Publications
Here is a selection of publications where different laminin isoforms were used to create more authentic cell culture systems.
Corneal integrins and their functions
Stepp M.A.Experimental Eye Research, 2005
There is a minimum of 12 different integrin heterodimers reported to be expressed by the major resident cells of the cornea: the corneal and limbal epithelial cells, keratocytes/fibroblasts, and corneal endothelial cells. These different integrin heterodimers play important and varied roles in maintaining the cornea and organizing how its cells interact with their surrounding extracellular matrix to maintain corneal clarity. In this review, an overview of the discovery and functions of integrins is provided along with a description of the current state of our knowledge of this large family of important proteins.
The functions of exogenous and endogenous laminin-5 on corneal epithelial cells
Ebihara N, Mizushima H, Miyazaki K, Watanabe Y, Ikawa S, Nakayasu K, Kanai A.Exp. Eye Res., 2000
In this study, the authors investigated the functions of laminin-332 on SV-40 transfected human corneal epithelial cells (HCE cells). We also revealed different functions between exogenous and endogenous laminin-332 on HCE cells. HCE cells themselves secreted laminin-5 endogenously. Exogenously added laminin-5 strongly promoted cell adhesion via a3b1 integrin, cell spreading, assembly of hemidesmosomes, and mildly inhibited cell migration. Using an anti-laminin-5 monoclonal antibody (mAb) or anti-integrin a3b1 mAbs, the blocking of the interaction between endogenously secreted laminin-5 and HCE cells caused strong inhibition of cell migration. Integrin a3b1 and a6b4 were expressed in HCE cells. These results indicated that endogenous (unprocessed) laminin-5 has a crucial role in cell migration on HCE cells via a3b1 integrin. In conclusion, structural differences between exogenous and endogenous laminin-5 regulated their functions on HCE cells. Exogenously added laminin-5 strongly promoted cell adhesion, cell spreading, and assembly of hemidesmosomes. Endogenously secreted laminin-5 had a crucial role in cell migration.
Highly sensitive droplet digital PCR method for detection of residual undifferentiated cells in cardiomyocytes derived from human pluripotent stem cells
Kuroda T., Yasuda S., Matsuyama S., Tano K., Kusakawa S., Sawa Y., Kawamata S., Sato Y. Regenerative Therapy, 2015
Human iPSCs were maintained on laminin-521 in Essential 8 medium. Cardiac differentiation on laminin-521 and Matrigel. Adult human cardiomyocytes (Promocell, Heidelberg) were cultured on laminin-211 in the Promocell myocyte growth medium. The authors have established a sensitive assay for the detection of the residual undifferentiated hiPSCs in cardiomyocytes, using droplet digital PCR (ddPCR). LIN28 was the most sensitive marker of residual undifferentiated cells in hiPSC-derived cardiomyocytes but also in other tissues such as liver, heart, pancreas, kidney, spinal cord, corneal epithelium, and lung. Hence, the LIN28/ddPCR method is applicable to the quality control of hiPSC-derived cell therapy products.
Xeno- and feeder-free differentiation of human pluripotent stem cells to two distinct ocular epithelial cell types using simple modifications of one method
Hongisto H., Ilmarinen T., Vattulainen M., Mikhailova A., Skottman H. Stem cell research and therapy, 2017
Here the authors describe a robust xeno- and feeder cell-free culture system for undifferentiated hPSCs along with efficient and scalable methods to derive high-quality retinal pigment epithelial (RPE) cells and corneal limbal epithelial stem cells (LESCs). Multiple genetically distinct hPSC lines were adapted to a robust, defined, xeno-, and feeder-free culture system of Essential 8™ medium and laminin-521 matrix. Thereafter, two-stage differentiation methods toward ocular epithelial cells were established utilizing xeno-free media and a combination of extracellular matrix proteins laminin-521 and Collagen IV. Derivative RPE formed functional epithelial monolayers with mature tight junctions and expression of RPE genes and proteins, as well as phagocytosis and key growth factor secretion capacity after 9 weeks of maturation on inserts. Efficient LESC differentiation led to cell populations expressing LESC markers such as p40/p63α by day 24. Finally, the authors established xeno-free cryobanking protocols for pluripotent hPSCs, hPSC-RPE cells, and hPSC-LESCs, and demonstrated successful recovery after thawing on laminin-521 and Collagen IV. The simple xeno-free methods described here could be upgraded to GMP-quality for future preclinical testing and safety and functional efficacy testing of the hPSC-RPE and hPSC LESCs produced with these protocols are currently ongoing in non-human primates and rabbit models of LSCD, respectively.
Laminin-511 is an epithelial message promoting dermal papilla development and function during early hair morphogenesis
Jing Gao, Mindy C. DeRouen, Chih-Hsin Chen, Michael Nguyen, Ngon T. Nguyen, Hiroyuki Ido, Kenji Harada, Kiyotoshi Sekiguchi, Bruce A. Morgan, Jeffery H. Miner, Anthony E. Oro, and M. Peter Marinkovich. Genes Dev, 2008
Abnormal adhesion of grafted corneal endothelial cells (CECs) affects the application of intracameral injection for corneal endothelial dysfunction therapy. The authors explored whether laminin 511 (LN511) improves the therapeutic function of the intracameral CEC injection. Injected LN511 was found to be able to settle and form a coating on the posterior surface of Descemet's membrane (DM). The data suggests that the strategy of LN511 precoating and CECs' intracameral injection could be a potential method for the therapy of corneal endothelial dysfunction.