title: Structure and barrier properties of human embryonic stem cell-derived retinal pigment epithelial cells are affected by extracellular matrix protein coating.
abstract: Extracellular matrix (ECM) interactions play a vital role in cell morphology, migration, proliferation, and differentiation of cells. We investigated the role of ECM proteins on the structure and function of human embryonic stem cell-derived retinal pigment epithelial (hESC-RPE) cells during their differentiation and maturation from hESCs into RPE cells in adherent differentiation cultures on several human ECM proteins found in native human Bruch's membrane, namely, collagen I, collagen IV, laminin, fibronectin, and vitronectin, as well as on commercial substrates of xeno-free CELLstart™ and Matrigel™. Cell pigmentation, expression of RPE-specific proteins, fine structure, as well as the production of basal lamina by hESC-RPE on different protein coatings were evaluated after 140 days of differentiation. The integrity of hESC-RPE epithelium and barrier properties on different coatings were investigated by measuring transepithelial resistance. All coatings supported the differentiation of hESC-RPE cells as demonstrated by early onset of cell pigmentation and further maturation to RPE monolayers after enrichment. Mature RPE phenotype was verified by RPE-specific gene and protein expression, correct epithelial polarization, and phagocytic activity. Significant differences were found in the degree of RPE cell pigmentation and tightness of epithelial barrier between different coatings. Further, the thickness of self-assembled basal lamina and secretion of the key ECM proteins found in the basement membrane of the native RPE varied between hESC-RPE cultured on compared protein coatings. In conclusion, this study shows that the cell culture substrate has a major effect on the structure and basal lamina production during the differentiation and maturation of hESC-RPE potentially influencing the success of cell integrations and survival after cell transplantation.
authors.affiliation: 1 Institute of Biomedical Technology, University of Tampere , Tampere, Finland .
authors.forename: Anni
authors.initials: A
authors.lastname: Sorkio
authors.forename: Heidi
authors.initials: H
authors.lastname: Hongisto
authors.forename: Kai
authors.initials: K
authors.lastname: Kaarniranta
authors.forename: Hannu
authors.initials: H
authors.lastname: Uusitalo
authors.forename: Kati
authors.initials: K
authors.lastname: Juuti-Uusitalo
authors.forename: Heli
authors.initials: H
authors.lastname: Skottman
mesh_terms: D015153:Blotting, Western; D002454:Cell Differentiation / Q000187:drug effects; D002460:Cell Line; D020099:Coated Materials, Biocompatible / Q000494:pharmacology*; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000187:drug effects / Q000648:ultrastructure; D004847:Epithelial Cells / Q000166:cytology* / Q000187:drug effects / Q000648:ultrastructure; D016326:Extracellular Matrix Proteins / Q000378:metabolism*; D006801:Humans; D010858:Pigmentation / Q000187:drug effects; D055213:Retinal Pigment Epithelium / Q000166:cytology* / Q000187:drug effects / Q000648:ultrastructure
chemical_list: D020099:Coated Materials, Biocompatible; D016326:Extracellular Matrix Proteins
references.citation: Stem Cells. 2009 Sep;27(9):2126-35
references.pmid: 19521979
references.citation: Exp Neurol. 2008 Dec;214(2):347-61
references.pmid: 18926821
references.citation: Mol Vis. 2011;17:2864-77
references.pmid: 22128234
references.citation: Restor Neurol Neurosci. 2010;28(4):589-603
references.pmid: 20714081
references.citation: Pathol Biol (Paris). 2012 Feb;60(1):15-9
references.pmid: 22265966
references.citation: Stem Cells Transl Med. 2013 May;2(5):384-93
references.pmid: 23599499
references.citation: J Cell Biol. 2012 Apr 30;197(3):351-60
references.pmid: 22547406
references.citation: Biomed Microdevices. 2012 Aug;14(4):659-67
references.pmid: 22391881
references.citation: Ophthalmologe. 2001 Dec;98(12):1181-5
references.pmid: 11799902
references.citation: Lancet. 2012 Feb 25;379(9817):713-20
references.pmid: 22281388
references.citation: In Vitro Cell Dev Biol Anim. 2010 Apr;46(3-4):206-9
references.pmid: 20177999
references.citation: Tissue Cell. 1983;15(6):993-1005
references.pmid: 6198782
references.citation: Cell Stem Cell. 2009 Oct 2;5(4):396-408
references.pmid: 19796620
references.citation: Invest Ophthalmol Vis Sci. 2011 Mar 10;52(3):1345-58
