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Main differences between corneal epithelium and hair-bearing epidermis. Note the location of the stem cells in each case and the different keratins expressed by differentiated cells. (Iconography: B. Peyrusse).
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Corneal epithelium transdifferentiation into a hair-bearing epidermis provides a particularly useful system for studying the possibility that transient amplifying (TA) cells are able to activate different genetic programs in response to a change in their fibroblast environment, as well as to follow the different steps of rebuilding an epidermis fro...
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... epithelia of the cornea and the skin display dis- tinct programs of differentiation: central corneal keratinocytes express the keratin pair K3/K12, epidermal keratinocytes the keratin pair K1-2/K10 ( Sun et al., 1983a;Sun et al., 1983b), whereas the basal layer of the epidermis and the basal layer of the limbus in the cornea, expresses K5/K14 (Fig. 1). Moreover, the epidermis forms cutaneous appendages, which express their own specific keratins. ...
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Citations
... Corneal epithelial basal cells, under the influence of Wnt and noggin signalling, can transdifferentiate into epidermis and mature hair [117]. Hair follicles are the main source of epidermal stem cells [118]. The regeneration of both the corneal epithelium and hair follicles is dependent upon cytokeratin-14-expressing stem cells residing in specific stem cell niches. ...
Destruction of the limbus and depletion of limbal stem cells (LSCs), the adult progenitors of the corneal epithelium, leads to limbal stem cell deficiency (LSCD). LSCD is a rare, progressive ocular surface disorder which results in conjunctivalisation and neovascularisation of the corneal surface. Many strategies have been used in the treatment of LSCD, the common goal of which is to regenerate a self-renewing, transparent, and uniform epithelium on the corneal surface. The development of these techniques has frequently resulted from collaboration between stem cell translational scientists and ophthalmologists. Direct transplantation of autologous or allogeneic limbal tissue from a healthy donor eye is regarded by many as the technique of choice. Expansion of harvested LSCs in vitro allows smaller biopsies to be taken from the donor eye and is considered safer and more acceptable to patients. This technique may be utilised in unilateral cases (autologous) or bilateral cases (living related donor). Recently developed, simple limbal epithelial transplant (SLET) can be performed with equally small biopsies but does not require in vitro cell culture facilities. In the case of bilateral LSCD, where autologous limbal tissue is not available, autologous oral mucosa epithelium can be expanded in vitro and transplanted to the diseased eye. Data on long-term outcomes (over 5 years of follow-up) for many of these procedures is needed, and it remains unclear how they produce a self-renewing epithelium without recreating the vital stem cell niche. Bioengineering techniques offer the ability to re-create the physical characteristics of the stem cell niche, while induced pluripotent stem cells offer an unlimited supply of autologous LSCs. In vivo confocal microscopy and anterior segment OCT will complement impression cytology in the diagnosis, staging, and follow-up of LSCD. In this review we analyse recent advances in the pathology, diagnosis, and treatment of LSCD.
... Nevertheless, the expression of K3/K12 IF also signifies the corneal type epithelial differentiation in rabbit (Wu et al., 1994;Zhu et al., 1992). Similarly, K5/K14 IF is associated with basal stem cells of corneal epithelium (Ferraris et al., 1994;Pearton et al., 2004). During wound heling, rabbit corneal epithelium transiently express K6/K16 similar to that of epidermal epithelium (Schermer et al., 1989). ...
Keratins are the forming units of intermediate filaments (IF) that provide mechanical support, and formation of desmosomes between cells and hemi desmosomes with basement membranes for epithelium integrity. Keratin IF are polymers of obligate heterodimer consisting one type I keratin and one type II keratin molecules. There are 54 functional keratin genes in human genome, which are classified into three major groups, i.e., epithelial keratins, hair follicle cell-specific epithelial keratins and hair keratins. Their expression is cell type-specific and developmentally regulated. Corneal epithelium expresses a subgroup of keratins similar to those of epidermal epithelium. Limbal basal stem cells express K5/K14 and K8/K18 and K8/K19 IF suggesting that there probably are two populations of limbal stem cells (LSCs). In human, LSCs at limbal basal layer can directly stratify and differentiate to limbal suprabasal cells that express K3/K12 IF, or centripetally migrate then differentiate to corneal basal transient amplifying cells (TAC) that co-express both K3/K12 and K5/K14 prior to moving upward and assuming suprabasal cells phenotype of only K3/K12 expression that signifies corneal type epithelium differentiation. In rodent, the differentiated cornea epithelial cells express K5/K12 in lieu of K3/K12, because K3 allele exists as a pseudogene and does not encode a functional K3 protein. The basal corneal cells of new-born mice originate from surface ectoderm during embryonic development slowly commit to differentiation of becoming TAC co-expressing K5/K12 and K5/K14 IF. However, the centripetal migration may still occur at a slower rate in young mice, which is accelerated during wound healing. In this review, we will discuss and compare the cornea–specific keratins expression patterns between corneal and epidermal epithelial cells during mouse development, and between human and mouse during development and homeostasis in adult, and pathology caused by a mutation of keratins.
