Sweet and bitter are two basic taste sensations perceivable thanks to taste receptor cells (TRCs) in the oral cavity. Interestingly, T1R3 sweet receptor and members of the bitter T2Rs family are also localized in different extra-oral tissues (e.g. airways system). In the context of this diffuse extra-oral chemosensory system, airways ciliated cells represent an interesting cellular population. Preliminary studies reported that their cilia exhibit bitter and sweet receptors and some elements of the chemoreception transduction pathway. To the best of our knowledge, no physiological data regarding ciliated cells sweet response in the airways have been reported in the literature. Therefore, to investigate the functional role of sweet receptors within the airways, we chose to explore the physiological behavior of murine tracheal ciliated cells stimulating them with sweet compounds by means of calcium-imaging technique. This method allows to follow the changes in intracellular calcium concentration thus showing possible calcium-mediated sweet responses. We planned to work with acute slices of mice tracheas, instead of using cells cultures, to better preserve the native conditions, avoiding excessive cells handling and artefacts introduction. We decided also to stimulate the ciliated cells with a bitter compound, yet investigated in cultured cells, to see the cellular behavior towards the two different stimulation qualities (sweet and bitter). In addition, we adopted immunohistochemical analysis to verify that the ciliated cells, used for the physiology experiments, retained the phenotypic expression of molecules of the taste transduction pathway normally expressed in native ciliated cells. Our experiments showed that some ciliated cells respond to sweet compounds. In particular, combining two different stimuli, glucose (sweet) first and then bitter (denatonium), 36% of the ciliated cells responded to both stimulations while 37% were responsive only to bitter, 2% only to sweet and 25% of the cells were unresponsive to both stimulations. Another important finding is that, when stimulating the cells with two artificial sweeteners (acesulfame K and sucralose), we observed that 31% of the cells responded to both the sweeteners, 24% only to acesulfame K and 5% only to sucralose. 40% of the cells were irresponsive to both the artificial sweeteners. Combining the artificial stimulus acesulfame K to the bitter one denatonium, 65% of the cells responded to both stimuli, 7% only to acesulfame K while 5% only to denatonium. Moreover, the immunohistochemical results confirmed the presence of α-gustducin and PLCβ2, two markers of taste signaling pathway, in the ciliated cells. The above described data are interesting and clearly demonstrate that the ciliated cells of murine trachea are able to perceive and respond to sweet compounds, natural and artificial, with possible implications in the glucose sensing mechanisms in the airways, especially in relationship with respiratory infections. Moreover, it is clear that the ciliated cells exhibit a different pattern of response suggesting their heterogeneity, according with the previous literature. These data might be important for considering this cellular population a new cellular model of extra-oral chemoreception investigation. Future research could unravel the roles of airways ciliated cells in health and pathological conditions with a possible therapeutic aim.Sweet and bitter are two basic taste sensations perceivable thanks to taste receptor cells (TRCs) in the oral cavity. Interestingly, T1R3 sweet receptor and members of the bitter T2Rs family are also localized in different extra-oral tissues (e.g. airways system). In the context of this diffuse extra-oral chemosensory system, airways ciliated cells represent an interesting cellular population. Preliminary studies reported that their cilia exhibit bitter and sweet receptors and some elements of the chemoreception transduction pathway. To the best of our knowledge, no physiological data regarding ciliated cells sweet response in the airways have been reported in the literature. Therefore, to investigate the functional role of sweet receptors within the airways, we chose to explore the physiological behavior of murine tracheal ciliated cells stimulating them with sweet compounds by means of calcium-imaging technique. This method allows to follow the changes in intracellular calcium concentration thus showing possible calcium-mediated sweet responses. We planned to work with acute slices of mice tracheas, instead of using cells cultures, to better preserve the native conditions, avoiding excessive cells handling and artefacts introduction. We decided also to stimulate the ciliated cells with a bitter compound, yet investigated in cultured cells, to see the cellular behavior towards the two different stimulation qualities (sweet and bitter). In addition, we adopted immunohistochemical analysis to verify that the ciliated cells, used for the physiology experiments, retained the phenotypic expression of molecules of the taste transduction pathway normally expressed in native ciliated cells. Our experiments showed that some ciliated cells respond to sweet compounds. In particular, combining two different stimuli, glucose (sweet) first and then bitter (denatonium), 36% of the ciliated cells responded to both stimulations while 37% were responsive only to bitter, 2% only to sweet and 25% of the cells were unresponsive to both stimulations. Another important finding is that, when stimulating the cells with two artificial sweeteners (acesulfame K and sucralose), we observed that 31% of the cells responded to both the sweeteners, 24% only to acesulfame K and 5% only to sucralose. 40% of the cells were irresponsive to both the artificial sweeteners. Combining the artificial stimulus acesulfame K to the bitter one denatonium, 65% of the cells responded to both stimuli, 7% only to acesulfame K while 5% only to denatonium. Moreover, the immunohistochemical results confirmed the presence of α-gustducin and PLCβ2, two markers of taste signaling pathway, in the ciliated cells. The above described data are interesting and clearly demonstrate that the ciliated cells of murine trachea are able to perceive and respond to sweet compounds, natural and artificial, with possible implications in the glucose sensing mechanisms in the airways, especially in relationship with respiratory infections. Moreover, it is clear that the ciliated cells exhibit a different pattern of response suggesting their heterogeneity, according with the previous literature. These data might be important for considering this cellular population a new cellular model of extra-oral chemoreception investigation. Future research could unravel the roles of airways ciliated cells in health and pathological conditions with a possible therapeutic aim.