references.pmid: 21398292
references.citation: BMC Dev Biol. 2008 Sep 22;8:90
references.pmid: 18808690
references.citation: J Cell Physiol. 2012 Feb;227(2):457-66
references.pmid: 21520078
references.citation: Exp Eye Res. 1997 Nov;65(5):603-8
references.pmid: 9367640
references.citation: Mol Vis. 2011 Feb 22;17:558-75
references.pmid: 21364903
references.citation: Invest Ophthalmol Vis Sci. 2011 Mar 01;52(3):1573-85
references.pmid: 21087957
references.citation: Proc Natl Acad Sci U S A. 2002 Nov 12;99(23):14682-7
references.pmid: 12391305
references.citation: Cloning Stem Cells. 2004;6(3):217-45
references.pmid: 15671670
references.citation: Prog Retin Eye Res. 2010 Jan;29(1):1-18
references.pmid: 19747980
references.citation: Stem Cells Dev. 2012 Aug 10;21(12):2262-72
references.pmid: 22145677
references.citation: Stem Cells. 2013 Feb;31(2):408-14
references.pmid: 23132794
references.citation: Exp Eye Res. 1998 Jun;66(6):807-15
references.pmid: 9657913
references.citation: Curr Eye Res. 1987 Oct;6(10):1211-21
references.pmid: 3677781
references.citation: J Exp Med. 2002 Jan 7;195(1):99-111
references.pmid: 11781369
references.citation: Neurosci Lett. 2009 Jul 24;458(3):126-31
references.pmid: 19379795
references.citation: Curr Opin Cell Biol. 1990 Oct;2(5):864-74
references.pmid: 2083086
references.citation: Curr Eye Res. 1996 Jul;15(7):739-48
references.pmid: 8670782
references.citation: Invest Ophthalmol Vis Sci. 1997 May;38(6):1110-8
references.pmid: 9152230
references.citation: PLoS One. 2009 Dec 03;4(12):e8152
references.pmid: 19997644
references.citation: Tokai J Exp Clin Med. 1998 Jun;23(3):147-51
references.pmid: 9972542
references.citation: Ophthalmic Res. 2012;48(4):186-91
references.pmid: 22868580
references.citation: Mol Vis. 2009;15:283-95
references.pmid: 19204785
references.citation: Prog Retin Eye Res. 1999 Mar;18(2):167-90
references.pmid: 9932282
references.citation: Biomaterials. 2012 Nov;33(32):8047-54
references.pmid: 22892561
references.citation: Pigment Cell Melanoma Res. 2012 Nov;25(6):804-14
references.pmid: 22883111
references.citation: Exp Eye Res. 2008 Jun;86(6):957-65
references.pmid: 18472095
references.citation: Stem Cells. 2009 Oct;27(10):2427-34
references.pmid: 19658190
references.citation: Exp Eye Res. 2006 Aug;83(2):315-21
references.pmid: 16579986
references.citation: Exp Eye Res. 2009 Nov;89(5):638-47
references.pmid: 19580809
references.citation: Prog Retin Eye Res. 2001 Nov;20(6):705-32
references.pmid: 11587915
references.citation: Ann Biomed Eng. 2011 Dec;39(12):3055-69
references.pmid: 21904797
references.citation: J Cell Biochem. 1999 Jul 1;74(1):135-43
references.pmid: 10381270
references.citation: Trans Am Ophthalmol Soc. 2003;101:499-520
references.pmid: 14971591
references.citation: Science. 2009 Nov 27;326(5957):1216-9
references.pmid: 19965464
references.citation: Methods. 2001 Dec;25(4):402-8
references.pmid: 11846609
references.citation: Neuron. 2012 Jul 12;75(1):26-39
references.pmid: 22794258
references.citation: J Biomed Biotechnol. 2010;2010:190724
references.pmid: 20182540
references.citation: Ann Med. 2006;38(7):450-71
references.pmid: 17101537
references.citation: Microsc Res Tech. 1998 Sep 1;42(5):311-6
references.pmid: 9766426
references.citation: Curr Biol. 2004 Apr 20;14(8):711-7
references.pmid: 15084287
references.citation: Cell Regul. 1991 Nov;2(11):939-49
references.pmid: 1667092
references.citation: J Tissue Eng Regen Med. 2013 Aug;7(8):642-53
references.pmid: 22514096
references.citation: Biomaterials. 2009 Jan;30(3):287-94
references.pmid: 18929407
references.citation: Mol Med. 2012 Dec 06;18:1312-9
references.pmid: 22895806
references.citation: Invest Ophthalmol Vis Sci. 1986 Nov;27(11):1615-21
references.pmid: 3771143
references.citation: FASEB J. 1999 Mar;13(3):477-84
references.pmid: 10064614
references.citation: Am J Physiol Cell Physiol. 2004 Jun;286(6):C1213-28
references.pmid: 15151915
references.citation: Prog Biophys Mol Biol. 2003 Jan;81(1):1-44
references.pmid: 12475568
references.citation: Surv Ophthalmol. 2006 Mar-Apr;51(2):137-52
references.pmid: 16500214
references.citation: Stem Cell Res. 2011 Jan;6(1):1-12
references.pmid: 21075697
references.citation: Stem Cells. 2010 Mar 31;28(3):390-8
references.pmid: 19998373
references.citation: Prog Retin Eye Res. 2008 Nov;27(6):579-95
references.pmid: 18775790
references.citation: Prog Retin Eye Res. 2007 Sep;26(5):516-54
references.pmid: 17532250