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Gurdal M, Baysal K, Durak I, Selver OB. Limbal explant cultures on amniotic membrane: The effects of passaging the explants on cell phenotype. Exp Eye Res 2025; 255:110392. [PMID: 40250725 DOI: 10.1016/j.exer.2025.110392] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/12/2025] [Revised: 04/04/2025] [Accepted: 04/15/2025] [Indexed: 04/20/2025]
Abstract
In vitro expansion of limbal epithelial stem cells (LESCs) while maintaining their characteristics has the potential to address the urgent need in ophthalmology clinics for the treatment of limbal stem cell deficiency (LSCD). Herein, we investigated the impact of explant passaging on the phenotype of LESCs cultured on human amniotic membrane (hAM). Following initial coverage of the hAM surface by cells (passage 0), the rabbit limbal explants underwent two additional passages. Expanded cells were then counted using a hemocytometer and examined by immunocytochemistry and RT-qPCR to assess markers associated with LESCs (ABCG2, P63, CK14, CXCR4, BMI-1, and vimentin) and differentiated LESCs (CK3 and connexin 43). The cell yield of passage 1 was the highest among all passages. Immunocytochemistry analysis revealed that the number of CK14-positive cells was similar across all passages; vimentin-positive cells were the lowest in passage 0, while vimentin-positive cells were the highest in passage 1; and CK3-positive cells were the highest in passage 0. RT-qPCR analysis revealed that CK3 and connexin 43 expression was significantly higher in passage 0 cells than in passage 2 cells; and CXCR4 and BMI-1 expressions were significantly higher in passage 1 cells than in passage 0 cells. Our data highlight that the passaging of limbal explant on hAM results in varying cell characteristics. The decrease in CK3 and increase in ABCG2 expression in cells obtained by passaging the limbal explant suggest that passaging could potentially enhance the stem cell population within the in vitro limbal explant culture on hAM.
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Affiliation(s)
- Mehmet Gurdal
- Dokuz Eylül University, Faculty of Medicine, Department of Medical Biochemistry, Izmir, Türkiye; Ege University, Faculty of Medicine, Ocular Surface Research Laboratory, Izmir, Türkiye; Limbustem R&D Medical Products Ltd., Ege University Technopark, Izmir, Türkiye.
| | - Kemal Baysal
- Dokuz Eylül University, Faculty of Medicine, Department of Medical Biochemistry, Izmir, Türkiye; Koc University, School of Medicine, Department of Biochemistry, Istanbul, Türkiye.
| | - Ismet Durak
- Dokuz Eylül University, Faculty of Medicine, Department of Ophthalmology, Izmir, Türkiye.
| | - Ozlem Barut Selver
- Ege University, Faculty of Medicine, Ocular Surface Research Laboratory, Izmir, Türkiye; Limbustem R&D Medical Products Ltd., Ege University Technopark, Izmir, Türkiye; Ege University, Faculty of Medicine, Department of Ophthalmology, Izmir, Türkiye; Department of Stem Cell, Institute of Health Sciences, Ege University, Izmir, Türkiye; Institute of Health Sciences, Department of Stem Cell, Ege University, Izmir, Türkiye; Cord Blood Cell-Tissue Application and Research Center, Ege University, Izmir, Türkiye.
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Tutas Günaydın N, Barut Selver O. Pediatric limbal stem cell deficiency: An overview of a rarely studied pathology. Eur J Ophthalmol 2025; 35:821-833. [PMID: 39473431 DOI: 10.1177/11206721241291995] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 04/22/2025]
Abstract
The cornea is the outermost transparent layer of the eye, and the continuous renewal of the corneal epithelium is important for its transparency. This process is primarily facilitated by corneal stem cells, most of which are found at the limbus. However, any deterioration or damage in this region leads to corneal conjunctivalization and consequent limbal stem cell deficiency (LSCD), which compromises corneal transparency. LSCD is an important condition, especially in the pediatric population, as it can lead to corneal vascularization, opacity and ultimately loss of vision and subsequent amblyopia, unlike adults. Although pediatric LSCD is often due to chemical injuries, as in adults, it is also caused by conditions such as congenital aniridia, Steven Johnson Syndrome and various other immunological disorders. Appropriate and timely treatment in pediatric LSCD is of particular importance in preventing progression to amblyopia, unlike adults. Accurate staging of the disease is essential for the necessary medical and surgical treatment decision. While medical approaches are at the forefront to eliminate the causative agent and improve the ocular surface in reversible cases, it is essential to replace the limbal stem cells lost in advanced disease. For these replacement procedures, it is noteworthy that there is a tendency for minimally invasive methods compared to adults to avoid possible complications due to long life expectancy in children. In conclusion, although there are various reviews on limbal stem cell deficiency (LSCD) in adults, we believe that this review for childhood LSCD will make an important contribution to the literature, since the relevant literature currently reported for the pediatric population is limited.
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Affiliation(s)
- Nesrin Tutas Günaydın
- Department of Ophtalmology, Istanbul Arel University, Bahçelievler Memorial Hospital, Istanbul, Turkey
| | - Ozlem Barut Selver
- Department of Ophthalmology, School of Medicine, Ege University, Izmir, Turkey
- Stem Cell Department, Graduate School of Health Sciences, Ege University, Izmir, Turkey
- Limbustem R&D Medical Products Ltd & Ocular Surface Research Laboratory, Ege University, Izmir, Turkey
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3
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Parekh M, Jurkunas UV. Cultivated Autologous Limbal Epithelial Cell Transplantation: A Comprehensive Review of Clinical Trials and Applications. Cornea 2025:00003226-990000000-00872. [PMID: 40315274 DOI: 10.1097/ico.0000000000003884] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/26/2025] [Accepted: 03/24/2025] [Indexed: 05/04/2025]
Abstract
PURPOSE Limbal stem cells (LSCs) are essential for corneal epithelial regeneration and ocular surface homeostasis. Dysfunction of LSCs results in LSC deficiency (LSCD), a leading cause of global blindness. Although ex vivo expansion and autologous transplantation of LSCs have demonstrated promising clinical outcomes, no Food and Drug Administration-approved therapies for LSCD are available in the United States. To address this gap, we developed a novel 2-step process for isolating and expanding LSCs on human amniotic membrane under good manufacturing practice conditions, using a xenobiotic-free, serum-free, and antibiotic-free environment. METHODS Autologous limbal biopsies were used to generate cultivated autologous limbal epithelial cell (CALEC) constructs, ensuring rigorous safety and efficacy measures. Furthermore, we compared the success rates of good manufacturing practice-manufactured cultivated limbal epithelial transplantation (CLET) products with those of CALEC. RESULTS Among 16 limbal biopsies harvested from 15 participants, 14 resulted in successful manufacturing of CALEC grafts. Phase I clinical trial demonstrated preliminary feasibility and no safety concerns. In the phase II trial, 92% of grafts showed partial or complete success at 18 months, with no safety issues. The success rate of CALEC grafts was comparable to currently available CLET products. CONCLUSIONS The findings underscore the safety and efficacy of CALEC transplantation as a promising therapeutic strategy for LSCD. The current review focuses on the manufacturing, quality control, and clinical performance of CALEC constructs in phase I/II trials for unilateral LSCD, paving the way for future trials in advancing LSC-based regenerative therapies in the United States.
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Affiliation(s)
- Mohit Parekh
- Schepens Eye Research Institute, Massachusetts Eye and Ear, Boston, MA; and
- Department of Ophthalmology, Harvard Medical School, Boston, MA
| | - Ula V Jurkunas
- Schepens Eye Research Institute, Massachusetts Eye and Ear, Boston, MA; and
- Department of Ophthalmology, Harvard Medical School, Boston, MA
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Wu KY, Kearn N, Truong D, Choulakian MY, Tran SD. Advances in Regenerative Medicine, Cell Therapy, and 3D Bioprinting for Corneal, Oculoplastic, and Orbital Surgery. ADVANCES IN EXPERIMENTAL MEDICINE AND BIOLOGY 2025. [PMID: 40131704 DOI: 10.1007/5584_2025_855] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Subscribe] [Scholar Register] [Indexed: 03/27/2025]
Abstract
Advances in regenerative medicine, cell therapy, and 3D bioprinting are reshaping the landscape of ocular surgery, offering innovative approaches to address complex conditions affecting the cornea, ocular adnexal structures, and the orbit. These technologies hold the potential to enhance treatment precision, improve functional outcomes, and address limitations in traditional surgical and therapeutic interventions.The cornea, as the eye's primary refractive and protective barrier, is particularly well-suited for regenerative approaches due to its avascular and immune-privileged nature. Cell-based therapies, including limbal stem cell transplantation as well as stromal keratocyte and corneal endothelial cell regeneration, are being investigated for their potential to restore corneal clarity and function in conditions such as limbal stem cell deficiency, keratoconus, and endothelial dysfunction. Simultaneously, 3D bioprinting technologies are enabling the development of biomimetic corneal constructs, potentially addressing the global shortage of donor tissues and facilitating personalized surgical solutions.In oculoplastic and orbital surgery, regenerative strategies and cell therapies are emerging as possible alternatives to conventional approaches for conditions such as eyelid defects, meibomian gland dysfunction, and Graves' orbitopathy. Stem cell-based therapies and bioengineered scaffolds are showing potential in restoring lacrimal glands' function as well as reconstructing complex ocular adnexal and orbital structures. Moreover, 3D-printed orbital implants and scaffolds offer innovative solutions for repairing traumatic, post-tumor resection, and congenital defects, with the potential for improved biocompatibility and precision.Molecular and gene-based therapies, including exosome delivery systems, nanoparticle-based interventions, and gene-editing techniques, are expanding the therapeutic arsenal for ophthalmic disorders. These approaches aim to enhance the efficacy of regenerative treatments by addressing underlying pathophysiological mechanisms of diseases. This chapter provides an overview of these advancements and the challenges of translating laboratory discoveries into effective therapies in clinical practice.
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Affiliation(s)
- Kevin Y Wu
- Department of Surgery, Division of Ophthalmology, University of Sherbrooke, Sherbrooke, QC, Canada
| | - Natalie Kearn
- Department of Medicine, School of Medicine, Queen's University, Kingston, ON, Canada
| | - Doanh Truong
- College of Arts & Science, Case Western Reserve University, Cleveland, OH, USA
| | - Mazen Y Choulakian
- Department of Surgery, Division of Ophthalmology, University of Sherbrooke, Sherbrooke, QC, Canada
| | - Simon D Tran
- Faculty of Dental Medicine and Oral Health Sciences, McGill University, Montreal, QC, Canada.
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Song S, Cheng Y, Li W, Yu H, Li Z, Li J, Li M, Huang Q, Liu Y, Ling S. Irradiated umbilical cord mesenchymal stem cell-coated high oxygen-permeable hydrogel lenses inhibit corneal inflammation and neovascularization after corneal alkali burns. Sci Rep 2025; 15:10401. [PMID: 40140459 PMCID: PMC11947097 DOI: 10.1038/s41598-025-95007-9] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/29/2024] [Accepted: 03/18/2025] [Indexed: 03/28/2025] Open
Abstract
Corneal alkali burns can cause persistent inflammation and corneal neovascularization. In this study, we divided corneal alkali burned rabbits into the untreated group, the blank lens group, the radiation-treated umbilical cord mesenchymal stem cells (UCMSC) lens group, and the UCMSC I.V. group, and then measured corneal inflammation, neovascularization and corneal injury repair via slit lamp microscopy, captured anterior segment optical coherence tomography (AS-OCT), and performed hematoxylin-eosin staining. Compared with those in the other experimental groups, radiation-treated UCMSC lenses significantly decreased inflammatory index (IF) scores, areas of corneal blood vessels and corneal epithelial injury. The expression of interleukin (IL)-17 in corneas treated with radiation-treated UCMSC lenses was lower than that in corneas treated with blank lenses, and radiation-treated UCMSC lenses exhibited greater expression of IL-4 and signal transducer and activator of transcription 1 (STAT1), while the expression of cluster of differentiation-3G (CD3G), a linker for the activation of T cells (LAT), IL-6, IL-1B, CC chemokine receptor 6 (CCR6) and IL-23 exhibited the opposite effects (all P < 0.05). Our findings demonstrated that irradiated UCMSC-coated high oxygen-permeable hydrogel lenses on the ocular surface inhibited corneal angiogenesis and inflammation after corneal alkaline burns. The downregulation of Th17 cell differentiation might be responsible for these effects.
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Affiliation(s)
- Siqi Song
- Department of Ophthalmology, The Third Affiliated Hospital, Sun Yat-Sen University, Guangzhou, P.R. China
| | - Yaqi Cheng
- Department of Ophthalmology, The Third Affiliated Hospital, Sun Yat-Sen University, Guangzhou, P.R. China
| | - Weihua Li
- State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Guangdong Provincial Key Laboratory of Ophthalmology and Visual Science, Sun Yat-sen University, Guangzhou, 510060, China
| | - Huan Yu
- State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Guangdong Provincial Key Laboratory of Ophthalmology and Visual Science, Sun Yat-sen University, Guangzhou, 510060, China
| | - Zhiquan Li
- State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Guangdong Provincial Key Laboratory of Ophthalmology and Visual Science, Sun Yat-sen University, Guangzhou, 510060, China
| | - Jianbing Li
- Department of Ophthalmology, The Third Affiliated Hospital, Sun Yat-Sen University, Guangzhou, P.R. China
| | - Meng Li
- Department of Ophthalmology, The Third Affiliated Hospital, Sun Yat-Sen University, Guangzhou, P.R. China
| | - Qunai Huang
- Department of Ophthalmology, The Third Affiliated Hospital, Sun Yat-Sen University, Guangzhou, P.R. China
| | - Yingjie Liu
- Department of Ophthalmology, The Third Affiliated Hospital, Sun Yat-Sen University, Guangzhou, P.R. China
| | - Shiqi Ling
- Department of Ophthalmology, The Third Affiliated Hospital, Sun Yat-Sen University, Guangzhou, P.R. China.
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Edel MJ, Casellas HS, Osete JR, Nieto-Nicolau N, Arnalich-Montiel F, De Miguel MP, McLenachan S, Roshandel D, Casaroli-Marano RP, Alvarez-Palomo B. An Optimized Method to Produce Human-Induced Pluripotent Stem Cell-Derived Limbal Stem Cells Easily Adaptable for Clinical Use. Stem Cells Dev 2025; 34:49-60. [PMID: 39689863 DOI: 10.1089/scd.2024.0172] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/19/2024] Open
Abstract
In adults, the limbal stem cells (LSC) reside in the limbal region of the eye, at the junction of the cornea and the sclera where they renew the outer epithelial layer of the cornea assuring transparency. LSC deficiencies (LSCD) due to disease or injury account for one of the major causes of blindness. Among current treatments for LSCD, cornea transparency can be restored by providing new LSC to the damaged eye and induced pluripotent stem cells (iPSC) holds great promise as a new advanced cell source. A synthetic mRNA-based protocol to produce human iPSC from bone marrow mesenchymal stem cells has been defined. The results demonstrate a standardizable method that can be easily adaptable for clinical-grade production standards, produce high-purity LSC-like cells in a relatively rapid timeframe of 12 days, and can be successfully seeded on amniotic membrane or a biodegradable fibrin gel for transplantation. In vivo data demonstrated it is feasible to transplant the iPSC-LSC fibrin patch. In conclusion, an efficient method has been developed to produce patient-specific LSC and seed them on a scaffold fibrin gel for future treatment of LSC-deficiency disease.
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Affiliation(s)
- Michael J Edel
- Autonomous University of Barcelona, Faculty of Medicine, Unit of Anatomy and Embryology, Barcelona, Spain
- Discipline of Medical Sciences and Genetics, School of Biomedical Sciences, University of Western Australia, Perth 6009, Australia
| | | | - Jordi Requena Osete
- Department of Medical Genetics, Oslo University Hospital, Oslo, Norway
- NORMENT, Institute of Clinical Medicine, University of Oslo, and Division of Mental Health and Addiction, Oslo University Hospital, Oslo, Norway
| | | | | | - María P De Miguel
- Cell Engineering Laboratory, La Paz Hospital Research Institute (IdiPAZ), Madrid, Spain
| | - Samuel McLenachan
- Centre for Ophthalmology and Visual Science, The University of Western Australia, Perth, Western Australia, Australia
- Lions Eye Institute (LEI), Perth, Western Australia, Australia
| | - Danial Roshandel
- Centre for Ophthalmology and Visual Science, The University of Western Australia, Perth, Western Australia, Australia
- Lions Eye Institute (LEI), Perth, Western Australia, Australia
| | - Ricardo P Casaroli-Marano
- Department of Surgery, Faculty of Medicine and Health Science & Hospital Clinic de Barcelona (IDIBAPS), Universitat de Barcelona, Spain
| | - Belén Alvarez-Palomo
- Cell Therapy Service, Banc de Sang i Teixits (BST), Passeig Taulat 116, 08005, Barcelona, Spain
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Genna VG, Maurizi E, Rama P, Pellegrini G. Biology and medicine on ocular surface restoration: Advancements and limits of limbal stem cell deficiency treatments. Ocul Surf 2025; 35:57-67. [PMID: 39580144 DOI: 10.1016/j.jtos.2024.11.005] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/17/2024] [Revised: 11/08/2024] [Accepted: 11/13/2024] [Indexed: 11/25/2024]
Abstract
Ocular vision can be hampered by corneal damages, sensibly reducing patients' quality of life and having important social and economic consequences. Ocular surface diseases, which often lead to corneal opacities with visual impairment are the most severe forms of the Limbal Stem Cell Deficiency (LSCD). The present review provides an updated perspective on the available treatments for LSCD, focusing on clinical and biological features, as well as critical points to monitor during clinical translation. Recently developed surgical treatments for LSCD are described, along with their benefits and limitations, with the aim of addressing the issue of correct patient selection. Autologous surgical approaches have been attempted, such as conjunctival limbal autograft (CLAU), simple limbal epithelial transplantation (SLET), and others. Allogeneic limbal stem cell transplantation represents an alternative but carries risk of rejection and requires immunosuppression. Other potential treatments are based on induced pluripotent stem cells (iPSCs), but they require further investigation. The development of advanced therapy medicinal products (ATMPs) such as cultivated limbal epithelial transplantation (CLET), or the use of other epithelia as cultivated oral mucosal epithelial cell transplantation (COMET), has opened additional therapeutic possibilities. Some common critical issues in clinical translation are described, such as patient selection, biopsy procurement, or the use of human/animal derived components, which require rigorous validation to ensure safety and efficacy. Personalized medicine is a promising field for ocular surface restoration, where long-term follow-up studies and standardized criteria are crucial to evaluate the efficacy of these treatments and their cost-effectiveness in providing high-value healthcare.
