure in Figure 2 indicates that they can mimic the structure of the intestinal epithelium. Some challenges to be addressed are that after day 7 organoids start to die and we plan to address this by incorporating the organoids on chips.

Conclusion: Overall, organoids are useful model system for studying the structural and functional properties of our intestine. We can see that over time these organoids develop this crypt-villus structure spontaneously. Organoid growth also increases over time, and this means that cells are proliferating, so the bigger the size of the organoid, the more cells are present and the more mature the tissue is. The budding structure of the organoids is significant because this shows they are more mature than the spherical organoids. The budding structure is an essential feature because this represents the crypt-villus architecture in our intestinal organ. Abnormal crypt-villus architecture can be linked to inflammatory bowel disease and other conditions. The surface area of the intestinal epithelium increases as they grow, which is vital for nutrient absorption.

Acknowledgements (Optional):