Abstract
Purpose :
The transisiton zone has been recently idenified as a stem cell niche for early corneal endothelial progenitors (CEP). It is a thin layer of tissue situated adjacent to the peripheral endothelium and the trabecular meshwork. Detailed analysis of the area has shown has shown expression of Sox2, Lgr5, CD34, Pitx2 in the inner TZ. In addition Lgr5 positive cells have been found to be streaming from the TZ into PE. The basement membrane (BM) is know to have an effect on stem cell differentiation. In this study we aimed to analyse the compositon of the BM of the TZ to the PE and central endothelium (CE).
Methods :
Descemet membrane from 6 donor corneal buttons were stripped from the trabecular meshwork to central corneal endothelium. They were placed in DNase/RNase to facilitate cell lysis. Following reconsitution the samples were analyzed using LC-MS/MS. Data was exported and analyzed using Metaboanlayst. Comparison amongst groups was done with Anova. PLS-DA plot, heatmap, dendorogram and volcano plots were generated using cut of values of >2 fold and <0.5 change between samples. Network maps were plotted with Metscape.
Results :
There were 90 proteins upregulated in the TZ compared to CE, but only 4 significant. There were 283 proteins down regulated in the TZ compared to the CE of which, 134 were significant. PLS-DA showed good seggregation of all 3 groups. The gene ontology showed 4 major groups of proteins affected; extracellular matrix, antigen binding, complement and growth factor/cytokine binding. There were 3 patterns of protein expression noted; i. Gradient reduction in protein from the CE to PE to TZ e.g. Fibroblast growth factor 2, Serine protease HTRA1, Collagen 8, Fibulin, Vitrin ii. Highest in TZ e.g. Matrillin, Mimecan, Collagen 1, Integrin a6, iii. Highest in PE e.g. ADP-ribosylation factor 4, Aggrecan.
Conclusions :
Significant differences were found in the basement membrane protein. structure of the TZ compared to PE and CE. The changes in the structural ECM components would indicate a less stiff BM in the TZ region.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.