Sunil K. Sukumaran
Date of this Version
A thesis presented to the faculty of the Graduate College at the University of Nebraska in partial fulfillment of requirements for the degree of Master of Science
Under the supervision of Professor Sunil K. Sukumaran
Lincoln, Nebraska, December 2023
Taste organoids generated from the adult taste stem cells in Matrigel embedded culture is a high-quality model for the study of taste signaling and development. The preliminary aim of this study was to generate organoids that have opposite basal/apical orientations, with cilia facing outward. For polarity reversal the Matrigel from the Organoid culture was removed after the first three weeks and allowed to grow in suspension in a low-attachment plate. The Immunostaining and qRT-PCR analyses determined the expression of taste marker genes and proteins to be comparable in both polarity-reversed and normal organoids. Calcium imaging experiments on this apical out organoids proved the functionality of the same and its potential contributions to the study of taste biology.
Another aim of this study was to identify the role of the surrounding fibroblast in the regeneration and development of taste buds. For this, we isolated the primary fibroblast from around the CV papillae and cocultured it with the taste organoids. Matrigel provided structural support whereas the fibroblast acted as the feeder layer providing the essential growth factors as in the native tissue. qPCR analysis of control and cocultured organoids showed higher expression of taste genes in the latter. This fact suggests the important role of Cell-to-Cell communication in the development of the taste buds.
The suitability of two organoid models for the study of taste signaling was determined using functional calcium imaging experiments. We believe that polarity-reversed taste organoids and co-cultured organoids represent two unique model systems that could prove particularly valuable for the study of taste signaling.
Advisor: Sunil K. Sukumaran