Abstract
Purpose :
Hyperevaporation of the tear film is the leading cause of Dry Eye Disease (DED), most commonly caused by a dysfunction of the Meibomian Gland (MG), which provides the lipid layer of the tear film. So far, there are no gold standards and only few pharmacological options for the treatment of DED. The cation permeable transient receptor potential (TRP) channels are new pharmacological targets in many different fields. They play an important role in the calcium regulation of cells and take part in several signaling pathways like nociception or thermosensation, pro-inflammatory cytokine release as well as apoptosis, cell growth and migration. Therefore, this study was undertaken to investigate the properties of temperature-sensitive TRPs in the MG.
Methods :
First, the expression of thermosensitive TRP channels was analysed in human and murine MG tissue and in an immortalized human meibomian gland epithelial cell line (hMGECs), by RT-PCR, western blot and immunofluorescence. Furthermore, the regulation of TRPV1 and TRPV4 gene expression in stimulated hMGECs was analysed by qPCR. Planar patch-clamping and fluorescence calcium imaging (fura-2) were performed for TRPV1 and TRPM8 to verify the functional activity of these channels in hMGECs. Oil Red O staining was performed on hMGECs to investigate a possible link between TRPV1 and TRPM8 activation and lipid synthesis
Results :
RT-PCR, Western blot, and immunofluorescence showed the expression of TRPV1, TRPV3, TRPV4, and TRPM8 channels in the MG at the gene and protein levels. RT-PCR also showed the expression of TRPV2 but not TRPA1. Treatment of hMGECs with media of different osmolarity showed a significant increase in the osmo-sensitive TRPV1 and TRPV4 gene expression between hypotonic and hypertonic stimulation. Treatment with proinflammatory mediators TNFα and IL-1ß did not result in significant changes in TRP expression. Planar patch-clamping and fura-2 calcium assays confirmed the functional activity of TRPV1 and TRPM8 in hMGECs.
Conclusions :
We demonstrated for the first time that thermosensitive TRP channels are expressed and functionally active in the human MG. They could be a promising new target for the treatment of DED.
This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.