The Expression Of The Notch Signaling Pathway In Human Corneal Endothelial Cells And The Preliminary Research On Its Participation In The Regulation Of EnMT

Objective: To investigate the expression of the receptors/ligands and thedownstream target genes named Hes1of the Notch signaling pathway in normalhuman corneal endothelial cells (hCECs), and explore the possibility of regulatinghuman corneal endothelial-to-mesenchymal transition(EnMT), then search a newmean to regulate corneal endothelial cells’differentiation.Methods: Take some corneal materials of normol donors and separate endothelialcell layer from descemet’s membrane, and fixed with4%paraformaldehyde.Wholemount Immunocytochemistry was used to examine receptors/ligands of theNotch signaling pathway, and the expression of downstream target genes namedHes1in human corneal endothelial cells (hCECs). Identify the Notch signalingpathways Whether exist in human corneal endothelial cells. corneal endothelial cellsin vitro culture was grouped into two parts,one with DAPT10uM and anotherwithout.Detect the Notch downstream target genes Hes1and α-SMA expression invitro culture cells by immunofluorescence and RT-PCR and explore therelationship between expression and activation process and corneal endothelial celldifferentiation.Result:(1)It was confirmed that except for Jagged1, the Notch signaling pathwayreceptors/ligands showed positive staining in normal human corneal endotheliumthrough immunofluorescence staining, and they distributed in different parts of thecells;Hes1expressed in the cell cytoplasm；(2)In the cultured human corneal endothelial cells, the cells cultured without DAPTappeared myofibroblast-like, immunofluorescence staining showed α-SMA(+);thecells cultured with10uM DAPT had a uniform hexagonal shape， immunofluorescence staining showed α-SMA(-)；(3) Immunofluorescence confirmed Hes1expressed in the cell nucleu and cytoplasmin DAPT (-) group of detected the stripe expression though RT-PCR；and it mainlyexpressed in the cell cytoplasm in10um DAPT (+) group;Hes1stripe were darker inDAPT (+) group than the DAPT (-) group in RT-PCR..Conclusion:(1)The Notch signaling pathways exist in normal human corneal endothelium and it isin the state of nonactivated;(2) The Notch signaling pathways are activated in cultured human corneal endothelialcells;(3)The Notch signaling pathways’ activation may be involved in the process ofpromoting human corneal endothelial-to-mesenchymal transition.