Sweet sensing in the ciliated cells of murine trachea

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 \u3b1-gustducin and PLC\u3b22, 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 \u3b1-gustducin and PLC\u3b22, 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

Molecules implicated in glucose homeostasis are differentially expressed in the trachea of lean and obese Zucker rats

Recent studies indicate that the processes mediated by the (T1R2/T1R3) glucose/sugar receptor of gustatory cells in the tongue, and hormones like leptin and ghrelin contribute to the regulation of glucose homeostasis. Altered plasma levels of leptin and ghrelin are associated with obesity both in humans and rodents. In the present study, we evaluated the ultrastructure of the mucosa, and the expression of molecules implicated in the regulation of glucose homeostasis (GLUT2, SGLT1, T1R3, ghrelin and its receptor) in the trachea of an animal model of obesity (Zucker rats). We found that the tracheal epithelium of obese animals was characterized by the presence of poorly differentiated cells. Ciliated and secretory cells were the cell lineages with greatest loss of differentiation. Severe epithelial alterations were associated with marked deposit of extracellular matrix in the lamina propria. The expression pattern of GLUT2 and SGLT1 glucose transporters was similar in the trachea of both the Zucker rat genotypes, whereas that of T1R3 was reduced in ciliated cells of obese rats. A different immunolocalization for ghrelin was also found in the trachea of obese rats. In conclusion, the tracheal morphological alterations in obese animals seem to compromise the expression of molecules involved in the homeostasis of glucose

Bitter tastants and artificial sweeteners activate a subset of epithelial cells in acute tissue slices of the rat trachea

Bitter and sweet receptors (T2Rs and T1Rs) are expressed in many extra-oral tissues including upper and lower airways. To investigate if bitter tastants and artificial sweeteners could activate physiological responses in tracheal epithelial cells we performed confocal Ca2+ imaging recordings on acute tracheal slices. We stimulated the cells with denatonium benzoate, a T2R agonist, and with the artificial sweeteners sucralose, saccharin and acesulfame-K. To test cell viability we measured responses to ATP. We found that 39% of the epithelial cells responding to ATP also responded to bitter stimulation with denatonium benzoate. Moreover, artificial sweeteners activated different percentages of the cells, ranging from 5% for sucralose to 26% for saccharin, and 27% for acesulfame-K. By using carbenoxolone, a gap junction blocker, we excluded that responses were mainly mediated by Ca2+ waves through cell-to-cell junctions. Pharmacological experiments showed that both denatonium and artificial sweeteners induced a PLC-mediated release of Ca2+ from internal stores. In addition, bitter tastants and artificial sweeteners activated a partially overlapping subpopulation of tracheal epithelial cells. Our results provide new evidence that a subset of ATP-responsive tracheal epithelial cells from rat are activated by both bitter tastants and artificial sweeteners

Hyperthermic superparamagnetic nanoparticles modulate adipocyte metabolism

Adipocytes are the principal cellular component in adipose tissue and their excessive hyperplasia or hypertrophy is actively involved in regulating physiologic and pathologic processes such as inflammation, cardiovascular disease, obesity and tumour. The main depot of energy in adipocytes is represented by lipid droplets, intracellular organelles that play fundamental roles in regulation of metabolic processes. An accumulation of such droplets could be a potential biomarker of disease caused by metabolic dysregulation. Recent studies have demonstrated that heat shock is associated with alteration in energy metabolism: the aim of this study is to modulate the energy metabolism of the adipocytes via controlled administration of thermal energy to reduce the number of lipid droplets. We have investigated the effect of controlled heating of adipocytes using an alternating magnetic field (AMF) on samples loaded with superparamagnetic nanoparticles (MNP) as heating agent

Innovative approach to safely induce controlled lipolysis by superparamagnetic iron oxide nanoparticles-mediated hyperthermic treatment

During last years, evidence has been provided on the involvement of overweight and obesity in the pathogenesis and aggravation of several life-threatening diseases. Here, we demonstrate that, under appropriate administration conditions, polyhedral iron oxide nanoparticles are efficiently and safely taken up by 3T3 cell line-derived adipocytes (3T3 adipocytes) in vitro. Since these nanoparticles proved to effectively produce heat when subjected to alternating magnetic field, 3T3 adipocytes were submitted to superparamagnetic iron oxide nanoparticles-mediated hyperthermia treatment (SMHT), with the aim of modulating their lipid content. Notably, the treatment resulted in a significant delipidation persisting for at least 24h, and in the absence of cell death, damage or dedifferentiation. Interestingly, transcript expression of adipose triglyceride lipase (ATGL), a key gene involved in canonical lipolysis, was not modulated upon SMHT, suggesting the involvement of a novel/alternative mechanism in the effective lipolysis observed. By applying the same experimental conditions successfully used for 3T3 adipocytes, SMHT was able to induce delipidation also in primary cultures of human adipose-derived adult stem cells. The success of this pioneering approach in vitro opens promising perspectives for the application of SMHT in vivo as an innovative safe and physiologically mild strategy against obesity, potentially useful in association with balanced diet and healthy lifestyle