Cell Surface Markers of Functional Phenotypic Corneal Endothelial Cells

Okumura, Naoki; Hirano, Hiroatsu; Numata, Ryohei; Nakahara, Makiko; Ueno, Masaki; Hamuro, Junji; Kinoshita, Shigeru; Koizumi, Noriko

doi:10.1167/iovs.14-14980

Cited by 66 publications

(73 citation statements)

References 41 publications

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“…with that of Okumura et al 11 who found that CD166 was expressed in both normal cells and in fibroblastic cells. CD markers effective for distinguishing distinct SPs were selected by analyzing those on established cHCECs with a small cell area and a high cell density.…”

Section: Subpopulation Proven As Effector Cells In the Context Of CD supporting

confidence: 73%

“…10 To date, no HCEC-specific cell surface markers have been elucidated, although two groups have reported the expression of CD markers. 11,12 The effector cell did not show any sign of surface CD200. Okumura et al 11 have reported that CD340, CD166, and CD98 are elevated in HCECs of nonfibroblastic phenotype, while CD9, CD49e, CD44, and CD73 are the indices of fibroblastic phenotypes.…”

Section: Discussionmentioning

confidence: 91%

“…11 Briefly, the Descemet membranes with the CECs were stripped from donor corneas and digested at 378C with 1 mg/mL collagenase A (Roche Applied Science, Penzberg, Germany) for 2 hours. The HCECs obtained from a single donor cornea were seeded in one well of a Type-I collagen-coated six-well plate (Corning, Inc., Corning, NY, USA).…”

Section: Cell Cultures Of Hcecsmentioning

confidence: 99%

“…Human corneal endothelial cells obtained from the corneas of five aged donors, one young donor (the age range of the aged and young donors was above 50 years and between 7 and 29 years, respectively), and two newborn donors were cultured according to the method described by Okumura et al, 11 and the surface expression of CD166, CD105, CD44, and CD24 was then characterized ( Table 1). The other set of data for cHCECs from other donor corneas for CD166, CD105, CD44, and LGR5, instead of CD24, is summarized in Table 1.…”

Section: Phenotypic Variations Among Culturesmentioning

confidence: 99%

“…In fact, cHCECs are heterogeneous from culture to culture in their morphology and in their surface markers, such as cluster of differentiation (CD) antigens. [10][11][12] Studies, [13][14][15][16][17][18][19][20] either directly or indirectly, and including those from Joyce et al, have shown that heterogeneity is present in HCEC cultures. Of particular and striking interest is the finding by Miyai et al 10 of the presence of frequent chromosomal aneuploidy in cHCECs, thus indicating the presence of heterogeneous subpopulations (SPs) with or without aneuploidy.…”

mentioning

confidence: 99%

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Cell Homogeneity Indispensable for Regenerative Medicine by Cultured Human Corneal Endothelial Cells

Hamuro

Toda

Asada

et al. 2016

Invest. Ophthalmol. Vis. Sci.

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Citation: Hamuro J, Toda M, Asada K, et al. Cell homogeneity indispensable for regenerative medicine by cultured human corneal endothelial cells. Invest Ophthalmol Vis Sci. 2016;57:4749-4761. DOI:10.1167/ iovs.16-19770 PURPOSE. To identify the subpopulation (SP) among heterogeneous cultured human corneal endothelial cells (cHCECs) devoid of cell-state transition applicable for cell-based therapy.METHODS. Subpopulation presence in cHCECs was confirmed via surface CD-marker expression level by flow cytometry. CD markers effective for distinguishing distinct SPs were selected by analyzing those on established cHCECs with a small cell area and high cell density. Contrasting features among three typical cHCEC SPs was confirmed by PCR array for extracellular matrix (ECM). Combined analysis of CD markers was performed to identify the SP (effector cells) applicable for therapy. ZO-1 and Na þ /K þ ATPase, CD200, and HLA expression were compared among heterogeneous SPs. RESULTS. Flow cytometry analysis identified the effector cell expressing CD166 and the presence of other SPs with CD166þþþ (CD26 and CD24, either þ or À) was confirmed. PCR array revealed three distinct ECM expression profiles. Some SPs expressed ZO-1 and Naþ ATPase at comparable levels with effector cells, while only one SP expressed CD200, but not on effector cells. Human leukocyte antigen expression was most reduced in the effector SP. The proportion of effector cells (E-ratio) inversely paralleled donor age and decreased during prolonged culture passages. The presence of Rho-associated protein kinase (ROCK) inhibitor increased the E-ratio in cHCECs. The average area of effector cells was approximately 200~220 lm 2 , and the density of cHCECs exceeded 2500 cells/mm 2 . CONCLUSIONS.A specified cultured effector cell population sharing the surface phenotypes with mature HCECs in corneal tissues may serve as an alternative to donor corneas for the treatment of corneal endothelial dysfunction.