Sweet sensing in the ciliated cells of murine trachea
LASCONI, CHIARA
2017-01-01
Abstract
Sweet and bitter are two basic taste sensations perceivable thanks to taste receptor cells (TRCs) in the oral cavity. Interestingly, T1R3 sweet receptor and members of the bitter T2Rs family are also localized in different extra-oral tissues (e.g. airways system). In the context of this diffuse extra-oral chemosensory system, airways ciliated cells represent an interesting cellular population. Preliminary studies reported that their cilia exhibit bitter and sweet receptors and some elements of the chemoreception transduction pathway. To the best of our knowledge, no physiological data regarding ciliated cells sweet response in the airways have been reported in the literature. Therefore, to investigate the functional role of sweet receptors within the airways, we chose to explore the physiological behavior of murine tracheal ciliated cells stimulating them with sweet compounds by means of calcium-imaging technique. This method allows to follow the changes in intracellular calcium concentration thus showing possible calcium-mediated sweet responses. We planned to work with acute slices of mice tracheas, instead of using cells cultures, to better preserve the native conditions, avoiding excessive cells handling and artefacts introduction. We decided also to stimulate the ciliated cells with a bitter compound, yet investigated in cultured cells, to see the cellular behavior towards the two different stimulation qualities (sweet and bitter). In addition, we adopted immunohistochemical analysis to verify that the ciliated cells, used for the physiology experiments, retained the phenotypic expression of molecules of the taste transduction pathway normally expressed in native ciliated cells. Our experiments showed that some ciliated cells respond to sweet compounds. In particular, combining two different stimuli, glucose (sweet) first and then bitter (denatonium), 36% of the ciliated cells responded to both stimulations while 37% were responsive only to bitter, 2% only to sweet and 25% of the cells were unresponsive to both stimulations. Another important finding is that, when stimulating the cells with two artificial sweeteners (acesulfame K and sucralose), we observed that 31% of the cells responded to both the sweeteners, 24% only to acesulfame K and 5% only to sucralose. 40% of the cells were irresponsive to both the artificial sweeteners. Combining the artificial stimulus acesulfame K to the bitter one denatonium, 65% of the cells responded to both stimuli, 7% only to acesulfame K while 5% only to denatonium. Moreover, the immunohistochemical results confirmed the presence of α-gustducin and PLCβ2, two markers of taste signaling pathway, in the ciliated cells. The above described data are interesting and clearly demonstrate that the ciliated cells of murine trachea are able to perceive and respond to sweet compounds, natural and artificial, with possible implications in the glucose sensing mechanisms in the airways, especially in relationship with respiratory infections. Moreover, it is clear that the ciliated cells exhibit a different pattern of response suggesting their heterogeneity, according with the previous literature. These data might be important for considering this cellular population a new cellular model of extra-oral chemoreception investigation. Future research could unravel the roles of airways ciliated cells in health and pathological conditions with a possible therapeutic aim.Sweet and bitter are two basic taste sensations perceivable thanks to taste receptor cells (TRCs) in the oral cavity. Interestingly, T1R3 sweet receptor and members of the bitter T2Rs family are also localized in different extra-oral tissues (e.g. airways system). In the context of this diffuse extra-oral chemosensory system, airways ciliated cells represent an interesting cellular population. Preliminary studies reported that their cilia exhibit bitter and sweet receptors and some elements of the chemoreception transduction pathway. To the best of our knowledge, no physiological data regarding ciliated cells sweet response in the airways have been reported in the literature. Therefore, to investigate the functional role of sweet receptors within the airways, we chose to explore the physiological behavior of murine tracheal ciliated cells stimulating them with sweet compounds by means of calcium-imaging technique. This method allows to follow the changes in intracellular calcium concentration thus showing possible calcium-mediated sweet responses. We planned to work with acute slices of mice tracheas, instead of using cells cultures, to better preserve the native conditions, avoiding excessive cells handling and artefacts introduction. We decided also to stimulate the ciliated cells with a bitter compound, yet investigated in cultured cells, to see the cellular behavior towards the two different stimulation qualities (sweet and bitter). In addition, we adopted immunohistochemical analysis to verify that the ciliated cells, used for the physiology experiments, retained the phenotypic expression of molecules of the taste transduction pathway normally expressed in native ciliated cells. Our experiments showed that some ciliated cells respond to sweet compounds. In particular, combining two different stimuli, glucose (sweet) first and then bitter (denatonium), 36% of the ciliated cells responded to both stimulations while 37% were responsive only to bitter, 2% only to sweet and 25% of the cells were unresponsive to both stimulations. Another important finding is that, when stimulating the cells with two artificial sweeteners (acesulfame K and sucralose), we observed that 31% of the cells responded to both the sweeteners, 24% only to acesulfame K and 5% only to sucralose. 40% of the cells were irresponsive to both the artificial sweeteners. Combining the artificial stimulus acesulfame K to the bitter one denatonium, 65% of the cells responded to both stimuli, 7% only to acesulfame K while 5% only to denatonium. Moreover, the immunohistochemical results confirmed the presence of α-gustducin and PLCβ2, two markers of taste signaling pathway, in the ciliated cells. The above described data are interesting and clearly demonstrate that the ciliated cells of murine trachea are able to perceive and respond to sweet compounds, natural and artificial, with possible implications in the glucose sensing mechanisms in the airways, especially in relationship with respiratory infections. Moreover, it is clear that the ciliated cells exhibit a different pattern of response suggesting their heterogeneity, according with the previous literature. These data might be important for considering this cellular population a new cellular model of extra-oral chemoreception investigation. Future research could unravel the roles of airways ciliated cells in health and pathological conditions with a possible therapeutic aim.
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SWEET SENSING IN THE CILIATED CELLS OF MURINE TRACHEA_ChiaraLasconi.pdf
Open Access dal 01/12/2018
Descrizione: Doctoral Thesis
Tipologia:
Tesi di dottorato
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2.31 MB
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