Abstract
The skin is susceptible to different injuries and diseases. One major obstacle in skin tissue engineering is how to develop functional three-dimensional (3D) substitute for damaged skin. Previous studies have proved a 3D dynamic simulated microgravity (SMG) culture system as a “stimulatory” environment for the proliferation and differentiation of stem cells. Here, we employed the NASA-approved rotary bioreactor to investigate the proliferation and differentiation of human epidermal stem cells (hEpSCs). hEpSCs were isolated from children foreskins and enriched by collecting epidermal stem cell colonies. Cytodex-3 micro-carriers and hEpSCs were co-cultured in the rotary bioreactor and 6-well dish for 15 days. The result showed that hEpSCs cultured in rotary bioreactor exhibited enhanced proliferation and viability surpassing those cultured in static conditions. Additionally, immunostaining analysis confirmed higher percentage of ki67 positive cells in rotary bioreactor compared with the static culture. In contrast, comparing with static culture, cells in the rotary bioreactor displayed a low expression of involucrin at day 10. Histological analysis revealed that cells cultured in rotary bioreactor aggregated on the micro-carriers and formed multilayer 3D epidermis structures. In conclusion, our research suggests that NASA-approved rotary bioreactor can support the proliferation of hEpSCs and provide a strategy to form multilayer epidermis structure.
Highlights
Skin is one of the major organs of the body and considered as the primary protective barrier against the external environment
Large colonies were formed and macroscopic after 12 days of growth with a colony formation efficiency (CFE) of 25% (Fig. 1M). To test whether these expanded cells still maintain characteristics of human epidermal stem cells (hEpSCs), we identified the expressions of hEpSCs markers after cell expanded
No significant differences of the expression level for b1-integrin, p63 or K14 were found between the isolated primary hEpSCs and the colony cells passageed three times from initial hEpSCs, and the majority of expanded hEpSCs were positive for these markers (Fig.2A)
Summary
Skin is one of the major organs of the body and considered as the primary protective barrier against the external environment. Tissue-engineered skin has been approved by the Food and Drug Administration in USA for use in wound healing, but the clinical results are far from satisfaction [5]. The application of this technique or method in therapy is presumably limited by the cultured epidermal and dermal autografts [6,7]. It is critical to improve the approaches to the isolation and culture of epidermal stem cells for their clinical utilization It remains a challenge for clinical application to improve the effect of wound healing and create a physiological three-dimensional (3D) tissue skin structure using EpSCs in-vitro before implantation
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