Publications
Here is a selection of publications where different laminin isoforms were used to create more authentic cell culture systems.
iPSC-derived human cardiac progenitor cells improve ventricular remodelling via angiogenesis and interstitial networking of infarcted myocardium
Ja K.P., Miao Q., Zhen Tee N.G., Lim S.Y., Nandihalli M, Ramachandra C.J.A, Mehta A, Shim W. J Cell Mol Med. 2015
Here they investigate the effects of myocardial transplantation of human induced pluripotent stem cell (iPSC)-derived progenitors and cardiomyocytes into acutely infarcted myocardium in immune deficient mice. iPSC cultured on Matrigel and differentiated in EB structures. Differences in integrin and laminin expression between cardiac progenitors and cardiomyocytes were observed. The a6 integrin was higher expressed in progenitors and integrin a1, a2, a3, a7 and b1 higher expression in cardiomyocytes. They observed a distinct swish in expression profile of laminin subunits during cardiac differentiation with laminin-411/421 pre-dominantly expressed early in progenitors and laminin-211/221 expressed later in cardiomyocytes. Improvements of myocardial function in the progenitor group corresponded to increased vascularization and coincided with augmented networking of cardiac telocytes in the interstitial space of the infarcted zone. Laminin-221/211-expressing cardiomyocytes only retained and engrafted around myofibres in the peri-infarct region.
Molecular Diversity of Midbrain Development in Mouse, Human, and Stem Cells
Manno, Gyllborg, Codeluppi, Nishimura, Salto, Zeisel, Borm, Stott, Toledo, Villaescusa, Lönnerberg, Ryge, Barker, Arenas, Linnarsson.Cell, 2016
Manno and colleagues used single-cell RNA sequencing to examine ventral midbrain development in humans and mice. They found that cell types and gene expression were generally conserved across species, but with clear differences in cell proliferation, developmental timing, and dopaminergic neuron development. Additionally, they quantitatively assessed the fidelity of dopaminergic neurons derived from human pluripotent stem cells, at a single-cell level. The study provides insight into the molecular programs controlling human midbrain development and provides a foundation for the development of cell replacement therapies.
Imaging-Based Screen Identifies Laminin 411 as a Physiologically Relevant Niche Factor with Importance for i-Hep Applications
Ong J., Paola Serra M., Segal J., Cujba A.-M., Seng Ng S., Butler R., Millar V., Hatch S., Zimri S., Koike H., Chan K., Bonham A., Walk M., Voss T., Heaton N., Mitry R., Dhawan A., Ebner D., Danovi D., Nakauchi H., Rashid S.T.Stem Cell Reports, 2018
Here, the authors show that extracellular niche factors likely play a critical role in bridging this gap in functional differences between hepatocytes derived from induced pluripotent stem cells (i-Heps) and primary cells. They defined a profile of healthy, freshly isolated primary hepatocytes (Hepatocyte Likeness Index; HLI) that cells of interest can be compared against for high-throughput screening. They applied this platform to screen hepatocyte niche factors for their ability to drive i-Heps closer to that target and validated their findings in a pharma-like screening environment. The HLI was applied in a targeted screen of extracellular niche factors to identify substrates driving i-Heps closer to the standard. Results from the screen performed highlighted the important role played by laminins where laminin 411 was identified as a key niche protein. Laminin 411 is a component of the hepatic niche. Laminin 411 advanced i-Heps toward functional significance and prolonged survival of hepatic progenitor cells, contributing to better i-Hep-based drug-screening applications. This paper underscores the importance of combining substrates, soluble factors, and HCA when developing iPSC applications.
Laminin-511 but not -332, -111, or -411 enables mouse embryonic stem cell self-renewal in vitro
Domogatskaya A., Rodin S., Boutaud A., Tryggvason K. Stem Cells, 2008
Different laminin isoforms, LN-511, -332, -411 and -111, as well as Matrigel, gelatin and poly-D-lysine are compared as substrates maintaining pluripotent mouse ES cells in vitro without the addition of leukemia inhibitory factor. Conclusions are that only LN-511 is able to sustain self-renewal for up to 169 days of culturing and cells maintain expression of pluripotency markers and can be used for generation of chimeric mice.
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.
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.
Laminin-511 expression is associated with the functionality of feeder cells in human embryonic stem cell culture
Hongisto H., Vuoristo S., Mikhailova A., Suuronen R., Virtanen I., Otonkoski T., Skottman H. Stem Cell Res., 2012
The authors of this study show that fibroblast feeders synthesize laminin-511 and that this is the main protein responsible for the maintenance of hESC pluripotency. This strengthens the already known biological function of laminin-511 and laminin-521 as the main cell-adhesion molecules for pluripotent stem cells.