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Affiliation(s)
| | - Eleonora Maurizi
- Centre for Regenerative Medicine ''S. Ferrari'', University of Modena and Reggio Emilia, Modena, Italy
| | - Paolo Rama
- Department of Ophthalmology, IRCCS Policlinico San Matteo, Pavia, Italy
| | - Graziella Pellegrini
- Centre for Regenerative Medicine ''S. Ferrari'', University of Modena and Reggio Emilia, Modena, Italy.
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8
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Setiawan AM, Kamarudin TA. Differentiation of Human Mesenchymal Stem Cells into Corneal Epithelial Cells: Current Progress. Curr Issues Mol Biol 2024; 46:13281-13295. [PMID: 39727920 DOI: 10.3390/cimb46120792] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/17/2024] [Revised: 11/13/2024] [Accepted: 11/17/2024] [Indexed: 12/28/2024] Open
Abstract
The limited availability of corneal tissue grafts poses significant challenges in the treatment of corneal blindness. Novel treatment utilizes stem cell grafts transplanted from the healthy side of the cornea to the damaged side. However, this procedure is only possible for those who have one-sided corneal blindness. Human stem cells offer promising potential for corneal tissue engineering, providing an alternative solution. Among the different types of stem cells, mesenchymal stem cells (MSCs) stand out due to their abundance and ease of isolation. Human MSCs can be derived from bone marrow, adipose, and umbilical cord tissues. Differentiating MSC toward corneal tissue can be achieved through several methods including chemical induction and co-culture with adult corneal cells such as human limbal epithelial stem cells (LESCs) and human corneal epithelial cells (hTCEpi). Adipose-derived stem cells (ADSCs) are the most common type of MSC that has been studied for corneal differentiation. Corneal epithelial cells are the most common corneal cell type targeted by researchers for corneal differentiation. Chemical induction with small molecules, especially bone morphogenetic protein 4 (BMP4), all-trans retinoic acid (ATRA), and epidermal growth factor (EGF), has gained more popularity in corneal epithelial cell differentiation. This review highlights the current progress in utilizing MSCs for corneal differentiation studies, showcasing their potential to revolutionize treatments for corneal blindness.
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Affiliation(s)
- Abdul Malik Setiawan
- Department of Anatomy, Faculty of Medicine, Universiti Kebangsaan Malaysia, Cheras, Kuala Lumpur 56000, Malaysia
- Department of Anatomy, Maulana Malik Ibrahim State Islamic University, Malang 65144, Indonesia
| | - Taty Anna Kamarudin
- Department of Anatomy, Faculty of Medicine, Universiti Kebangsaan Malaysia, Cheras, Kuala Lumpur 56000, Malaysia
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Wang F, Xu Y, Zhou Q, Xie L. Biomolecule-based hydrogels as delivery systems for limbal stem cell transplantation: A review. Int J Biol Macromol 2024; 280:135778. [PMID: 39304050 DOI: 10.1016/j.ijbiomac.2024.135778] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/25/2024] [Revised: 08/25/2024] [Accepted: 09/17/2024] [Indexed: 09/22/2024]
Abstract
Limbal stem cell deficiency (LSCD) is a complex disease of the cornea resulting from dysfunction and/or loss of limbal stem cells (LSCs) and their niche. Most patients with LSCD cannot be treated by conventional corneal transplants because the donor tissue lacks the LSCs necessary for corneal epithelial regeneration. Successful treatment of LSCD depends on effective stem cell transplantation to the ocular surface for replenishment of the LSC reservoir. Thus, stem cell therapies employing carrier substrates for LSCs have been widely explored. Hydrogel biomaterials have many favorable characteristics, including hydrophilicity, flexibility, cytocompatibility, and optical properties suitable for the transplantation of LSCs. Therefore, due to these properties, along with the necessary signals for stem cell proliferation and differentiation, hydrogels are ideal carrier substrates for LSCD treatment. This review summarizes the use of different medical-type hydrogels in LSC transplantation from 2001 to 2024. First, a brief background of LSCD is provided. Then, studies that employed various hydrogel scaffolds as LSC carriers are highlighted to provide a multimodal strategic reference for LSCD treatment. Finally, an analysis of prospective future developments and challenges in the field of hydrogels as LSC carriers for treating LSCD is presented.
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Affiliation(s)
- Fuyan Wang
- Eye Institute of Shandong First Medical University, Qingdao Eye Hospital of Shandong First Medical University, State Key Laboratory Cultivation Base, Shandong Provincial Key Laboratory of Ophthalmology, School of Ophthalmology, Shandong First Medical University, Qingdao 266071, China
| | - Yuehe Xu
- Eye Institute of Shandong First Medical University, Qingdao Eye Hospital of Shandong First Medical University, State Key Laboratory Cultivation Base, Shandong Provincial Key Laboratory of Ophthalmology, School of Ophthalmology, Shandong First Medical University, Qingdao 266071, China
| | - Qingjun Zhou
- Eye Institute of Shandong First Medical University, Qingdao Eye Hospital of Shandong First Medical University, State Key Laboratory Cultivation Base, Shandong Provincial Key Laboratory of Ophthalmology, School of Ophthalmology, Shandong First Medical University, Qingdao 266071, China.
| | - Lixin Xie
- Eye Institute of Shandong First Medical University, Qingdao Eye Hospital of Shandong First Medical University, State Key Laboratory Cultivation Base, Shandong Provincial Key Laboratory of Ophthalmology, School of Ophthalmology, Shandong First Medical University, Qingdao 266071, China.
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10
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Sharma N, Venugopal R, Mohanty S, Priyadarshini K, Nagpal R, Singhal D, Bari A, Dada T, Maharana PK, Agarwal T, Upadhyay AD. Simple limbal epithelial transplantation versus cultivated limbal epithelial transplantation in ocular burns. Ocul Surf 2024; 34:504-509. [PMID: 39442877 DOI: 10.1016/j.jtos.2024.10.007] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/09/2024] [Revised: 09/18/2024] [Accepted: 10/20/2024] [Indexed: 10/25/2024]
Abstract
PURPOSE To compare the outcomes of simple limbal epithelial transplantation (SLET) with cultivated limbal epithelial transplantation (CLET) for the management of total limbal stem cell deficiency (LSCD) in eyes with unilateral ocular burns. DESIGN Randomized controlled trial. METHODS 100 patients (100 eyes) with unilateral total LSCD following ocular burns undergoing autologous Limbal Stem Cell Transplantation (LSCT) were enrolled and randomized into SLET and CLET groups. Restoration of an epithelized ocular surface was the primary outcome measure. Occurrences of progressive conjunctivalization and persistent epithelial defects postoperatively were considered surgical failures. RESULTS Mean age was 20.2 ± 13.1 years (SLET) and 22.6 ± 14.3 years (CLET) (p = 0.363). Alkali burn was the most common causative factor in both groups and had comparable mean logMAR BCVA at presentation [SLET: 2.33 ± 0.5, CLET: 2.23 ± 1.48 (p = 0.652)]. Median time interval between injury and surgical intervention was 18 months (SLET) and 12 months (CLET) (p = 0.06). 88 % eyes in SLET group maintained a stable ocular surface at 1 year period versus CLET group (86 %) (p = 0.999). Mean logMAR BCVA significantly improved in both groups with SLET having significantly better BCVA versus CLET at 6 months (p = 0.0390), 1 year (p = 0.0001), 2 year (p = 0.0001) and 3 years (p = 0.0001) follow up. Kaplan-Meier survival analysis was statistically insignificant amongst the 2 groups (p = 0.590). CONCLUSIONS Compared to CLET, SLET is equally efficacious in restoring and maintaining a stable ocular surface in eyes with total LSCD due to ocular burns, with the added advantage of providing superior visual outcomes.
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Affiliation(s)
- Namrata Sharma
- Cornea, Cataract & Refractive Surgery Services, Dr. Rajendra Prasad Centre for Ophthalmic Sciences, All India Institute of Medical Sciences, New Delhi, India.
| | - Renu Venugopal
- Cornea, Cataract & Refractive Surgery Services, Dr. Rajendra Prasad Centre for Ophthalmic Sciences, All India Institute of Medical Sciences, New Delhi, India
| | - Sujata Mohanty
- Stem Cell Facility, All India Institute of Medical Sciences, New Delhi, India.
| | - K Priyadarshini
- Cornea, Cataract & Refractive Surgery Services, Dr. Rajendra Prasad Centre for Ophthalmic Sciences, All India Institute of Medical Sciences, New Delhi, India
| | - Ritu Nagpal
- Cornea, Cataract & Refractive Surgery Services, Dr. Rajendra Prasad Centre for Ophthalmic Sciences, All India Institute of Medical Sciences, New Delhi, India
| | - Deepali Singhal
- Cornea, Cataract & Refractive Surgery Services, ASG Eye Hospitals, Bhubaneswar, Odisha, India
| | - Aafreen Bari
- Cornea, Cataract & Refractive Surgery Services, Dr. Rajendra Prasad Centre for Ophthalmic Sciences, All India Institute of Medical Sciences, New Delhi, India
| | - Tanuj Dada
- Cornea, Cataract & Refractive Surgery Services, Dr. Rajendra Prasad Centre for Ophthalmic Sciences, All India Institute of Medical Sciences, New Delhi, India
| | - Prafulla Kumar Maharana
- Cornea, Cataract & Refractive Surgery Services, Dr. Rajendra Prasad Centre for Ophthalmic Sciences, All India Institute of Medical Sciences, New Delhi, India
| | - Tushar Agarwal
- Cornea, Cataract & Refractive Surgery Services, Dr. Rajendra Prasad Centre for Ophthalmic Sciences, All India Institute of Medical Sciences, New Delhi, India
| | - Ashish Dutt Upadhyay
- Department of Biostatistics, All India Institute of Medical Sciences, New Delhi, India
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11
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Suanno G, Genna VG, Maurizi E, Dieh AA, Griffith M, Ferrari G. Cell therapy in the cornea: The emerging role of microenvironment. Prog Retin Eye Res 2024; 102:101275. [PMID: 38797320 DOI: 10.1016/j.preteyeres.2024.101275] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/11/2023] [Revised: 05/22/2024] [Accepted: 05/23/2024] [Indexed: 05/29/2024]
Abstract
The cornea is an ideal testing field for cell therapies. Its highly ordered structure, where specific cell populations are sequestered in different layers, together with its accessibility, has allowed the development of the first stem cell-based therapy approved by the European Medicine Agency. Today, different techniques have been proposed for autologous and allogeneic limbal and non-limbal cell transplantation. Cell replacement has also been attempted in cases of endothelial cell decompensation as it occurs in Fuchs dystrophy: injection of cultivated allogeneic endothelial cells is now in advanced phases of clinical development. Recently, stromal substitutes have been developed with excellent integration capability and transparency. Finally, cell-derived products, such as exosomes obtained from different sources, have been investigated for the treatment of severe corneal diseases with encouraging results. Optimization of the success rate of cell therapies obviously requires high-quality cultured cells/products, but the role of the surrounding microenvironment is equally important to allow engraftment of transplanted cells, to preserve their functions and, ultimately, lead to restoration of tissue integrity and transparency of the cornea.
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Affiliation(s)
- Giuseppe Suanno
- Vita-Salute San Raffaele University, Milan, Italy; Eye Repair Unit, Division of Neuroscience, IRCCS San Raffaele Scientific Institute, Milan, Italy
| | | | - Eleonora Maurizi
- Centre for Regenerative Medicine ''S. Ferrari'', University of Modena and Reggio Emilia, Modena, Italy
| | - Anas Abu Dieh
- Maisonneuve-Rosemont Hospital Research Centre, Montreal, Quebec, Canada
| | - May Griffith
- Maisonneuve-Rosemont Hospital Research Centre, Montreal, Quebec, Canada.
| | - Giulio Ferrari
- Vita-Salute San Raffaele University, Milan, Italy; Eye Repair Unit, Division of Neuroscience, IRCCS San Raffaele Scientific Institute, Milan, Italy; Ophthalmology Unit, IRCCS San Raffaele Scientific Institute, Milan, Italy.
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Korkmaz I, Palamar M, Timarci I, Egrilmez S, Yagci A, Barut Selver O. Limbal graft transplantation: a rare implementation in pediatric limbal stem cell deficiency. Int Ophthalmol 2024; 44:337. [PMID: 39093517 DOI: 10.1007/s10792-024-03269-5] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/24/2024] [Accepted: 07/29/2024] [Indexed: 08/04/2024]
Abstract
PURPOSE To evaluate limbal graft transplantation success in pediatric patients with chemical injury-induced limbal stem cell deficiency (LSCD) using the 'LSCD Working Group' staging system. METHODS Medical records of 11 eyes of 11 children who underwent limbal graft transplantation (limbal autograft/limbal allograft) were included. Surgical success was defined as improvement in the post-operative 1st year LSCD stage. RESULTS The mean age was 12 ± 5 (4-17) years. Causative agent was alkaline in 4(36.4%) and acid in 3(27.2%) patients. Limbal autograft was performed in 9 (81.8%) eyes with unilateral LSCD, and allograft transplantation was performed in 2 (18.2%) eyes with bilateral LSCD. The mean follow-up time was 33.89 ± 30.73 (12-102.33) months. The overall limbal graft transplantation success rate was 72.7%. Among 9 patients who receive limbal autograft, 8 had improvement in post-operative LSCD stage, 1 had stable LSCD stage. Of the 2 patients who receive limbal allograft, post-operative LSCD stage remained the same in 1 and worsened in 1 patient. The mean time between injury and the surgery was 30.47 ± 30.08 (7-108.47) months. Penetrating keratoplasty was performed in 3 (27.2%) of 11 patients following limbal graft transplantation. CONCLUSION Management of LSCD in children is challenging and appears to be somewhat different from that of adults. Limited data in the literature indicate that cultivated or simple limbal epithelial transplantations (CLET/SLET) are primarily preferred in children. Although the tendency to take small tissue from the healthy eye is noteworthy, conventional limbal allograft and autograft transplantations also show promising results without any further complications in at least 1 year follow-up period.
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Affiliation(s)
- Ilayda Korkmaz
- Department of Ophthalmology, Ege University Medical Faculty Hospital, 35100, Izmir, Turkey
| | - Melis Palamar
- Department of Ophthalmology, Ege University Medical Faculty Hospital, 35100, Izmir, Turkey
| | - Ilgin Timarci
- Department of Public Health, Katip Celebi University School of Medicine, Izmir, Turkey
| | - Sait Egrilmez
- Department of Ophthalmology, Ege University Medical Faculty Hospital, 35100, Izmir, Turkey
| | - Ayse Yagci
- Department of Ophthalmology, Ege University Medical Faculty Hospital, 35100, Izmir, Turkey
| | - Ozlem Barut Selver
- Department of Ophthalmology, Ege University Medical Faculty Hospital, 35100, Izmir, Turkey.
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13
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Li Z, Böhringer D, Stachon T, Nastaranpour M, Fries FN, Seitz B, Ulrich M, Munteanu C, Langenbucher A, Szentmáry N. Culturing Limbal Epithelial Cells of Long-term Stored Corneal Donors (Organ Culture) In Vitro - A Stepwise Linear Regression Algorithm. Klin Monbl Augenheilkd 2024; 241:964-971. [PMID: 37130569 DOI: 10.1055/a-2084-7168] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 05/04/2023]
Abstract
PURPOSE To assess various potential factors on human limbal epithelial cell (LEC) outgrowth in vitro using corneal donor tissue following long-term storage (organ culture) and a stepwise linear regression algorithm. METHODS Of 215 donors, 304 corneoscleral rings were used for our experiments. For digestion of the limbal tissue and isolation of the limbal epithelial cells, the tissue pieces were incubated with 4.0 mg/mL collagenase A at 37 °C with 95% relative humidity and a 5% CO2 atmosphere overnight. Thereafter, limbal epithelial cells were separated from limbal keratocytes using a 20-µm CellTricks filter. The separated human LECs were cultured in keratinocyte serum-free medium medium, 1% penicillin/streptomycin (P/S), 0.02% epidermal growth factor (EGF), and 0.3% bovine pituitary extract (BPE). The potential effect of donor age (covariate), postmortem time (covariate), medium time (covariate), size of the used corneoscleral ring (360°, 270°180°, 120°, 90°, less than 90°) (covariate), endothelial cell density (ECD) (covariate), gender (factor), number of culture medium changes during organ culture (factor), and origin of the donor (donating institution and storing institution, factor) on the limbal epithelial cell outgrowth was analyzed with a stepwise linear regression algorithm. RESULTS The rate of successful human LEC outgrowth was 37.5%. From the stepwise linear regression algorithm, we found out that the relevant influencing parameters on the LEC growth were intercept (p < 0.001), donor age (p = 0.002), number of culture medium changes during organ culture (p < 0.001), total medium time (p = 0.181), and size of the used corneoscleral ring (p = 0.007), as well as medium time × size of the corneoscleral ring (p = 0.007). CONCLUSIONS The success of LEC outgrowth increases with lower donor age, lower number of organ culture medium changes during storage, shorter medium time in organ culture, and smaller corneoscleral ring size. Our stepwise linear regression algorithm may help us in optimizing LEC cultures in vitro.
