Abstract
Following injury, corneal stromal keratocytes transform into repair-phenotype of activated stromal fibroblasts (SFs) and participate in wound repair. Simultaneously, ongoing bi-directional communications between corneal stromal-epithelial cells also play a vital role in mediating the process of wound healing. Factors produced by stromal cells are known to induce proliferation, differentiation, and motility of corneal epithelial cells, which are also subsequently the main processes that occur during wound healing. In this context, the present study aims to investigate the effect of SFs conditioned medium (SFCM) on corneal epithelial cell function along with substance P (SP). Antibody microarrays were employed to profile differentially expressed cell surface markers and cytokines in the presence of SFCM and SP. Antibody microarray data revealed enhanced expression of the ITGB1 in corneal epithelial cells following stimulation with SP whereas SFCM induced abundant expression of IL-8, ITGB1, PD1L1, PECA1, IL-15, BDNF, ICAM1, CD8A, CD44 and NTF4. All these proteins have either direct or indirect roles in epithelial cell growth, movement and adhesion related signaling cascades during tissue regeneration. We also observed activation of MAPK signaling pathway along with increased expression of focal adhesion kinase (FAK), paxillin, vimentin, β-catenin and vasodilator-stimulated phosphoprotein (VASP) phosphorylation. Additionally, epithelial-to-mesenchymal transition (EMT) regulating transcription factors Slug and ZEB1 expression were enhanced in the presence of SFCM. SP enriched the expression of integrin subunits α4, α5, αV, β1 and β3 whereas SFCM increased α4, α5, αV, β1 and β5 integrin subunits. We also observed increased expression of Serpin E1 following SP and SFCM treatment. Wound healing scratch assay revealed enhanced migration of epithelial cells following the addition of SFCM. Taken together, we conclude that SFCM-mediated sustained activation of ZEB1, Slug in combination with upregulated migration-associated integrins and ERK (Extracellular signal-regulated kinase)-FAK-paxillin axis, may lead to induce type 2 EMT-like changes during corneal epithelial wound healing.
Highlights
Corneal stromal keratocytes are neural crest-derived, quiescent, mesenchymal cells of the stroma and are capable of transforming into repair-phenotype of activated stromal fibroblasts (SFs) following injury [1]
Stromal keratocytes produce hepatocyte and keratinocyte growth factors that are in turn induce proliferation, differentiation, and motility of corneal epithelial cells by paracrine mechanisms, which are subsequently the main processes that occur during corneal tissue repair [3,4,5]
The presence of substance P (SP) in incubation medium suppressed the level of interferon-α1 (IFNA1) and potently stimulated the expression of integrin β1 (ITGB1) in comparison with control cells (Figure 1A; Table 1)
Summary
Corneal stromal keratocytes are neural crest-derived, quiescent, mesenchymal cells of the stroma and are capable of transforming into repair-phenotype of activated stromal fibroblasts (SFs) following injury [1]. Stromal keratocytes produce hepatocyte and keratinocyte growth factors that are in turn induce proliferation, differentiation, and motility of corneal epithelial cells by paracrine mechanisms, which are subsequently the main processes that occur during corneal tissue repair [3,4,5]. Conditioned media (CM) collected from SFs (SFCM) has been shown to contain epidermal, basic fibroblast growth factors and contribute to the activation of corneal epithelial cells [3,7,8]. Activated SFs contribute to nerve regeneration following injury. Proteomic analysis of the SFCM from activated SFs reported more than 130 proteins which are predicted to regulate focal adhesion, nerve and tissue regeneration [9]
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
