title: Photo-cross-linking of amniotic membranes for limbal epithelial cell cultivation.
abstract: In the present study, we developed photo-cross-linked amniotic membrane (AM) as a limbal stem cell niche. After ultraviolet (UV) irradiation for varying time periods, the biological tissues were studied by determinations of cross-linking structure, degradability, and nutrient permeation ability. Our results showed that the number of cross-links per unit mass of AM significantly increased with increasing illumination time from 5 to 50 min. However, the cross-link formation was inhibited by longer irradiation time (i.e., 150 min), probably due to the scission of tissue collagen chains through irradiation. The biological stability and matrix permeability of photo-cross-linked AM materials strongly depended on their cross-linking densities affected by the UV irradiation. In vitro biocompatibility studies including cell viability and pro-inflammatory gene expression analyses demonstrated that, irrespective of the irradiation time employed, the physically cross-linked biological tissues exhibited negligible cytotoxicity and similar interleukin-6 (IL-6) mRNA levels. The data clearly indicate that these AM matrices do not cause potential harm to the corneal epithelial cells. After the growth of limbal epithelial cells (LECs) on AM substrates, Western blot analyses were conducted to examine the expression of ABCG2. It was found that the ability of UV-irradiated AM to maintain the undifferentiated precursor cell phenotype was significantly enhanced with increasing extent of photo-cross-linking. In summary, the UV irradiation time may have a profound influence on the fabrication of photo-cross-linked AM matrices for LEC cultivation.
authors.affiliation: Institute of Biochemical and Biomedical Engineering, Chang Gung University, Taoyuan 33302, Taiwan, ROC; Biomedical Engineering Research Center, Chang Gung University, Taoyuan 33302, Taiwan, ROC; Molecular Medicine Research Center, Chang Gung University, Taoyuan 33302, Taiwan, ROC. Electronic address: jylai@mail.cgu.edu.tw.
authors.forename: Jui-Yang
authors.initials: JY
authors.lastname: Lai
mesh_terms: D000650:Amnion / Q000737:chemistry*; D001672:Biocompatible Materials / Q000737:chemistry; D018929:Cell Culture Techniques; D002460:Cell Line; D002470:Cell Survival; D003094:Collagen / Q000378:metabolism; D003432:Cross-Linking Reagents / Q000737:chemistry; D004847:Epithelial Cells / Q000166:cytology*; D006801:Humans; D015850:Interleukin-6 / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D055153:Stem Cell Niche; D014466:Ultraviolet Rays
chemical_list: D001672:Biocompatible Materials; D003432:Cross-Linking Reagents; D015850:Interleukin-6; D012333:RNA, Messenger; D003094:Collagen
keywords: Amniotic membrane; Biological tissue matrices; Limbal epithelial cells; Photo-cross-linking; Ultraviolet irradiation time