mTOR Regulates the Differentiation of Corneal Schwann cells into Myofibroblasts.

Abstract

Purpose : Sustained activation of the extracellular signal-regulated kinases 1 and 2 (ERK1/2) in corneal Schwann cells (cSCs) temporally drives corneal neurofibromas [J. Neurosci. Res. 95:1712-1729]. In the central nervous system, ERK-mediated effects on myelin formation activates the mammalian target of rapamycin (mTOR) in an AKT-independent manner [Journal of Neuroscience 37: 2931-2946]. This is the premise that mTOR activation can also be driven by ERK1/2 in the peripheral nervous system. Here we tested if deletion of mTOR in cSCs overexpressing ERK1/2 prevents their differentiation into α-SMA-expressing myofibroblasts and prevents corneal fibrosis.

Methods : We investigated two transgenic lines, CNPCreMek^DD/⁺ (overexpression of MEK1 in CNP-expressing SCs) and CNPCre;mTOR-KO/Mek^DD/⁺ (constitutive activation of MEK1 with simultaneous deletion of mTOR in SCs). Transgenic mouse corneas were analyzed for fibrosis by biomicroscopy at 6.0 months and tissue sections by immunohistochemistry (IHC). WT C57Bl/6 mice were also subjected to a penetrating corneal stromal injury. Corneal tissue sections were stained using the following antibodies: Glial Fibrillary Acidic Protein (GFAP), Adhesion molecule L1 (L1), Sry-related HMg-Box gene 10 (SOX-10) (markers of cSCs), α-Smooth Muscle Actin (myofibroblast marker), p-ERK1/2 (activation pathway), β-III tubulin (axonal marker).

Results : In the transgenic CNPCre;mTOR-KO/Mek^DD/⁺ mice, the deletion of mTOR in cSCs prevents the development of corneal fibrosis and neurofibromas that were observed in 6-month-old CNPCreMek^DD/⁺ mice, overriding sustained ERK1/2 activation and resulting in prevention of corneal fibrosis. This was confirmed by absence of α-SMA⁺ cells and showing of an intact corneal structure in CNPCre;mTOR-KO/Mek^DD/⁺ mice. In injured WT corneas, GFAP⁺/L1⁺ and GFAP⁺/SOX10⁺ cSCs temporally activated and sustained ERK1/2 expression and differentiated into α-SMA⁺ cells at the wound edge.

Conclusions : We discovered that cSCs have plasticity, and they actively respond to injury in a setting of deep corneal injury to differentiate into myofibroblasts. For the first time, we show that genetic deletion of mTOR in cSCs prevents their differentiation into myofibroblasts, and prevents ERK1/2-driven corneal fibrosis. This is of clinical relevance to how cSCs can contribute to fibrosis in disease, as well as, in severe corneal injuries that have poor prognosis and often lead to blindness.

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.