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Zubaidah Zakaria - One of the best experts on this subject based on the ideXlab platform.
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Ex vivo expanded SSEA-4+ human limbal stromal cells are multipotent and do not express other embryonic stem cell markers
2016Co-Authors: Rahman Jamal, Zubaidah ZakariaAbstract:Purpose: The presence of multipotent human limbal stromal cells resembling mesenchymal stromal cells (MSC) provides new insights to the characteristic of these cells and its therapeutic potential. However, little is known about the expression of stage-specific embryonic antigen 4 (SSEA-4) and the embryonic stem cell (ESC)-like properties of these cells. We studied the expression of SSEA-4 surface protein and the various ESC and MSC markers in the ex vivo cultured limbal stromal cells. The phenotypes and multipotent differentiation potential of these cells were also evaluated. Methods: Limbal stromal cells were derived from corneoscleral rims. The SSEA-4+ and SSEA-4- limbal stromal cells were sorted by fluorescence-activated cells sorting (FACS). Isolated cells were expanded and reanalyzed for their expression of SSEA-4. Expression of MSC and ESC markers on these cells were also analyzed by FACS. In addition, expression of limbal epithelial and corneal stromal proteins such as ATP-binding cassette sub-family G member 2 (ABCG2), tumour protein p63 (p63), paired box 6 (Pax6), Cytokeratin 3 (AE5), Cytokeratin 10, and keratocan sulfate were evaluated either by immunofluorecence staining or reverse transcription polymerase chain reaction. Appropriate induction medium was used to differentiate these cells into adipocytes, osteocytes, and chondrocytes. Results: Expanded limbal stromal cells expressed the majority of mesenchymal markers. These cells were negative for ABCG2, p63, Pax6, AE-5, and keratocan sulfate. After passaged, a subpopulation of these cells showed low expression of SSEA-4 but were negative for other important ESC surface markers such as Tra-1–60, Tra-1–81, and transcriptio
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ex vivo expanded ssea 4 human limbal stromal cells are multipotent and do not express other embryonic stem cell markers
Molecular Vision, 2012Co-Authors: Moon Nian Lim, Noor Hamidah Hussin, Ainoon Othman, Thiageswari Umapathy, Puteri Baharuddin, Rahman A A Jamal, Zubaidah ZakariaAbstract:Purpose: The presence of multipotent human limbal stromal cells resembling mesenchymal stromal cells (MSC) provides new insights to the characteristic of these cells and its therapeutic potential. However, little is known about the expression of stage-specific embryonic antigen 4 (SSEA-4) and the embryonic stem cell (ESC)-like properties of these cells. We studied the expression of SSEA-4 surface protein and the various ESC and MSC markers in the ex vivo cultured limbal stromal cells. The phenotypes and multipotent differentiation potential of these cells were also evaluated. Methods: Limbal stromal cells were derived from corneoscleral rims. The SSEA-4+ and SSEA-4- limbal stromal cells were sorted by fluorescence-activated cells sorting (FACS). Isolated cells were expanded and reanalyzed for their expression of SSEA-4. Expression of MSC and ESC markers on these cells were also analyzed by FACS. In addition, expression of limbal epithelial and corneal stromal proteins such as ATP-binding cassette sub-family G member 2 (ABCG2), tumour protein p63 (p63), paired box 6 (Pax6), Cytokeratin 3 (AE5), Cytokeratin 10, and keratocan sulfate were evaluated either by immunofluorecence staining or reverse transcription polymerase chain reaction. Appropriate induction medium was used to differentiate these cells into adipocytes, osteocytes, and chondrocytes. Results: Expanded limbal stromal cells expressed the majority of mesenchymal markers. These cells were negative for ABCG2, p63, Pax6, AE-5, and keratocan sulfate. After passaged, a subpopulation of these cells showed low expression of SSEA-4 but were negative for other important ESC surface markers such as Tra-1–60, Tra-1–81, and transcription factors like octamer-binding transcription factor 4 (Oct4), SRY(sex determining region Y)-box 2 (Sox2), and Nanog. Early passaged cells when induced were able to differentiate into adipocytes, osteocytes and chondrocytes. Conclusions: The expanded limbal stromal cells showed features of multipotent MSC. Our study confirmed the expression of SSEA-4 by a subpopulation of cultured limbal stromal cells. However, despite the expression of SSEA-4, these cells did not express any other markers of ESC. Therefore, we conclude that the cells did not show properties of ESC. The cornea is the major refractive element of the adult eye. It consists primarily of three layers: an outer layer of epithelium, a middle stromal layer of collagen-rich extracellular matrix (ECM) interspersed with keratocytes and an inner layer of endothelial cells. The stroma comprises 90% of the thickness of the cornea. It consists of dense, regularly packed collagen fibrils arrange as orthogonal layers or lamellae. The corneal stroma is unique in having a homogeneous distribution of fibrils with small diameter (25– 30 nm) that are regularly packed within lamellae and this arrangement minimizes light scattering permitting transparency. When an incisional wound penetrating into stroma occurs, the keratocytes become hypercellular myofibroblasts. These cells can later become wound
Inkeun Jang - One of the best experts on this subject based on the ideXlab platform.
