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Published on: November 22, 2024
Down-regulation of Notch signaling during corneal epithelial proliferation
A R Djalilian1, A Namavari, A Ito
1Department of Ophthalmology, University of Illinois at Chicago, Chicago, IL 60612, USA. adjalili@uic.edu <adjalili@uic.edu>
Purpose:
We evaluated the expression and activation of Notch pathway genes in the adult human and murine corneal epithelium during proliferation.
Methods:
The expression of Notch pathway genes in the limbal and central human corneal epithelium was compared by reverse transcription polymerase chain reaction (RT-PCR). Their expression pattern was examined by immunofluorescence and in situ hybridization. The temporal expression of Notch1 during murine wound healing was assessed by RT-PCR. Notch activity was determined using western blot for the Notch intracellular domain (NotchIC). The expression of Hes1 was evaluated in cell culture.
Results:
The expression of Notch1 and Jagged1 was higher in the human limbal epithelium while the expression of Hes1 and Hes5 was higher in the central cornea. Expression of Notch1, Jagged1, and Hes1 was found predominantly in the basal and immediate suprabasal cells. During neonatal corneal development, NotchIC was detected in occasional cells at P10 while at P15 and P90, it was found in the basal and early suprabasal layers. NotchIC was found to be lower in the limbal compared to central corneal epithelium. The expression of Notch1 was lower at 24 h post-wounding but was completely restored in six days. The levels of NotchIC were decreased at 24 h post-wounding and after application of topical phorbol myristate. In vitro, the expression of Hes1 was higher in confluent cells maintained under high calcium conditions.
Conclusions:
The inverse correlation between Notch signaling and the proliferative status of the corneal epithelium is consistent with the idea that Notch plays a role in corneal epithelial differentiation.
Insights
Notch pathway genes are expressed in the corneal epithelium, with differing levels in limbal versus central areas. Notch signaling inversely correlates with corneal epithelial proliferation, suggesting a role in differentiation.
Area of Science:
- Ophthalmology
- Developmental Biology
- Cell Biology
Background:
- The corneal epithelium maintains transparency and undergoes continuous renewal.
- Notch signaling is a critical pathway regulating cell fate and differentiation in various tissues.
Purpose of the Study:
- To investigate the expression and activation of Notch pathway genes in the adult human and murine corneal epithelium during proliferation.
- To understand the role of Notch signaling in corneal epithelial homeostasis and differentiation.
Main Methods:
- Reverse transcription polymerase chain reaction (RT-PCR) to assess gene expression.
- Immunofluorescence and in situ hybridization to examine expression patterns.
- Western blot for Notch intracellular domain (NotchIC) to determine Notch activity.
- Murine wound healing models and in vitro cell culture experiments.
Main Results:
- Notch1 and Jagged1 expression was higher in human limbal epithelium, while Hes1 and Hes5 were higher in the central cornea.
- Notch pathway gene expression was localized to basal and suprabasal cells.
- Notch intracellular domain levels decreased during wound healing and with phorbol myristate application.
- Hes1 expression increased in confluent cells under high calcium conditions in vitro.
Conclusions:
- Notch signaling exhibits an inverse correlation with the proliferative status of the corneal epithelium.
- These findings support the role of Notch signaling in regulating corneal epithelial differentiation.
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