generation-culture-lingual-organoids-derived-from-adult-mouse-taste

Generation and Culture of Lingual Organoids Derived from Adult Mouse Taste Stem Cells

The sense of taste is mediated by taste buds on the tongue, which are composed of rapidly renewing taste receptor cells (TRCs). This continual turnover is ...

University of Colorado Anschutz Medical Campus

Wnt/β-catenin signaling initiates taste papilla development in mouse embryos, however, its involvement in taste cell turnover in adult mice has not been explored. Here we used the BATGAL reporter mouse model, which carries an engineered allele in which the LacZ gene is expressed in the presence of activated β-catenin, to determine the responsiveness of adult taste bud cells to canonical Wnt signaling. Double immunostaining with markers of differentiated taste cells revealed that a subset of Type I, II, and III taste cells express β-galactosidase. Using in situ hybridization, we showed that β-catenin activates the transcription of the LacZ gene mainly in intragemmal basal cells that are immature taste cells, identified by their expression of Sonic Hedgehog (Shh). Finally, we showed that β-catenin activity is significantly reduced in taste buds of 25-week-old mice compared with 10-week-old animals. Our data suggest that Wnt/β-catenin signaling may influence taste cell turnover by regulating cell differentiation. Reduced canonical Wnt signaling in older mice could explain in part the loss of taste sensitivity with aging, implicating a possible deficiency in the rate of taste cell renewal. More investigations are now necessary to understand if and how Wnt signaling regulates adult taste cell turnover.

Taste bud cells of adult mice are responsive to Wnt/β-catenin signaling: implications for the renewal of mature taste cells.

Continuous taste bud cell renewal is essential to maintain taste function in adults; however, the molecular mechanisms that regulate taste cell turnover are unknown. Using inducible Cre-lox technology, we show that activation of β-catenin signaling in multipotent lingual epithelial progenitors outside of taste buds diverts daughter cells from a general epithelial to a taste bud fate. Moreover, while taste buds comprise 3 morphological types, β-catenin activation drives overproduction of primarily glial-like Type I taste cells in both anterior fungiform (FF) and posterior circumvallate (CV) taste buds, with a small increase in Type II receptor cells for sweet, bitter and umami, but does not alter Type III sour detector cells. Beta-catenin activation in post-mitotic taste bud precursors likewise regulates cell differentiation; forced activation of β-catenin in these Shh+ cells promotes Type I cell fate in both FF and CV taste buds, but likely does so non-cell autonomously. Our data are consistent with a model where β-catenin signaling levels within lingual epithelial progenitors dictate cell fate prior to or during entry of new cells into taste buds; high signaling induces Type I cells, intermediate levels drive Type II cell differentiation, while low levels may drive differentiation of Type III cells. 

β-Catenin Signaling Biases Multipotent Lingual Epithelial Progenitors to Differentiate and Acquire Specific Taste Cell Fates.

The natural like and dislike of foods based on taste is one of the most easily observed behaviors in animals. Animals eat palatable foods and reject aversive foods, which makes measurement of taste perception possible using various behavioral techniques. Three different methods to accurately measure taste behavior are described here. First, two-bottle preference tests evaluate whether a taste compound (tastant) is preferred over water. Second, lickometer tests quantify the like and dislike for multiple concentrations of the same tastant or multiple tastants at the same time. Finally, conditioned taste aversion tests accurately determine the perceived taste threshold for palatable tastants. Together, these diverse methods enable researchers to observe and measure behavioral taste responses in mice to any tastant. © 2016 by John Wiley & Sons, Inc.

Measurement of Behavioral Taste Responses in Mice: Two-Bottle Preference, Lickometer, and Conditioned Taste-Aversion Tests.

Taste stimuli are transduced by taste buds and transmitted to the brain via afferent gustatory fibers. Renewal of taste receptor cells from actively dividing progenitors is finely tuned to maintain taste sensitivity throughout life. We show that conditional β-catenin deletion in mouse taste progenitors leads to rapid depletion of progenitors and Shh+ precursors, which in turn causes taste bud loss, followed by loss of gustatory nerve fibers. In addition, our data suggest LEF1, TCF7 and Wnt3 are involved in a Wnt pathway regulatory feedback loop that controls taste cell renewal in the circumvallate papilla epithelium. Unexpectedly, taste bud decline is greater in the anterior tongue and palate than in the posterior tongue. Mutant mice with this regional pattern of taste bud loss were unable to discern sweet at any concentration, but could distinguish bitter stimuli, albeit with reduced sensitivity. Our findings are consistent with published reports wherein anterior taste buds have higher sweet sensitivity while posterior taste buds are better tuned to bitter, and suggest β-catenin plays a greater role in renewal of anterior versus posterior taste buds.

β-catenin is required for taste bud cell renewal and behavioral taste perception in adult mice.

New evidence for fat as a primary taste quality.

