title: An epidermal stem cells niche microenvironment created by engineered human amniotic membrane.
abstract: How to amplify epidermal stem cells (ESCs) rapidly is a challenging crux in skin tissue engineering research. The present study describes the preparation of 3D micronized (300-600 μm) amniotic membrane (mAM) by means of repeated freeze-thawing cycles to deplete cell components and homogenized with a macrohomogenizer in liquid nitrogen. This newly prepared mAM not only possessed the characteristics of a microcarrier but completely retained the basement membrane structure and abundant active substances such as NGF, HGF, KGF, bFGF, TGF-β1 and EGF in the AM matrix. The result showed that mAM combined with rotary cell culture system (RCCS) was able to amplify ESCs quickly. The relative cell viability at day 7 and 14 was significantly higher than that of the conventional 2D plate culture (326 ± 28% and 535 ± 47% versus 232 ± 21% and 307 ± 32%, P < 0.05). In addition, the new method was able to prevent cell differentiation effectively and retain the characteristics of stem cells. When mAM loaded with ESCs (ESC-mAM) was further transplanted to full-thickness skin defects in nude mice, ESCs survived well and formed a new epidermis. Four weeks after transplantation, papilla-like structures were observed, and collagen fibers were well and regularly arranged in the newly formed dermal layer. In conclusion, the mAM as a novel natural microcarrier possesses an intact basement membrane structure and bioactivities. It not only provides the microenvironment similar to the stem cell niche within the human body favorable for ex vivo culture and amplification of ESCs but can be used as the dermal scaffold in constructing a skin substitute containing ESCs for the repair of full-thickness skin defects.
authors.affiliation: Burns Institute of People's Liberation Army, Changhai Hospital, The Second Military Medical University, Shanghai 200433, People's Republic of China.
authors.forename: Shi-zhao
authors.initials: SZ
authors.lastname: Ji
authors.forename: Shi-chu
authors.initials: SC
authors.lastname: Xiao
authors.forename: Peng-fei
authors.initials: PF
authors.lastname: Luo
authors.forename: Guo-feng
authors.initials: GF
authors.lastname: Huang
authors.forename: Guang-yi
authors.initials: GY
authors.lastname: Wang
authors.forename: Shi-hui
authors.initials: SH
authors.lastname: Zhu
authors.forename: Min-juan
authors.initials: MJ
authors.lastname: Wu
authors.forename: Zhao-fan
authors.initials: ZF
authors.lastname: Xia
mesh_terms: D000650:Amnion / Q000166:cytology / Q000378:metabolism / Q000637:transplantation* / Q000648:ultrastructure; D000818:Animals; D015415:Biomarkers / Q000378:metabolism; D015153:Blotting, Western; D002454:Cell Differentiation; D049109:Cell Proliferation; D004247:DNA / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D000078404:Epidermal Cells; D004817:Epidermis / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry; D036341:Intercellular Signaling Peptides and Proteins / Q000378:metabolism; D008422:Materials Testing; D051379:Mice; D008819:Mice, Nude; D055153:Stem Cell Niche; D033581:Stem Cell Transplantation; D023822:Tissue Engineering / Q000379:methods*; D014945:Wound Healing
chemical_list: D015415:Biomarkers; D036341:Intercellular Signaling Peptides and Proteins; D004247:DNA