Generation of Corneal Epithelial Cells from Induced Pluripotent Stem Cells Derived from Human Dermal Fibroblast and Corneal Limbal Epithelium

Abstract

Induced pluripotent stem (iPS) cells can be established from somatic cells. However, there is currently no established strategy to generate corneal epithelial cells from iPS cells. In this study, we investigated whether corneal epithelial cells could be differentiated from iPS cells. We tested 2 distinct sources: human adult dermal fibroblast (HDF)-derived iPS cells (253G1) and human adult corneal limbal epithelial cells (HLEC)-derived iPS cells (L1B41). We first established iPS cells from HLEC by introducing the Yamanaka 4 factors. Corneal epithelial cells were successfully induced from the iPS cells by the stromal cell-derived inducing activity (SDIA) differentiation method, as Pax6+/K12+ corneal epithelial colonies were observed after prolonged differentiation culture (12 weeks or later) in both the L1B41 and 253G1 iPS cells following retinal pigment epithelial and lens cell induction. Interestingly, the corneal epithelial differentiation efficiency was higher in L1B41 than in 253G1. DNA methylation analysis revealed that a small proportion of differentially methylated regions still existed between L1B41 and 253G1 iPS cells even though no significant difference in methylation status was detected in the specific corneal epithelium-related genes such as K12, K3, and Pax6. The present study is the first to demonstrate a strategy for corneal epithelial cell differentiation from human iPS cells, and further suggests that the epigenomic status is associated with the propensity of iPS cells to differentiate into corneal epithelial cells.

Generating minicorneal organoids from human induced pluripotent stem cells

“…Cell replacement therapy using autologous or allogeneic adult limbal grafts has been the standard treatment for patients with severe limbal stem cell deficiency (LSCD) (Rama et al, 2010;Sangwan et al, 2011;Basu et al, 2016). However, in the case of bilateral epithelial defects and for the treatment of conditions affecting the stromal and endothelial cell layers, alternative stem cell sources such as embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) have been explored with a view to generating the various corneal cell types (Ahmad et al, 2007;Shalom-Feuerstein et al, 2012;Hayashi et al, 2012;Sareen et al, 2014;Mikhailova et al, 2014;Chan et al, 2013;Zhang et al, 2014;Chen et al, 2015;McCabe et al, 2015).…”

Wound Repair and Regeneration

“…Given this, the authors suggest that adipose derived pericytes present a more preferable option for transplantation than BM-MSCs as they can be isolated 205 …”

Regenerative Therapies for the Ocular Surface

“…Another group has reported the presence of stromal stem cells at the limbal niche and demonstrated the usefulness of these autologous, mesenchymal-like stem cells in the treatment of corneal stromal scars (Li et al 2012 ;Basu et al 2014 ). Apart from various adult stem cell sources, pluripotent stem cell (PSCs) sources such as the human embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) have been tested for their ability of to generate corneal epithelial, endothelial and stromal cells (Ahmad et al 2007 ;Yoshida et al 2011 ;Hayashi et al 2012 ;Chan et al 2013 ;Sareen et al 2014 ;Mikhailova et al 2014 ;Zhang et al 2014 ). Spontaneous differentiation of PSCs results in the generation of heterogenous cell types representing all three germ layers of the body.…”

“…Also, the epigenetic memory of parental somatic cells is known to infl uence lineage differentiation propensity of iPSCs. To address this problem, a couple of recent studies have attempted to generate iPSCs from limbal fi broblasts and epithelial cells and shown that these iPSCs could effi ciently differentiate into corneal cells (Hayashi et al 2012 ;Sareen et al 2014 ). These stem cell sources are amenable for large scale expansion and are suitable for genetic manipulations in vitro.…”