... 12 Finally, corneal epithelium can transdifferentiate into a hair-bearing epidermis under the appropriate conditions. 13,14 Therefore, we hypothesized that the function of these cells might be regulated by similar mechanisms. ...
Purpose:
In contact with the external environment, the cornea can easily be injured. Although corneal wounds generally heal rapidly, the pain and increased risk of infection associated with a damaged cornea, as well as the impaired healing observed in some individuals, emphasize the need for novel treatments to accelerate corneal healing. We previously demonstrated in epidermal keratinocytes that the glycerol channel aquaporin-3 (AQP3) interacts with phospholipase D2 (PLD2) to produce the signaling phospholipid phosphatidylglycerol (PG), which has been shown to accelerate skin wound healing in vivo. We hypothesized that the same signaling pathway might be operational in corneal epithelial cells.
Methods:
We used co-immunoprecipitation, immunohistochemistry, scratch wound healing assays in vitro, and corneal epithelial wound healing assays in vivo to determine the role of the AQP3/PLD2/PG signaling pathway in corneal epithelium.
Results:
AQP3 was present in human corneas in situ, and AQP3 and PLD2 were co-immunoprecipitated from corneal epithelial cell lysates. The two proteins could also be co-immunoprecipitated from insect cells simultaneously infected with AQP3- and PLD2-expressing baculoviruses, suggesting a likely direct interaction. A particular PG, dioleoylphosphatidylglycerol (DOPG), enhanced scratch wound healing of a corneal epithelial monolayer in vitro. DOPG also accelerated corneal epithelial wound healing in vivo, both in wild-type mice and in a mouse model exhibiting impaired corneal wound healing (AQP3 knockout mice).
Conclusions:
These results indicate the importance of the AQP3/PLD2/PG signaling pathway in corneal epithelial cells and suggest the possibility of developing DOPG as a pharmacologic therapy to enhance corneal wound healing in patients.
... The third main characteristic is the plasticity, ability to give rise to different cell types in a culture. The stem cells in culture have the ability to differentiate into different cell types of the same germ lineage or the ability to transdifferentiate, which is the ability to differentiate into cell types across the lineage [6][7][8][9][10][11] . The characteristics of stem cells noted in the above paragraph have been derived from studies have done on the animal cells. ...
... Pax6 deficiency results in no eye development [3], while Pax6 haploinsufficiency and ectopic expression of Pax6 result in severe eye and corneal defects [4,5], reminiscent of the phenotype of aniridia patients that suffer from a mutation in the PAX6 allele [6,7]. Interestingly, rabbit corneal epithelium transdifferentiated into a hair-forming epidermis when transplanted on mouse dermis, and this sequential process was accompanied by loss of Pax6 expression [8,9]. Similarly, a reduction in Pax6 expression was observed in corneal diseases in which the cornea gains epidermal characteristics and becomes opaque [10]. ...
