Il "metabolismo" del sapore

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Taste buds and solitary chemoreceptor cells (SCCs) in the oral cavity, larynx and nasopharynx must both monitor potential foods as well as guard against intake of toxic materials into the alimentary and respiratory passageways. All of these sensory cells utilize TR receptors to initiate a signaling cascade, which utilizes TrpM5 as a requisite downstream element. Accordingly, TrpM5 is a useful marker of likely chemoreceptor cells whether they are in taste buds or SCCs. To understand the role of these cells, we set out to determine which taste signal transduction elements they express. For sweet and umami, the taste signaling cascade is initiated in the taste receptor cells (TRCs) by activation T1R receptors (good markers of chemoreceptors responsive to appetitive qualities), and for bitter and sour, the taste signalling cascade is initiated in the TRCs by activation T2R receptors and PKD2L1/2 candidate receptors (good markers of chemoreceptors responsive against to intake of toxic compounds). The downstream elements for sweet, umami and bitter taste include a- gustducin, PLCb2, IP3R3 and Trpm5. I utilized in situ hybridization, RT-PCR, two lines of transgenic mice, TrpM5-GFP and T1R3-GFP, immunoblotting and immunohistochemistry, to assess the distribution within the oropharynx and respiratory tract, of solitary and taste buds chemoreceptor cells expressing these different markers. TrpM5-GFP receptor cells occur within all taste buds, including those on the epiglottis and larynx. In addition, numerous, scattered TrpM5- GFP SCCs lie within the nasopharynx, larynx and trachea. a-gustducin, PLCb2 and Trpm5 are expressed in taste buds cells of the oropharynx as well as SCCs throughout the epiglottis, oropharynx, larynx and trachea of mice, rat and bovine. By RT-PCR studies on rats, T1Rs receptors were localized predominantly in lingual, palatal and laryngeal tissue. Comparatively, T1R3-GFP cells occur predominantly in lingual and palatal taste buds, few T1R3-GFP cells are present in laryngeal taste buds and essentially no T1R3-positive SCCs are present within the airways. These results from transgenic mice T1R3-GFP are largely confirmed by in situ hybridization studies on rat T1Rs: laryngeal taste buds exhibit low expression of T1Rs compared to lingual and palatal taste buds, and no T1Rs-positive SCCs are present within the airways. Thus, it seems that laryngeal taste buds and SCCs do not display molecular features associated with detection of sweet or umami compounds. By RT-PCR studies on rats, bitter and sour receptors occur either in lingual, palatal, nasal and laryngeal tissue, but only T2Rs were present in trachea and bronchi. Thus, it seems that T2Rs play an important role as guard against intake of toxic materials and in the protective chemoreflex of respiratory passageways. These findings are consistent with electrophysiological studies on the superior laryngeal nerve (188), showing minimal responsiveness to sweet and umami substances, but large responses to bitter and sour-tasting stimuli. Moreover, by gene microarray experiments, I demonstrate that the action of AHLs, quorum sensing gram negative bacterial pheromones, is not only limited to activate conspecific bacteria, but also modify innate defence mechanisms of the airway. AHLs induce an early mucosal response related to the length of their acylic chain. The chondrioma of ciliate and secretory cells is the main cytological target of AHL action. However, the airways present a population of cells, the SCCs, that resists to the AHL-induced lesion. An important question is if some of the responses detected after treatment with AHL are part of a defensive mechanism operated by the mucosa. This might be the case of the secretory response elicited by C12-HSL. It is possible to speculate that the increased secretion could dilute the bacterial infochemicals, blocking the quorum sensing based strategy. In the response of the airway to AHL, the most interesting new finding is the high resistance of the SCCs, which do not display the mitochondrial alterations detectable in ciliate or secretory elements. The SCCs are a part of a diffuse chemosensory system (DCS) that control the whole airway, and are located in ideal positions to evaluate the mucus composition into the airways or the presence of infochemicals generated by resident micro-organisms. Recent studies demonstrated that SCCs are sentinels active against irritants (105). These data suggest that SCCs could have a role in bacterial infections, because ultrastructural analysis demonstrated that they are activated by AHL. Considering the chemoreceptorial capability of SCCs, it seems probable that they have a role in innate mechanism of defences that operates in presence of bacterial infochemicals. In conclusion, if the main taste system plays a role in food sampling, the DCS seems rather to be an alarm system. In the respiratory apparatus it could provide information about the luminal microenvironment. The DCS is probably the afferent branch of intrinsic mechanisms, which might involve gland secretion and muscle contraction. Local reflexes could be generated by interaction between the DCS and surrounding elements. The transmission of information from the DCS to the central nervous system is possible, but appears to be limited to the chemosensory cells innervated by afferent axons. Such cells seem to be a minority and are mainly localized in the larynx. For the elements not innervated, a paracrine action seems probable. Finally, the DCS seems to be a potential new drug target because several elements indicate that information obtained by this system induces secretory reflexes. Therefore, modulation of the respiratory apparatus by substances acting on their chemoreceptors could be important in the treatment of diseases such as cystic fibrosis and asthma, and might open a new frontier in drug discovery. Finally, by RT-PCR studies on humans and rats, I showed that a new form of P5CDh enzyme, involved in proline catabolism, exist in rat liver. LRRP Ba1-651 is probably a Delta-1-pyrroline-5- carboxylate dehydrogenase, fused with the extramembrane receptor domain of sweet receptor T1R2, activated by changes in the concentration of sweet molecules. After partial epatectomy, constitutive P5CDh and inducible P5CDh-T1R2 co-exist in rat liver tissue. This finding demonstrates as part of a taste receptor sequence, the receptor domain, could be used for a completely different function and in no taste organs. Type II hyperprolinemia (HPII) is an inborn error of metabolism due to a deficiency of P5CDh enzyme, thus it will be very interesting and important carry out further studies that contribute to understand the biological significance of this P5CDh-ANF enzyme, especially for the potential applications in the treatment of HPII, other liver diseases and in liver transplantation.