Chenying Guo

Ephrin-A3 Suppresses Wnt Signaling to Control Retinal Stem Cell Potency†‡§

The ciliary epithelium (CE) of adult mammals has been reported to provide a source of retinal stem cells (RSCs) that can give rise to all retinal cell types in vitro. A recent study, however, suggests that CE‐derived cells possess properties of pigmented ciliary epithelial cells and display little neurogenic potential. Here we show that the neurogenic potential of CE‐derived cells is negatively regulated by ephrin‐A3, which is upregulated in the CE of postnatal mice and presents a strong prohibitory niche for adult RSCs. Addition of ephrin‐A3 inhibits proliferation of CE‐derived RSCs and increases pigment 349 cell 359. In contrast, absence of ephrin‐A3 promotes proliferation and increases expression of neural progenitor cell markers and photoreceptor progeny. The negative effects of ephrin‐A3 on CE‐derived RSCs are mediated through activation of an EphA4 receptor and suppression of Wnt3a/β‐catenin signaling. Together, our data suggest that CE‐derived RSCs contain the intrinsic machinery to generate photoreceptors and other retinal neurons, while the CE of adult mice expresses negative regulators that prohibit the proliferation and neural differentiation of RSCs. Manipulating ephrin and Wnt/β‐catenin signaling may, thus, represent a viable approach in activating the endogenous neurogenic potential of CE‐derived RSCs for treating photoreceptor damage and retinal degenerative disorders. STEM CELLS2013;31:349–359

Postnatal onset of retinal degeneration by loss of embryonic Ezh2 repression of Six1.

Some adult-onset disorders may be linked to dysregulated embryonic development, yet the mechanisms underlying this association remain poorly understood. Congenital retinal degenerative diseases are blinding disorders characterized by postnatal degeneration of photoreceptors, and affect nearly 2 million individuals worldwide, but ∼50% do not have a known mutation, implicating contributions of epigenetic factors. We found that embryonic deletion of the histone methyltransferase (HMT) Ezh2 from all retinal progenitors resulted in progressive photoreceptor degeneration throughout postnatal life, via derepression of fetal expression of Six1 and its targets. Forced expression of Six1 in the postnatal retina was sufficient to induce photoreceptor degeneration. Ezh2, although enriched in the embryonic retina, was not present in the mature retina; these data reveal an Ezh2-mediated feed-forward pathway that is required for maintaining photoreceptor homeostasis in the adult and suggest novel targets for retinal degeneration therapy.

IGFBPL1 Regulates Axon Growth through IGF-1-mediated Signaling Cascades

Activation of axonal growth program is a critical step in successful optic nerve regeneration following injury. Yet the molecular mechanisms that orchestrate this developmental transition are not fully understood. Here we identified a novel regulator, insulin-like growth factor binding protein-like 1 (IGFBPL1), for the growth of retinal ganglion cell (RGC) axons. Expression of IGFBPL1 correlates with RGC axon growth in development, and acute knockdown of IGFBPL1 with shRNA or IGFBPL1 knockout in vivo impaired RGC axon growth. In contrast, administration of IGFBPL1 promoted axon growth. Moreover, IGFBPL1 bound to insulin-like growth factor 1 (IGF-1) and subsequently induced calcium signaling and mammalian target of rapamycin (mTOR) phosphorylation to stimulate axon elongation. Blockage of IGF-1 signaling abolished IGFBPL1-mediated axon growth, and vice versa, IGF-1 required the presence of IGFBPL1 to promote RGC axon growth. These data reveal a novel element in the control of RGC axon growth and suggest an unknown signaling loop in the regulation of the pleiotropic functions of IGF-1. They suggest new therapeutic target for promoting optic nerve and axon regeneration and repair of the central nervous system.

Ezh2 does not mediate retinal ganglion cell homeostasis or their susceptibility to injury

Epigenetic predisposition is thought to critically contribute to adult-onset disorders, such as retinal neurodegeneration. The histone methyltransferase, enhancer of zeste homolog 2 (Ezh2), is transiently expressed in the perinatal retina, particularly enriched in retinal ganglion cells (RGCs). We previously showed that embryonic deletion of Ezh2 from retinal progenitors led to progressive photoreceptor degeneration throughout life, demonstrating a role for embryonic predisposition of Ezh2-mediated repressive mark in maintaining the survival and function of photoreceptors in the adult. Enrichment of Ezh2 in RGCs leads to the question if Ezh2 also mediates gene expression and function in postnatal RGCs, and if its deficiency changes RGC susceptibility to cell death under injury or disease in the adult. To test this, we generated mice carrying targeted deletion of Ezh2 from RGC progenitors driven by Math5-Cre (mKO). mKO mice showed no detectable defect in RGC development, survival, or cell homeostasis as determined by physiological analysis, live imaging, histology, and immunohistochemistry. Moreover, RGCs of Ezh2 deficient mice revealed similar susceptibility against glaucomatous and acute optic nerve trauma-induced neurodegeneration compared to littermate floxed or wild-type control mice. In agreement with the above findings, analysis of RNA sequencing of RGCs purified from Ezh2 deficient mice revealed few gene changes that were related to RGC development, survival and function. These results, together with our previous report, support a cell lineage-specific mechanism of Ezh2-mediated gene repression, especially those critically involved in cellular function and homeostasis.

Chapter 20 – Mobilizing Endogenous Stem Cells for Retinal Repair

Irreversible vision loss is often caused by the loss of retinal neurons, and one reason that retinal degenerative diseases are devastating is that, once retinal neurons are lost, they do not grow back. Stem cell-based neuron replacement strategy for retinal degenerative diseases are leading the way in clinical trials of transplantation therapy, and the exciting findings in both human and animal models point to the possibility of restoring vision through a cell replacement approach. Presumably, a less invasive method of retinal regeneration by mobilizing endogenous stem cells to replace those lost is highly desirable and promising for restoring vision. However, many obstacles remain, including mobilization of endogenous stem cells for restoration of lost cells, directed differentiation of progenitor cells, and establishment of functional connectivity in order to regain vision. The field of retinal self-repair is progressing at a rapid pace and encouraging outcomes.

The Intrinsic Determinants of Axon Regeneration in the Central Nervous System

Abstract Neurons in the mammalian central nervous system (CNS) undergo a developmental loss in their ability to grow axons as they mature, which is a critical contributing factor to the failure of nerve regeneration and repair after injury. This growth failure can be attributed, at least in part, to the induction of molecular programs preventing cellular overgrowth and termination of axonal growth on maturation. Key intracellular signals and transcription factors, including Bcl-2, cyclic adenine monophospate (cAMP), mammalian target of rapamycin (mTOR), and Kruppel-like transcription factors (KLFs), have been identified as playing central roles in this process. This chapter will provide an overview introducing the functions of these intracellular signals in their regulation of CNS axon regeneration.