Department of Pathology, Faculty of Medicine, Universiti Kebangsaan Malaysia, Jalan Yaacob Latif, Bandar Tun Razak, Cheras, Kuala Lumpur, Malaysia.
Cytotherapy. 2013 Aug;15(8):1030-41. doi: 10.1016/j.jcyt.2013.05.003.
The aim of the present study was to evaluate the effects of air-liquid interface on the differentiation potential of human amnion epithelial cells (HAECs) to skin-like substitute in organotypic culture.
HAECs at passage 1-2 were seeded onto a fibrin layer populated with human amnion mesenchymal cells to form the organotypic cultures. The organotypic HAECs were then cultured for 7, 14 and 21 d in two types of culture system: the submerged culture and the air-liquid interface culture. Cell morphogenesis was examined under the light and electron microscopes (transmission and scanning) and analyzed by immunohistochemistry.
Organotypic HAECs formed a single layer epithelium after 3 wk in submerged as well as air-liquid interface cultures. Ultrastructurally, desmosomes were observed in organotypic HAECs cultured in the air-liquid interface but not in the submerged culture. The presence of desmosomes marked the onset of early epidermal differentiation. Organotypic HAECs were positive against anti-CK18 and anti-CK14 in both the submerged and the air-liquid interface cultures. The co-expression of CK14 and CK18 suggested that differentiation of HAECs into skin may follow the process of embryonic skin development. However, weak expression of CK14 was observed after 2 and 3 wk of culture in air-liquid interface. CK10, involucrin, type IV collagen and laminin-5 expression was absent in organotypic HAECs. This observation reflects the initial process of embryonic epidermal differentiation and stratification.
Results from the present study suggest that the air-liquid interface could stimulate early differentiation of organotypic HAECs to epidermal cells, with a potential use for skin regeneration.
本研究旨在评估气液界面对人羊膜上皮细胞（HAECs）在器官型培养中向皮肤样替代物分化潜能的影响。
第 1-2 代 HAECs 接种到纤维蛋白层上，该纤维蛋白层上种植有人羊膜间充质细胞，以形成器官型培养物。然后，将器官型 HAECs 在两种培养系统中培养 7、14 和 21 天：浸没培养和气液界面培养。在光镜和电子显微镜（透射和扫描）下检查细胞形态发生，并通过免疫组织化学进行分析。
在浸没和气液界面培养中，器官型 HAECs 在 3 周后形成单层上皮。超微结构观察到，在气液界面培养的器官型 HAECs 中存在桥粒，但在浸没培养中不存在。桥粒的存在标志着早期表皮分化的开始。器官型 HAECs 在浸没和气液界面培养中均对 CK18 和 CK14 呈阳性。CK14 和 CK18 的共表达表明 HAECs 的分化可能遵循胚胎皮肤发育的过程。然而，在气液界面培养 2 和 3 周后，观察到 CK14 的表达较弱。CK10、内披蛋白、IV 型胶原和层粘连蛋白-5 在器官型 HAECs 中均不表达。这一观察结果反映了胚胎表皮分化和分层的初始过程。
本研究结果表明，气液界面可刺激器官型 HAECs 早期向表皮细胞分化，有望用于皮肤再生。