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Affiliation(s)
- Zhen Li
- Dr. Rolf M. Schwiete Center for Limbal Stem Cell and Congenital Aniridia Research, Saarland University, Homburg, Germany
| | - Daniel Böhringer
- Department of Ophthalmology, University of Freiburg, Freiburg im Breisgau, Germany
| | - Tanja Stachon
- Dr. Rolf M. Schwiete Center for Limbal Stem Cell and Congenital Aniridia Research, Saarland University, Homburg, Germany
| | - Mahsa Nastaranpour
- Dr. Rolf M. Schwiete Center for Limbal Stem Cell and Congenital Aniridia Research, Saarland University, Homburg, Germany
| | - Fabian Norbert Fries
- Dr. Rolf M. Schwiete Center for Limbal Stem Cell and Congenital Aniridia Research, Saarland University, Homburg, Germany
- Department of Ophthalmology, Saarland University, Homburg, Germany
| | - Berthold Seitz
- Department of Ophthalmology, Saarland University, Homburg, Germany
| | - Myriam Ulrich
- Dr. Rolf M. Schwiete Center for Limbal Stem Cell and Congenital Aniridia Research, Saarland University, Homburg, Germany
| | | | | | - Nóra Szentmáry
- Dr. Rolf M. Schwiete Center for Limbal Stem Cell and Congenital Aniridia Research, Saarland University, Homburg, Germany
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Bian Y, Jurkunas U. Ocular Chemical Injuries and Limbal Stem Cell Deficiency (LSCD): An Update on Management. Int Ophthalmol Clin 2024; 64:31-48. [PMID: 38525980 DOI: 10.1097/iio.0000000000000487] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 03/26/2024]
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Lee YF, Yong DWW, Manotosh R. A Review of Contact Lens-Induced Limbal Stem Cell Deficiency. BIOLOGY 2023; 12:1490. [PMID: 38132316 PMCID: PMC10740976 DOI: 10.3390/biology12121490] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 09/19/2023] [Revised: 11/11/2023] [Accepted: 11/24/2023] [Indexed: 12/23/2023]
Abstract
Limbal stem cell deficiency (LSCD) is a pathologic condition caused by the dysfunction and destruction of stem cells, stem cell precursors and limbal cell niche in the corneal epithelium, leading to severe conjunctivalization of the cornea. Etiologies for LSCD span from congenital (aniridia), traumatic (chemical or thermal injuries), autoimmune (Stevens-Johnson syndrome) and iatrogenic disease to contact lens (CL) wear. Of these, CL wear is the least understood and is often a subclinical cause of LSCD. Even with recent advances in LSCD research, limitations persist in establishing the pathogenesis and treatment guidelines for CL-induced LSCD. A literature search was conducted to include original articles containing patients with CL-induced LSCD. This review will critically discuss the complex pathophysiology behind CL-induced LSCD, the underlying risk factors and epidemiology of the disease as well as methods to obtain a diagnosis. Various treatment options will be reviewed based on proposed treatment strategies.
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Affiliation(s)
- Yhu Fhei Lee
- Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117597, Singapore
| | - Dayna Wei Wei Yong
- Department of Ophthalmology, National University Hospital, Singapore 119074, Singapore
| | - Ray Manotosh
- Department of Ophthalmology, National University Hospital, Singapore 119074, Singapore
- Department of Ophthalmology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 119228, Singapore
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16
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Chaudhary S, Kate A, Chappidi K, Basu S, Shanbhag SS. Safety and Efficacy of Contact Lenses in Eyes After Simple Limbal Epithelial Transplantation. Cornea 2023; 42:1513-1519. [PMID: 36728263 DOI: 10.1097/ico.0000000000003228] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/02/2022] [Accepted: 11/28/2022] [Indexed: 02/03/2023]
Abstract
PURPOSE The aim of this study was to evaluate the safety and efficacy of contact lenses (CLs) in eyes after simple limbal epithelial transplantation (SLET) for limbal stem cell deficiency (LSCD). METHODS This retrospective study included 61 eyes with partial or total LSCD which underwent SLET and were fitted with corneal or scleral rigid gas-permeable CLs. The primary outcome measure was best-corrected visual acuity (BCVA) with CLs. RESULTS The median age at presentation was 22 years. The most common cause of LSCD was chemical injury [47/61 eyes (77%)]. Twenty-seven eyes (44%) were fitted with corneal rigid gas-permeable lenses, while 34 eyes (56%) were prescribed scleral lenses. The median duration of interval between SLET and CL trial was 10 months (interquartile range: 4-17). The median preoperative BCVA was logarithm of minimal angle of resolution (logMAR) 1.8. This improved to logMAR 1 ( P < 0.001) after SLET and to logMAR 0.6 ( P < 0.001) with CLs. Eyes with chemical injury (logMAR 1 vs. 0.6, P = 0.0001), grade 1 (logMAR 0.8 vs. 0.4, P < 0.0001), and grade 2 (logMAR 0.9 vs. 0.6, P = 0.004) corneal scarring had better improvement in BCVA with CLs. No complications were noted until the last follow-up visit. CONCLUSIONS CLs, both corneal and scleral lenses, provide significant visual improvement in eyes after SLET with residual corneal scarring. These can be safely prescribed in such eyes without any adverse effects. This option can be considered before offering keratoplasty in these eyes.
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Affiliation(s)
- Simmy Chaudhary
- The Cornea Institute, KAR Campus, L V Prasad Eye Institute, Hyderabad, India
- Bausch & Lomb Contact Lens Centre, L V Prasad Eye Institute, Hyderabad, India
| | - Anahita Kate
- The Cornea Institute, L V Prasad Eye Institute, Vijaywada, India
| | - Kiranmayi Chappidi
- The Cornea Institute, L V Prasad Eye Institute, Vijaywada, India
- Brien Holden Institute of Optometry and Vision Sciences, L V Prasad Eye Institute, Hyderabad, India
| | - Sayan Basu
- The Cornea Institute, KAR Campus, L V Prasad Eye Institute, Hyderabad, India
- Centre for Ocular Regeneration (CORE), L V Prasad Eye Institute, Hyderabad, India; and
- Brien Holden Eye Research Centre (BHERC), L V Prasad Eye Institute, Hyderabad, India
| | - Swapna S Shanbhag
- The Cornea Institute, KAR Campus, L V Prasad Eye Institute, Hyderabad, India
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17
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Korkmaz I, Palamar M, Egrilmez S, Gurdal M, Yagci A, Barut Selver O. Evaluation of Limbal Stem Cell Transplant Success in Ocular Chemical Injury. EXP CLIN TRANSPLANT 2023; 21:684-690. [PMID: 34981716 DOI: 10.6002/ect.2021.0393] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/05/2022]
Abstract
OBJECTIVES We evaluated limbal stem cell transplant success in limbal stem cell deficiency due to chemical injury at a tertiary eye care center in Turkey with a novel system for describing limbal stem cell deficiency, as developed by the Limbal Stem Cell Working Group. MATERIALS AND METHODS Medical records of 80 eyes of 80 patients after limbal stem cell transplant for limbal stem cell deficiency secondary to chemical injury were included, with patients grouped according to surgical procedure, ie, limbal autograft, limbal allograft, and cultivated limbal epithelial cell transplant. Surgical success was defined as improvement in postoperative year 1 of limbal stem cell deficiency stage. RESULTS Patients' mean age was 37.9 ± 15.7 years (range, 4-71 years). Male/female ratio was 2.4. Forty-five patients (56.3%) were injured with alkaline substance, and 16 (20%) with acid substance. Mean follow-up time was 60.3 ± 30.6 months (range, 6-118.6 months). Limbal autograft, allograft, and cultivated limbal epithelial cell transplants were performed in 58 (72.5%), 12 (15%), and 10 (12.5%) eyes, respectively. Intervals between injury and surgery in limbal autograft, limbal allograft, and cultivated limbal epithelial cell transplants were 43.3 ± 94.1 months (range, 0.5-592 months), 14.5 ± 10.6 months (range, 2.4-32.5 months), and 122.8 ± 158.9 months (range, 21.1-504 months),respectively (P = .02); and surgical success rates in each group were 65.5%, 41.7%, and 90%, respectively (P = .03). Overall surgical success rate was 65%. CONCLUSIONS Accurate determination of the limbal stem cell deficiency stage is crucial for proper evaluation of surgical success. Surgery type and interval between injury and surgery were the most important factors associated with higher surgical success rates. Despite the limited number of patients in the subgroups, the results were remarkable to emphasize the significance of a novel limbal stem cell deficiency scoring system.
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Affiliation(s)
- Ilayda Korkmaz
- From the Department of Ophthalmology, Ege University, Izmir, Turkey
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18
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Tran LT, Li JY. The role of eye banking with cell-based therapies. Curr Opin Ophthalmol 2023; 34:334-338. [PMID: 37097197 DOI: 10.1097/icu.0000000000000962] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 04/26/2023]
Abstract
PURPOSE OF REVIEW Cell-based therapies are an exciting new frontier in managing corneal diseases. The introduction of these novel therapies may provide new alternatives to corneal transplantation and decrease the dependence on donor corneal tissue. These changes have the potential to significantly impact eye banking in the future. RECENT FINDINGS The current article reviews current research involving cell-based therapy for treating corneal disorders, including cultivated limbal stem cell transplantation, limbal mesenchymal stem cells for stromal regeneration, and the use of human-cultivated endothelial cells. We will look at barriers to the development and implementation of these therapies. SUMMARY As corneal surgery expands to include cell-based therapies; eye banks will need to redefine their role to support the everchanging landscape of corneal surgery and the decreased demand for corneal donor tissue.
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Affiliation(s)
- Lillian T Tran
- Department of Ophthalmology & Vision Science, University of California Davis Health, Sacramento, California, USA
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19
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Tomczak W, Winkler-Lach W, Tomczyk-Socha M, Misiuk-Hojło M. Advancements in Ocular Regenerative Therapies. BIOLOGY 2023; 12:biology12050737. [PMID: 37237549 DOI: 10.3390/biology12050737] [Citation(s) in RCA: 2] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 04/07/2023] [Revised: 05/12/2023] [Accepted: 05/16/2023] [Indexed: 05/28/2023]
Abstract
The use of stem cells (SCs) has emerged as a promising avenue in ophthalmology, offering potential therapeutic solutions for various vision impairments and degenerative eye diseases. SCs possess the unique ability to self-renew and differentiate into specialised cell types, making them valuable tools for repairing damaged tissues and restoring visual function. Stem cell-based therapies hold significant potential for addressing conditions such as age-related macular degeneration (AMD), retinitis pigmentosa (RP), corneal disorders, and optic nerve damage. Therefore, researchers have explored different sources of stem cells, including embryonic stem cells (ESC), induced pluripotent stem cells (iPSCs), and adult stem cells, for ocular tissue regeneration. Preclinical studies and early-phase clinical trials have demonstrated promising outcomes, with some patients experiencing improved vision following stem cell-based interventions. However, several challenges remain, including optimising the differentiation protocols, ensuring transplanted cells' safety and long-term viability, and developing effective delivery methods. The field of stem cell research in ophthalmology witnesses a constant influx of new reports and discoveries. To effectively navigate these tons of information, it becomes crucial to summarise and systematise these findings periodically. In light of recent discoveries, this paper demonstrates the potential applications of stem cells in ophthalmology, focusing on their use in various eye tissues, including the cornea, retina, conjunctiva, iris, trabecular meshwork, lens, ciliary body, sclera, and orbital fat.
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Affiliation(s)
| | | | | | - Marta Misiuk-Hojło
- Department of Ophthalmology, Wroclaw Medical University, 50556 Wroclaw, Poland
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20
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Lee JY, Knight RJ, Deng SX. Future regenerative therapies for corneal disease. Curr Opin Ophthalmol 2023; 34:267-272. [PMID: 36602407 DOI: 10.1097/icu.0000000000000938] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/06/2023]
Abstract
PURPOSE OF REVIEW To highlight the progress and future direction of stem-cell based regenerative therapies for the treatment of corneal disease. RECENT FINDINGS Corneal stem cell-based therapies, such as limbal stem cell transplantation, corneal stromal stem cell transplantation, endothelial stem cell transplantation, and stem cell-derived extracellular vesicles have demonstrated promising results in the laboratory. Although most are still in preclinical development or early phase clinical trials, these stem cell-based therapies hold potential to facilitate tissue regeneration, restore native function, and inhibit pathologic disease processes such as fibrosis, inflammation, and neovascularization. SUMMARY Stem cell-based therapy offers a promising therapeutic option that can circumvent several of the challenges and limitations of traditional surgical treatment. This concise review summarizes the progress in stem-cell based therapies for corneal diseases along with their history, underlying mechanisms, limitations, and future areas for development.
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Affiliation(s)
- John Y Lee
- Stein Eye Institute, Department of Ophthalmology, David Geffen School of Medicine
| | - Robert J Knight
- Stein Eye Institute, Department of Ophthalmology, David Geffen School of Medicine
| | - Sophie X Deng
- Stein Eye Institute, Department of Ophthalmology, David Geffen School of Medicine
- Molecular Biology Institute, University of California, Los Angeles, California, USA
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21
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Cadenas-Martin M, Arnalich-Montiel F, Miguel MPD. Derivation of Limbal Stem Cells from Human Adult Mesenchymal Stem Cells for the Treatment of Limbal Stem Cell Deficiency. Int J Mol Sci 2023; 24:ijms24032350. [PMID: 36768672 PMCID: PMC9916480 DOI: 10.3390/ijms24032350] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/24/2022] [Revised: 01/18/2023] [Accepted: 01/23/2023] [Indexed: 01/27/2023] Open
Abstract
Approximately 10 million individuals have blindness due to limbal stem cell (LSCs) deficiency, one of the most challenging problems in ophthalmology. To replenish the LSC pool, an autologous extraocular cell source is appropriate, thereby avoiding the risk of immune rejection, the need for immunosuppression and the risk of damaging the contralateral eye. In recent years, adipose-derived mesenchymal stem cells (ADSCs) have been a key element in ocular regenerative medicine. In this study, we developed a protocol for deriving human LSCs from ADSCs compatible with the standard carrier human amniotic membrane, helping provide a stem cell pool capable of maintaining proper corneal epithelial homeostasis. The best protocol included an ectodermal induction step by culturing ADSCs with media containing fetal bovine serum, transforming growth factor-β inhibitor SB-505124, Wnt inhibitor IWP-2 and FGF2 for 7 days, followed by an LSC induction step of culture in modified supplemental hormonal epithelial medium supplemented with pigment epithelium-derived factor and keratinocyte growth factor for 10 additional days. The optimal differentiation efficiency was achieved when cells were cultured in this manner over vitronectin coating, resulting in up to 50% double-positive αp63/BMI-1 cells. The results of this project will benefit patients with LSC deficiency, aiding the restoration of vision.
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Affiliation(s)
- Marta Cadenas-Martin
- Cell Engineering Laboratory, La Paz University Hospital Health Research Institute, IdiPAZ, 28046 Madrid, Spain
| | - Francisco Arnalich-Montiel
- Ophthalmology Department, Ramón y Cajal University Hospital, Ramón y Cajal Health Research Institute, 28034 Madrid, Spain
| | - Maria P De Miguel
- Ophthalmology Department, Ramón y Cajal University Hospital, Ramón y Cajal Health Research Institute, 28034 Madrid, Spain
- Correspondence: ; Tel.: +34-912-071458
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22
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Tavakkoli F, Eleiwa TK, Elhusseiny AM, Damala M, Rai AK, Cheraqpour K, Ansari MH, Doroudian M, H Keshel S, Soleimani M, Djalilian AR, Sangwan VS, Singh V. Corneal stem cells niche and homeostasis impacts in regenerative medicine; concise review. Eur J Ophthalmol 2023:11206721221150065. [PMID: 36604831 DOI: 10.1177/11206721221150065] [Citation(s) in RCA: 4] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/07/2023]
Abstract
The limbal stem cells niche (LSCN) is an optimal microenvironment that provides the limbal epithelial stem cells (LESCs) and strictly regulates their proliferation and differentiation. Disturbing the LSCN homeostasis can lead to limbal stem cell dysfunction (LSCD) and subsequent ocular surface aberrations, such as corneal stromal inflammation, persistent epithelial defects, corneal neovascularisation, lymphangiogenesis, corneal opacification, and conjunctivalization. As ocular surface disorders are considered the second main cause of blindness, it becomes crucial to explore different therapeutic strategies for restoring the functions of the LSCN. A major limitation of corneal transplantation is the current shortage of donor tissue to meet the requirements worldwide. In this context, it becomes mandatory to find an alternative regenerative medicine, such as using cultured limbal epithelial/stromal stem cells, inducing the production of corneal like cells by using other sources of stem cells, and using tissue engineering methods aiming to produce the three-dimensional (3D) printed cornea. Limbal epithelial stem cells have been considered the magic potion for eye treatment. Epithelial and stromal stem cells in the limbal niche hold the responsibility of replenishing the corneal epithelium. These stem cells are being used for transplantation to maintain corneal epithelial integrity and ultimately sustain optimal vision. In this review, we summarised the characteristics of the LSCN and their current and future roles in restoring corneal homeostasis in eyes with LSCD.
