Influence of Biochemical Cues in Human Corneal Stromal Cell Phenotype

Fernández‐Pérez, Julia; Ahearne, Mark

doi:10.1080/02713683.2018.1536216

Cited by 34 publications

(21 citation statements)

References 48 publications

(63 reference statements)

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“…As shown in Figure 5C, AA concentration in the corneal stroma significantly increases after a topical application of AA. According to Fernández-Pérez J, AA induces the dendritic morphology, increases the expression of keratocyte markers, and prevents myofibroblast differentiation [41]. Therefore, topical use of AA is not likely to produce side effects on keratocytes.…”

Section: Discussionmentioning

confidence: 99%

Topical Ascorbic Acid Ameliorates Oxidative Stress-Induced Corneal Endothelial Damage via Suppression of Apoptosis and Autophagic Flux Blockage

Hsueh

Meir

Yeh

et al. 2020

Cells

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Compromised pumping function of the corneal endothelium, due to loss of endothelial cells, results in corneal edema and subsequent visual problems. Clinically and experimentally, oxidative stress may cause corneal endothelial decompensation after phacoemulsification. Additionally, in vitro and animal studies have demonstrated the protective effects of intraoperative infusion of ascorbic acid (AA). Here, we established a paraquat-induced cell damage model, in which paraquat induced reactive oxygen species (ROS) production and apoptosis in the B4G12 and ARPE-19 cell lines. We demonstrate that oxidative stress triggered autophagic flux blockage in corneal endothelial cells and that addition of AA ameliorated such oxidative damage. We also demonstrate the downregulation of Akt phosphorylation in response to oxidative stress. Pretreatment with ascorbic acid reduced the downregulation of Akt phosphorylation, while inhibition of the PI3K/Akt pathway attenuated the protective effects of AA. Further, we establish an in vivo rabbit model of corneal endothelial damage, in which an intracameral infusion of paraquat caused corneal opacity. Administration of AA via topical application increased its concentration in the corneal stroma and reduced oxidative stress in the corneal endothelium, thereby promoting corneal clarity. Our findings indicate a perioperative strategy of topical AA administration to prevent oxidative stress-induced damage, particularly for those with vulnerable corneal endothelia.

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Section: Discussionmentioning

confidence: 99%

Topical Ascorbic Acid Ameliorates Oxidative Stress-Induced Corneal Endothelial Damage via Suppression of Apoptosis and Autophagic Flux Blockage

Hsueh

Meir

Yeh

et al. 2020

Cells

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“…The utilization of hCFs in this model over the quiescent, native keratocyte must also be carefully considered when applying this model in the study of corneal wound healing. While hCFs are easily isolated from the human cornea due to their high proliferative and migratory properties [51], the cellular and morphological properties of hCFs are distinct from the keratocyte with lower expression of the classic keratocyte markers, aldehyde dehydrogenase 3a [52], and keratocan [53]. However, studies have suggested that transfer of hCFs to a serum-free environment may recover the keratocyte phenotype [54].…”

Section: Discussionmentioning

confidence: 99%

Corneal Epithelial–Stromal Fibroblast Constructs to Study Cell–Cell Communication in Vitro

et al. 2019

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Cell–cell communication plays a fundamental role in mediating corneal wound healing following injury or infection. Depending on the severity of the wound, regeneration of the cornea and the propensity for scar development are influenced by the acute resolution of the pro-fibrotic response mediated by closure of the wound via cellular and tissue contraction. Damage of the corneal epithelium, basement membrane, and anterior stroma following a superficial keratectomy is known to lead to significant provisional matrix deposition, including secretion of fibronectin and thrombospondin-1, as well as development of a corneal scar. In addition, corneal wounding has previously been shown to promote release of extracellular vesicles from the corneal epithelium, which, in addition to soluble factors, may play a role in promoting tissue regeneration. In this study, we report the development and characterization of a co-culture system of human corneal epithelial cells and corneal stromal fibroblasts cultured for 4 weeks to allow extracellular matrix deposition and tissue maturation. The secretion of provisional matrix components, as well as small and large extracellular vesicles, was apparent within the constructs, suggesting cell–cell communication between epithelial and stromal cell populations. Laminin-1β was highly expressed by the corneal epithelial layer with the presence of notable patches of basement membrane identified by transmission electron microscopy. Interestingly, we identified expression of collagen type III, fibronectin, and thrombospondin-1 along the epithelial–stromal interface similar to observations seen in vivo following a keratectomy, as well as expression of the myofibroblast marker, α-smooth muscle actin, within the stroma. Our results suggest that this corneal epithelial–stromal model may be useful in the study of the biochemical phenomena that occur during corneal wound healing.

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“…This medium was chosen as it has been shown to promote a keratocyte-like phenotype. 10 Constructs were fed every second day for 3 weeks.…”

Section: Construction Of An Engineered Corneal Stromamentioning

confidence: 99%

Engineering a Corneal Stromal Equivalent Using a Novel Multilayered Fabrication Assembly Technique

Fernández‐Pérez

Madden

Ahearne

2020

Tissue Engineering Part A

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To overcome the serious shortage of donor corneas for transplantation, alternatives based on tissue engineering need to be developed. Decellularized corneas are one potential alternative, but their densely packed collagen architecture inhibits recellularization in vitro. Therefore, a new rapid method of recellularizing these constructs to ensure high cellularity throughout the collagen scaffold is needed. In this study, we developed a novel method for fabricating corneal constructs by using decellularized porcine corneal sheets assembled using a bottom-up approach by layering multiple sheets between cell-laden collagen I hydrogel. Corneal lenticules were cut from porcine corneas by cryosectioning, then decellularized with detergents and air-dried for storage as sheets. Human corneal stromal cells were encapsulated in collagen I hydrogel and cast between the dried sheets. Constructs were cultured in serum-free medium supplemented with ascorbic acid and insulin for 2 weeks. Epithelial cells were then seeded on the surface and cultured for an additional week. Transparency, cell viability, and phenotype were analyzed by qPCR, histology, and immunofluorescence. Constructs without epithelial cells were sutured onto an ex vivo porcine cornea and cultured for 1 week. Lenticules were successfully decellularized, achieving dsDNA values of 13-1.2 ng/mg dry tissue, and were more resistant to degradation than the collagen I hydrogels. Constructs maintained high cell viability with a keratocyte-like phenotype with upregulation of keratocan, decorin, lumican, collagen I, ALDH3A1, and CD34 and the corneal epithelial cells stratified with a cobblestone morphology. The construct was amenable to surgical handling and no tearing occurred during suturing. After 7 days ex vivo, constructs were covered by a neoepithelium from the host porcine cells and integration into the host stroma was observed. This study describes a novel approach toward fabricating anterior corneal substitutes in a simple and rapid manner, obtaining mature and suturable constructs using only tissue-derived materials.

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Influence of Biochemical Cues in Human Corneal Stromal Cell Phenotype

Cited by 34 publications

References 48 publications

Topical Ascorbic Acid Ameliorates Oxidative Stress-Induced Corneal Endothelial Damage via Suppression of Apoptosis and Autophagic Flux Blockage

Topical Ascorbic Acid Ameliorates Oxidative Stress-Induced Corneal Endothelial Damage via Suppression of Apoptosis and Autophagic Flux Blockage

Corneal Epithelial–Stromal Fibroblast Constructs to Study Cell–Cell Communication in Vitro

Engineering a Corneal Stromal Equivalent Using a Novel Multilayered Fabrication Assembly Technique

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