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Differentiation Induction of Human Stem Cells for Corneal Epithelial

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Differentiation Induction of Human Stem Cells for Corneal Epithelial International Journal of Molecular Sciences Review Differentiation Induction of Human Stem Cells for Corneal Epithelial Regeneration 1, 1, 1 Kasem Theerakittayakorn y , Hong Thi Nguyen y, Jidapa Musika , Hataiwan Kunkanjanawan 2, Sumeth Imsoonthornruksa 1, Sirilak Somredngan 1, Mariena Ketudat-Cairns 1 and Rangsun Parnpai 1,* 1 Embryo Technology and Stem Cell Research Center, School of Biotechnology, Institute of Agricultural Technology, Suranaree University of Technology, Nakhon Ratchasima 30000, Thailand; [email protected] (K.T.); [email protected] (H.T.N.); [email protected] (J.M.); [email protected] (S.I.); [email protected] (S.S.); [email protected] (M.K.-C.) 2 Medeze Research and Development Co., Ltd. 28/9 Moo 8, Phutthamonthon Sai 4 Rd., Krathum Lom, Sam Phran, Nakhon Pathom 73220, Thailand; [email protected] * Correspondence: [email protected]; Tel.: +66-442-242-34 These authors contributed equally to this work. y Received: 25 September 2020; Accepted: 17 October 2020; Published: 22 October 2020 Abstract: Deficiency of corneal epithelium causes vision impairment or blindness in severe cases. Transplantation of corneal epithelial cells is an effective treatment but the availability of the tissue source for those cells is inadequate. Stem cells can be induced to differentiate to corneal epithelial cells and used in the treatment. Multipotent stem cells (mesenchymal stem cells) and pluripotent stem cells (embryonic stem cells and induced pluripotent stem cells) are promising cells to address the problem. Various protocols have been developed to induce differentiation of the stem cells into corneal epithelial cells. The feasibility and efficacy of both human stem cells and animal stem cells have been investigated for corneal epithelium regeneration. However, some physiological aspects of animal stem cells are different from those of human stem cells, the protocols suited for animal stem cells might not be suitable for human stem cells. Therefore, in this review, only the investigations of corneal epithelial differentiation of human stem cells are taken into account. The available protocols for inducing the differentiation of human stem cells into corneal epithelial cells are gathered and compared. Also, the pathways involving in the differentiation are provided to elucidate the relevant mechanisms. Keywords: mesenchymal stem cell; pluripotent stem cell; differentiation; corneal epithelium 1. Introduction Cornea is the transparent anterior part of the eye. Its transparency allows light to enter into the eye and the visual sensation will be initiated once the light touches the photoreceptor cells in the retina. The curvature of the cornea plays an important role in light refraction. It has a refractive power of approximately 60 diopters or two-thirds of the total eye refractive power [1]. The cornea is the first part of the eye that touches the light and it is exposed to the external environment, therefore it is easily damaged more than other parts of the eye. It can be injured by chemicals, heat, and forces from accidents. The outer surface of the cornea is covered by epithelial tissue in which the corneal epithelial cells are generated from the stem cells at the corneoscleral junction called the limbus. Limbal stem cells, the stem cells residing in the limbus, generate new corneal epithelial cells to replace the shed old cells or damaged cells to maintain the corneal epithelial population. Severe damage, severe microbial infections, or some diseases such as Stevens–Johnson syndrome and ocular cicatricial pemphigoid [2,3] may cause the depletion of limbal stem cells. Such a condition is called limbal stem cell deficiency Int. J. Mol. Sci. 2020, 21, 7834; doi:10.3390/ijms21217834 www.mdpi.com/journal/ijms Int. J. Mol. Sci. 2020, 21, 7834 2 of 20 (LSCD). Once the limbal stem cells are lost, corneal epithelial cells are unable to be generated and it will result in visual impairment and finally vision loss. Many medical treatments have been investigated to restore the corneal epithelium. The common strategy is the transplantation of autologous epithelial cells from the contralateral cornea [4,5]. In this strategy, limbal tissue is biopsied from the healthy cornea. Then, epithelial cells are isolated and expanded in laboratory until enough cells are obtained for transplantation. Even though this procedure is effective and has high success rate of 70–80% [4], it is unable to be applied to patients with bilateral LSCD. Transplantation of allogeneic limbal epithelial cells is an optional strategy but it has a risk of graft rejection [6–8]. To avoid the lack of autologous limbal epithelium and high risk of graft failure from