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Affiliation(s)
- Fatemeh Tavakkoli
- Department of Community Health, College of Health Technology, Cihan University, Erbil, Iraq.,SSR Stem Cell Biology Laboratory, Brien Holden Eye Research Centre, Centre for Ocular Regeneration, Hyderabad Eye Research Foundation, L.V. Prasad Eye Institute, Hyderabad, India.,Centre for Genetic Disorders, Banaras Hindu University, Varanasi, India
| | - Taher K Eleiwa
- Department of Ophthalmology, Benha University, Benha, Egypt
| | - Abdelrahman M Elhusseiny
- Department of Ophthalmology, Harvey and Bernice Jones Eye Institute, University of Arkansas for Medical Sciences, Little Rock, AR, USA
| | - Mukesh Damala
- SSR Stem Cell Biology Laboratory, Brien Holden Eye Research Centre, Centre for Ocular Regeneration, Hyderabad Eye Research Foundation, L.V. Prasad Eye Institute, Hyderabad, India.,School of Life Sciences, University of Hyderabad, Hyderabad, India
| | - Amit K Rai
- Centre for Genetic Disorders, Banaras Hindu University, Varanasi, India
| | - Kasra Cheraqpour
- Translational Eye Research Center, Farabi Eye Hospital, 48439Tehran University of Medical Sciences, Tehran, Iran
| | - Mohammad H Ansari
- Ophthalmic Research Center, Department of Ophthalmology, Labbafinejad Medical Center, 556492Shahid Beheshti University of Medical Sciences, Tehran, Iran
| | - Mohammad Doroudian
- Department of Cell and Molecular Biology, Faculty of Biological Sciences, 145440Kharazmi University, Tehran, Iran
| | - Saeed H Keshel
- Department of Tissue Engineering and Applied Cell Sciences, 556492Shahid Beheshti University of Medical Sciences, Tehran, Iran
| | - Mohammad Soleimani
- Department of Ophthalmology, 159636Illinois Eye and Ear Infirmary, University of Illinois at Chicago, Chicago, IL, USA
| | - Ali R Djalilian
- Department of Ophthalmology, 159636Illinois Eye and Ear Infirmary, University of Illinois at Chicago, Chicago, IL, USA
| | | | - Vivek Singh
- SSR Stem Cell Biology Laboratory, Brien Holden Eye Research Centre, Centre for Ocular Regeneration, Hyderabad Eye Research Foundation, L.V. Prasad Eye Institute, Hyderabad, India
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23
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Li Z, Yang K, Zhou Y, Wu T, Zhang H, Yang Q, Wang Q, Huang Y, Wang L. Outcome of keratolimbal allograft transplantation with deep anterior lamellar keratoplasty for bilateral limbal stem cell deficiency. Front Med (Lausanne) 2022; 9:986194. [PMID: 36457564 PMCID: PMC9705574 DOI: 10.3389/fmed.2022.986194] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/04/2022] [Accepted: 10/21/2022] [Indexed: 01/09/2024] Open
Abstract
OBJECTIVES To evaluate and compare the outcome of keratolimbal allograft (KLAL) transplantation with or without deep anterior lamellar keratoplasty (DALK) for bilateral severe limbal stem cell deficiency (LSCD). METHODS This retrospective review included 49 eyes of 46 patients who underwent KLAL transplantation at the Department of Ophthalmology of Chinese PLA general hospital, 2009-2020, for bilateral severe LSCD were examined for corneal clarity and corneal scarring to determine whether to combine DALK with KLAL transplantation. Preoperative information, surgical decision tree, surgical procedures, and postoperative data were collected for each eye. RESULTS All patients had preoperative severe or total LSCD. Twenty-four eyes underwent KLAL transplantation only, 25 KLAL transplantation plus DALK. The mean follow-up was 46.80 ± 31.22 months (18-158 months). Overall KLAL survival (with or without DALK) was 71.43% at the final follow-up (KLAL-only 66.67%, KLAL-DALK 76%). Kaplan-Meier survival analysis showed that the 3-year survival probability of all grafts was 70.53 ± 10.89% (KLAL-only 64.86 ± 10.11%, KLAL-DALK 75.79 ± 8.62%). The proportion of BCVA ≥ 20/200 eyes among all KLAL transplantations increased from 11 eyes (22.45%) preoperatively to 25 eyes (51.02%) after 1 year and 24 eyes (48.98%) at the last follow-up (P = 0.01). The proportion of BCVA ≥ 20/200 eyes in the KLAL-DALK group increased significantly (P = 0.04), from 16.0% at baseline to 48.0% after 1 year to 44.0% at the last follow-up. Seventeen eyes (34.69%) had postoperative complications. CONCLUSION KLAL-DALK is an effective option to restore a stable ocular surface and visual acuity rapidly in patients with bilateral, late-stage, severe LSCD.
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Affiliation(s)
- Zongyuan Li
- Medical School of Chinese PLA, Beijing, China
- Department of Ophthalmology, The First Medical Center, Chinese PLA General Hospital, Beijing, China
| | - Kunkun Yang
- Medical School of Chinese PLA, Beijing, China
- Department of Ophthalmology, The First Medical Center, Chinese PLA General Hospital, Beijing, China
| | - Yannan Zhou
- Medical School of Chinese PLA, Beijing, China
- Department of Ophthalmology, The First Medical Center, Chinese PLA General Hospital, Beijing, China
| | - Tengyun Wu
- Medical School of Chinese PLA, Beijing, China
- Department of Ophthalmology, The First Medical Center, Chinese PLA General Hospital, Beijing, China
| | - Hongtao Zhang
- Senior Department of Ophthalmology, The Third Medical Center, Chinese PLA General Hospital, Beijing, China
| | - Qinghua Yang
- Senior Department of Ophthalmology, The Third Medical Center, Chinese PLA General Hospital, Beijing, China
| | - Qun Wang
- Senior Department of Ophthalmology, The Third Medical Center, Chinese PLA General Hospital, Beijing, China
| | - Yifei Huang
- Department of Ophthalmology, The First Medical Center, Chinese PLA General Hospital, Beijing, China
- Senior Department of Ophthalmology, The Third Medical Center, Chinese PLA General Hospital, Beijing, China
| | - Liqiang Wang
- Department of Ophthalmology, The First Medical Center, Chinese PLA General Hospital, Beijing, China
- Senior Department of Ophthalmology, The Third Medical Center, Chinese PLA General Hospital, Beijing, China
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Menzel-Severing J, Spaniol K, Groeber-Becker F, Geerling G. [Regenerative medicine for the corneal epithelium : Cell therapy from bench to bedside]. DIE OPHTHALMOLOGIE 2022; 119:891-901. [PMID: 35925345 DOI: 10.1007/s00347-022-01674-8] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Accepted: 05/31/2022] [Indexed: 06/15/2023]
Abstract
In the case of thermal or caustic burns of the ocular surface, loss of limbal epithelial stem cells leads to compromised self-renewal of the corneal epithelium. This results in permanent loss of vision. In these situations, transplantation of cultured limbal epithelial cells on an amniotic membrane or fibrin gel as substrate (Holoclar®) can help to regenerate the corneal surface. The required cells are obtained from the healthy partner eye, if available. Adult stem cells from other parts of the body potentially serve as alternative cell sources: hair follicles, oral mucosa, mesenchymal stromal cells, or induced pluripotent stem cells (originally, e.g., skin fibroblasts). The reprogramming of such cells can be achieved with the help of transcription factors. In addition, work is being done on biosynthetic or synthetic matrices, which not only serve as substrate material for the transplantation but also support the functional properties of these cells (self-renewal, corneal epithelial-typical phenotype).
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Affiliation(s)
- Johannes Menzel-Severing
- Klinik für Augenheilkunde, Universitätsklinikum Düsseldorf, Moorenstr. 5, 40225, Düsseldorf, Deutschland.
| | - Kristina Spaniol
- Klinik für Augenheilkunde, Universitätsklinikum Düsseldorf, Moorenstr. 5, 40225, Düsseldorf, Deutschland
| | - Florian Groeber-Becker
- Translationszentrum Regenerative Therapien | TLZ-RT, Leitung In-vitro-Testsysteme, Fraunhofer-Institut für Silicatforschung ISC, Würzburg, Deutschland
| | - Gerd Geerling
- Klinik für Augenheilkunde, Universitätsklinikum Düsseldorf, Moorenstr. 5, 40225, Düsseldorf, Deutschland
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Le-Bel G, Desjardins P, Gross C, Cortez Ghio S, Couture C, Germain L, Guérin SL. Influence of the Postmortem/Storage Time of Human Corneas on the Properties of Cultured Limbal Epithelial Cells. Cells 2022; 11:cells11172716. [PMID: 36078126 PMCID: PMC9455001 DOI: 10.3390/cells11172716] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/01/2022] [Revised: 08/25/2022] [Accepted: 08/26/2022] [Indexed: 11/25/2022] Open
Abstract
Besides being a powerful model to study the mechanisms of corneal wound healing, tissue-engineered human corneas (hTECs) are sparking interest as suitable substitutes for grafting purposes. To ensure the histological and physiological integrity of hTECs, the primary cultures generated from human cornea (identified as human limbal epithelial cells (hLECs) that are used to produce them must be of the highest possible quality. The goal of the present study consisted in evaluating the impact of the postmortem/storage time (PM/ST) on their properties in culture. hLECs were isolated from the entire cornea comprising the limbus and central cornea. When grown as monolayers, short PM/ST hLECs displayed increased daily doublings and generated more colonies per seeded cells than long PM/ST hLECs. Moreover, hLECs with a short PM/ST exhibited a markedly faster wound closure kinetic both in scratch wound assays and hTECs. Collectively, these results suggest that short PM/ST hLECs have a greater number of highly proliferative stem cells, exhibit a faster and more efficient wound healing response in vitro, and produce hTECs of a higher quality, making them the best candidates to produce biomaterial substitutes for clinical studies.
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Affiliation(s)
- Gaëtan Le-Bel
- Centre de Recherche en Organogénèse Expérimentale de l’Université Laval/LOEX, and Centre de Recherche du CHU de Québec-Université Laval, Axe Médecine Régénératrice, Québec, QC G1J 1Z4, Canada
- Centre Universitaire d’Ophtalmologie (CUO)-Recherche, Centre de Recherche du CHU de Québec-Université Laval, Axe Médecine Régénératrice, Québec, QC G1S 4L8, Canada
- Département de Chirurgie, Faculté de Médecine, Université Laval, Québec, QC G1V 0A6, Canada
- Département d’Ophtalmologie, Faculté de Médecine, Université Laval, Québec, QC G1V 0A6, Canada
| | - Pascale Desjardins
- Centre de Recherche en Organogénèse Expérimentale de l’Université Laval/LOEX, and Centre de Recherche du CHU de Québec-Université Laval, Axe Médecine Régénératrice, Québec, QC G1J 1Z4, Canada
- Centre Universitaire d’Ophtalmologie (CUO)-Recherche, Centre de Recherche du CHU de Québec-Université Laval, Axe Médecine Régénératrice, Québec, QC G1S 4L8, Canada
- Département de Chirurgie, Faculté de Médecine, Université Laval, Québec, QC G1V 0A6, Canada
- Département d’Ophtalmologie, Faculté de Médecine, Université Laval, Québec, QC G1V 0A6, Canada
| | - Christelle Gross
- Centre Universitaire d’Ophtalmologie (CUO)-Recherche, Centre de Recherche du CHU de Québec-Université Laval, Axe Médecine Régénératrice, Québec, QC G1S 4L8, Canada
- Département d’Ophtalmologie, Faculté de Médecine, Université Laval, Québec, QC G1V 0A6, Canada
| | - Sergio Cortez Ghio
- Centre de Recherche en Organogénèse Expérimentale de l’Université Laval/LOEX, and Centre de Recherche du CHU de Québec-Université Laval, Axe Médecine Régénératrice, Québec, QC G1J 1Z4, Canada
- Département de Chirurgie, Faculté de Médecine, Université Laval, Québec, QC G1V 0A6, Canada
| | - Camille Couture
- Centre de Recherche en Organogénèse Expérimentale de l’Université Laval/LOEX, and Centre de Recherche du CHU de Québec-Université Laval, Axe Médecine Régénératrice, Québec, QC G1J 1Z4, Canada
- Centre Universitaire d’Ophtalmologie (CUO)-Recherche, Centre de Recherche du CHU de Québec-Université Laval, Axe Médecine Régénératrice, Québec, QC G1S 4L8, Canada
- Département de Chirurgie, Faculté de Médecine, Université Laval, Québec, QC G1V 0A6, Canada
- Département d’Ophtalmologie, Faculté de Médecine, Université Laval, Québec, QC G1V 0A6, Canada
| | - Lucie Germain
- Centre de Recherche en Organogénèse Expérimentale de l’Université Laval/LOEX, and Centre de Recherche du CHU de Québec-Université Laval, Axe Médecine Régénératrice, Québec, QC G1J 1Z4, Canada
- Centre Universitaire d’Ophtalmologie (CUO)-Recherche, Centre de Recherche du CHU de Québec-Université Laval, Axe Médecine Régénératrice, Québec, QC G1S 4L8, Canada
- Département de Chirurgie, Faculté de Médecine, Université Laval, Québec, QC G1V 0A6, Canada
- Département d’Ophtalmologie, Faculté de Médecine, Université Laval, Québec, QC G1V 0A6, Canada
| | - Sylvain L. Guérin
- Centre de Recherche en Organogénèse Expérimentale de l’Université Laval/LOEX, and Centre de Recherche du CHU de Québec-Université Laval, Axe Médecine Régénératrice, Québec, QC G1J 1Z4, Canada
- Centre Universitaire d’Ophtalmologie (CUO)-Recherche, Centre de Recherche du CHU de Québec-Université Laval, Axe Médecine Régénératrice, Québec, QC G1S 4L8, Canada
- Département d’Ophtalmologie, Faculté de Médecine, Université Laval, Québec, QC G1V 0A6, Canada
- Correspondence: ; Tel.: +1-418-682-7565
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Jurkunas U, Johns L, Armant M. Cultivated Autologous Limbal Epithelial Cell Transplantation: New Frontier in the Treatment of Limbal Stem Cell Deficiency. Am J Ophthalmol 2022; 239:244-268. [PMID: 35314191 DOI: 10.1016/j.ajo.2022.03.015] [Citation(s) in RCA: 10] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/04/2020] [Revised: 01/27/2022] [Accepted: 03/08/2022] [Indexed: 11/19/2022]
Abstract
PURPOSE Taking into consideration prior human experience with treating limbal stem cell deficiency (LSCD) with cultivated limbal epithelial cells (CLEC) from other countries, we have set a goal to optimize and standardize the techniques of CLEC preparation (called CALEC by our group) for the clinical trial in the United States. METHODS We performed an extensive literature review of all human trials, case series, and reports involving autologous cultivated limbal epithelial cell transplantation. Allogeneic cultivated limbal epithelial cell transplantations were reported only when combined with autologous studies. We also searched prior animal data aiding in detailing regulatory toxicology requirements. RESULTS Between 1997 and 2020, the analysis of human trials revealed 21 studies on autologous grafts, and 13 studies analyzing both autologous grafts and allogeneic grafts. Of a total of 34 studies, 6 studies used good manufacturing process (GMP) facilities, and 11 studies had no animal-derived products or murine feeder layers, whereas only 1 study had both. Overall, the treatment with autologous CLEC grafts was 68.9% successful. In total there were 6 preclinical studies using rabbits, serving as surrogate studies to assess the safety and toxicity of cultivated limbal epithelial cells for human trials. Based on prior human experience, we further optimized the manufacturing conditions with GMP-grade and serum and animal-free reagents, and developed cell characterization assays for the CALEC product release. CONCLUSIONS These data were used to develop a novel and consistent manufacturing process using only qualified and validated reagents for performing the first clinical trial on CALEC transplantation to treat LSCD in the United States.
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Affiliation(s)
- Ula Jurkunas
- From the Schepens Eye Research Institute (U.J., L.J.), Massachusetts Eye and Ear, Boston, Massachusetts, USA.
| | - Lynette Johns
- From the Schepens Eye Research Institute (U.J., L.J.), Massachusetts Eye and Ear, Boston, Massachusetts, USA
| | - Myriam Armant
- TransLab (M.A.), Translational Research Program, Boston Children's Hospital, Boston, Massachusetts, USA
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27
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Nguyen HT, Theerakittayakorn K, Somredngan S, Ngernsoungnern A, Ngernsoungnern P, Sritangos P, Ketudat-Cairns M, Imsoonthornruksa S, Assawachananont J, Keeratibharat N, Wongsan R, Rungsiwiwut R, Laowtammathron C, Bui NX, Parnpai R. Signaling Pathways Impact on Induction of Corneal Epithelial-like Cells Derived from Human Wharton’s Jelly Mesenchymal Stem Cells. Int J Mol Sci 2022; 23:ijms23063078. [PMID: 35328499 PMCID: PMC8949174 DOI: 10.3390/ijms23063078] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/08/2022] [Revised: 03/07/2022] [Accepted: 03/10/2022] [Indexed: 02/05/2023] Open
Abstract
Corneal epithelium, the outmost layer of the cornea, comprises corneal epithelial cells (CECs) that are continuously renewed by limbal epithelial stem cells (LESCs). Loss or dysfunction of LESCs causes limbal stem cell deficiency (LSCD) which results in corneal epithelial integrity loss and visual impairment. To regenerate the ocular surface, transplantation of stem cell-derived CECs is necessary. Human Wharton’s jelly derived mesenchymal stem cells (WJ-MSCs) are a good candidate for cellular therapies in allogeneic transplantation. This study aimed to test the effects of treatments on three signaling pathways involved in CEC differentiation as well as examine the optimal protocol for inducing corneal epithelial differentiation of human WJ-MSCs. All-trans retinoic acid (RA, 5 or 10 µM) inhibited the Wnt signaling pathway via suppressing the translocation of β-catenin from the cytoplasm into the nucleus. SB505124 downregulated the TGF-β signaling pathway via reducing phosphorylation of Smad2. BMP4 did not increase phosphorylation of Smad1/5/8 that is involved in BMP signaling. The combination of RA, SB505124, BMP4, and EGF for the first 3 days of differentiation followed by supplementing hormonal epidermal medium for an additional 6 days could generate corneal epithelial-like cells that expressed a CEC specific marker CK12. This study reveals that WJ-MSCs have the potential to transdifferentiate into CECs which would be beneficial for further applications in LSCD treatment therapy.
