Laminin-511 and -521 enable efficient in vitro expansion of human corneal endothelial cells
| Autor: | Ursula Schlötzer-Schrehardt, Ryohei Numata, Friedrich E. Kruse, Naoki Okumura, Kazuya Kakutani, Shigeru Kinoshita, Makiko Nakahara, Noriko Koizumi |
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| Rok vydání: | 2015 |
| Předmět: |
Corneal endothelium
Endothelium Integrin Cell Culture Techniques Gene Expression Tissue engineering Laminin medicine Cell Adhesion Humans Protein Isoforms Cell adhesion Descemet Membrane Cell Proliferation biology Integrin alpha6beta1 Tissue Engineering Chemistry Endothelium Corneal Integrin alpha3beta1 Endothelial Cells Cell Differentiation Adhesion eye diseases Cell biology Culture Media medicine.anatomical_structure Phenotype Cell culture biology.protein sense organs |
| Zdroj: | Investigative ophthalmologyvisual science. 56(5) |
| ISSN: | 1552-5783 |
| Popis: | Purpose The purpose of this study was to investigate the usefulness of laminin isoforms as substrates for culturing human corneal endothelial cells (HCECs) for clinical application of tissue engineering therapy. Methods Expression of specific laminin chains in human corneal endothelium and Descemet's membrane was analyzed at the mRNA and protein levels. The effect of laminin-511 and -521 on cell adhesion and proliferation was evaluated. Recombinant laminin E8 fragments (E8s), which represent functionally minimal forms of laminins, were also evaluated for their effects on cell density and cellular phenotype. The potential involvement of α3β1 and α6β1 integrins in laminin signal transduction was also investigated using neutralizing antibodies. Results Laminin-511 and -521 were expressed in Descemet's membrane and corneal endothelium. These laminin isoforms significantly enhanced the in vitro adhesion and proliferation, and differentiation of HCECs. A cell density of 2200 to 2400 cells/mm2 was achieved when HCECs were cultured on laminin-511 or -521, whereas the density was only 1100 cells/mm2 on an uncoated control. E8s also supported HCEC cultivation with a similar efficacy to that obtained with full-length laminin. Functional blocking of α3β1 and α6β1 integrins suppressed the adhesion of HCECs even in the presence of laminin-511. Conclusions Laminin-511 and -521 were the laminin isoforms present in Descemet's membrane, and these laminins modulate the adhesion and proliferation of CECs. Laminin E8s represent an ideal xeno-free defined substrate for the culture of CECs for clinical applications. |
| Databáze: | OpenAIRE |
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