allogeneic sources, autologous stem cells become a promising option for the regeneration of corneal epithelial tissue. With the differentiation capacity, stem cells can be induced to be corneal epithelial cells and used in medical treatments. Various kinds of stem cells have been studied for corneal epithelial tissue regeneration [9–11]. Mesenchymal stem cells (MSCs), which possess multipotent differentiation capacity, can be isolated from patients and used in the treatments without the concern of immune rejection [12]. By using minimally invasive procedures and uncomplicated techniques, MSCs can be obtained from diverse tissues, including bone marrow, adipose tissue, dental pulp, umbilical cord, and amniotic fluid [13–15]. However, it is difficult to induce MSCs from mesoderm to undergo transdifferentiation into corneal epithelial cells which originally derived from ectoderm. Pluripotent stem cells (PSCs) such as embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) are also of interest. They are unlimited self-renewal and have the potential to differentiate into any cell type of the adult organs [16,17]. However, complicated techniques and expensive materials are required for obtaining PSCs [18,19]. Several publications reported about the capability of MSCs and PSCs to differentiate into corneal epithelial cells [20–25]. However, the methods used in the differentiation induction are very diverse. Until now, no standard differentiation protocol is available and most published researches studied stem cells from animals. Because animal stem cells are different from human stem cells in many physiological aspects [26–28], the protocols developed from the studies of animal stem cells may not be suitable for human stem cells [29,30]. Therefore, herein, we review the progress of the corneal epithelial differentiation of only human stem cells. We also provide fundamental knowledge related to the differentiation methods which include the nature of corneal epithelial cells and the relevant differentiation pathways. 2. Corneal Epithelial Cell The cornea consists of three cellular layers: epithelium, stroma, and endothelium [31]. Bowman’s membrane is the thin matrix layer between epithelium and stroma whereas Descemet’s membrane is the matrix layer between stroma and endothelium [32]. The corneal epithelium covers the outermost surface of the cornea and protects the cornea from pathogen invasion [33]. The corneal epithelium is derived from surface ectoderm at 5–6 weeks of gestation [34]. There are 5–6 layers of non-keratinized stratified epithelial cells [35,36] in the corneal epithelial tissue which has a thickness of approximately 50 µm [37]. The corneal epithelium consists of three types of epithelial cells: basal cells, wing cells, and superficial cells [38]. The basal cells are arranged in a single cell layer adhered to the underlying Bowman’s membrane. They have cuboidal or columnar shape and have the ability for mitotic cell division [34]. Wing cells are in the suprabasal layers covering the layer of basal cells. They are polyhedral cells stratified 1–3 layers [39]. The superficial cells cover the outer surface of cornea. They are squamous cells with flattened nuclei. Terminal differentiation of the superficial cells results in cells sloughing off into the tears [40,41]. The integrity of corneal epithelium requires the balance between cellular proliferation and differentiation. While corneal epithelial cells are continuously desquamated from the corneal surface into the tear film, the new corneal epithelial cells are continually generated from the basal Int. J. Mol. Sci. 2020, 21, 7834 3 of 20 cells [37,42] and the stem cells residing at the limbus, the transitional area between the cornea and the sclera [43]. The regeneration of new corneal epithelial cells is achieved by the proliferation, migration, and differentiation of the corneal stem cells. The stem cells will divide into transient amplifying cells. The transient amplifying cells, which have rapid mitotic activity, will proliferate to get enough cell number before terminal differentiation [44]. The newly generated cells from the basal cells migrate vertically to the corneal surface and those from limbal stem cells migrate centripetally to the central cornea [45]. The differentiation process requires 7–10 days before they undergo apoptosis and desquamation [34]. By the delicate control, the rate of cell proliferation equals the rate of cell desquamation. Consequently, the mass of the corneal epithelium remains constant. The epithelial cells of cornea, limbus, and conjunctiva possess distinct characteristics. Cytokeratins (CKs), which are intermediate filaments expressed in a tissue-specific manner [46], are usually used to distinguish corneal epithelial cells from other cell types. There are about 20 types of CKs in humans [47]. The expressions of CK1–CK10
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