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Affiliation(s)
- Hong Thi Nguyen
- Embryo Technology and Stem Cell Research Center, School of Biotechnology, Institute of Agricultural Technology, Suranaree University of Technology, Nakhon Ratchasima 30000, Thailand; (H.T.N.); (K.T.); (S.S.)
- Laboratory of Embryo Technology, Institute of Biotechnology, Vietnam Academy of Science and Technology, Hanoi 100000, Vietnam
| | - Kasem Theerakittayakorn
- Embryo Technology and Stem Cell Research Center, School of Biotechnology, Institute of Agricultural Technology, Suranaree University of Technology, Nakhon Ratchasima 30000, Thailand; (H.T.N.); (K.T.); (S.S.)
| | - Sirilak Somredngan
- Embryo Technology and Stem Cell Research Center, School of Biotechnology, Institute of Agricultural Technology, Suranaree University of Technology, Nakhon Ratchasima 30000, Thailand; (H.T.N.); (K.T.); (S.S.)
| | - Apichart Ngernsoungnern
- School of Preclinical Sciences, Institute of Science, Suranaree University of Technology, Nakhon Ratchasima 30000, Thailand; (A.N.); (P.N.); (P.S.)
| | - Piyada Ngernsoungnern
- School of Preclinical Sciences, Institute of Science, Suranaree University of Technology, Nakhon Ratchasima 30000, Thailand; (A.N.); (P.N.); (P.S.)
| | - Pishyaporn Sritangos
- School of Preclinical Sciences, Institute of Science, Suranaree University of Technology, Nakhon Ratchasima 30000, Thailand; (A.N.); (P.N.); (P.S.)
| | - Mariena Ketudat-Cairns
- School of Biotechnology, Institute of Agricultural Technology, Suranaree University of Technology, Nakhon Ratchasima 30000, Thailand; (M.K.-C.); (S.I.)
| | - Sumeth Imsoonthornruksa
- School of Biotechnology, Institute of Agricultural Technology, Suranaree University of Technology, Nakhon Ratchasima 30000, Thailand; (M.K.-C.); (S.I.)
| | - Juthaporn Assawachananont
- School of Ophthalmology, Institute of Medicine, Suranaree University of Technology, Nakhon Ratchasima 30000, Thailand;
| | - Nattawut Keeratibharat
- School of Surgery, Institute of Medicine, Suranaree University of Technology, Nakhon Ratchasima 30000, Thailand;
| | - Rangsirat Wongsan
- The Center for Scientific and Technological Equipment, Suranaree University of Technology, Nakhon Ratchasima 30000, Thailand;
| | - Ruttachuk Rungsiwiwut
- Department of Anatomy, Faculty of Medicine, Srinakharinwirot University, Bangkok 10000, Thailand;
| | - Chuti Laowtammathron
- Siriraj Center of Excellence for Stem Cell Research, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok 10000, Thailand;
| | | | - Rangsun Parnpai
- Embryo Technology and Stem Cell Research Center, School of Biotechnology, Institute of Agricultural Technology, Suranaree University of Technology, Nakhon Ratchasima 30000, Thailand; (H.T.N.); (K.T.); (S.S.)
- Correspondence: ; Tel.: +66-442-242-34
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Lasagni Vitar R, Triani F, Barbariga M, Fonteyne P, Rama P, Ferrari G. Substance P/neurokinin-1 receptor pathway blockade ameliorates limbal stem cell deficiency by modulating mTOR pathway and preventing cell senescence. Stem Cell Reports 2022; 17:849-863. [PMID: 35334220 PMCID: PMC9023781 DOI: 10.1016/j.stemcr.2022.02.012] [Citation(s) in RCA: 9] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/06/2021] [Revised: 02/18/2022] [Accepted: 02/21/2022] [Indexed: 11/01/2022] Open
Abstract
Severe ocular surface diseases can lead to limbal stem cell deficiency (LSCD), which is accompanied by defective healing. We aimed to evaluate the role of the substance P (SP)/neurokinin-1 receptor (NK1R) pathway in corneal epithelium wound healing in a pre-clinical model of LSCD. SP ablation or NK1R blockade significantly increased epithelial wound healing (p < 0.001) and corneal transparency (p < 0.001), compared with wild type (WT). In addition, a reduced number of infiltrating goblet and conjunctival cells (p < 0.05) and increased number of epithelial stem cells (p < 0.01), which also expressed NK1R, was observed. The mammalian target of rapamycin (mTOR) pathway was significantly inhibited (p < 0.05) and expression of γH2AX was significantly reduced (p < 0.05) after SP ablation. These results suggest that excessive expression of SP is associated with LSCD and results in accelerated senescence and exhaustion of residual stem cells. Topical treatment with NK1R antagonist ameliorates clinical signs associated with LSCD and could be used as an adjuvant treatment in LSCD.
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Affiliation(s)
- Romina Lasagni Vitar
- Cornea and Ocular Surface Disease Unit, Eye Repair Lab, IRCCS San Raffaele Scientific Institute, Via Olgettina 60, 20132 Milan, Italy
| | - Francesca Triani
- Cornea and Ocular Surface Disease Unit, Eye Repair Lab, IRCCS San Raffaele Scientific Institute, Via Olgettina 60, 20132 Milan, Italy
| | - Marco Barbariga
- Cornea and Ocular Surface Disease Unit, Eye Repair Lab, IRCCS San Raffaele Scientific Institute, Via Olgettina 60, 20132 Milan, Italy
| | - Philippe Fonteyne
- Cornea and Ocular Surface Disease Unit, Eye Repair Lab, IRCCS San Raffaele Scientific Institute, Via Olgettina 60, 20132 Milan, Italy
| | - Paolo Rama
- Cornea and Ocular Surface Disease Unit, Eye Repair Lab, IRCCS San Raffaele Scientific Institute, Via Olgettina 60, 20132 Milan, Italy
| | - Giulio Ferrari
- Cornea and Ocular Surface Disease Unit, Eye Repair Lab, IRCCS San Raffaele Scientific Institute, Via Olgettina 60, 20132 Milan, Italy.
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Mohanna S, Elahi S, Panthier C, Gatinel D. Surgical approaches to autologous limbal stem cell transplantation (LSCT) following severe corneal chemical burns. J Fr Ophtalmol 2022; 45:352-357. [DOI: 10.1016/j.jfo.2021.08.003] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/07/2021] [Revised: 08/21/2021] [Accepted: 08/27/2021] [Indexed: 11/27/2022]
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Evaluating the clinical translational relevance of animal models for limbal stem cell deficiency: A systematic review. Ocul Surf 2021; 23:169-183. [PMID: 34583088 DOI: 10.1016/j.jtos.2021.09.006] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/07/2021] [Revised: 09/20/2021] [Accepted: 09/23/2021] [Indexed: 12/13/2022]
Abstract
PURPOSE Animal models are pivotal for elucidating pathophysiological mechanisms and evaluating novel therapies. This systematic review identified studies that developed or adapted animal models of limbal stem cell deficiency (LSCD), assessed their reporting quality, summarized their key characteristics, and established their clinical translational relevance to human disease. METHODS The protocol was prospectively registered (PROSPERO CRD42020203937). Searches were conducted in PubMed, Ovid EMBASE and Web of Science in August 2020. Two authors screened citations, extracted data, assessed the reporting quality of eligible studies using the ARRIVE guidelines, and judged the clinical translational relevance of each model using a custom matrix. RESULTS 105 studies were included. Rabbits were the most common animal species. Overall, 97% of studies recapitulated LSCD to a clinical etiology, however 62% did not provide sufficient methodological detail to enable independent reproduction of the model. Adverse events and/or exclusion of animals were infrequently (20%) reported. Approximately one-quarter of studies did not produce the intended severity of LSCD; 34% provided insufficient information to assess the fidelity of disease induction. Adjunctive diagnostic confirmation of LSCD induction was performed in 13% of studies. CONCLUSIONS This is the first systematic review to assess the reporting quality and clinical translational relevance of animal models of LSCD. Models of LSCD have evolved over time, resulting in variable reporting of the characteristics of animals, experimental procedures and adverse events. In most studies, validation of LSCD was made using clinical tests; newer adjunctive techniques would enhance diagnostic validation. As most studies sought to evaluate novel therapies for LSCD, animal models should ideally recapitulate all features of the condition that develop in patients.
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Goals and Challenges of Stem Cell-Based Therapy for Corneal Blindness Due to Limbal Deficiency. Pharmaceutics 2021; 13:pharmaceutics13091483. [PMID: 34575560 PMCID: PMC8466237 DOI: 10.3390/pharmaceutics13091483] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/30/2021] [Revised: 09/09/2021] [Accepted: 09/13/2021] [Indexed: 12/13/2022] Open
Abstract
Corneal failure is a highly prevalent cause of blindness. One special cause of corneal failure occurs due to malfunction or destruction of the limbal stem cell niche, upon which the superficial cornea depends for homeostatic maintenance and wound healing. Failure of the limbal niche is referred to as limbal stem cell deficiency. As the corneal epithelial stem cell niche is easily accessible, limbal stem cell-based therapy and regenerative medicine applied to the ocular surface are among the most highly advanced forms of this novel approach to disease therapy. However, the challenges are still great, including the development of cell-based products and understanding how they work in the patient's eye. Advances are being made at the molecular, cellular, and tissue levels to alter disease processes and to reduce or eliminate blindness. Efforts must be coordinated from the most basic research to the most clinically oriented projects so that cell-based therapies can become an integrated part of the therapeutic armamentarium to fight corneal blindness. We undoubtedly are progressing along the right path because cell-based therapy for eye diseases is one of the most successful examples of global regenerative medicine.
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Corneal Epithelial Stem Cells-Physiology, Pathophysiology and Therapeutic Options. Cells 2021; 10:cells10092302. [PMID: 34571952 PMCID: PMC8465583 DOI: 10.3390/cells10092302] [Citation(s) in RCA: 32] [Impact Index Per Article: 8.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/01/2021] [Revised: 08/27/2021] [Accepted: 08/28/2021] [Indexed: 12/12/2022] Open
Abstract
In the human cornea, regeneration of the epithelium is regulated by the stem cell reservoir of the limbus, which is the marginal region of the cornea representing the anatomical and functional border between the corneal and conjunctival epithelium. In support of this concept, extensive limbal damage, e.g., by chemical or thermal injury, inflammation, or surgery, may induce limbal stem cell deficiency (LSCD) leading to vascularization and opacification of the cornea and eventually vision loss. These acquired forms of limbal stem cell deficiency may occur uni- or bilaterally, which is important for the choice of treatment. Moreover, a variety of inherited diseases, such as congenital aniridia or dyskeratosis congenita, are characterized by LSCD typically occurring bilaterally. Several techniques of autologous and allogenic stem cell transplantation have been established. The limbus can be restored by transplantation of whole limbal grafts, small limbal biopsies or by ex vivo-expanded limbal cells. In this review, the physiology of the corneal epithelium, the pathophysiology of LSCD, and the therapeutic options will be presented.
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Romo-Valera C, Pérez-Garrastachu M, Hernáez-Moya R, Rodriguez-Astigarraga M, Romano-Ruiz P, Etxebarria J, Arluzea J, Andollo N. Characterisation of corneas following different time and storage methods for their use as a source of stem-like limbal epithelial cells. Exp Eye Res 2021; 211:108720. [PMID: 34389315 DOI: 10.1016/j.exer.2021.108720] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/29/2021] [Revised: 07/21/2021] [Accepted: 08/05/2021] [Indexed: 12/13/2022]
Abstract
The transplantation of expansions of limbal epithelial stem cells (LESC) remains one of the most efficient therapies for the treatment of limbal stem cell deficiency (LSCD) to date. However, the available donor corneas are scarce, and the corneas conserved for long time, under hypothermic conditions (after 7 days) or in culture (more than 28 days), are usually discarded due to poor viability of the endothelial cells. To establish an objective criterion for the utilisation or discarding of corneas as a source of LESC, we characterized, by immunohistochemistry analysis, donor corneas conserved in different conditions and for different periods of time. We also studied the potency of LESCs isolated from these corneas and maintained in culture up to 3 cell passages. We hoped that the study of markers of LESCs present in both the corneoscleral histological sections and the cell cultures would show the adequacy of the methods used for cell isolation and how fit the LESC enrichment of the obtained cell populations to be expanded was. Thus, the expressions of markers of the cells residing in the human limbal and corneal epithelium (cytokeratin CK15 and CK12, vimentin, Collagen VII, p63α, ABCG2, Ki67, Integrin β4, ZO1, and melan A) were analysed in sections of corneoscleral tissues conserved in hypothermic conditions for 2-9 days with post-mortem time (pmt) < 8 h or for 1 day with pmt > 16 h, and in sclerocorneal rims maintained in an organ culture medium for 29 days. Cell populations isolated from donor corneoscleral tissues were also assessed based on these markers to verify the adequacy of isolation methods and the potential of expanding LESCs from these tissues. Positivity for several putative stem cell markers such as CK15 and p63α was detected in all corneoscleral tissues, although a decrease was recorded in the ones conserved for longer times. The barrier function and the ability to adhere to the extracellular matrix were maintained in all the analysed tissues. In limbal epithelial cell cultures, a simultaneous decrease in the melan A melanocyte marker and the putative stem cell markers was detected, suggesting a close relationship between the melanocytes and the limbal stem cells of the niche. Holoclones stained with putative stem cell markers were obtained from long-term, hypothermic, stored sclerocorneal rims. The results showed that the remaining sclerocorneal rims after corneal transplantation, which were conserved under hypothermic conditions for up to 7 days and would have been discarded at a first glance, still maintained their potential as a source of LESC cultures.
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Affiliation(s)
- Cristina Romo-Valera
- Department of Cell Biology and Histology, School of Medicine and Nursing, University of the Basque Country, Sarriena, S/N, 48940, Leioa, Spain
| | - Miguel Pérez-Garrastachu
- Department of Cell Biology and Histology, School of Medicine and Nursing, University of the Basque Country, Sarriena, S/N, 48940, Leioa, Spain
| | - Raquel Hernáez-Moya
- Department of Cell Biology and Histology, School of Medicine and Nursing, University of the Basque Country, Sarriena, S/N, 48940, Leioa, Spain
| | - Maddalen Rodriguez-Astigarraga
- Department of Cell Biology and Histology, School of Medicine and Nursing, University of the Basque Country, Sarriena, S/N, 48940, Leioa, Spain
| | - Paula Romano-Ruiz
- Department of Cell Biology and Histology, School of Medicine and Nursing, University of the Basque Country, Sarriena, S/N, 48940, Leioa, Spain
| | - Jaime Etxebarria
- Department of Cell Biology and Histology, School of Medicine and Nursing, University of the Basque Country, Sarriena, S/N, 48940, Leioa, Spain; Department of Ophthalmology, University Hospital of Cruces, Cruces Plaza S/N, 48903, Barakaldo, Spain; BioCruces Bizkaia Health Research Institute, Begiker, Cruces Plaza S/N, 48903, Barakaldo, Spain
| | - Jon Arluzea
- Department of Cell Biology and Histology, School of Medicine and Nursing, University of the Basque Country, Sarriena, S/N, 48940, Leioa, Spain
| | - Noelia Andollo
- Department of Cell Biology and Histology, School of Medicine and Nursing, University of the Basque Country, Sarriena, S/N, 48940, Leioa, Spain; BioCruces Bizkaia Health Research Institute, Begiker, Cruces Plaza S/N, 48903, Barakaldo, Spain.
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Singh V, Tiwari A, Kethiri AR, Sangwan VS. Current perspectives of limbal-derived stem cells and its application in ocular surface regeneration and limbal stem cell transplantation. Stem Cells Transl Med 2021; 10:1121-1128. [PMID: 33951336 PMCID: PMC8284782 DOI: 10.1002/sctm.20-0408] [Citation(s) in RCA: 19] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/31/2020] [Revised: 02/11/2021] [Accepted: 03/04/2021] [Indexed: 12/13/2022] Open
Abstract
Limbal stem cells are involved in replenishing and maintaining the epithelium of the cornea. Damage to the limbus due to chemical/physical injury, infections, or genetic disorders leads to limbal stem cell deficiency (LSCD) with partial or total vision loss. Presently, LSCD is treated by transplanting limbal stem cells from the healthy eye of the recipient, living-related, or cadaveric donors. This review discusses limbal-derived stem cells, the importance of extracellular matrix in stem cell niche maintenance, the historical perspective of treating LSCD, including related advantages and limitations, and our experience of limbal stem cell transplantation over the decades.