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transplantation of reconstructed corneal layer composed of corneal epithelium and fibroblasts on a lyophilized amniotic membrane to severely alkali burned cornea
Artificial Organs, 2006Co-Authors: Inkeun Jang, Jun Seop Shin, Youngsam Kwon, Jungkeug Park, Kyeyong Song, Eunkyung YangAbstract:Abstract: The purpose of this article was to evaluate the graft efficacy of reconstructed corneal layer, composed of autologous corneal epithelium and fibroblasts on a lyophilized amniotic membrane (LAM), in a severely alkali-burned corneal model. After biopsy specimens were obtained from the left eyes of 24 rabbits, the corneal epithelial cells and fibroblasts were expanded in vitro and the corneal layer was reconstructed on LAM. Thirty-six eyes of rabbits underwent alkali burn (1 N NaOH, 30 s) to create a limbal deficiency and a deeply damaged corneal stroma. Four weeks later, group 1 underwent a graft of the reconstructed corneal layer composed of autologous corneal epithelium and fibroblasts on LAM. Group 2 was transplanted with a graft of the reconstructed autologous corneal epithelium, and group 3 served as a control without surgery. Wound healing and stabilization of the ocular surfaces occurred much faster in group 1 than in groups 2 and 3. The eyes in group 3 revealed typical limbal deficiencies with conjuctivalization and persistent corneal epithelial defects. However, the corneas in group 1 developed only mild peripheral neovascularization. Immunohistochemical staining in group 1 demonstrated that p63, Cytokeratin 3, E-cadherin, transforming growth factor (TGF)-β1, and collagen IV were expressed strongly in the corneal epithelium and basement membrane. On the basis of these results, transplantation of the reconstructed corneal layer, composed of autologous corneal epithelium and fibroblasts on LAM, partially accelerated the recovery of the alkali-injured rabbit ocular surface, and might be useful therapeutically for the treatment of patients with severely damaged cornea.
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transplantation of reconstructed corneal layer composed of corneal epithelium and fibroblasts on a lyophilized amniotic membrane to severely alkali burned cornea
Artificial Organs, 2006Co-Authors: Inkeun Jang, Jun Seop Shin, Youngsam Kwon, Jungkeug Park, Kyeyong Song, Eunkyung Yang, Jaeil Ahn, Yanghwan Ryu, Jeong Kyu Lee, Jae Chan KimAbstract:The purpose of this article was to evaluate the graft efficacy of reconstructed corneal layer, composed of autologous corneal epithelium and fibroblasts on a lyophilized amniotic membrane (LAM), in a severely alkali-burned corneal model. After biopsy specimens were obtained from the left eyes of 24 rabbits, the corneal epithelial cells and fibroblasts were expanded in vitro and the corneal layer was reconstructed on LAM. Thirty-six eyes of rabbits underwent alkali burn (1 N NaOH, 30 s) to create a limbal deficiency and a deeply damaged corneal stroma. Four weeks later, group 1 underwent a graft of the reconstructed corneal layer composed of autologous corneal epithelium and fibroblasts on LAM. Group 2 was transplanted with a graft of the reconstructed autologous corneal epithelium, and group 3 served as a control without surgery. Wound healing and stabilization of the ocular surfaces occurred much faster in group 1 than in groups 2 and 3. The eyes in group 3 revealed typical limbal deficiencies with conjuctivalization and persistent corneal epithelial defects. However, the corneas in group 1 developed only mild peripheral neovascularization. Immunohistochemical staining in group 1 demonstrated that p63, Cytokeratin 3, E-cadherin, transforming growth factor (TGF)-beta1, and collagen IV were expressed strongly in the corneal epithelium and basement membrane. On the basis of these results, transplantation of the reconstructed corneal layer, composed of autologous corneal epithelium and fibroblasts on LAM, partially accelerated the recovery of the alkali-injured rabbit ocular surface, and might be useful therapeutically for the treatment of patients with severely damaged cornea.
