While the involvement of thermosensitive transient receptor potential channels (TRPs) in dry eye disease (DED) has been known for years, their expression in the meibomian gland (MG) has never been investigated. This study aims to show their expression and involvement in the lipogenesis of the MG, providing a possible new drug target in the treatment of DED. Our RT-PCR, Western blot and immunofluorescence analysis showed the expression of TRPV1, TRPV3, TRPV4 and TRPM8 in the MG at the gene and the protein level. RT-PCR also showed gene expression of TRPV2 but not TRPA1. Calcium imaging and planar patch-clamping performed on an immortalized human meibomian gland epithelial cell line (hMGECs) demonstrated increasing whole-cell currents after the application of capsaicin (TRPV1) or icilin (TRPM8). Decreasing whole-cell currents could be registered after the application of AMG9810 (TRPV1) or AMTB (TRPM8). Oil red O staining on hMGECs showed an increase in lipid expression after TRPV1 activation and a decrease after TRPM8 activation. We conclude that thermo-TRPs are expressed at the gene and the protein level in MGs. Moreover, TRPV1 and TRPM8\'s functional expression and their contribution to their lipid expression could be demonstrated. Therefore, TRPs are potential drug targets and their clinical relevance in the therapy of meibomian gland dysfunction requires further investigation.

Temperature-sensitive ion channels, such as those from the TRP family (thermo-TRPs) present in all animal cells, serve to perceive heat and cold sensations. A considerable number of protein structures have been reported for these ion channels, providing a solid basis for revealing their structure-function relationship. Previous functional studies suggest that the thermosensing ability of TRP channels is primarily determined by the properties of their cytosolic domain. Despite their importance in sensing and wide interests in the development of suitable therapeutics, the precise mechanisms underlying acute and steep temperature-mediated channel gating remain enigmatic. Here, we propose a model in which the thermo-TRP channels directly sense external temperature through the formation and dissociation of metastable cytoplasmic domains. An open-close bistable system is described in the framework of equilibrium thermodynamics, and the middle-point temperature T½ similar to the V½ parameter for a voltage-gating channel is defined. Based on the relationship between channel opening probability and temperature, we estimate the change in entropy and enthalpy during the conformational change for a typical thermosensitive channel. Our model is able to accurately reproduce the steep activation phase in experimentally determined thermal-channel opening curves, and thus should greatly facilitate future experimental verification.

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It is believed that the biological systems perceiving temperature and light daily cycles were subjected to the simultaneous selective pressures, which resulted in their co-evolutionary association. We investigated the influence of 1h 33°C heat shock on the expression of clock and heat shock protein genes, as well as the role of the thermo-TRP channel, TRPV1, in ZEM-2S cells of the teleost Danio rerio, in constant dark (DD) or light-dark cycles (LD). After heat shock, we observed an acute increase of hsp90 aa1 levels in both DD and LD conditions. Interestingly, the expression of hsp90 aa1 was two-fold lower in LD than in DD, what suggests an antagonistic effect of white light on heat shock action. Regarding clock genes, no effect was found in cells subjected to the heat shock in DD. When cells were kept in LD, the expression of per1, per2, cry1a, and cry1b increased in response to heat shock, indicating that heat shock only affects clock core of LD-synchronized ZEM-2S cells. We then evaluated whether TRPV1 played a role in heat-mediated hsp90 aa1 and per2 responses: hsp90 aa1 increase was unaffected whereas per2 increase was partially blocked by TRPV1 inhibitor, demonstrating the channel participation in clock gene regulation by heat shock. Taken together, our results open a novel investigative perspective regarding the relationship between temperature and clock genes, placing a new player in the regulation of this phenomenon: the TRPV1 channel.