Clonal culturing of human embryonic stem cells on laminin-521/E-cadherin matrix in defined and xeno-free environment.
Lack of robust methods for establishment and expansion of pluripotent human embryonic stem (hES) cells still hampers development of cell therapy. Laminins (LN) are a family of highly cell-type specific basement membrane proteins important for cell adhesion, differentiation, migration and phenotype stability. Here we produce and isolate a human recombinant LN-521 isoform and develop a cell culture matrix containing LN-521 and E-cadherin, which both localize to stem cell niches in vivo. This matrix allows clonal derivation, clonal survival and long-term self-renewal of hES cells under completely chemically defined and xeno-free conditions without ROCK inhibitors. Neither LN-521 nor E-cadherin alone enable clonal survival of hES cells. The LN-521/E-cadherin matrix allows hES cell line derivation from blastocyst inner cell mass and single blastomere cells without a need to destroy the embryo. This method can facilitate the generation of hES cell lines for development of different cell types for regenerative medicine purposes.
Division of Matrix Biology, Department of Medical Biochemistry and Biophysics, Karolinska Institute, Stockholm SE-171 77, Sweden.
Sergey
S
Rodin
Division of Obstetrics and Gynecology, Department of Clinical Sciences, Intervention and Technology, Karolinska Institute and Karolinska University Hospital, Huddinge, Stockholm SE-141 86, Sweden.
Liselotte
L
Antonsson
Vascular Biology, Department of Medical Biochemistry and Biophysics, Karolinska Institute Stockholm SE-171 77, Sweden.
Colin
C
Niaudet
Division of Cardiothoracic Surgery and Anaesthesiology, Department of Molecular Medicine and Surgery, Advanced Center for Translational Regenerative Medicine (ACTREM), Karolinska Institute and Karolinska University Hospital, Stockholm SE-17177, Sweden.
Oscar E
OE
Simonson
1] Research Programs Unit, Molecular Neurology, University of Helsinki, Helsinki FI-00014, Finland [2] Folkhälsan Institute of Genetics, Helsinki FI-00014, Finland.
Elina
E
Salmela
Cardiovascular Research Center, Massachusetts General Hospital, Charles River Plaza/CPZN 3200, 185 Cambridge Street, Boston, Massachusetts 02114-2790, USA.
Emil M
EM
Hansson
Division of Matrix Biology, Department of Medical Biochemistry and Biophysics, Karolinska Institute, Stockholm SE-171 77, Sweden.
Anna
A
Domogatskaya
Division of Matrix Biology, Department of Medical Biochemistry and Biophysics, Karolinska Institute, Stockholm SE-171 77, Sweden.
Zhijie
Z
Xiao
Division of Obstetrics and Gynecology, Department of Clinical Sciences, Intervention and Technology, Karolinska Institute and Karolinska University Hospital, Huddinge, Stockholm SE-141 86, Sweden.
Pauliina
P
Damdimopoulou
Division of Obstetrics and Gynecology, Department of Clinical Sciences, Intervention and Technology, Karolinska Institute and Karolinska University Hospital, Huddinge, Stockholm SE-141 86, Sweden.
Mona
M
Sheikhi
Division of Obstetrics and Gynecology, Department of Clinical Sciences, Intervention and Technology, Karolinska Institute and Karolinska University Hospital, Huddinge, Stockholm SE-141 86, Sweden.
José
J
Inzunza
Division of Matrix Biology, Department of Medical Biochemistry and Biophysics, Karolinska Institute, Stockholm SE-171 77, Sweden.
Ann-Sofie
AS
Nilsson
Sheffield Diagnostic Genetic Services, Sheffield Children's NHS Trust, Sheffield S102TH, UK.
Duncan
D
Baker
Department of Laboratory Medicine, FENO, Karolinska Institute, Huddinge, Karolinska Hospital, Huddinge, Stockholm SE-141 86, Sweden.
Raoul
R
Kuiper
BioLamina AB, Löfströms Allé 5A, Sundbyberg 17266, Sweden.
Yi
Y
Sun
Department of Molecular Medicine and Surgery and Center for Molecular Medicine Karolinska Institutet, Karolinska University Hospital, Stockholm 171 76, Sweden.
Elisabeth
E
Blennow
Department of Molecular Medicine and Surgery and Center for Molecular Medicine Karolinska Institutet, Karolinska University Hospital, Stockholm 171 76, Sweden.
Magnus
M
Nordenskjöld
Division of Cardiothoracic Surgery and Anaesthesiology, Department of Molecular Medicine and Surgery, Advanced Center for Translational Regenerative Medicine (ACTREM), Karolinska Institute and Karolinska University Hospital, Stockholm SE-17177, Sweden.
Karl-Henrik
KH
Grinnemo
1] Research Programs Unit, Molecular Neurology, University of Helsinki, Helsinki FI-00014, Finland [2] Folkhälsan Institute of Genetics, Helsinki FI-00014, Finland [3] Department of Biosciences and Nutrition and Science for Life Laboratory, Karolinska Institute, Stockholm SE-141 83 Sweden.
Juha
J
Kere
Vascular Biology, Department of Medical Biochemistry and Biophysics, Karolinska Institute Stockholm SE-171 77, Sweden.
Christer
C
Betsholtz
Division of Obstetrics and Gynecology, Department of Clinical Sciences, Intervention and Technology, Karolinska Institute and Karolinska University Hospital, Huddinge, Stockholm SE-141 86, Sweden.
Outi
O
Hovatta
1] Division of Matrix Biology, Department of Medical Biochemistry and Biophysics, Karolinska Institute, Stockholm SE-171 77, Sweden [2] Cardiovascular and Metabolic Disorders Program, Duke-NUS, Singapore City 169857, Singapore.
Karl
K
Tryggvason
D015820:Cadherins; D018929:Cell Culture Techniques; D053595:Embryonic Stem Cells / Q000502:physiology*; D006801:Humans; D039121:Integrin alpha6beta1 / Q000378:metabolism; D007621:Karyotyping; D007797:Laminin; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism
D015820:Cadherins; D039121:Integrin alpha6beta1; D007797:Laminin; C474559:laminin 11 protein, human; D019869:Phosphatidylinositol 3-Kinases; D051057:Proto-Oncogene Proteins c-akt