Eunkyung Yang - One of the best experts on this subject based on the ideXlab platform.
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transplantation of reconstructed corneal layer composed of corneal epithelium and fibroblasts on a lyophilized amniotic membrane to severely alkali burned cornea
Artificial Organs, 2006Co-Authors: Inkeun Jang, Jun Seop Shin, Youngsam Kwon, Jungkeug Park, Kyeyong Song, Eunkyung YangAbstract:Abstract: The purpose of this article was to evaluate the graft efficacy of reconstructed corneal layer, composed of autologous corneal epithelium and fibroblasts on a lyophilized amniotic membrane (LAM), in a severely alkali-burned corneal model. After biopsy specimens were obtained from the left eyes of 24 rabbits, the corneal epithelial cells and fibroblasts were expanded in vitro and the corneal layer was reconstructed on LAM. Thirty-six eyes of rabbits underwent alkali burn (1 N NaOH, 30 s) to create a limbal deficiency and a deeply damaged corneal stroma. Four weeks later, group 1 underwent a graft of the reconstructed corneal layer composed of autologous corneal epithelium and fibroblasts on LAM. Group 2 was transplanted with a graft of the reconstructed autologous corneal epithelium, and group 3 served as a control without surgery. Wound healing and stabilization of the ocular surfaces occurred much faster in group 1 than in groups 2 and 3. The eyes in group 3 revealed typical limbal deficiencies with conjuctivalization and persistent corneal epithelial defects. However, the corneas in group 1 developed only mild peripheral neovascularization. Immunohistochemical staining in group 1 demonstrated that p63, Cytokeratin 3, E-cadherin, transforming growth factor (TGF)-β1, and collagen IV were expressed strongly in the corneal epithelium and basement membrane. On the basis of these results, transplantation of the reconstructed corneal layer, composed of autologous corneal epithelium and fibroblasts on LAM, partially accelerated the recovery of the alkali-injured rabbit ocular surface, and might be useful therapeutically for the treatment of patients with severely damaged cornea.
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transplantation of reconstructed corneal layer composed of corneal epithelium and fibroblasts on a lyophilized amniotic membrane to severely alkali burned cornea
Artificial Organs, 2006Co-Authors: Inkeun Jang, Jun Seop Shin, Youngsam Kwon, Jungkeug Park, Kyeyong Song, Eunkyung Yang, Jaeil Ahn, Yanghwan Ryu, Jeong Kyu Lee, Jae Chan KimAbstract:The purpose of this article was to evaluate the graft efficacy of reconstructed corneal layer, composed of autologous corneal epithelium and fibroblasts on a lyophilized amniotic membrane (LAM), in a severely alkali-burned corneal model. After biopsy specimens were obtained from the left eyes of 24 rabbits, the corneal epithelial cells and fibroblasts were expanded in vitro and the corneal layer was reconstructed on LAM. Thirty-six eyes of rabbits underwent alkali burn (1 N NaOH, 30 s) to create a limbal deficiency and a deeply damaged corneal stroma. Four weeks later, group 1 underwent a graft of the reconstructed corneal layer composed of autologous corneal epithelium and fibroblasts on LAM. Group 2 was transplanted with a graft of the reconstructed autologous corneal epithelium, and group 3 served as a control without surgery. Wound healing and stabilization of the ocular surfaces occurred much faster in group 1 than in groups 2 and 3. The eyes in group 3 revealed typical limbal deficiencies with conjuctivalization and persistent corneal epithelial defects. However, the corneas in group 1 developed only mild peripheral neovascularization. Immunohistochemical staining in group 1 demonstrated that p63, Cytokeratin 3, E-cadherin, transforming growth factor (TGF)-beta1, and collagen IV were expressed strongly in the corneal epithelium and basement membrane. On the basis of these results, transplantation of the reconstructed corneal layer, composed of autologous corneal epithelium and fibroblasts on LAM, partially accelerated the recovery of the alkali-injured rabbit ocular surface, and might be useful therapeutically for the treatment of patients with severely damaged cornea.
Gervaise Mosser - One of the best experts on this subject based on the ideXlab platform.