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Affiliation(s)
- Vivek Singh
- Stem Cell Biology Laboratory, Center for Ocular Regeneration, L V Prasad Eye Institute, Hyderabad, India
| | - Anil Tiwari
- Department of Ophthalmology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, USA
- Department of Cornea and Uveitis, Dr. Shroff's Charity Eye Hospital, New Delhi, India
| | - Abhinav Reddy Kethiri
- Stem Cell Biology Laboratory, Center for Ocular Regeneration, L V Prasad Eye Institute, Hyderabad, India
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Prabhasawat P, Chirapapaisan C, Ngowyutagon P, Ekpo P, Tangpagasit W, Lekhanont K, Sikarinkul R, Matamnan S, Boonwong C, Pinitpuwadol W, Thamphithak R, Sukon N, Neti N. Efficacy and outcome of simple limbal epithelial transplantation for limbal stem cell deficiency verified by epithelial phenotypes integrated with clinical evaluation. Ocul Surf 2021; 22:27-37. [PMID: 34214675 DOI: 10.1016/j.jtos.2021.06.012] [Citation(s) in RCA: 12] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/21/2021] [Revised: 06/13/2021] [Accepted: 06/28/2021] [Indexed: 11/17/2022]
Abstract
PURPOSE To evaluate the efficacy and outcome of simple limbal epithelial transplantation (SLET) for limbal stem cell deficiency (LSCD) using epithelial phenotype detection integrated with clinical manifestation. METHODS This prospective multicenter study included patients with LSCD who underwent autologous SLET (autoSLET) and living-related allogenic SLET (Lr-alloSLET). All patients were assessed by slit-lamp biomicroscopy, in vivo confocal microscopy (IVCM), and impression cytology with immunofluorescence staining (ICIF) before and after surgery. The criteria for success were the presence of a clinically non-conjunctivalized cornea and corneal epithelium detected by IVCM or ICIF. Otherwise, the case would be considered a failure. Visual improvement and risk factors for SLET failure were analyzed. RESULTS A total of 28 eyes of 26 patients (11 autoSLET and 17 Lr-alloSLET) were included. The median age was 53 years (range, 35-63), and the follow-up time was 29.5 months (range, 17.5-39.8). The overall survival rate was 89.3% at 2 years and 75.6% at 3 years with no difference between autoSLET and Lr-alloSLET (p = 0.24). Seven eyes subsequently underwent penetrating keratoplasty. Immunohistochemistry analysis showed that all corneal buttons had corneal epithelium and limbal stem cell markers. Visual improvement was achieved in both SLET groups (p < 0.001). Failed SLET developed between 5 and 32 months postoperatively. However, absolute risk factors for SLET failure were unidentified. CONCLUSION The efficacy of autoSLET and Lr-alloSLET for LSCD was excellent. Limbal explants can regenerate and restore the corneal surface while maintaining the characteristics of limbal stem cells as shown by epithelial phenotype detection and immunohistochemistry integrated with clinical evaluation.
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Affiliation(s)
- Pinnita Prabhasawat
- Department of Ophthalmology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand
| | - Chareenun Chirapapaisan
- Department of Ophthalmology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand.
| | - Panotsom Ngowyutagon
- Department of Ophthalmology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand
| | - Pattama Ekpo
- Department of Immunology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand; Research Division, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand
| | - Wimolwan Tangpagasit
- Department of Ophthalmology, Faculty of Medicine, Thammasat University, Prathumthani, Thailand
| | - Kaevalin Lekhanont
- Department of Ophthalmology, Faculty of Medicine Ramathibodi Hospital, Mahidol University, Thailand
| | - Rosanun Sikarinkul
- Department of Ophthalmology, Golden Jubilee Medical Center, Mahidol University, Thailand
| | - Sutthicha Matamnan
- Research Division, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand
| | - Chawikan Boonwong
- Research Division, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand
| | - Warinyupa Pinitpuwadol
- Department of Ophthalmology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand
| | - Ranida Thamphithak
- Department of Ophthalmology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand
| | - Nutchaya Sukon
- Department of Ophthalmology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand
| | - Nutnicha Neti
- Department of Ophthalmology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand
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Nieto-Nicolau N, Martínez-Conesa EM, Fuentes-Julián S, Arnalich-Montiel F, García-Tuñón I, De Miguel MP, Casaroli-Marano RP. Priming human adipose-derived mesenchymal stem cells for corneal surface regeneration. J Cell Mol Med 2021; 25:5124-5137. [PMID: 33951289 PMCID: PMC8178265 DOI: 10.1111/jcmm.16501] [Citation(s) in RCA: 28] [Impact Index Per Article: 7.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/24/2019] [Revised: 03/17/2021] [Accepted: 03/18/2021] [Indexed: 12/13/2022] Open
Abstract
Limbal stem cells (LSC) maintain the transparency of the corneal epithelium. Chemical burns lead the loss of LSC inducing an up-regulation of pro-inflammatory and pro-angiogenic factors, triggering corneal neovascularization and blindness. Adipose tissue-derived mesenchymal stem cells (AT-MSC) have shown promise in animal models to treat LSC deficiency (LSCD), but there are not studies showing their efficacy when primed with different media before transplantation. We cultured AT-MSC with standard medium and media used to culture LSC for clinical application. We demonstrated that different media changed the AT-MSC paracrine secretion showing different paracrine effector functions in an in vivo model of chemical burn and in response to a novel in vitro model of corneal inflammation by alkali induction. Treatment of LSCD with AT-MSC changed the angiogenic and inflammatory cytokine profile of mice corneas. AT-MSC cultured with the medium that improved their cytokine secretion, enhanced the anti-angiogenic and anti-inflammatory profile of the treated corneas. Those corneas also presented better outcome in terms of corneal transparency, neovascularization and histologic reconstruction. Priming human AT-MSC with LSC specific medium can potentiate their ability to improve corneal wound healing, decrease neovascularization and inflammation modulating paracrine effector functions in an in vivo optimized rat model of LSCD.
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Affiliation(s)
- Núria Nieto-Nicolau
- CellTec-UB, Department of Cell Biology, University of Barcelona, Barcelona, Spain.,Barcelona Tissue Bank (BTB), Banc de Sang I Teixits (BST), Barcelona, Spain.,Institute of Biomedical Research IIB-Sant Pau (SGR1113), Barcelona, Spain
| | - Eva M Martínez-Conesa
- Barcelona Tissue Bank (BTB), Banc de Sang I Teixits (BST), Barcelona, Spain.,Institute of Biomedical Research IIB-Sant Pau (SGR1113), Barcelona, Spain
| | | | | | - Ignacio García-Tuñón
- Cell Engineering Laboratory, La Paz Hospital Research Institute (IdiPAZ), Madrid, Spain
| | - María P De Miguel
- Cell Engineering Laboratory, La Paz Hospital Research Institute (IdiPAZ), Madrid, Spain
| | - Ricardo P Casaroli-Marano
- CellTec-UB, Department of Cell Biology, University of Barcelona, Barcelona, Spain.,Barcelona Tissue Bank (BTB), Banc de Sang I Teixits (BST), Barcelona, Spain.,Institute of Biomedical Research IIB-Sant Pau (SGR1113), Barcelona, Spain.,Department of Surgery & Hospital Clinic de Barcelona, School of Medicine, University of Barcelona, Barcelona, Spain
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37
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Collin J, Queen R, Zerti D, Bojic S, Dorgau B, Moyse N, Molina MM, Yang C, Dey S, Reynolds G, Hussain R, Coxhead JM, Lisgo S, Henderson D, Joseph A, Rooney P, Ghosh S, Clarke L, Connon C, Haniffa M, Figueiredo F, Armstrong L, Lako M. A single cell atlas of human cornea that defines its development, limbal progenitor cells and their interactions with the immune cells. Ocul Surf 2021; 21:279-298. [PMID: 33865984 PMCID: PMC8343164 DOI: 10.1016/j.jtos.2021.03.010] [Citation(s) in RCA: 117] [Impact Index Per Article: 29.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/15/2020] [Revised: 03/05/2021] [Accepted: 03/25/2021] [Indexed: 02/08/2023]
Abstract
Purpose Single cell (sc) analyses of key embryonic, fetal and adult stages were performed to generate a comprehensive single cell atlas of all the corneal and adjacent conjunctival cell types from development to adulthood. Methods Four human adult and seventeen embryonic and fetal corneas from 10 to 21 post conception week (PCW) specimens were dissociated to single cells and subjected to scRNA- and/or ATAC-Seq using the 10x Genomics platform. These were embedded using Uniform Manifold Approximation and Projection (UMAP) and clustered using Seurat graph-based clustering. Cluster identification was performed based on marker gene expression, bioinformatic data mining and immunofluorescence (IF) analysis. RNA interference, IF, colony forming efficiency and clonal assays were performed on cultured limbal epithelial cells (LECs). Results scRNA-Seq analysis of 21,343 cells from four adult human corneas and adjacent conjunctivas revealed the presence of 21 cell clusters, representing the progenitor and differentiated cells in all layers of cornea and conjunctiva as well as immune cells, melanocytes, fibroblasts, and blood/lymphatic vessels. A small cell cluster with high expression of limbal progenitor cell (LPC) markers was identified and shown via pseudotime analysis to give rise to five other cell types representing all the subtypes of differentiated limbal and corneal epithelial cells. A novel putative LPCs surface marker, GPHA2, expressed on the surface of 0.41% ± 0.21 of the cultured LECs, was identified, based on predominant expression in the limbal crypts of adult and developing cornea and RNAi validation in cultured LECs. Combining scRNA- and ATAC-Seq analyses, we identified multiple upstream regulators for LPCs and demonstrated a close interaction between the immune cells and limbal progenitor cells. RNA-Seq analysis indicated the loss of GPHA2 expression and acquisition of proliferative limbal basal epithelial cell markers during ex vivo LEC expansion, independently of the culture method used. Extending the single cell analyses to keratoconus, we were able to reveal activation of collagenase in the corneal stroma and a reduced pool of limbal suprabasal cells as two key changes underlying the disease phenotype. Single cell RNA-Seq of 89,897 cells obtained from embryonic and fetal cornea indicated that during development, the conjunctival epithelium is the first to be specified from the ocular surface epithelium, followed by the corneal epithelium and the establishment of LPCs, which predate the formation of limbal niche by a few weeks. Conclusions Our scRNA-and ATAC-Seq data of developing and adult cornea in steady state and disease conditions provide a unique resource for defining genes/pathways that can lead to improvement in ex vivo LPCs expansion, stem cell differentiation methods and better understanding and treatment of ocular surface disorders.
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Affiliation(s)
- Joseph Collin
- Biosciences Institute, Faculty of Medical Sciences, Newcastle University, UK
| | - Rachel Queen
- Biosciences Institute, Faculty of Medical Sciences, Newcastle University, UK
| | - Darin Zerti
- Biosciences Institute, Faculty of Medical Sciences, Newcastle University, UK
| | - Sanja Bojic
- Biosciences Institute, Faculty of Medical Sciences, Newcastle University, UK
| | - Birthe Dorgau
- Biosciences Institute, Faculty of Medical Sciences, Newcastle University, UK
| | - Nicky Moyse
- Newcastle Cellular Therapies Facility, Newcastle University and Newcastle Upon Tyne Hospitals NHS Foundation Trust, UK
| | - Marina Moya Molina
- Biosciences Institute, Faculty of Medical Sciences, Newcastle University, UK
| | - Chunbo Yang
- Biosciences Institute, Faculty of Medical Sciences, Newcastle University, UK
| | - Sunanda Dey
- Department of Genetics and Developmental Biology, The Ruth and Bruce Rappaport Faculty of Medicine, Technion - Israel Institute of Technology, Israel
| | - Gary Reynolds
- Biosciences Institute, Faculty of Medical Sciences, Newcastle University, UK
| | - Rafiqul Hussain
- Biosciences Institute, Faculty of Medical Sciences, Newcastle University, UK
| | - Jonathan M Coxhead
- Biosciences Institute, Faculty of Medical Sciences, Newcastle University, UK
| | - Steven Lisgo
- Biosciences Institute, Faculty of Medical Sciences, Newcastle University, UK
| | - Deborah Henderson
- Biosciences Institute, Faculty of Medical Sciences, Newcastle University, UK
| | - Agatha Joseph
- NHS Blood and Transplant Tissue and Eye Services, Liverpool, UK
| | - Paul Rooney
- NHS Blood and Transplant Tissue and Eye Services, Liverpool, UK
| | - Saurabh Ghosh
- Sunderland Eye Infirmary, South Tyneside and Sunderland NHS Foundation Trust, Sunderland, UK
| | - Lucy Clarke
- UK Department of Ophthalmology, Royal Victoria Infirmary and Newcastle University, Newcastle, UK
| | - Che Connon
- Biosciences Institute, Faculty of Medical Sciences, Newcastle University, UK
| | - Muzlifah Haniffa
- Biosciences Institute, Faculty of Medical Sciences, Newcastle University, UK
| | - Francisco Figueiredo
- Biosciences Institute, Faculty of Medical Sciences, Newcastle University, UK; UK Department of Ophthalmology, Royal Victoria Infirmary and Newcastle University, Newcastle, UK
| | - Lyle Armstrong
- Biosciences Institute, Faculty of Medical Sciences, Newcastle University, UK.
| | - Majlinda Lako
- Biosciences Institute, Faculty of Medical Sciences, Newcastle University, UK.
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Adil MT, Henry JJ. Understanding cornea epithelial stem cells and stem cell deficiency: Lessons learned using vertebrate model systems. Genesis 2021; 59:e23411. [PMID: 33576188 DOI: 10.1002/dvg.23411] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/05/2020] [Revised: 01/08/2021] [Accepted: 01/09/2021] [Indexed: 12/13/2022]
Abstract
Animal models have contributed greatly to our understanding of human diseases. Here, we focus on cornea epithelial stem cell (CESC) deficiency (commonly called limbal stem cell deficiency, LSCD). Corneal development, homeostasis and wound healing are supported by specific stem cells, that include the CESCs. Damage to or loss of these cells results in blindness and other debilitating ocular conditions. Here we describe the contributions from several vertebrate models toward understanding CESCs and LSCD treatments. These include both mammalian models, as well as two aquatic models, Zebrafish and the amphibian, Xenopus. Pioneering developments have been made using stem cell transplants to restore normal vision in patients with LSCD, but questions still remain about the basic biology of CESCs, including their precise cell lineages and behavior in the cornea. We describe various cell lineage tracing studies to follow their patterns of division, and the fates of their progeny during development, homeostasis, and wound healing. In addition, we present some preliminary results using the Xenopus model system. Ultimately, a more thorough understanding of these cornea cells will advance our knowledge of stem cell biology and lead to better cornea disease therapeutics.
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Affiliation(s)
- Mohd Tayyab Adil
- Department of Cell & Developmental Biology, University of Illinois at Urbana-Champaign, Urbana, Illinois, USA
| | - Jonathan J Henry
- Department of Cell & Developmental Biology, University of Illinois at Urbana-Champaign, Urbana, Illinois, USA
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Le Q, Chauhan T, Yung M, Tseng CH, Deng SX. Outcomes of Limbal Stem Cell Transplant: A Meta-analysis. JAMA Ophthalmol 2021; 138:660-670. [PMID: 32324211 DOI: 10.1001/jamaophthalmol.2020.1120] [Citation(s) in RCA: 46] [Impact Index Per Article: 11.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/19/2023]
Abstract
Importance Limbal stem cell transplant (LSCT) can be categorized as direct autologous limbal transplant (AULT), direct allogenic limbal transplant (ALLT), cultivated autologous limbal stem cells transplant (cAULT), and cultivated allogenic limbal stem cells transplant (cALLT). To our knowledge, there is no study directly comparing the outcomes and complications of these procedures. Objective To evaluate the outcomes of different LSCT procedures. Data Source We searched PubMed, EMBASE, Web of Science, and Cochrane without language filter for peer-reviewed articles about LSCT. The latest search was performed on June 30, 2019. Study Selection Clinical studies with the outcome of at least 20 eyes after LSCT were included. Animal studies and studies of other surgical interventions were excluded. Data Extraction and Synthesis Two reviewers independently abstracted the data from each study. Heterogeneity was evaluated with the I2 statistic, and a meta-analysis was performed using the random-effects model. Main Outcomes and Measures Outcome measures included the improvement of ocular surface, visual acuity (VA), and adverse events of recipient eyes and donor eyes. Results Forty studies (2202 eyes) with a mean (SD) follow-up of 31.3 (20.9) months met the inclusion criteria. The mean (SD) age of study participants was 38.4 (13.1) years, and men accounted for 74%. The number of eyes that underwent AULT, ALLT, cAULT, and cALLT were 505, 742, 771, and 184, respectively. Improvement of the ocular surface was achieved in 74.5% of all eyes, 85.7% of eyes after AULT (95% CI, 79.5%-90.3%), 84.7% after cAULT (95% CI, 77.2%-90.0%), 57.8% after ALLT (95% CI, 49.0%-66.1%), and 63.2% after cALLT (95% CI, 49.3%-75.2%). Autologous limbal transplantation resulted in a greater VA improvement rate (76%) than did the other 3 procedures (cAULT: 56.4%; ALLT: 52.3%; cALLT: 43.3%; all P < .001). The most common adverse events in all recipient eyes were recurrent/persistent epithelial erosion (10.5%; 95% CI, 7.2%-23.3%) and elevated intraocular pressure (intraocular pressure, 1.7%; 95% CI, 0.5%-7.8%). Patients who underwent ALLT had the highest rate of recurrent epithelial erosion (27.8%; 95% CI, 17.1%-41.9%) and intraocular pressure elevation (6.3%; 95% CI, 1.8%-19.4%). Conclusions and Relevance These findings suggest LSCT can improve or stabilize the corneal surface with a low rate of severe ocular complications and that autologous LSCT may have a higher success rate and fewer complications than allogenic LSCT.