Head and neck cancer patients receiving conventional repeated, low dose radiotherapy (fractionated IR) suffer from taste dysfunction that can persist for months and often years after treatment. To understand the mechanisms underlying functional taste loss, we established a fractionated IR mouse model to characterize how taste buds are affected. Following fractionated IR, we found as in our previous study using single dose IR, taste progenitor proliferation was reduced and progenitor cell number declined, leading to interruption in the supply of new taste receptor cells to taste buds. However, in contrast to a single dose of IR, we did not encounter increased progenitor cell death in response to fractionated IR. Instead, fractionated IR induced death of cells within taste buds. Overall, taste buds were smaller and fewer following fractionated IR, and contained fewer differentiated cells. In response to fractionated IR, expression of Wnt pathway genes, Ctnnb1, Tcf7, Lef1 and Lgr5 were reduced concomitantly with reduced progenitor proliferation. However, recovery of Wnt signaling post-IR lagged behind proliferative recovery. Overall, our data suggest carefully timed, local activation of Wnt/β-catenin signaling may mitigate radiation injury and/or speed recovery of taste cell renewal following fractionated IR.

Fractionated head and neck irradiation impacts taste progenitors, differentiated taste cells, and Wnt/β-catenin signaling in adult mice.

We have previously shown that the tyrosine kinase inhibitors (TKIs) dasatinib and imatinib can protect salivary glands from irradiation (IR) damage without impacting tumor therapy. However, how they induce this protection not unknown. Here we show that TKIs mediate radioprotection by increasing the repair of DNA double stranded breaks. DNA repair in IR-treated parotid cells, but not oral cancer cells, occurs more rapidly following pretreatment with imatinib or dasatinib, and is accompanied by faster formation of DNA damage-induced foci. Similar results were observed in the parotid glands of mice pretreated with imatinib prior to IR, suggesting that TKIs "prime" cells for DNA repair. Mechanistically, we observed that TKIs increased IR-induced activation of DNA-PK, but not ATM. Pretreatment of parotid cells with the DNA-PK inhibitor NU7441 reversed the increase in DNA repair induced by TKIs. Reporter assays specific for homologous recombination (HR) or non-homologous end joining (NHEJ) verified TKIs functionally regulate both DNA repair pathways. Moreover, TKIs also increased basal and IR-induced expression of genes associated with NHEJ (DNA ligase 4, Artemis, XLF) and HR (Rad50, Rad51 and BRCA1); depletion of DNA ligase 4 or BRCA1 reversed the increase in DNA repair mediated by TKIs. In addition, TKIs increased activation of the ERK survival pathway in parotid cells, and ERK was required for the increased survival of TKI treated cells. Our studies demonstrate a dual mechanism by which TKIs provide radioprotection of salivary gland tissues and support exploration of TKIs clinically in head and neck cancer patients undergoing IR therapy.

Tyrosine kinase inhibitors protect the salivary gland from radiation damage by increasing DNA double strand break repair.

Since the early 20 th century, progress in cancer therapies has significantly improved disease prognosis. Nonetheless, cancer treatments are often associated with side effects that can negatively affect patient wellbeing and disrupt the course of treatment. Among the main side effects, taste impairment is associated with depression, malnutrition and morbid weight loss. Although relatively common, taste disruption associated with cancer therapies remains poorly understood. Here, we review the current knowledge related to the molecular mechanisms underlying taste maintenance and disruption in the context of cancer therapies.

Article	Year
Taste bud cells of adult mice are responsive to Wnt/β-catenin signaling: implications for the renewal of mature taste cells. Genesis (New York, N.Y. : 2000)\| PubMed ID: 21328519	2011
β-Catenin Signaling Biases Multipotent Lingual Epithelial Progenitors to Differentiate and Acquire Specific Taste Cell Fates. PLoS genetics\| PubMed ID: 26020789	2015
Measurement of Behavioral Taste Responses in Mice: Two-Bottle Preference, Lickometer, and Conditioned Taste-Aversion Tests. Current protocols in mouse biology\| PubMed ID: 27906463	2016
β-catenin is required for taste bud cell renewal and behavioral taste perception in adult mice. PLoS genetics\| PubMed ID: 28846687	2017
New evidence for fat as a primary taste quality. Acta physiologica (Oxford, England)\| PubMed ID: 30588748	2019
Fractionated head and neck irradiation impacts taste progenitors, differentiated taste cells, and Wnt/β-catenin signaling in adult mice. Scientific reports\| PubMed ID: 31784592	2019
Tyrosine kinase inhibitors protect the salivary gland from radiation damage by increasing DNA double strand break repair. The Journal of biological chemistry\| PubMed ID: 33571522	2021
A mechanistic overview of taste bud maintenance and impairment in cancer therapies. Chemical senses\| PubMed ID: 33693542	2021

Dany Gaillard

Biography

JoVE Journal Publications

Other Publications