Publications
Here is a selection of publications where different laminin isoforms were used to create more authentic cell culture systems.
Changes in Corneal Basal Epithelial Phenotypes in an Altered Basement Membrane
Wang I-J., Jui-Fang Tsai R., Yeh L-K., Yao-Nien Tsai R., Hu F-R., Kao W.W.Y.PLOS ONE, 2011
Analysis of human healthy donors shows that the BM of the limbal epithelium differs from that of the central cornea. Aside from laminin-111 and laminin-332, the limbal BM also contains laminin α2β2 chains, while the corneal BM does not.
Adhesion, Migration, and Proliferation of Cultured Human Corneal Endothelial Cells by Laminin-5
Yamaguchi M., Ebihara N., Shima N., Kimoto M., Funaki T., Yokoo S., Murakami A., Yamagami S. Investigative Ophthalmology and Visual Science, 2011
Here, the authors investigate the expression of laminin-332 and its receptors by human corneal endothelial cells (HCECs) and the effect on adhesion, proliferation, and migration of cultured HCECs. Adult HCECs expressed the laminin-332 receptor a3B1 integrin, but not laminin-332 itself. Laminin-332 is expressed in the basement membrane of the corneal epithelium, but not in the corneal endothelium. Significantly more HCEC cells became adherent to recombinant laminin-332-coated dishes than to uncoated dishes in the cell adhesion assay. The proliferation of cultured HCECs was moderately promoted by laminin-332. A significantly higher percentage of wound closure was obtained with medium containing soluble laminin-332 than with the control medium in the wound-healing assay. To conclude, recombinant laminin-332 promotes adhesion, migration, and moderate proliferation of cultured HCECs. The results suggest that immature or undifferentiated HCECs express laminin-332, whereas it is suppressed during development or differentiation.
The Different Binding Properties of Cultured Human Corneal Endothelial Cell Subpopulations to Descemet’s Membrane Components
Toda M., Ueno M., Yamada J., Hiraga A., Tanaka H., Schlötzer-Schrehardt U., Sotozono C., Kinoshita S., Hamuro J.Invest Ophthalmol Vis Sci. 2016
In culture, human corneal endothelial cell (cHCEC) tend to enter into cell-state transition (CST), such as epithelial-to-mesenchymal transition (EMT) or fibrosis, thus resulting in the production of different subpopulations. In this study, the authors examined the binding ability of cHCECs subpopulations to major Descemet’s membrane components that distribute to the endothelial face; that is, laminin-511, -411, Type-IV collagen, and proteoglycans. Each subpopulation was prepared by controlling the culture conditions or by using magnetic cell separation and then confirmed by staining with several cell-surface markers. Binding abilities of HCEC subpopulations were examined by adding the cells to culture plates immobilized with collagens, laminins, or proteoglycans, and then centrifuging the plates. The cHCECs showed the best attachment to laminin laminin-521 and -511. The cells showed a weaker binding to laminin-411, laminin-332, Type-IV collagen. The minimum concentrations necessary for the observed cell binding in this study were as follows: laminin-521 and -511, 3 ng/mL; laminin-411, 2.85 ug/mL; Type-IV collagen, 250 ng/mL. Cells suspended in Opti-MEM-I or Opeguard-MA were bound to laminin, yet no binding was observed in cells suspended in BSS-Plus. Both the fully differentiated, mature cHCEC subpopulations and the epithelial-to-mesenchymal– transitioned (EMT)-phenotype subpopulation were found to attach to laminin- or collagen-coated plates. Interestingly, the binding properties to laminins differed among those subpopulations. Although the level of cells adhered to the laminin-411–coated plate was the same among the cHCEC subpopulations, the fully differentiated, mature cHCEC subpopulations were significantly more tightly bound to laminin-511 than was the EMT-phenotype subpopulations. These findings suggest that the binding ability of cHCECs to major Descemet’s membrane components is distinct among cHCEC subpopulations and that Opti-MEM-I and Opeguard-MA are useful cell-suspension vehicles for cell-injection therapy. This research group focused on developing a novel medical approach, termed cell-injection therapy, for the treatment of patients with endothelial dysfunction.
Laminin synthesis and the adhesion characteristics of immortalized human corneal epithelial cells to laminin isoforms
Filenius S., Hormia M., Rissanen J., Burgeson R.E., Yamada Y., Araki-Sasaki K., Nakamura M., Virtanen I., Tervo T. Exp. Eye Res., 2001
The authors studied the synthesis of laminins and determined the specific integrins mediating the adhesion of immortalized human corneal epithelial cells to mouse laminin-111, and human laminin-332 and laminin-511. The cells produced a3, b3 and g2 chains of laminin-332, but not laminin-111 and laminin-511. Integrin a3B1 complex-mediated the adhesion of corneal epithelial cells to human laminin-332 and laminin-511. Integrin complex a3B, as well as laminin a3 chain, was also shown to mediate cell adhesion to newly produced endogenous laminin-332. The results also show that among corneal basement membrane laminins, laminin-332 is synthesized by epithelial cells while laminin-511 may be a product of keratocytes.
