title: A feeder-free, defined three-dimensional polyethylene glycol-based extracellular matrix niche for culture of human embryonic stem cells.
abstract: We report optimization of a serum- and feeder-free, three-dimensional (3D) niche created with a synthetic polyethylene glycol (PEG)-based extracellular matrix for self-renewal of human embryonic stem cells (hESCs). Three hESC lines (H9, H1 and Novo) were cultured in hydrogels of different mechanical properties, and cellular morphology and activity were compared to culture in feeder-free or feeder-containing two-dimensional (2D) niches. Significant effects of PEG concentration (5, 7.5, 10, 12.5 or 15%) and vinyl sulfone-functionalized PEG multiarm number (3, 4 or 8) on hESC morphology were detected in the H9 line. Cell growth was maximal with an 8 multiarm architecture of any PEG concentration, which yielded the highest expression of stemness-related genes. Alkaline phosphatase activity in cultured H9 cells was similar between the optimized feeder-free 3D and the feeder-containing 2D systems. However, increased expression of the KLF4, CDH1, TERT, SOX2, and UTF1 genes and expression of pluripotency-specific SSEA-4, Oct3/4, Nanog, Tra-1-60 and Tra-1-81 were detected in the 3D-cultured hESC clumps. H1 and Novo cell lines also expanded in the optimized 3D system, which maintain stemness properties. Although different proliferation activities were detected among three lines, the difference was decreased after the 3D culture. These results demonstrate that chemically defined, acellular niches created using PEG-based hydrogels have the potential to support hESC self-renewal. Modulation of 3D properties can create various models for cell transformation and differentiation.
authors: WCU Biomodulation Program, Seoul National University, Seoul, Republic of Korea.
authors: Mi
authors: M
authors: Jang
authors: Seung Tae
authors: ST
authors: Lee
authors: Jae Won
authors: JW
authors: Kim
authors: Ji Hye
authors: JH
authors: Yang
authors: Jung Ki
authors: JK
authors: Yoon
authors: Joo-Cheol
authors: JC
authors: Park
authors: Hyun-Mo
authors: HM
authors: Ryoo
authors: André J
authors: AJ
authors: van der Vlies
authors: Ji Yeon
authors: JY
authors: Ahn
authors: Jeffrey A
authors: JA
authors: Hubbell
authors: Yong Sang
authors: YS
authors: Song
authors: Gene
authors: G
authors: Lee
authors: Jeong Mook
authors: JM
authors: Lim
mesh_terms: D001696:Biomechanical Phenomena; D018929:Cell Culture Techniques; D002454:Cell Differentiation; D049109:Cell Proliferation; D002478:Cells, Cultured; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000378:metabolism; D005109:Extracellular Matrix / Q000737:chemistry*; D006801:Humans; D020136:Hydrogel, Polyethylene Glycol Dimethacrylate / Q000737:chemistry; D007621:Karyotyping; D000090062:Kruppel-Like Factor 4; D011092:Polyethylene Glycols / Q000737:chemistry*
chemical_list: C000715467:KLF4 protein, human; D000090062:Kruppel-Like Factor 4; D020136:Hydrogel, Polyethylene Glycol Dimethacrylate; D011092:Polyethylene Glycols