Construction of a Defined Biomimetic Matrix for Long-Term Maintenance of Mouse Induced Pluripotent Stem Cells.
The existing in vitro culture systems often use undefined and animal-derived components for the culture of pluripotent stem cells. Artificial bioengineered peptides have the potential to become alternatives to these components of extracellular matrix (ECM). Integrins and cadherins are two cell adhesion proteins important for stem cell self-renewal, differentiation, and phenotype stability. In the present study, we sought to mimic the physico-biochemical properties of natural ECMs that allow self-renewal of mouse induced pluripotent stem cells (iPSCs). We develop a genetically engineered ECM protein (ERE-CBP) that contains (i) an integrin binding peptide sequence (RGD/R), (ii) an E-/N-cadherin binding peptide sequence (SWELYYPLRANL/CBP), and (iii) 12 repeats of APGVGV elastin-like polypeptides (ELPs/E).While ELPs allow efficient coating by binding to nontreated hydrophobic tissue culture plates, RGD/R and CBP support integrin- and cadherin-dependent cell attachment, respectively. Mouse iPSCs on this composite matrix exhibit a more compact phenotype compared to cells on control gelatin substrate. We also demonstrated that the ERE-CBP supports proliferation and long-term self-renewal of mouse iPSCs for up to 17 passages without GSK3β (CHIR99021) and Erk (PD0325901) inhibitors. Overall, our engineered ECM protein, which is cost-effective to produce in prokaryotic origin and flexible to modify with other cell adhesion peptides or growth factors, provides a novel approach for expansion of mouse iPSCs in vitro.
Department of Environmental Chemistry and Engineering, Interdisciplinary Graduate School of Science and Engineering, Tokyo Institute of Technology , Yokohama, 226-8502, Japan.
Nihad
N
Adnan
Department of Environmental Chemistry and Engineering, Interdisciplinary Graduate School of Science and Engineering, Tokyo Institute of Technology , Yokohama, 226-8502, Japan.
Masayasu
M
Mie
Department of Biomedical Engineering, University of California Davis , Davis, California 95616, United States.
Amranul
A
Haque
Biomaterials Center for Regenerative Medical Engineering, Foundation for Advancement of International Science , Tsukuba, 305-0821, Japan.
Sharif
S
Hossain
Department of Environmental Chemistry and Engineering, Interdisciplinary Graduate School of Science and Engineering, Tokyo Institute of Technology , Yokohama, 226-8502, Japan.
Yasumasa
Y
Mashimo
Biomaterials Center for Regenerative Medical Engineering, Foundation for Advancement of International Science , Tsukuba, 305-0821, Japan.
Toshihiro
T
Akaike
Department of Environmental Chemistry and Engineering, Interdisciplinary Graduate School of Science and Engineering, Tokyo Institute of Technology , Yokohama, 226-8502, Japan.
Eiry
E
Kobatake
D000327:Adsorption; D000595:Amino Acid Sequence; D000818:Animals; D032701:Biomimetics / Q000379:methods*; D018929:Cell Culture Techniques / Q000379:methods*; D002454:Cell Differentiation / Q000187:drug effects; D000066673:Cell Self Renewal / Q000187:drug effects; D016326:Extracellular Matrix Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D057927:Hydrophobic and Hydrophilic Interactions; D057026:Induced Pluripotent Stem Cells / Q000166:cytology* / Q000187:drug effects*; D016023:Integrins / Q000378:metabolism; D051379:Mice; D015202:Protein Engineering
D016326:Extracellular Matrix Proteins; D016023:Integrins