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development of human corneal epithelium on organized fibrillated transparent collagen matrices synthesized at high concentration
Acta Biomaterialia, 2015Co-Authors: Aurelien Tidu, Djida Ghoubaybenallaoua, Barbara Lynch, Bernard Haye, Corinne Illoul, Jeanmarc Allain, V Borderie, Gervaise MosserAbstract:Several diseases can lead to opacification of cornea requiring transplantation of donor tissue to restore vision. In this context, transparent collagen I fibrillated matrices have been synthesized at 15, 30, 60 and 90 mg/mL. The matrices were evaluated for fibril organizations, transparency, mechanical properties and ability to support corneal epithelial cell culture. The best results were obtained with 90 mg/mL scaffolds. At this concentration, the fibril organization presented some similarities to that found in corneal stroma. Matrices had a mean Young's modulus of 570 kPa and acellular scaffolds had a transparency of 87% in the 380-780 nm wavelength range. Human corneal epithelial cells successfully colonized the surface of the scaffolds and generated an epithelium with characteristics of corneal epithelial cells (i.e. expression of Cytokeratin 3 and presence of desmosomes) and maintenance of stemness during culture (i.e. expression of Delta Np63 alpha and formation of holoclones in colony formation assay). Presence of cultured epithelium on the matrices was associated with increased transparency (89%). (C) 2015 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.
Moon Nian Lim - One of the best experts on this subject based on the ideXlab platform.
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ex vivo expanded ssea 4 human limbal stromal cells are multipotent and do not express other embryonic stem cell markers
Molecular Vision, 2012Co-Authors: Moon Nian Lim, Noor Hamidah Hussin, Ainoon Othman, Thiageswari Umapathy, Puteri Baharuddin, Rahman A A Jamal, Zubaidah ZakariaAbstract:Purpose: The presence of multipotent human limbal stromal cells resembling mesenchymal stromal cells (MSC) provides new insights to the characteristic of these cells and its therapeutic potential. However, little is known about the expression of stage-specific embryonic antigen 4 (SSEA-4) and the embryonic stem cell (ESC)-like properties of these cells. We studied the expression of SSEA-4 surface protein and the various ESC and MSC markers in the ex vivo cultured limbal stromal cells. The phenotypes and multipotent differentiation potential of these cells were also evaluated. Methods: Limbal stromal cells were derived from corneoscleral rims. The SSEA-4+ and SSEA-4- limbal stromal cells were sorted by fluorescence-activated cells sorting (FACS). Isolated cells were expanded and reanalyzed for their expression of SSEA-4. Expression of MSC and ESC markers on these cells were also analyzed by FACS. In addition, expression of limbal epithelial and corneal stromal proteins such as ATP-binding cassette sub-family G member 2 (ABCG2), tumour protein p63 (p63), paired box 6 (Pax6), Cytokeratin 3 (AE5), Cytokeratin 10, and keratocan sulfate were evaluated either by immunofluorecence staining or reverse transcription polymerase chain reaction. Appropriate induction medium was used to differentiate these cells into adipocytes, osteocytes, and chondrocytes. Results: Expanded limbal stromal cells expressed the majority of mesenchymal markers. These cells were negative for ABCG2, p63, Pax6, AE-5, and keratocan sulfate. After passaged, a subpopulation of these cells showed low expression of SSEA-4 but were negative for other important ESC surface markers such as Tra-1–60, Tra-1–81, and transcription factors like octamer-binding transcription factor 4 (Oct4), SRY(sex determining region Y)-box 2 (Sox2), and Nanog. Early passaged cells when induced were able to differentiate into adipocytes, osteocytes and chondrocytes. Conclusions: The expanded limbal stromal cells showed features of multipotent MSC. Our study confirmed the expression of SSEA-4 by a subpopulation of cultured limbal stromal cells. However, despite the expression of SSEA-4, these cells did not express any other markers of ESC. Therefore, we conclude that the cells did not show properties of ESC. The cornea is the major refractive element of the adult eye. It consists primarily of three layers: an outer layer of epithelium, a middle stromal layer of collagen-rich extracellular matrix (ECM) interspersed with keratocytes and an inner layer of endothelial cells. The stroma comprises 90% of the thickness of the cornea. It consists of dense, regularly packed collagen fibrils arrange as orthogonal layers or lamellae. The corneal stroma is unique in having a homogeneous distribution of fibrils with small diameter (25– 30 nm) that are regularly packed within lamellae and this arrangement minimizes light scattering permitting transparency. When an incisional wound penetrating into stroma occurs, the keratocytes become hypercellular myofibroblasts. These cells can later become wound