Identification and Potential Application of Human Corneal Endothelial Progenitor Cells
Hara S., Hayashi R., Soma T., Kageyama T., Duncan T., Tsujikawa M., Nishida K.Stem Cells Dev. 2014
This article demonstrates for the first time that Laminin-511 is an optimal, human matrix for the isolation and expansion of corneal endothelial progenitors. The authors show that the proliferative capacity of these endothelial progenitors is very high on Laminin-511 compared to conventional methods. Laminin-511 can be used to rapidly isolate and expand a homogenous population of endothelial progenitor cells that can be differentiated to endothelial cells in a biorelevant environment. The authors demonstrate that the proliferative capacity of these endothelial progenitors is very high on Laminin-511 compared to conventional methods. Laminin-511 can thus be used to rapidly isolate and expand a homogenous population of endothelial progenitors that can be differentiated to endothelial cells in a biorelevant environment. Main points of the article are: 1) High proliferative capacity in serum-free media compared to standard methods, 2) Large numbers of cells generated, 3) Facilitates rapid isolation of a homogenous population of endothelial progenitors, 4) Enables differentiation to endothelial cells in a biorelevant environment, 5) Cells can be subcultured for at least 5 passages.
Laminin-511 and -521 Enable Efficient In Vitro Expansion of Human Corneal Endothelial Cells
Okumura N., Kakutani K., Numata R., Nakahara M., Schlötzer-Schrehardt U., Kruse F., Kinoshita S., Koizumi N. IVOS Cornea, 2015
The purpose of this study was to investigate the usefulness of laminin isoforms as substrates for culturing human corneal endothelial cells (HCECs) for clinical applications. 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 compared to uncoated control, fibronectin, and collagen I. iMatrix also supported HCEC cultivation with similar efficacy to that obtained with full-length laminin. Functional blocking of a3b1 and a6b1 integrins suppressed the adhesion of HCECs even in the presence of laminin-511.
Molecular interactions in the retinal basement membrane system: A proteomic approach
Manimalha Balasubramani, Emanuel M Schreiber, Joseph Candiello, G K Balasubramani, Justin Kurtz, Willi Halfter. Matrix Biol, 2010
The authors describe a direct analysis of an in vivo basement membrane (BM) system using a mass spectrometry (MS) based proteomics approach. Nidogens-1 and -2, laminin subunits α1, α5, β2, and γ1, agrin, collagen XVIII, perlecan, FRAS1 and FREM2 were the most abundant BM protein components.
Endothelial basement membrane limits tip cell formation by inducing Dll4/Notch signaling in vivo
Stenzel D., Franco C.A., Estrach S., Mettouchi A., Sauvaget D., Rosewell I., Schertel A., Armer H., Domogatskaya A., Rodin S., Tryggvason K., Collinson L., Sorokin L., Gerhardt H. EMBO reports, 2011
Here the authors show that laminin α4 regulates tip cell numbers and vascular density by inducing endothelial Dll4/Notch signaling in vivo deficiency leads to reduced Dll4 expression, excessive filopodia and tip cell formation in the mouse retina, phenocopying the effects of Dll4/Notch inhibition. Lama4-mediated Dll4 expression requires a combination of integrins in vitro and integrin β1 in vivo. The authors conclude that appropriate laminin/integrin‐induced signaling is necessary to induce physiologically functional levels of Dll4 expression and regulate branching frequency during sprouting angiogenesis.
Laminin Expression in Adult and Developing Retinae: Evidence of Two Novel CNS Laminins
Libby R.T., Champliaud M-F, Claudepierre T., Xu Y., Gibbons E.P., Koch M., Burgeson R.E., Hunter D.D, Brunken W.J. The Journal of Neuroscience, 2000
Here, they examine the expression of all known laminin chains within the retina. The interphotoreceptor matrix (and, during early development, the subretinal space) contains the laminin a3, a4, a5, b2, b3, g2, and g3 chains. This suggests the presence of three laminins: laminin-332, laminin-423, and laminin-523. These laminin isoforms could exert important effects on photoreceptor development and may play a role in photoreceptor production, stability and synaptic organization.
Retinal Pigment Epithelial Cells Synthesize Laminins, Including Laminin 5, and Adhere to Them through a3- and a6-Containing Integrins
Aisenbrey S., Zhang M., Bacher D., Yee J., Brunken W.J., Hunter D.D.Invest Ophthalmol Vis Sci., 2006
The multilayered extracellular matrix underlying the retina is Bruch’s membrane (BM). Here they show that BM contains laminin chains that could form laminin-111, -332, -511, and -521. RPE cells synthesized these laminin chains in vitro, hence, RPE cells may synthesize BM laminins. The RPE cells adhered to the BM component collagen IV, but preferentially adhered to laminins. Of the laminins tested, the RPE cells adhered preferentially to laminin 332. The RPE cells interacted with these laminins via a3 and a6 containing integrins.