Approximately 6 million people worldwide are suffering from severe visual impairments or blindness due to corneal diseases. Corneal allogeneic transplantation is often required to restore vision; however, shortage in corneal grafts and immunorejections remain major challenges. The molecular basis of corneal diseases is poorly understood largely due to lack of appropriate cellular models. Here, we described a robust differentiation of human-induced pluripotent stem cells (hiPSCs) derived from hair follicles or skin fibroblasts into corneal epithelial-like cells. We found that BMP4, coupled with corneal fibroblast-derived conditioned medium and collagen IV allowed efficient corneal epithelial commitment of hiPSCs in a manner that recapitulated corneal epithelial lineage development with high purity. Organotypic reconstitution assays suggested the ability of these cells to stratify into a corneal-like epithelium. This model allowed us identifying miR-450b-5p as a molecular switch of Pax6, a major regulator of eye development. miR-450b-5p and Pax6 were reciprocally distributed at the presumptive epidermis and ocular surface, respectively. miR-450b-5p inhibited Pax6 expression and corneal epithelial fate in vitro, altogether, suggesting that by repressing Pax6, miR-450b-5p triggers epidermal specification of the ectoderm, while its absence allows ocular epithelial development. Additionally, miR-184 was detectable in early eye development and corneal epithelial differentiation of hiPSCs. The knockdown of miR-184 resulted in a decrease in Pax6 and K3, in line with recent findings showing that a point mutation in miR-184 leads to corneal dystrophy. Altogether, these data indicate that hiPSCs are valuable for modeling corneal development and may pave the way for future cell-based therapy.
... Elegant tissue recombination studies using rabbit limbal and central corneal epithelial sheets placed on either limbal or corneal stroma demonstrated that the limbal stroma maintained ''stemness,'' while the corneal stroma promoted cell differentiation, proliferation, and apoptosis [57]. In other tissue recombination studies, adult central corneal epithelium responded to specific signals derived from embryonic dermis to undergo transdifferentiation into epidermal cells [58]. Conversely, a corneal limbal microenvironment facilitated the reprogramming of HFSCs into corneal epithelium [59]. ...
The cornea contains a reservoir of self-regenerating epithelial cells that are essential for maintaining its transparency and good vision. The study of stem cells in this functionally important organ has grown over the past four decades, partly due to the ease with which this tissue is visualized, its accessibility with minimally invasive instruments, and the fact that its stem cells are segregated within a transitional zone between two functionally diverse epithelia. While human, animal, and ex vivo models have been instrumental in progressing the corneal stem cell field, there is still much to be discovered about this exquisitely sensitive window for sight. This review will provide an overview of the human cornea, where its stem cells reside and how components of the microenvironment including extracellular matrix proteins and their integrin receptors are thought to govern corneal stem cell homeostasis.
Disclosure of potential conflicts of interest is found at the end of this article.
... In addition, similar stratification and differentiation was observed ex vivo on mouse corneas as well (Figure 5). Re-direction of corneal epithelial cells [30] and thymic epithelial cells [31] into epidermal cells by graft on mouse skin has been reported. In addition, Blazejewska et al. have reported the transdifferentiation of hair follicle stem cells into cornea epithelial-like cells using corneal limbal fibroblasts [32]. ...
Application of induced pluripotent stem (iPS) cells in regenerative medicine will bypass ethical issues associated with use of embryonic stem cells. In addition, patient-specific IPS cells can be useful to elucidate the pathophysiology of genetic disorders, drug screening, and tailor-made medicine. However, in order to apply iPS cells to mitotic tissue, induction of tissue stem cells that give rise to progeny of the target organ is required.
We induced stratified epithelial cells from mouse iPS cells by co-culture with PA6 feeder cells (SDIA-method) with use of BMP4. Clusters of cells positive for the differentiation markers KRT1 or KRT12 were observed in KRT14-positive colonies. We successfully cloned KRT14 and p63 double-positive stratified epithelial progenitor cells from iPS-derived epithelial cells, which formed stratified epithelial sheets consisting of five- to six-polarized epithelial cells in vitro. When these clonal cells were cultured on denuded mouse corneas, a robust stratified epithelial layer was observed with physiological cell polarity including high levels of E-cadherin, p63 and K15 expression in the basal layer and ZO-1 in the superficial layer, recapitulating the apico-basal polarity of the epithelium in vivo.
These results suggest that KRT14 and p63 double-positive epithelial progenitor cells can be cloned from iPS cells in order to produce polarized multilayer epithelial cell sheets.
... When recombined with embryonic mouse hair-forming dermis, adult corneal epithelial cells retain the ability to be reprogrammed into an epidermis and to produce HFs with associated sebaceous glands [39]. Corneal epithelium differentiation into a hair-bearing epidermis provides evidence that transient amplifying cells are able to activate different genetic programs in response to a change in their fibroblast environment [40]. ...
