The cornea is exposed daily to a number of mechanical stresses including shear stress from tear film and blinking. Over time, these stressors can lead to changes in the extracellular matrix that alter corneal stiffness, cell-substrate structures, and the integrity of cell-cell junctions. We hypothesized that changes in tissue stiffness of the cornea with age may alter calcium signaling between cells after injury, and the downstream effects of this signaling on cellular motility and wound healing. Nanoindentation studies revealed that there were significant differences in the stiffness of the corneal epithelium and stroma between corneas of 9- and 27-week mice. These changes corresponded to differences in the timeline of wound healing and in cell signaling. Corneas from 9-week mice were fully healed within 24 h. However, the wounds on corneas from 27-week mice remained incompletely healed. Furthermore, in the 27-week cohort there was no detectable calcium signaling at the wound in either apical or basal corneal epithelial cells. This is in contrast to the young cohort, where there was elevated basal cell activity relative to background levels. Cell culture experiments were performed to assess the roles of P2Y2, P2X7, and pannexin-1 in cellular motility during wound healing. Inhibition of P2Y2, P2X7, or pannexin-1 all significantly reduce wound closure. However, the inhibitors all have different effects on the trajectories of individual migrating cells. Together, these findings suggest that there are several significant differences in the stiffness and signaling that underlie the decreased wound healing efficacy of the cornea in older mice.
Wnt signaling regulates cell fate decisions in diverse contexts during development, and loss of Wnt signaling in the cranial mesenchyme results in a robust and binary cell fate switch from cranial bone to ectopic cartilage. The Extracellular signal-regulated protein kinase 1 and 2 (ERK1/2) and Wnt signaling pathways are activated during calvarial osteoblast cell fate selection. Here, we test the hypothesis that ERK signaling is a mediator of Wnt-dependent cell fate decisions in the cranial mesenchyme. First, we show that loss of Erk1/2 in the cranial mesenchyme results in a diminished domain of osteoblast marker expression and increased expression of cartilage fate markers and ectopic cartilage formation in the frontal bone primordia. Second, we show that mesenchyme Wnt/β-catenin signaling and Wntless are required for ERK activation in calvarial osteoblasts. Third, we demonstrate that Wnt and ERK signaling pathways function together to repress SOX9 expression in mouse cranial mesenchyme. Our results demonstrate an interaction between the Wnt and ERK signaling pathways in regulating lineage selection in a subset of calvarial cells and provide new insights into Wnt-dependent cell fate decisions.
Wnt signaling regulates cell fate decisions in diverse contexts during development, and loss of Wnt signaling in the cranial mesenchyme results in a robust and binary cell fate switch from cranial bone to ectopic cartilage. The Extracellular signal-regulated protein kinase 1 and 2 (ERK1/2) and Wnt signaling pathways are activated during calvarial osteoblast cell fate selection. Here, we test the hypothesis that ERK signaling is a mediator of Wnt-dependent cell fate decisions in the cranial mesenchyme. First, we show that loss ofErk1/2 in the cranial mesenchyme results in a diminished domain of osteoblast marker expression and increased expression of cartilage fate markers and ectopic cartilage formation in the frontal bone primordia. Second, we show that mesenchyme Wnt/β-catenin signaling andWntlessare required for ERK activation in calvarial osteoblasts. Third, we demonstrate that Wnt and ERK signaling pathways function together to repress Sox9 expression in mouse cranial mesenchyme. Our results demonstrate a link between the Wnt and ERK signaling pathways in regulating lineage selection in a subset of calvarial cells and provide new insights into Wnt-dependent cell fate decisions.
Wnt signaling regulates cell fate decisions in diverse contexts during development and disease. In the mouse cranial mesenchyme (CM) Wnt signaling pathway components and reporters are spatially distributed during calvarial osteoblast fate selection. Within 24 hours, loss of Wnt signaling in the mouse embryonic CM results in a robust and binary cell fate switch, from calvarial bone to ectopic cartilage. The mechanism by which Wnt signaling regulates this cell fate switch is not clear. Extracellular signal‐regulated protein kinase 1 and 2 (ERK1/2) activation is required for bone‐cartilage cell fate decisions in long bone perichondrium, and we have demonstrated that ERK1/2 activation is present in calvarial osteoblasts. Here, we test the hypothesis that ERK signaling is a mediator of binary Wnt‐dependent bone‐cartilage cell fate decisions. First, we used three distinct Wnt signaling loss‐of‐function mouse models to demonstrate that Wnt signaling is required for activation of ERK signaling in calvarial bone progenitors. Second, we showed that loss of ERK signaling precedes formation of ectopic cartilage in CM‐Wnt signaling mutants and ERK activation is highly sensitive to levels of Wnt signaling within the cranial mesenchyme. Third, loss of Erk1/2 in the CM results in elevated levels of the master cartilage determinant, SOX9, by E13.5 and loss of calvarial bone by E16.5. These results demonstrate a link between the Wnt and ERK signaling pathways in regulating calvarial bone cell fate decisions in vivo. We propose a new model whereby reciprocal regulation of both canonical and non‐canonical Wnt pathways in the CM generates a gradient of Wnt signaling and utilizes ERKs to reinforce binary cell fate decisions in vivo. This offers a new opportunity for therapeutic targeting of Wnt signaling in craniofacial skeletal defects and disease.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.