Safety and Feasibility of Intrastromal Injection of Cultivated Human Corneal Stromal Keratocytes as Cell-Based Therapy for Corneal Opacities

Yam, Gary Hin‐Fai; Fuest, Matthias; Yusoff, Nur Zahirah Binte M; Goh, Tze-Wei; Bandeira, Francisco; Setiawan, Melina; Seah, Xin-Yi; Lwin, Nyein-Chan; Stanzel, Tisha Prabriputaloong; Ong, Hon Shing; Mehta, Jodhbir S.

doi:10.1167/iovs.17-23575

Cited by 36 publications

(47 citation statements)

References 54 publications

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“…A major hurdle to the development of clinically relevant tissue constructs or cell-based therapy is the lack of a widely acceptable source of corneal stromal keratocytes (CSKs), which are crucial for maintaining corneal transparency, refractivity and mechanical strength [32,33]. Ex vivo culture of human CSKs is now possible with the use of AME and supplements [12,18], but the safety concerns regarding use of human amnion, such as disease transmission and screening limitations, make this approach potentially difficult. Currently, the complete maternal screening for infectious diseases (e.g., HIV, HBV) takes 6 months post-partum [34,35].…”

Section: Discussionmentioning

confidence: 99%

“…Our group has previously reported on the use of intrastromal injection of CSKs to arrest corneal haze development and restore corneal clarity in a rat model of early corneal opacities [12]. Stromal cell-based strategies require ex vivo propagation of CSKs.…”

Section: Introductionmentioning

confidence: 99%

“…This irreversible change has created obstacles to the application of cultured stromal cells as a medically useful corneal stromal replacement. SF injection to normal rodent corneas deposited fibrotic ECM proteins to increase light scattering, resulting in haze development [12,17]. The elevated metalloproteinase levels released by fibroblasts also triggered neovascularization [12].…”

Section: Introductionmentioning

confidence: 99%

“…SF injection to normal rodent corneas deposited fibrotic ECM proteins to increase light scattering, resulting in haze development [12,17]. The elevated metalloproteinase levels released by fibroblasts also triggered neovascularization [12]. Hence, the use of correct stromal cell type (i.e., CSKs) is crucial for corneal stromal therapy.…”

Section: Introductionmentioning

confidence: 99%

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A cellular and proteomic approach to assess proteins extracted from cryopreserved human amnion in the cultivation of corneal stromal keratocytes for stromal cell therapy

et al. 2019

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Background: Human corneal stromal keratocytes propagated in culture media supplemented with human amnion extract (AME) can correct early corneal haze in an animal model. Clinical application of cultivated keratocytes is limited by infectious disease screening before amnion products can be used in humans. It remains unclear if AME from cryopreserved versus fresh human amnion can support human keratocyte propagation, and which components of the extract promote keratocyte growth. Methods: Three placentas were collected for the preparation of fresh and cryopreserved amnion tissues followed by homogenization and protein extraction. AME protein profiles were studied using isobaric tagging for relative and absolute quantitation (iTRAQ) proteomics. Enriched gene ontology (GO) terms and functional classes were identified. Primary human keratocytes from 4 donor corneas were cultured in media supplemented with fresh AME (F-AME) or cryopreserved AME (C-AME). Cell viability, proliferation and keratocyte marker expression were examined by confocal immunofluorescence and flow cytometry. Results: AME proteomics revealed 1385 proteins with similar expression levels (between 0.5-and 2-fold) between Fand CAME , while 286 proteins were reduced (less than 0.5-fold) in CAME. Enriched GO term and biological pathway analysis showed that those proteins with comparable expression between FAME and CAME were involved in cell metabolism, epithelial-mesenchymal transition, focal adhesion, cell-extracellular matrix interaction, cell stress regulation and complement cascades. Human corneal stromal keratocytes cultured with FAME or CAME showed similar morphology and viability, while cell proliferation was mildly suppressed with CAME (P > 0.05). Expression of aldehyde dehydrogenase 3A1 (ALDH3A1) and CD34 was similar in both cultures. Conclusion: AME from cryopreserved amnion had limited influence on keratocyte culture. It is feasible to use protein extract from cryopreserved amnion to propagate human keratocytes for potential translational applications.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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A cellular and proteomic approach to assess proteins extracted from cryopreserved human amnion in the cultivation of corneal stromal keratocytes for stromal cell therapy

et al. 2019

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“…On the other hand, the presence of serum has been reported to convert the keratocytes to fibroblast phenotype and enhances its viability at the cost of ECM production [ 60 , 65 ]. The possibility to differentiate hiPSCs to bona fide human CSKs has significant implications for modeling corneal diseases and for cell replacement therapy, where CSKs have shown robust potentials in animal studies [ 75 ]. However, the characteristic cellular plasticity of CSKs in culture is to be taken into consideration while developing strategies involving these cells in human cell therapy.…”

