Here is a selection of publications where different laminin isoforms were used to create more authentic cell culture systems.
Direct Reprogramming of Human Fetal- and Stem Cell-Derived Glial Progenitor Cells into Midbrain Dopaminergic Neurons
Nolbrant S., Giacomoni J., Hoban D.B, Bruzelius A., Birtele M., Chandler-Militello D., Pereira M., Rylander Ottosson D., Goldman S.A., Parmar M. Stem Cell Reports, 2020
Human glial progenitor cells (hGPCs) are promising cellular substrates to explore for the in situ production of new neurons for brain repair. Proof of concept for direct neuronal reprogramming of glial progenitors using human cells has been difficult to perform since hGPCs are born late during human fetal development, with limited accessibility for in vitro culture. In this study, the authors provide evidence that hGPCs isolated from both the human fetal brain and differentiated from hESCs can be successfully reprogrammed into functional iNs, including induced DA neurons (iDANs). They also establish a renewable and reproducible stem cell-based hGPC system for direct neural conversion in vitro. Using this system, they have identified optimal combinations of fate determinants for the efficient dopaminergic (DA) conversion of hGPCs, thereby yielding a therapeutically relevant cell type that selectively degenerates in Parkinson’s disease.
Highly efficient reprogramming to pluripotency and directed differentiation of human cells using synthetic modified mRNA
Warren L., Manos P.D., Ahfeldt T., Loh Y-H., Li H., Lau F., Ebina W., Mandal P., Smith Z.D., Meissner A., Daley g.Q., Brack A.S., Collins J.J, Cowan C., Schlaeger T.M, Rossi D.J. Cell Stem Cell., 2010
In this article, the authors describe a simple, non-integrating strategy for reprogramming cell fate based on the administration of synthetic mRNA modified to overcome innate antiviral responses. Laminin-521 is used as a matrix. They show that this approach can reprogram multiple human cell types to pluripotency with efficiencies that greatly surpass established protocols and that the same technology can be used to efficiently direct the differentiation of RNA-induced pluripotent stem (RiPS) cells into terminally differentiated myogenic cells.
Chemically defined and xeno-free culture condition for human extended pluripotent stem cells
Bei Liu, Shi Chen, Yaxing Xu, Yulin Lyu, Jinlin Wang, Yuanyuan Du, Yongcheng Sun, Heming Liu, Haoying Zhou, Weifeng Lai, Anqi Xue, Ming Yin, Cheng Li, Yun Bai, Jun Xu & Hongkui Deng. Nature Communications, 2021
This study shows the significant benefit of culturing and deriving extended pluripotent stem cells on Biolaminin 521 (LN521) compared to using Matrigel, Geltrex, vitronectin, extracted laminin, collagen, fibronectin, or laminin-511. In addition to enabling xeno-free and chemically defined culture conditions, the study clearly shows that laminin-521 promotes attachment (measured at 1.5 h), survival (24h), and proliferation (72h). Human EPS cells were long-term and genetically stably maintained in vitro, preserving their embryonic and extraembryonic developmental potentials. The study also showed efficient derivation from human fibroblast through reprogramming.
Directed differentiation of human iPSC into insulin-producing cells is improved by induced expression of PDX1 and NKX6.1 factors in IPC progenitors
Walczak M. P., Drozd A. M., Stoczynska-Fidelus E., Rieske P., Grzela D.P.Journal of Translational Medicine, 2016
Here, the authors show that the highest efficiencies of reprogramming of fibroblasts were obtained on Laminin-511 and Laminin-521, compared to other coatings. iPSC cell lines were created with stably integrated PDX1 and NKX6.1 transgenes under the transcriptional control of the doxycycline-inducible promoter. These cells were differentiated into insulin-producing cells. Generated cells displayed molecular markers characteristic for respective steps of the differentiation. The obtained IPC secreted insulin and produced C-peptide with a significantly higher hormone release level in the case of the combined expression of PDX1 and NKX6.1 induced at the last stage of the differentiation. The efficiency of differentiation of iPSC to IPC can be increased by concurrent expression of PDX1 and NKX6.1 during progenitor cell maturation.
