Abstract
Purpose :
Investigate how factors in the microenvironment determine whether mesenchymal leader cells activated in response to wounding function as directors of regenerative repair or if their fate is altered to the myofibroblasts responsible for fibrosis.
Methods :
Our ex vivo mock cataract surgery culture model is used to study, concurrently, mechanisms of wound healing and fibrosis. Cultures are fixed, immunolabeled and imaged by high-resolution confocal microscopy, or extracted and analyzed by immunoblot. To analyze the coordinate impact of the extracellular matrix (ECM) environment and TGFβ signaling on leader cell fate cultures in defined ECM environments are exposed to inhibitors of TGFβR1.
Results :
There is a fine balance between outcomes of regenerative repair or fibrosis/scarring in response to wounding. At the wound edge created by the removal of the lens fiber cells, mesenchymal leader cells activated upon wounding direct collective migration of the lens epithelium across the endogenous lens basement membrane (BM) capsule to close the wound. These mesenchymal cells also migrate to the outside cut edges of the explant created to flatten the lens capsular bag onto the tissue culture platform where they direct the injured epithelium to migrate off the lens capsule onto and across the adjacent culture platform, the extracapsular zone (ECZ). Here leader cells are signaled to differentiate to αSMA+ myofibroblasts that persist to cause fibrosis. We now show that the leader cells in the ECZ, but not on the endogenous lens capsule, produce TN-C and FN-EDA, which they assemble as a provisional matrix on the substrate surface. We find that signaling by endogenous TGFβ produced by the cataract-surgery wounded cells is required for expression and organization of the FN-EDA ECM in the ECZ and for emergence of myofibroblasts in this environment. While exposure to the same injury-induced endogenous TGFβ the leader cells on the lens capsule are not induced to alter their matrix environment and inhibition of TGFβ signaling does not alter their role in wound repair.
Conclusions :
Determination of whether TGFβ expressed in response to wounding leads to fibrosis is dependent on the matrix microenvironment of the mesenchymal leader cells activated to direct the repair process.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.