Section: Derivation Of Corneal Keratocytes From Ipscsmentioning

confidence: 99%

Corneal cell therapy: with iPSCs, it is no more a far-sight

2018

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Human-induced pluripotent stem cells (hiPSCs) provide a personalized approach to study conditions and diseases including those of the eye that lack appropriate animal models to facilitate the development of novel therapeutics. Corneal disease is one of the most common causes of blindness. Hence, significant efforts are made to develop novel therapeutic approaches including stem cell-derived strategies to replace the diseased or damaged corneal tissues, thus restoring the vision. The use of adult limbal stem cells in the management of corneal conditions has been clinically successful. However, its limited availability and phenotypic plasticity necessitate the need for alternative stem cell sources to manage corneal conditions. Mesenchymal and embryonic stem cell-based approaches are being explored; nevertheless, their limited differentiation potential and ethical concerns have posed a significant hurdle in its clinical use. hiPSCs have emerged to fill these technical and ethical gaps to render clinical utility. In this review, we discuss and summarize protocols that have been devised so far to direct differentiation of human pluripotent stem cells (hPSCs) to different corneal cell phenotypes. With the summarization, our review intends to facilitate an understanding which would allow developing efficient and robust protocols to obtain specific corneal cell phenotype from hPSCs for corneal disease modeling and for the clinics to treat corneal diseases and injury.

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Biomimetic Corneal Stroma for Scarless Corneal Wound Healing via Structural Restoration and Microenvironment Modulation

Huang,

Jiang,

et al. 2023

Adv Healthcare Materials

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Corneal injury‐induced stromal scarring causes the most common subtype of corneal blindness, and there is an unmet need to promote scarless corneal wound healing. Herein, a biomimetic corneal stroma with immunomodulatory properties is bioengineered for scarless corneal defect repair. First, a fully defined serum‐free system is established to derive stromal keratocytes (hAESC‐SKs) from a current Good Manufacturing Practice (cGMP)‐grade human amniotic epithelial stem cells (hAESCs), and RNA‐seq is used to validate the phenotypic transition. Moreover, hAESC‐SKs are shown to possess robust immunomodulatory properties in addition to the keratocyte phenotype. Inspired by the corneal stromal extracellular matrix (ECM), a photocurable gelatin‐based hydrogel is fabricated to serve as a scaffold for hAESC‐SKs for bioengineering of a biomimetic corneal stroma. The rabbit corneal defect model is used to confirm that this biomimetic corneal stroma rapidly restores the corneal structure, and effectively reshapes the tissue microenvironment via proteoglycan secretion to promote transparency and inhibition of the inflammatory cascade to alleviate fibrosis, which synergistically reduces scar formation by ≈75% in addition to promoting wound healing. Overall, the strategy proposed here provides a promising solution for scarless corneal defect repair.

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Safety and Feasibility of Intrastromal Injection of Cultivated Human Corneal Stromal Keratocytes as Cell-Based Therapy for Corneal Opacities

Abstract: We demonstrated the safety, feasibility, and therapeutic efficacy of intrastromal CSK injection. The cultivated CSKs can be a reliable cell source for potential cell-based therapy for corneal opacities.

Cited by 36 publications

References 54 publications

A cellular and proteomic approach to assess proteins extracted from cryopreserved human amnion in the cultivation of corneal stromal keratocytes for stromal cell therapy

A cellular and proteomic approach to assess proteins extracted from cryopreserved human amnion in the cultivation of corneal stromal keratocytes for stromal cell therapy

Corneal cell therapy: with iPSCs, it is no more a far-sight

Biomimetic Corneal Stroma for Scarless Corneal Wound Healing via Structural Restoration and Microenvironment Modulation

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