Integration Free Derivation of Human Induced Pluripotent Stem Cells Using Laminin 521 Matrix
Uhlin E., Marin Navarro A., Rönnholm H., Day K., Kele M., Falk A. J Vis Exp. 2017
In this video publication, the authors describe a consistent, highly reproducible and easy-to-use protocol, providing a robust and practical way to generate defined and xeno-free human hiPS cells from fibroblasts. It also offers a user-friendly culture system for the maintenance of established hiPS cells. Xeno-free and fully defined conditions are key parameters for robust and reproducible generation of homogenous human induced pluripotent stem (hiPS) cells. Utilizing the defined recombinant human laminin 521 (LN-521) matrix in combination with xeno-free and defined media formulations reduces variability and allows for the consistent generation of hiPS cells. The Sendai virus (SeV) vector is a non-integrating RNA-based system, thus circumventing concerns associated with the potential disruptive effect on genome integrity integrating vectors can have. Furthermore, these vectors have demonstrated relatively high efficiency in the reprogramming of dermal fibroblasts. In addition, enzymatic single-cell passaging of cells facilitates homogeneous maintenance of hiPS cells without substantial prior experience of stem cell culture. This protocol has been extensively tested and used to derive more than 300 hiPS cell lines in the Swedish national human iPS Core facility at Karolinska Institutet of which some lines have previously been described.
Generation of human iPS cell line CTL07-II from human fibroblasts, under defined and xeno-free conditions
Kele M., Day K., Rönnholm H., Schuster J., Dahl N., Falk A.Stem cell research, 2016
CTL07-II is a healthy feeder-free and characterized human induced pluripotent stem (iPS) cell line cultured under xeno-free and defined conditions. iPS cell coating during derivation and expansion was human recombinant Laminin-521. The line is generated from healthy human fibroblasts with non-integrating Sendai virus vectors encoding the four Yamanaka factors, OCT4, SOX2, KLF4, and cMYC. The generated iPS cells are free from reprogramming vectors and their purity, karyotypic stability and pluripotent capacity are confirmed.
A deﬁned xeno-free and feeder-free culture system for the derivation, expansion and direct differentiation of transgene-free patient-speciﬁc induced pluripotent stem cells
Lu H.F., Chai C., Lim T.C., Leong M.F., Lim J.K., Gao S., Lim K.L., Wan A.C. Biomaterials 2014
Reprogramming of iPSCs on LN-521 and direct differentiation to dopaminergic cells on Laminin-521. This article demonstrates LN-521 as an optimal defined, xeno- and feeder-free matrix for the reprogramming of human iPS cells. Laminin-521 achieves high-efficiency reprogramming in different media, fast and easy expansion as well as direct differentiation to dopaminergic neurons on LN-521. The authors conclude that the efficient transgene-free hiPSC derivation and expansion on LN-521 enables clinical applications useful for human patient iPSCs and derivatives for cellular therapy.
Derivation of human iPS cell lines from monozygotic twins in defined and xeno-free conditions
Uhlin E., Rönnholm H., Day K., Kele M., Tammimies K., Bölte S., Falk A.Stem Cell Res., 2017
Human-induced pluripotent stem (hiPS) cell lines CTRL-9-II and CTRL-10-I were derived from healthy monozygotic twin donors using non-integrating RNA based Sendai virus reprogramming and cultured in a xeno-free chemically defined condition on the laminin-521 cell culture substrate. The established hiPS cell lines, CTRL-9-II and CTRL-10-I, are karyotypically normal, free from reprogramming vectors, display endogenously expression of pluripotency factors at levels similar to embryonic stem cells. The generated iPS cell lines demonstrate pluripotency bypassing bioinformatics assay PluriTest and by embryonic body assay.