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EMBRYONIC STEM CELLS/INDUCED PLURIPOTENT STEM CELLS

Laminin-511 but Not -332, -111, or -411 Enables Mouse Embryonic Stem Cell Self-Renewal In Vitro

^aDivision of Matrix Biology, Department of Medical Biochemistry and Biophysics, Karolinska Institute, Stockholm, Sweden;
^bBioStratum, Durham, North Carolina, USA

Key Words. Laminins • Mouse embryonic stem cells • Self-renewal • Extracellular matrix • Adhesion • Proliferation

Correspondence: Correspondence: Karl Tryggvason, M.D., Ph.D., Division of Matrix Biology, Department of Medical Biochemistry and Biophysics, Karolinska Institute, 171 77 Stockholm, Sweden. Telephone: +46 8 5248 7720; Fax: +46 8 316 165; e-mail: Karl.Tryggvason{at}ki.se

Received on May 21, 2007; accepted for publication on August 14, 2008.

First published online in STEM CELLS EXPRESS  August 28, 2008.

We tested specific laminin (LN) isoforms for their ability to serve as substrata for maintaining mouse embryonic stem (ES) cells pluripotent in vitro in the absence of leukemia inhibitory factor or any other differentiation inhibitors or feeder cells. Recombinant human LN-511 alone was sufficient to enable self-renewal of mouse ES cells for up to 169 days (31 passages). Cells cultured on LN-511 maintained expression of pluripotency markers, such as Oct4, Sox2, Tert, UTF1, and Nanog, during the entire period, and cells cultured for 95 days (17 passages) were used to generate chimeric mice. LN-332 enabled ES cells proliferation but not pluripotency. In contrast, under the same conditions LN-111, Matrigel, and gelatin caused rapid differentiation, whereas LN-411 and poly-D-lysine did not support survival. ES cells formed a thin monolayer on LN-511 that differed strikingly from typical dense cluster ES cell morphology. However, expression of pluripotency markers was not affected by morphological changes. The effect was achieved at low ES cell density (<200 cell/mm²). The ability of LN-511 and LN-332 to support ES cell proliferation correlated with increased cell contact area with those adhesive substrata. ES cells interacted with LN-511 via β1-integrins, mostly 6β1 and Vβ1. This is the first demonstration that certain extracellular matrix molecules can support ES cell self-renewal in the absence of differentiation inhibitors and at low cell density. The results suggest that recombinant laminin isoforms can provide a basis for defined surface coating systems for feeder-free maintenance of undifferentiated mammalian ES cells in vitro.

Disclosure of potential conflicts of interest is found at the end of this article.