Endres Michaela, Leinhase Iris, Kaps Christian, Wentges Marek, Unger Manja, Olze Heidi, Ringe Jochen, Sittinger Michael, Rotter Nicole
Tissue Engineering Laboratory, University Medical Center Charité, Tucholskystr. 2, 10117 Berlin, Germany.
Eur Arch Otorhinolaryngol. 2005 May;262(5):390-6. doi: 10.1007/s00405-004-0842-9. Epub 2004 Nov 11.
The replacement of extensive tracheal defects resulting from intensive care medicine, trauma or large resections is still challenged by the re-epithelialization of an autologous or alloplastic trachea replacement. Therefore, this study was performed to investigate the potential of culture-expanded human respiratory epithelial cells (hREC) to regenerate a functional epithelium for tracheal tissue engineering. hREC from seven male nasal turbinates were freshly isolated, expanded on a collagenous matrix and subsequently cultured in high-density multi-layers to allow epithelial differentiation. The composition of epithelial cells in native respiratory epithelial tissue and culture-expanded hREC was analyzed by histological staining with Alcian blue and by immunohistochemical staining of cytokeratin pairs CK1/10 and CK5/14 with the antibodies 34betaE12 and CD44v6. Differentiation of culture-expanded hREC was further characterized by gene expression analysis of cytokeratins CK5, CK13, CK14 and CK18 using semi-quantitative real-time RT-PCR technique. Histological and immunohistochemical staining of culture-expanded hREC demonstrated basal cells covering the collagenous matrix. These cells formed a cellular multi-layer, which was composed of a basal layer of undifferentiated basal cells and an upper layer of cells differentiating along the squamous metaplasia and ciliated cell lineage. Lineage development of culture-expanded hREC was further documented by the induction of cytokeratins CK13 and CK18. Our results suggest that culture-expanded hREC have the potential to colonize collagen-coated biomaterials and to regenerate epithelial cell types for tracheal tissue engineering.
重症监护医学、创伤或大型切除术后导致的广泛气管缺损的修复，仍然面临着自体或异体气管替代物重新上皮化的挑战。因此，本研究旨在探讨培养扩增的人呼吸道上皮细胞（hREC）在气管组织工程中再生功能性上皮的潜力。从7名男性鼻甲中新鲜分离出hREC，在胶原基质上进行扩增，随后在高密度多层培养中诱导上皮分化。通过阿尔辛蓝组织学染色以及使用抗体34βE12和CD44v6对细胞角蛋白对CK1/10和CK5/14进行免疫组化染色，分析天然呼吸道上皮组织和培养扩增的hREC中上皮细胞的组成。使用半定量实时RT-PCR技术对细胞角蛋白CK5、CK13、CK14和CK18进行基因表达分析，进一步表征培养扩增的hREC的分化情况。培养扩增的hREC的组织学和免疫组化染色显示，基底细胞覆盖胶原基质。这些细胞形成细胞多层结构，由未分化的基底细胞的基底层和沿鳞状化生及纤毛细胞谱系分化的上层细胞组成。细胞角蛋白CK13和CK18的诱导进一步证明了培养扩增的hREC的谱系发育。我们的结果表明，培养扩增的hREC有潜力在胶原包被的生物材料上定植，并为气管组织工程再生上皮细胞类型。