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Section: Subpopulation Proven As Effector Cells In the Context Of CD supporting

confidence: 73%

Section: Discussionmentioning

confidence: 91%

Section: Cell Cultures Of Hcecsmentioning

confidence: 99%

Section: Phenotypic Variations Among Culturesmentioning

confidence: 99%

mentioning

confidence: 99%

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Cell Homogeneity Indispensable for Regenerative Medicine by Cultured Human Corneal Endothelial Cells

Hamuro

Toda

Asada

et al. 2016

Invest. Ophthalmol. Vis. Sci.

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Ex Vivo Functionality of 3D Bioprinted Corneal Endothelium Engineered with Ribonuclease 5‐Overexpressing Human Corneal Endothelial Cells

Kim

Lee

Park

et al. 2018

Adv Healthcare Materials

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Human corneal endothelial cells (HCECs) are scarcely proliferative in vivo. The cultured HCECs engineered to overexpress ribonuclease (RNase) 5 (R5-HCECs) are prepared after transient transfection with RNase 5 plasmid vector. As candidate targets of R5-HCECs for enhancement of cellular proliferation and survival of R5-HCECs, programmed cell death protein 4 is inhibited, and cyclin D1 and cyclin E1 are activated. The cultured R5-HCECs and control HCECs on lyophilized amniotic membrane (AM) are deposited as a carrier by extrusion-based 3D bioprinting to prepare transplantable RNase 5 vector-transfected HCECs-laden AM graft (R5-Graft) and the control HCECs-laden AM graft (Ct-Graft), respectively. The ready-to-use R5-Graft shows clearer basolateral expression of Na -K ATPase pump and higher cell confluency than Ct-Graft. From 2 weeks after graft transplantation, both R5-Graft and Ct-Graft start restoring clarity of the rabbit corneas, and their central corneal edema are much less than those in the control group at 3 and 4 weeks. The ex vivo expression of corneal endothelial phenotypical markers is clear in R5-Grafs rather than in Ct-Grafts at 4 weeks. In conclusion, the fabricated corneal endothelium with cultured HCECs easily survives and functions as corneal endothelium in vivo. Furthermore, the use of the cultured HCECs engineered to overexpress RNase 5 (R5-HCECs) may be an option to obtain higher graft cellularity and to enhance the function of transplanted grafts.

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Translational issues for human corneal endothelial tissue engineering

Soh

Peh

2016

J Tissue Eng Regen Med

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Corneal endothelial disorders collectively represent a significant healthcare burden in most developed nations, and corneal transplantation is currently the only treatment available for patients with poor visual acuity and corneal blindness secondary to endothelial failure. Although vision in these patients can be restored by transplantation, the global demand for donor human corneas is far in excess of what can be provided for by eye banks around the world, and this deficit is set to increase with an ageing global population. As such, there has been a pressing need to explore novel and more sustainable options for the treatment of corneal endothelial diseases. In recent years, significant progress has been made not only in the development of corneal endothelial cell culture techniques, but also in the exploration of various translational strategies. Considered together, we are now much closer to attaining success in the treatment of corneal endothelial diseases via a cell-based, tissue-engineering approach. The aim of this review article is to provide an update of the translational issues currently facing human corneal endothelial cell therapy. Copyright © 2016 John Wiley & Sons, Ltd.

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Cell Surface Markers of Functional Phenotypic Corneal Endothelial Cells

Abstract: These markers will be useful for quality control to characterize the phenotype of cells destined for tissue engineering-based therapy. In addition, this selection protocol will provide a novel method for purification of functional cells.

Cited by 66 publications

References 41 publications

Cell Homogeneity Indispensable for Regenerative Medicine by Cultured Human Corneal Endothelial Cells

Cell Homogeneity Indispensable for Regenerative Medicine by Cultured Human Corneal Endothelial Cells

Ex Vivo Functionality of 3D Bioprinted Corneal Endothelium Engineered with Ribonuclease 5‐Overexpressing Human Corneal Endothelial Cells

Translational issues for human corneal endothelial tissue engineering

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