Archives

  • 2018-07
  • 2018-10
  • 2018-11
  • 2019-04
  • 2019-05
  • 2019-06
  • 2019-07
  • 2019-08
  • 2019-09
  • 2019-10
  • 2019-11
  • 2019-12
  • 2020-01
  • 2020-02
  • 2020-03
  • 2020-04
  • 2020-05
  • 2020-06
  • 2020-07
  • 2020-08
  • 2020-09
  • 2020-10
  • 2020-11
  • 2020-12
  • 2021-01
  • 2021-02
  • 2021-03
  • 2021-04
  • 2021-05
  • 2021-06
  • 2021-07
  • 2021-08
  • 2021-09
  • 2021-10
  • 2021-11
  • 2021-12
  • 2022-01
  • 2022-02
  • 2022-03
  • 2022-04
  • 2022-05
  • 2022-06
  • 2022-07
  • 2022-08
  • 2022-09
  • 2022-10
  • 2022-11
  • 2022-12
  • 2023-01
  • 2023-02
  • 2023-03
  • 2023-04
  • 2023-05
  • 2023-06
  • 2023-08
  • 2023-09
  • 2023-10
  • 2023-11
  • 2023-12
  • 2024-01
  • 2024-02
  • 2024-03
  • 2024-04

    • The following are the supplementary data

    2018-10-20

    The following are the supplementary data related to this article.
    Acknowledgment This work was supported by the Ministry of Science and Technology 973 program of China (2011CB504402, 2012CB966802 and 2012CB966304), and the \"Strategic Priority Research Program\" of the Chinese Academy of Sciences, Grant No XDA01020401, the National Natural Science Foundation of China (81071901), Ministry of Science and Technology International Technology Cooperation Program (2012DFH30050), the National S&T Major Special Project on Major New Drug Innovation, Grant No. 2011ZX09102-010-01. This study was also supported by multi-year research grant, University of Macau (MYRG122-ICMS12-SHX) and the Spinal Cord Injury Foundation of the University of Hong Kong, and One Hundred Person Project of The Chinese Academy of Sciences to D Pei.
    Introduction Recent advances in developmental and stem cell biology have spurred the rapid progression of cell-based therapies that are becoming more efficient and attractive especially in clinical scenarios that were traditionally deemed as intractable. On the forefront of this exciting revolution is the field of ophthalmology, where corneal burns were treated by transplantation of ex vivo expanded limbal epithelial stem HZ-1157 (LESCs) as early as in 1997 (Pellegrini et al., 1997). The LESCs are tissue-specific unipotent stem cells that reside in the corneal limbus, and are capable of regenerating the corneal epithelium throughout the lifetime of an individual (Cotsarelis et al., 1989; Davanger and Evensen, 1971; Huang and Tseng, 1991; Lehrer et al., 1998; Mann, 1944; Nagasaki and Zhao, 2003; Shortt et al., 2007; Thoft and Friend, 1983). From their location in distinctive stem cell niches in the basal limbus (Dua, 2005; Shortt et al., 2007), the LESCs migrate superficially towards central cornea (Thoft and Friend, 1983). During this migration, the LESCs are believed to differentiate into transit amplifying cells (TACs) before maturing first into postmitotic cells (PMCs) and finally into terminally differentiated cells (TDCs). Depletion or dysfunction of corneal epithelial stem cells gives rise to the potentially blinding condition termed limbal stem cell deficiency (LSCD). Since the pioneering work by Pellegrini et al. (1997), the preferred way to treat patients suffering from LSCD has been by transplantation of ex vivo expanded LESCs, a procedure also known as cultured limbal epithelial transplantation (CLET). This protocol adopts a technique originating from the keratinocyte research (Rheinwald and Green, 1975a, 1975b) and is based on graft sheets using inactivated 3T3 cells as a supportive feeder layer. Many studies have attempted to optimize different aspects of the culture conditions in order to improve the outcome of the therapeutic intervention. The common denominator of the different paradigms remains the selective enhancement of LESC growth. Recently, systems have been devised that enable expansion of limbal epithelial cells (LECs) without the use of feeder cells, thereby diminishing the risk of xenotoxicity and facilitating the clinical translation (Lekhanont et al., 2009). It is interesting to note that relatively little effort has been invested into investigating the significance of atmospheric culture conditions, even though LESCs, migrating and differentiating from the niches to the ocular surface, experience steep gradient of oxygen (Kwan et al., 1972). That the oxygen concentration may have significant influence on the biology of LESCs is furthermore indirectly inferred from in vivo and in vitro observations with several other types of stem cells (Mohyeldin et al., 2010; Toussaint et al., 2011; Zachar et al., 2011). Hence, it is important to investigate whether the same also holds true for LESCs. To our knowledge, only two studies have examined the effect of oxygen on in vitro growth and differentiation of LESCs (Miyashita et al., 2007; O\'Callaghan et al., 2011). These reports provided incongruent conclusions regarding maintenance of LESCs in hypoxia, which was probably due to the use of cells from different species and technical issues regarding the control of culture atmosphere. The current investigation is the first attempt to comprehensively compare growth and maintenance of human LESCs in a classic and a serum-free system across a variety of physiological O2 concentrations. Emphasis was put on the stability of atmospheric conditions so that the cells were grown in hypoxia without the effect of deleterious reoxygenation.