Limbal stem cell deficiency (LSCD) leads to severe ocular surface abnormalities that can result in the loss of vision. The most successful therapy currently being used is transplantation of limbal epithelial cell sheets cultivated from a limbal biopsy obtained from the patient's healthy, contralateral eye or cadaveric tissue. In this study, we investigated the therapeutic potential of murine vibrissae hair follicle bulge-derived stem cells (HFSCs) as an autologous stem cell (SC) source for ocular surface reconstruction in patients bilaterally affected by LSCD. This study is an expansion of our previously published work showing transdifferentiation of HFSCs into cells of a corneal epithelial phenotype in an in vitro system. In this study, we used a transgenic mouse model, K12(rtTA/rtTA) /tetO-cre/ROSA(mTmG) , which allows for HFSCs to change color, from red to green, once differentiation to corneal epithelial cells occurs and Krt12, the corneal epithelial-specific differentiation marker, is expressed. HFSCs were isolated from transgenic mice, amplified by clonal expansion on a 3T3 feeder layer, and transplanted on a fibrin carrier to the eye of LSCD wild-type mice (n = 31). The HFSC transplant was able to reconstruct the ocular surface in 80% of the transplanted animals; differentiating into cells with a corneal epithelial phenotype, expressing Krt12, and repopulating the corneal SC pool while suppressing vascularization and conjunctival ingrowth. These data highlight the therapeutic properties of using HFSC to treat LSCD in a mouse model while demonstrating a strong translational potential and points to the niche as a key factor for determining stem cell differentiation.
... The stem cells proliferate, migrate centripetally , and differentiate into the terminal corneal epithelial cells that replace the cells shed in the central cornea. When Pearton and colleagues (2004, 2005) performed the recombination between wild-type mouse embryonic dermis and adult rabbit central corneal epithelium, they found that the differentiated cells of the corneal epithelium dedifferentiated and reverted to a limbal basal cell phenotype. This process was confirmed to be triggered by dermal developmental signals, such as Wnt/β-catenin and BMP (bone morphogenetic protein)/Noggin signaling. ...
Dedifferentiation is an important biological phenomenon whereby cells regress from a specialized function to a simpler state
reminiscent of stem cells. Stem cells are self-renewing cells capable of giving rise to differentiated cells when supplied
with the appropriate factors. Stem cells that are derived by dedifferentiation of one's own cells could be a new resource
for regenerative medicine, one that poses no risk of genetic incompatibility or immune rejection and provokes fewer ethical
debates than the use of stem cells derived from embryonic tissue. Until now, it has not been quite clear why some differentiated
cell types can dedifferentiate and proliferate, whereas others cannot. A better understanding of the mechanisms involved in
dedifferentiation may enable scientists to control and possibly alter the plasticity of the differentiated state, which may
lead to benefits not only in stem cell research but also in regenerative medicine and even tumor biology. If so, dedifferentiation
will offer an ethically acceptable alternative route to obtain an abundant source of stem cells. Dedifferentiation is likely
to become a new focus of stem cell research. Here we compile recent advances in this emerging but significant research, highlighting
its central concepts, research findings, possible signaling pathways, and potential applications.
... K10 has been previously observed in diseased or injured corneal tissues in patients with Stevens-Johnson syndrome, ocular cicatricial pemphigoid, and chemical injuries (Elder et al., 1997;Nakamura et al., 2001), as well as in a rat model of severe dry eyes (Toshino et al., 2005). It can also be induced in animal tissue experimentally by recombination with embryonic dermis (Ferraris et al., 1994;Pearton et al., 2004Pearton et al., , 2005 or in transgenic mice expressing a dominant negative co-factor of LIM domain (Xu et al., 2007) or Dkk2 null mice (Mukhopadhyay et al., 2006). Isolated corneal epithelial cells cultured in defined media also express K10, which is expedited under high Ca 2þ conditions (Kawakita et al., 2004). ...
The corneal epithelium is continuously replaced by epithelial stem cells located in the basal layer of the limbus, located at the margin of the cornea. Studying how the stem cell niche is established at the limbus during development of the eye may lead to better understanding and treatments for diseases associated with limbal deficiencies. Using two highly specific commercially available antibodies, K10 was consistently detected suprabasally throughout the developing limbal epithelium of late gestation (20.5 dpc) and neonatal rat corneas, with interrupted expression in adult rat limbal epithelium. RT-PCR confirmed K10 expression at the transcript level in embryonic, neonatal and adult rat eyes. We have identified a time point where early stages of limbal development may be facilitated by the suprabasal expression of K10.