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Affiliation(s)
- Qihua Le
- David Geffen School of Medicine, Stein Eye Institute, Cornea Division, University of California, Los Angeles.,Eye, Ear, Nose, and Throat Hospital of Fudan University, Shanghai, China
| | - Tulika Chauhan
- David Geffen School of Medicine, Stein Eye Institute, Cornea Division, University of California, Los Angeles.,Mahatma Gandhi Memorial Medical College, DAVV University, Indore, MP, India
| | - Madeline Yung
- David Geffen School of Medicine, Stein Eye Institute, Cornea Division, University of California, Los Angeles
| | - Chi-Hong Tseng
- David Geffen School of Medicine, Division of General Internal Medicine and Health Services Research, University of California, Los Angeles
| | - Sophie X Deng
- David Geffen School of Medicine, Stein Eye Institute, Cornea Division, University of California, Los Angeles
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40
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Galindo S, de la Mata A, López-Paniagua M, Herreras JM, Pérez I, Calonge M, Nieto-Miguel T. Subconjunctival injection of mesenchymal stem cells for corneal failure due to limbal stem cell deficiency: state of the art. Stem Cell Res Ther 2021; 12:60. [PMID: 33441175 PMCID: PMC7805216 DOI: 10.1186/s13287-020-02129-0] [Citation(s) in RCA: 27] [Impact Index Per Article: 6.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/21/2020] [Accepted: 12/28/2020] [Indexed: 12/13/2022] Open
Abstract
Mesenchymal stem cells (MSCs) have unique and beneficial properties and are currently used to treat a broad variety of diseases. These properties include the potential for differentiation into other cell types, secretion of different trophic factors that promote a regenerative microenvironment, anti-inflammatory actions, selective migration to damaged tissues, and non-immunogenicity. MSCs are effective for the treatment of ocular surface diseases such as dry eye, corneal burns, and limbal stem cell deficiency (LSCD), both in experimental models and in humans. LSCD is a pathological condition in which damage occurs to the limbal epithelial stem cells, or their niche, that are responsible for the continuous regeneration of the corneal epithelium. If LSCD is extensive and/or severe, it usually causes corneal epithelial defects, ulceration, and conjunctival overgrowth of the cornea. These changes can result in neovascularization and corneal opacity, severe inflammation, pain, and visual loss. The effectiveness of MSCs to reduce corneal opacity, neovascularization, and inflammation has been widely studied in different experimental models of LSCD and in some clinical trials; however, the methodological disparity used in the different studies makes it hard to compare outcomes among them. In this regard, the MSC route of administration used to treat LSCD and other ocular surface diseases is an important factor. It should be efficient, minimally invasive, and safe. So far, intravenous and intraperitoneal injections, topical administration, and MSC transplantation using carrier substrata like amniotic membrane (AM), fibrin, or synthetic biopolymers have been the most commonly used administration routes in experimental models. However, systemic administration carries the risk of potential side effects and transplantation requires surgical procedures that could complicate the process. Alternatively, subconjunctival injection is a minimally invasive and straightforward technique frequently used in ophthalmology. It enables performance of local treatments using high cell doses. In this review, we provide an overview of the current status of MSC administration by subconjunctival injection, analyzing the convenience, safety, and efficacy for treatment of corneal failure due to LSCD in different experimental models. We also provide a summary of the clinical trials that have been completed, are in progress, or being planned.
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Affiliation(s)
- Sara Galindo
- Instituto de Oftalmobiología Aplicada (IOBA), Universidad de Valladolid, Edificio IOBA, Campus Miguel Delibes, Paseo de Belén 17, 47011, Valladolid, Spain.,Centro de Investigación Biomédica en Red de Bioingeniería, Biomateriales y Nanomedicina (CIBER-BBN), Instituto de Salud Carlos III, Madrid, Spain.,Centro en Red de Medicina Regenerativa y Terapia Celular de Castilla y León, Valladolid, Spain
| | - Ana de la Mata
- Instituto de Oftalmobiología Aplicada (IOBA), Universidad de Valladolid, Edificio IOBA, Campus Miguel Delibes, Paseo de Belén 17, 47011, Valladolid, Spain. .,Centro de Investigación Biomédica en Red de Bioingeniería, Biomateriales y Nanomedicina (CIBER-BBN), Instituto de Salud Carlos III, Madrid, Spain. .,Centro en Red de Medicina Regenerativa y Terapia Celular de Castilla y León, Valladolid, Spain.
| | - Marina López-Paniagua
- Instituto de Oftalmobiología Aplicada (IOBA), Universidad de Valladolid, Edificio IOBA, Campus Miguel Delibes, Paseo de Belén 17, 47011, Valladolid, Spain.,Centro de Investigación Biomédica en Red de Bioingeniería, Biomateriales y Nanomedicina (CIBER-BBN), Instituto de Salud Carlos III, Madrid, Spain.,Centro en Red de Medicina Regenerativa y Terapia Celular de Castilla y León, Valladolid, Spain
| | - Jose M Herreras
- Instituto de Oftalmobiología Aplicada (IOBA), Universidad de Valladolid, Edificio IOBA, Campus Miguel Delibes, Paseo de Belén 17, 47011, Valladolid, Spain.,Centro de Investigación Biomédica en Red de Bioingeniería, Biomateriales y Nanomedicina (CIBER-BBN), Instituto de Salud Carlos III, Madrid, Spain.,Centro en Red de Medicina Regenerativa y Terapia Celular de Castilla y León, Valladolid, Spain
| | - Inmaculada Pérez
- Instituto de Oftalmobiología Aplicada (IOBA), Universidad de Valladolid, Edificio IOBA, Campus Miguel Delibes, Paseo de Belén 17, 47011, Valladolid, Spain.,Centro en Red de Medicina Regenerativa y Terapia Celular de Castilla y León, Valladolid, Spain
| | - Margarita Calonge
- Instituto de Oftalmobiología Aplicada (IOBA), Universidad de Valladolid, Edificio IOBA, Campus Miguel Delibes, Paseo de Belén 17, 47011, Valladolid, Spain.,Centro de Investigación Biomédica en Red de Bioingeniería, Biomateriales y Nanomedicina (CIBER-BBN), Instituto de Salud Carlos III, Madrid, Spain.,Centro en Red de Medicina Regenerativa y Terapia Celular de Castilla y León, Valladolid, Spain
| | - Teresa Nieto-Miguel
- Instituto de Oftalmobiología Aplicada (IOBA), Universidad de Valladolid, Edificio IOBA, Campus Miguel Delibes, Paseo de Belén 17, 47011, Valladolid, Spain. .,Centro de Investigación Biomédica en Red de Bioingeniería, Biomateriales y Nanomedicina (CIBER-BBN), Instituto de Salud Carlos III, Madrid, Spain. .,Centro en Red de Medicina Regenerativa y Terapia Celular de Castilla y León, Valladolid, Spain.
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Dadkhah Tehrani F, Firouzeh A, Shabani I, Shabani A. A Review on Modifications of Amniotic Membrane for Biomedical Applications. Front Bioeng Biotechnol 2021; 8:606982. [PMID: 33520961 PMCID: PMC7839407 DOI: 10.3389/fbioe.2020.606982] [Citation(s) in RCA: 50] [Impact Index Per Article: 12.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/16/2020] [Accepted: 11/25/2020] [Indexed: 12/18/2022] Open
Abstract
The amniotic membrane (AM) is the innermost layer of the fetal placenta, which surrounds and protects the fetus. Its unique structure, in addition to its physical and biological properties, makes it a useful substance in many applications related to regenerative medicine. The use of this fantastic substance with a century-old history has produced remarkable results in vivo, in vitro, and even in clinical studies. While the intact or preserved AM is widely used for these purposes, the addition of further modifications to AM can be considered as a relatively new subject in its applications. These modifications are applied to improve AM properties, ease of handling, and durability. Here, we will discuss the cases in which AM has undergone additional modifications besides the required processes for sterilization and preservation. In this article, we have categorized these modifications and discussed their applications and results.
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Affiliation(s)
- Fatemeh Dadkhah Tehrani
- Cell Engineering and Bio-microsystems Laboratory, Biomedical Engineering Faculty, Amirkabir University of Technology, Tehran, Iran
| | - Arezoo Firouzeh
- Cell Engineering and Bio-microsystems Laboratory, Biomedical Engineering Faculty, Amirkabir University of Technology, Tehran, Iran
| | - Iman Shabani
- Cell Engineering and Bio-microsystems Laboratory, Biomedical Engineering Faculty, Amirkabir University of Technology, Tehran, Iran
| | - Azadeh Shabani
- Preventative Gynecology Research Center, Department of Gynecology and Obstetrics, School of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran
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42
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Figueiredo FC, Glanville JM, Arber M, Carr E, Rydevik G, Hogg J, Okonkwo A, Figueiredo G, Lako M, Whiter F, Wilson K. A systematic review of cellular therapies for the treatment of limbal stem cell deficiency affecting one or both eyes. Ocul Surf 2021; 20:48-61. [PMID: 33412337 DOI: 10.1016/j.jtos.2020.12.008] [Citation(s) in RCA: 21] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/04/2020] [Revised: 12/15/2020] [Accepted: 12/28/2020] [Indexed: 01/18/2023]
Abstract
PURPOSE This systematic review (SR) assessed the efficacy, safety and cost-effectiveness of cell-based therapy to manage limbal stem cell deficiency (LSCD), a sight-threatening orphan condition most frequently associated with severe chemical or thermal burns. LSCD has historically been treated by transplanting limbal tissue. In 1997, a new treatment, cultured limbal epithelial autografts, was described for unilateral LSCD. In cases of bilateral disease cultured autologous oral mucosa stem cells have been used. The relative efficacy of different cultured tissue procedures is unknown. METHODS A protocol was registered with PROSPERO (CRD42017081117). Searches were conducted in 14 databases and 6 conference websites. Two reviewers independently selected studies, conducted data extraction and assessed risk of bias. One reviewer extracted individual patient data (IPD); a second checked extracted data. Data were assessed to determine the feasibility of statistical analysis, with Bayesian synthesis used to estimate improvement achieved by different treatments. RESULTS Fifty-two studies were eligible for inclusion (1113 eyes); 41 studies (716 eyes) reported IPD. No evidence was identified on cost-effectiveness. This SR was unable to confirm that any of the types of ex vivo cultured stem cell transplants identified for LSCD treatment were statistically superior when assessed against the outcomes of interest. CONCLUSIONS We believe this SR is the first to include IPD analysis of LSCD data. There is no evidence for the superiority of any method of limbal stem cell transplant. Confirmation of the safety and efficacy of this treatment modality is challenging due to heterogeneity within and between the studies identified. Therefore, recommendations for future research are proposed.
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Affiliation(s)
- F C Figueiredo
- Department of Ophthalmology, Royal Victoria Infirmary and University of Newcastle, Queen Victoria Road, Newcastle Upon Tyne, NE1 4LP, UK.
| | - J M Glanville
- York Health Economics Consortium, Enterprise House, Innovation Way, University of York, York, YO10 5NQ, UK
| | - M Arber
- York Health Economics Consortium, Enterprise House, Innovation Way, University of York, York, YO10 5NQ, UK
| | - E Carr
- York Health Economics Consortium, Enterprise House, Innovation Way, University of York, York, YO10 5NQ, UK
| | - G Rydevik
- Quantics Biostatistics, West End House, 28 Drumsheugh Gardens, Edinburgh, EH3 7RN, UK
| | - J Hogg
- Department of Ophthalmology, Royal Victoria Infirmary and University of Newcastle, Queen Victoria Road, Newcastle Upon Tyne, NE1 4LP, UK
| | - A Okonkwo
- Department of Ophthalmology, Royal Victoria Infirmary and University of Newcastle, Queen Victoria Road, Newcastle Upon Tyne, NE1 4LP, UK
| | - G Figueiredo
- Department of Ophthalmology, Royal Victoria Infirmary and University of Newcastle, Queen Victoria Road, Newcastle Upon Tyne, NE1 4LP, UK
| | - M Lako
- Biosciences Institute, Newcastle University, International Centre for Life, Newcastle, NE1 3BZ, UK
| | - F Whiter
- York Health Economics Consortium, Enterprise House, Innovation Way, University of York, York, YO10 5NQ, UK
| | - K Wilson
- York Health Economics Consortium, Enterprise House, Innovation Way, University of York, York, YO10 5NQ, UK
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Hristova R, Zdravkov Y, Markov G, Borroni D, Oscar A, Petkova I. Comparison of amniotic membrane transplantation with and without cultured limbal epithelium for persistent corneal ulcers. BIOTECHNOL BIOTEC EQ 2021. [DOI: 10.1080/13102818.2021.1924860] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/21/2022] Open
Affiliation(s)
- Rozaliya Hristova
- Department of Ophthalmology, Medical University of Sofia, University Hospital Alexandrovska, Sofia, Bulgaria
| | - Yani Zdravkov
- Department of Ophthalmology, Medical University of Sofia, University Hospital Alexandrovska, Sofia, Bulgaria
| | - Georgi Markov
- Department of Ophthalmology, Medical University of Sofia, University Hospital Alexandrovska, Sofia, Bulgaria
| | | | - Alexander Oscar
- Department of Ophthalmology, Medical University of Sofia, University Hospital Alexandrovska, Sofia, Bulgaria
| | - Iva Petkova
- Department of Ophthalmology, Medical University of Sofia, University Hospital Alexandrovska, Sofia, Bulgaria
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Pedrotti E, Chierego C, Cozzini T, Merz T, Lagali N, De Gregorio A, Fasolo A, Bonacci E, Bonetto J, Marchini G. In Vivo Confocal Microscopy of the Corneal-Conjunctival Transition in the Evaluation of Epithelial Renewal after SLET. J Clin Med 2020; 9:jcm9113574. [PMID: 33171960 PMCID: PMC7694659 DOI: 10.3390/jcm9113574] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/12/2020] [Revised: 10/26/2020] [Accepted: 11/04/2020] [Indexed: 12/13/2022] Open
Abstract
Examination of the corneal surface by in vivo confocal microscopy (IVCM) allows for objective identification of corneal and conjunctival cell phenotypes to evaluate different epithelialization patterns. Detection of a corneal-conjunctival epithelial transition could be considered as a sign of restored epithelial function following simple limbal epithelial transplantation (SLET). This is a prospective, interventional case series. We assessed patients with limbal stem cell deficiency (LSCD) by IVCM, preoperatively and at monthly intervals following SLET. Sectors in the central and peripheral cornea were scanned. Immediately upon detection of multi-layered cells with the epithelial phenotype in the central cornea and confirmation of epithelial transition in all corneal sectors, the decision for keratoplasty was taken. Ten patients were enrolled. After SLET, epithelial phenotype in the central cornea and an epithelial transition were identified within six and nine months in seven and one patients, respectively. One patient was a partial success and one failed. Five patients underwent keratoplasty, with stable results up to 12 months. Identification of the epithelial transition zone by IVCM permits assessment of the efficacy of SLET, enabling subsequent planning of keratoplasty for visual rehabilitation. The stability of the corneal surface following keratoplasty confirms that the renewal of the corneal epithelium was effectively retained.
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Affiliation(s)
- Emilio Pedrotti
- Ophthalmic Unit, Department of Neurosciences, Biomedicine and Movement Sciences, University of Verona, Policlinico G.B. Rossi, P.le L.A. Scuro 10, 37134 Verona, Italy; (E.P.); (C.C.); (T.M.); (A.F.); (E.B.); (J.B.); (G.M.)
| | - Chiara Chierego
- Ophthalmic Unit, Department of Neurosciences, Biomedicine and Movement Sciences, University of Verona, Policlinico G.B. Rossi, P.le L.A. Scuro 10, 37134 Verona, Italy; (E.P.); (C.C.); (T.M.); (A.F.); (E.B.); (J.B.); (G.M.)
| | - Tiziano Cozzini
- Ophthalmic Unit, Department of Neurosciences, Biomedicine and Movement Sciences, University of Verona, Policlinico G.B. Rossi, P.le L.A. Scuro 10, 37134 Verona, Italy; (E.P.); (C.C.); (T.M.); (A.F.); (E.B.); (J.B.); (G.M.)
- Correspondence: ; Tel.: +39-045-812-6129; Fax: +39-045-812-6175
| | - Tommaso Merz
- Ophthalmic Unit, Department of Neurosciences, Biomedicine and Movement Sciences, University of Verona, Policlinico G.B. Rossi, P.le L.A. Scuro 10, 37134 Verona, Italy; (E.P.); (C.C.); (T.M.); (A.F.); (E.B.); (J.B.); (G.M.)
| | - Neil Lagali
- Division of Ophthalmology, Department of Biomedical and Clinical Sciences, BKV, Linköping University, SE-581 83 Linköping, Sweden;
| | - Alessandra De Gregorio
- Ophthalmic Unit, San Bassiano Hospital, Via dei Lotti, 40, 36061 Bassano del Grappa, Italy;
| | - Adriano Fasolo
- Ophthalmic Unit, Department of Neurosciences, Biomedicine and Movement Sciences, University of Verona, Policlinico G.B. Rossi, P.le L.A. Scuro 10, 37134 Verona, Italy; (E.P.); (C.C.); (T.M.); (A.F.); (E.B.); (J.B.); (G.M.)
- The Veneto Eye Bank Foundation, Padiglione G. Rama, Via Paccagnella 11, 30174 Zelarino Venezia, Italy
| | - Erika Bonacci
- Ophthalmic Unit, Department of Neurosciences, Biomedicine and Movement Sciences, University of Verona, Policlinico G.B. Rossi, P.le L.A. Scuro 10, 37134 Verona, Italy; (E.P.); (C.C.); (T.M.); (A.F.); (E.B.); (J.B.); (G.M.)
| | - Jacopo Bonetto
- Ophthalmic Unit, Department of Neurosciences, Biomedicine and Movement Sciences, University of Verona, Policlinico G.B. Rossi, P.le L.A. Scuro 10, 37134 Verona, Italy; (E.P.); (C.C.); (T.M.); (A.F.); (E.B.); (J.B.); (G.M.)
| | - Giorgio Marchini
- Ophthalmic Unit, Department of Neurosciences, Biomedicine and Movement Sciences, University of Verona, Policlinico G.B. Rossi, P.le L.A. Scuro 10, 37134 Verona, Italy; (E.P.); (C.C.); (T.M.); (A.F.); (E.B.); (J.B.); (G.M.)
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Ghareeb AE, Lako M, Figueiredo FC. Recent Advances in Stem Cell Therapy for Limbal Stem Cell Deficiency: A Narrative Review. Ophthalmol Ther 2020; 9:809-831. [PMID: 32970311 PMCID: PMC7708613 DOI: 10.1007/s40123-020-00305-2] [Citation(s) in RCA: 36] [Impact Index Per Article: 7.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/18/2020] [Accepted: 09/15/2020] [Indexed: 12/13/2022] Open
Abstract
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.