Human embryonic stem cell dispersion in electrospun PCL fiber scaffolds by coating with laminin-521 and E-cadherin-Fc
Leino M., Astrand C., Hughes-Brittain N., Robb B., McKean R., Chotteau V.J Biomed Mater Res Part B 2017
Here, the authors describe a nonaggregate culture system of human embryonic stem cells inside electrospun polycaprolactone (PCL) fiber scaffolds combined with the defined extracellular proteins laminin-521, naturally occurring in the stem cell niche. PCL fiber scaffolds coated with recombinant human laminin-521 readily supported initial stem cell attachment and growth from a single-cell suspension. The combination of recombinant E-cadherin-Fc and laminin-521 slightly improved cell dispersion rendering a uniform cell population. Finally, we showed that the cells cultured in E-cadherin-Fc- and laminin-521-coated PCL scaffolds could differentiate into all three germ layers. Importantly, we provided a chemically defined 3-D system in which pluripotent stem cells grew and differentiated avoiding the formation of cell aggregates.
Dynamic blebbing: A bottleneck to human embryonic stem cell culture that can be overcome by Laminin-Integrin signaling
Weng J-H., Cheung C., Talbot P.Stem Cell Research, 2018
This study characterizes dynamic and apoptotic blebbing in human embryonic stem cells (hESC), identifies dynamic blebbing as a bottleneck to successful cell attachment during passaging, and demonstrates that dynamic blebbing can be rapidly stopped by plating cells on recombinant human laminin. In freshly plated hESC, dynamic and apoptotic blebbing differed in time of occurrence, bleb retraction rate, mitochondrial membrane potential, and caspase 3&7 activation. While dynamic blebbing can be controlled with drugs that inhibit myosin II, these methods have off-target effects and are not suitable for clinical applications. Recombinant human laminin-521 or addition of laminin-111 to Matrigel provided a safe method to drastically decrease dynamic blebbing and improve cell attachment with proteins normally found in the inner cell mass. Inhibition of focal adhesion kinase, which is activated by binding of integrins to laminin, prolonged dynamic blebbing and inhibited attachment. These data show that hESC binds rapidly to laminins through an integrin, which activates focal adhesion kinase that in turn downregulates dynamic blebbing. Laminins enabled hESC to rapidly attach during passaging, improved plating efficiency, enabled passaging of single pluripotent stem cells, and avoided the use of inhibitors that have non-specific off-target effects. These data provide a strategy for improving hESC culture using biologically safe recombinant human proteins.
Laminin 521 stabilizes the pluripotency expression pattern of human embryonic stem cells initially derived on feeder cells
Albalushi H., Kurek M., Karlsson L., Landreh L., Rós Kjartansdóttir K., Söder O., Hovatta O., Stukenborg J-B.Stem Cell International, 2017
In this article, the authors tested the effect of the laminin-521 substrate on cultured hES cells. Five male hES cell lines, originally derived on human foreskin fibroblasts (hFFs), were cultured on hFF, Matrigel, laminin-521 or laminin-121 for nine passages. Variations in gene expression related to pluripotency, stemness, and male germ and somatic gonadal cells at different passages and different culture methods were evaluated. All cell lines expressed pluripotency markers at protein and gene level and were able to differentiate into cell types of the three germ layers after being cultured on laminin-521 for nine passages. Laminin 521 had no obvious effect on the expression of genes related to male germ cells and somatic gonadal cells when used as a matrix for hES cell cultures. Importantly, reduction in variation of pluripotency marker expression was observed after culturing the cells on laminin-521 compared to culture on hFFs, and could be reduced further with an increased culture period on laminin-521. Changing the cell culture medium of hES cells on hFFs from Dulbecco's Modified Eagle Medium (DMEM) to NutriStem, which is commonly used for culturing the cells on matrices, did not affect the variation of pluripotency genes and genes related to stemness expression. hES cells cultured on laminin-521 were more homogenous, attached better and grew faster compared to hES cells cultured on matrigel. The results show that laminin-521 provides optimal culture conditions for adaptation to feeder and xeno-free conditions of hES cells derived and cultured on feeder cells. Moreover, laminin-521 has a positive effect on stabilizing and homogenizing pluripotent gene expression profiles between hES cell lines provides the first step towards more controllable and robust culture conditions for hES cells.