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Affiliation(s)
- Ali E Ghareeb
- Biosciences Institute, Newcastle University, Newcastle-upon-Tyne, UK.,Department of Ophthalmology, Royal Victoria Infirmary, Newcastle Hospitals NHS Foundation Trust, Newcastle-upon-Tyne, UK
| | - Majlinda Lako
- Biosciences Institute, Newcastle University, Newcastle-upon-Tyne, UK
| | - Francisco C Figueiredo
- Biosciences Institute, Newcastle University, Newcastle-upon-Tyne, UK. .,Department of Ophthalmology, Royal Victoria Infirmary, Newcastle Hospitals NHS Foundation Trust, Newcastle-upon-Tyne, UK.
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Smith AB, Retzler J, Taylor MJ. Standard Gamble to Derive Utility Health States for Limbal Stem Cell Deficiency. CLINICOECONOMICS AND OUTCOMES RESEARCH 2020; 12:535-546. [PMID: 32982343 PMCID: PMC7501967 DOI: 10.2147/ceor.s251918] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/29/2020] [Accepted: 07/31/2020] [Indexed: 11/23/2022] Open
Abstract
Purpose To generate UK health-related quality-of-life (HRQoL) data for adult patients with moderate-to-severe limbal stem cell deficiency (LSCD), unilateral or bilateral, due to physical or chemical ocular burns to help inform economic evaluations of treatments. Patients and Methods EQ-5D-3L with vision bolt-on scores was prospectively measured for one of five clinical scenarios of LSCD described in vignettes in a demographically representative population of 520 UK adults. These were converted to health state utilities using three different UK value sets. A standard gamble (SG) was then undertaken using 12 SG scenarios to examine the component drivers of health utility for the treatment of LSCD. Results For the EQ-5D-3L scenarios, the mean disutility for LSCD with poor visual acuity, pain and disfigurement in both eyes compared to one eye was −0.084 (range=−0.156 to −0.045 across the value sets). The mean disutility of bilateral LSCD with pain, disfigurement, and poor visual acuity compared to unilateral LSCD with only poor visual acuity in one eye was −0.104 (range=−0.151 to −0.078). Similarly, where one eye was affected, pain and disfigurement in combination were associated with a greater mean disutility than improvements in visual acuity alone: −0.011 (range=−0.04 to 0.005). Mean SG utilities were within a narrow range (0.682–0.765). Where one eye was affected, the main driver was disfigurement: mean utility was 0.731 (0.709–0.753) compared to 0.682 (0.659–0.704) when disfigurement was removed compared to vision restored to normal. For bilateral LSCD, mean utilities were 0.693 (0.672–0.715) for normal vision and 0.75 (0.73–0.771) when disfigurement and pain were removed. Conclusion Improvements in pain and disfigurement appeared to be the main factors driving differences in health utilities associated with symptom profiles in LSCD, with improvements in visual acuity having lesser impact.
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Affiliation(s)
- Adam B Smith
- York Health Economics Consortium Ltd, University of York, York YO10 5NQ, UK
| | - Jenny Retzler
- York Health Economics Consortium Ltd, University of York, York YO10 5NQ, UK
| | - Matthew J Taylor
- York Health Economics Consortium Ltd, University of York, York YO10 5NQ, UK
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Hernáez-Moya R, González S, Urkaregi A, Pijoan JI, Deng SX, Andollo N. Expansion of Human Limbal Epithelial Stem/Progenitor Cells Using Different Human Sera: A Multivariate Statistical Analysis. Int J Mol Sci 2020; 21:ijms21176132. [PMID: 32854428 PMCID: PMC7503296 DOI: 10.3390/ijms21176132] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/31/2020] [Revised: 08/23/2020] [Accepted: 08/24/2020] [Indexed: 12/13/2022] Open
Abstract
Transplantation of human cultured limbal epithelial stem/progenitor cells (LESCs) has demonstrated to restore the integrity and functionality of the corneal surface in about 76% of patients with limbal stem cell deficiency. However, there are different protocols for the expansion of LESCs, and many of them use xenogeneic products, being a risk for the patients’ health. We compared the culture of limbal explants on the denuded amniotic membrane in the culture medium—supplemental hormone epithelial medium (SHEM)—supplemented with FBS or two differently produced human sera. Cell morphology, cell size, cell growth rate, and the expression level of differentiation and putative stem cell markers were examined. Several bioactive molecules were quantified in the human sera. In a novel approach, we performed a multivariate statistical analysis of data to investigate the culture factors, such as differently expressed molecules of human sera that specifically influence the cell phenotype. Our results showed that limbal cells cultured with human sera grew faster and contained similar amounts of small-sized cells, higher expression of the protein p63α, and lower of cytokeratin K12 than FBS cultures, thus, maintaining the stem/progenitor phenotype of LESCs. Furthermore, the multivariate analysis provided much data to better understand the obtaining of different cell phenotypes as a consequence of the use of different culture methodologies or different culture components.
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Affiliation(s)
- Raquel Hernáez-Moya
- Department of Cell Biology and Histology, School of Medicine and Nursing, Biocruces Bizkaia Health Research Institute, University of the Basque Country UPV/EHU, 48940 Leioa, Bizkaia, Spain;
| | - Sheyla González
- Cornea Division, Stein Eye Institute, University of California, Los Angeles, CA 90095, USA; (S.G.); (S.X.D.)
| | - Arantza Urkaregi
- Department of Applied Mathematics and Statistics and Operational Research, Biocruces Bizkaia Health Research Institute, University of the Basque Country UPV/EHU, 48940 Leioa, Bizkaia, Spain;
| | - Jose Ignacio Pijoan
- Clinical Epidemiology Unit, Cruces University Hospital, Biocruces Bizkaia Health Research Institute, 48903 Barakaldo, Bizkaia, Spain;
| | - Sophie X. Deng
- Cornea Division, Stein Eye Institute, University of California, Los Angeles, CA 90095, USA; (S.G.); (S.X.D.)
| | - Noelia Andollo
- Department of Cell Biology and Histology, School of Medicine and Nursing, Biocruces Bizkaia Health Research Institute, University of the Basque Country UPV/EHU, 48940 Leioa, Bizkaia, Spain;
- Correspondence: ; Tel.: +34-94-601-3295
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Ozer MD, Altınkurt E, Yilmaz YC, Gedik AC, Alparslan N. The Surgical Outcomes of Limbal Allograft Transplantation in Eyes Having Limbal Stem Cell Deficiency. J Curr Ophthalmol 2020; 32:132-141. [PMID: 32775793 PMCID: PMC7337021 DOI: 10.4103/joco.joco_91_20] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/27/2019] [Revised: 12/08/2019] [Accepted: 12/23/2019] [Indexed: 12/15/2022] Open
Abstract
Purpose: To report the limbal allograft transplantation and penetrating keratoplasty (PK) results in limbal stem cell deficiency (LSCD)-developed eyes because of chemical or thermal injury. Methods: Medical records of 18 eyes of 14 patients who had undergone keratolimbal allograft (KLAL) or living-related conjunctival limbal allograft (lr-CLAL) with or without PK and followed up at least 1 year postoperatively were evaluated retrospectively. The preoperative LSCD grade was noted in all patients. Rejection incidents, recurrence of LSCD, and corneal graft clarity along with a visual improvement during the follow-up were noted. The complications rate due to surgery or injury itself, for instance, glaucoma and cataract, were evaluated. The limbal allograft tissue survival analysis and corneal allograft survival analysis were done to reveal the differences in both the procedures. The existence of normal corneal epithelium and improvement in visual acuity were accepted as the surgical success criteria. Results: In the limbal allograft transplantation group, the survival rates of the allograft tissue were 65 ± 10.7% at 1 year and 36.6 ± 11.4% at 3 years in lr-CLAL and 66.7 ± 15.7% at 12 months and 53.3 ± 17.3% at 18 months in KLAL-transplanted eyes. The survival rate of corneal allograft at the 5th postoperative year was lower in the simultaneous procedure compared to the staged procedure, but it was not statistically significant (25.7 ± 25.8% vs. 62.5 ± 17.1%, P = 0.75). The ambulatory vision was achieved in 10 eyes (56%) after a mean follow-up time of 93.8 ± 37.8 months. The visual acuity level has increased in 12 eyes (67%) in which the limbal allograft transplantation was applied. The ambulatory visual acuity level was achieved (≤1.0 logMar [20/200]) in 10 eyes (56%). In addition, two or more Snellen lines’ gain in the best corrected visual acuity was observed in 12 eyes of 18 (67%) at the last follow-up, and there was not any significant difference between the KLAL and lr-CLAL. Conclusions: Ocular surface integrity was longer in KLAL than in lr-CLAL transplantation, but it was not statistically significant. The staged procedure was more convenient than the simultaneous procedure in terms of corneal allograft clarity maintenance in limbal allograft-employed eyes.
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Affiliation(s)
- Muhammet Derda Ozer
- Department of Ophthalmology, Medical Faculty, Van Yuzuncu Yil University, Van, Turkey
| | - Emre Altınkurt
- Department of Ophthalmology, Istanbul Faculty of Medicine, Istanbul University, İstanbul, Turkey
| | | | - Ali Ceyhun Gedik
- Department of Ophthalmology, Luleburgaz State Hospital, Kirklareli, Turkey
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Park GW, Heo J, Kang JY, Yang JW, Kim JS, Kwon KD, Yu BC, Lee SJ. Topical cell-free conditioned media harvested from adipose tissue-derived stem cells promote recovery from corneal epithelial defects caused by chemical burns. Sci Rep 2020; 10:12448. [PMID: 32709896 PMCID: PMC7381646 DOI: 10.1038/s41598-020-69020-z] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/07/2019] [Accepted: 06/29/2020] [Indexed: 02/07/2023] Open
Abstract
Corneal chemical burns can lead to blindness following serious complications. As most of these complications are caused by failure of reepithelization during the acute phase, treatment at this stage is critical. Although there have been some studies on corneal injury recovery using adipose tissue-derived stem cells (ADSCs), none has reported the effect of topical cell-free conditioned culture media (CM) derived from ADSCs on corneal epithelial regeneration. Here, the best conditions for CM were selected and used for in vitro and in vivo experiments. Corneal burn in rats was induced using 100% alcohol. The chosen CM was administered to corneal burn rats (CM-treated [CT] group) four times a day for three days and this group was compared with the normal control and corneal burn (CB) groups. Biomicroscopic fluorescence images and the actual physical corneas were taken over time and used for analysis. mRNA levels of hepatocyte growth factor and epidermal growth factor (EGF) were significantly increased, whereas those of vascular endothelial growth factor, interleukin (IL)-1β, IL-6, IL-10, and matrix metalloproteinase-9 were significantly decreased in the CT group compared with those in the CB group. The numbers of proliferating cell nuclear antigen- and zonular occludens-1-positive cells in the CT group were significantly higher than those in the CB group. The macrophage-infiltrating corneas in the CT group expressed significantly more of the M2 marker arginase than corneas in the CB group. Optimal CM (× 0.5 concentration) treatment significantly accelerated the migration of corneal epithelial cells and induced upregulation of the expression of IL-6, EGF, and C-X-C chemokine receptor type 4 mRNAs. Overall, in this study, topical administration of cell-free CM promoted regeneration of the corneal epithelium after induction of chemical burns.
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Affiliation(s)
| | - Jeonghoon Heo
- Department of Molecular Biology and Immunology, College of Medicine, Kosin University, Busan, Korea
| | | | - Ji Won Yang
- Stem Bank Company, Busan, Korea
- Department of Ophthalmology, College of Medicine, Kosin University, #34 Amnam-dong, Suh-ku, Busan, 602-702, Korea
| | - Jong Sik Kim
- Department of Anatomy, College of Medicine, Kosin University, Busan, Korea
| | - Ki Dong Kwon
- Department of Ophthalmology, College of Medicine, Kosin University, #34 Amnam-dong, Suh-ku, Busan, 602-702, Korea
| | - Byung Chul Yu
- Department of Preventive Medicine, College of Medicine, Kosin University, Busan, Korea
| | - Sang Joon Lee
- Department of Ophthalmology, College of Medicine, Kosin University, #34 Amnam-dong, Suh-ku, Busan, 602-702, Korea.
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50
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He J, Ou S, Ren J, Sun H, He X, Zhao Z, Wu H, Qu Y, Liu T, Jeyalatha V, Zhang L, Li Q, Reinach PS, Quantock A, Hao J, Liu Z, Li W. Tissue engineered corneal epithelium derived from clinical-grade human embryonic stem cells. Ocul Surf 2020; 18:672-680. [PMID: 32710961 DOI: 10.1016/j.jtos.2020.07.009] [Citation(s) in RCA: 19] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/14/2020] [Revised: 07/10/2020] [Accepted: 07/12/2020] [Indexed: 02/07/2023]
Abstract
PURPOSE To construct tissue engineered corneal epithelium from a clinical-grade human embryonic stem cells (hESCs) and investigate the dynamic gene profile and phenotypic transition in the process of differentiation. METHODS A stepwise protocol was applied to induce differentiation of clinical-grade hESCs Q-CTS-hESC-1 and construct tissue engineered corneal epithelium. Single cell RNA sequencing (scRNA-seq) analysis was performed to monitor gene expression and phenotypic changes at different differentiation stages. Immunostaining, real-time quantitative PCR and Western blot analysis were conducted to detect gene and protein expressions. After subcutaneous transplantation into nude mice to test the biosafety, the epithelial construct was transplanted in a rabbit corneal limbal stem cell deficiency (LSCD) model and followed up for eight weeks. RESULTS The hESCs were successfully induced into epithelial cells. scRNA-seq analysis revealed upregulation of ocular surface epithelial cell lineage related genes such as TP63, Pax6, KRT14, and activation of Wnt, Notch, Hippo, and Hedgehog signaling pathways during the differentiation process. Tissue engineered epithelial cell sheet derived from hESCs showed stratified structure and normal corneal epithelial phenotype with presence of clonogenic progenitor cells. Eight weeks after grafting the cell sheet onto the ocular surface of LSCD rabbit model, a full-thickness continuous corneal epithelium developed to fully cover the damaged areas with normal limbal and corneal epithelial phenotype. CONCLUSION The tissue engineered corneal epithelium generated from a clinical-grade hESCs may be feasible in the treatment of limbal stem cell deficiency.
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Affiliation(s)
- Jia He
- Eye Institute of Xiamen University, Fujian Provincial Key Laboratory of Ophthalmology and Visual Science, School of Medicine, Xiamen University, China
| | - Shangkun Ou
- Eye Institute of Xiamen University, Fujian Provincial Key Laboratory of Ophthalmology and Visual Science, School of Medicine, Xiamen University, China
| | - Jun Ren
- School of Informatics, Xiamen University, Xiamen, Fujian, China; National Institute for Data Science in Health and Medicine, Xiamen University, Xiamen, Fujian, China
| | - Huimin Sun
- Eye Institute of Xiamen University, Fujian Provincial Key Laboratory of Ophthalmology and Visual Science, School of Medicine, Xiamen University, China
| | - Xin He
- Eye Institute of Xiamen University, Fujian Provincial Key Laboratory of Ophthalmology and Visual Science, School of Medicine, Xiamen University, China
| | - Zhongyang Zhao
- Eye Institute of Xiamen University, Fujian Provincial Key Laboratory of Ophthalmology and Visual Science, School of Medicine, Xiamen University, China
| | - Han Wu
- Eye Institute of Xiamen University, Fujian Provincial Key Laboratory of Ophthalmology and Visual Science, School of Medicine, Xiamen University, China
| | - Yangluowa Qu
- Eye Institute of Xiamen University, Fujian Provincial Key Laboratory of Ophthalmology and Visual Science, School of Medicine, Xiamen University, China
| | - Tingting Liu
- Eye Institute of Xiamen University, Fujian Provincial Key Laboratory of Ophthalmology and Visual Science, School of Medicine, Xiamen University, China
| | - Vimalin Jeyalatha
- Eye Institute of Xiamen University, Fujian Provincial Key Laboratory of Ophthalmology and Visual Science, School of Medicine, Xiamen University, China
| | - Liying Zhang
- Eye Institute of Xiamen University, Fujian Provincial Key Laboratory of Ophthalmology and Visual Science, School of Medicine, Xiamen University, China
| | - Qiyuan Li
- Eye Institute of Xiamen University, Fujian Provincial Key Laboratory of Ophthalmology and Visual Science, School of Medicine, Xiamen University, China
| | - Peter Sol Reinach
- Wenzhou Medical University, Department of Ophthalmology, Wenzhou, Zhejiang, China; Wenzhou Medical University, Department of Optometry, Wenzhou, Zhejiang, China
| | - Andrew Quantock
- Structural Biophysics Group, School of Optometry and Vision Sciences, Cardiff University, Cardiff, Wales, United Kingdom
| | - Jie Hao
- State Key Laboratory of Stem Cell and Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing, China; Institute for Stem Cell and Regeneration, Chinese Academy of Sciences, Beijing, China; National Stem Cell Resource Center, Chinese Academy of Sciences, Beijing, China.
| | - Zuguo Liu
- Eye Institute of Xiamen University, Fujian Provincial Key Laboratory of Ophthalmology and Visual Science, School of Medicine, Xiamen University, China.; Department of Ophthalmology, Xiang'an Hospital of Xiamen University, Xiamen, Fujian, China; Xiamen University Affiliated Xiamen Eye Center, Xiamen, Fujian, China.
| | - Wei Li
- Eye Institute of Xiamen University, Fujian Provincial Key Laboratory of Ophthalmology and Visual Science, School of Medicine, Xiamen University, China.; Department of Ophthalmology, Xiang'an Hospital of Xiamen University, Xiamen, Fujian, China; Xiamen University Affiliated Xiamen Eye Center, Xiamen, Fujian, China.
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