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The reconstruction of lung alveolus-like structure in collagen-matrigel/microcapsules scaffolds in vitro
Periodical: J Cell Mol Med ISBN: 1582-4934 (Electronic)
1582-1838 (Linking)
Number: 9
Date: 2010/10/30
Language: eng
Pages: 1878-86
Authors:Zhang, W. J., Lin, Q. X., Zhang, Y., Liu, C. T., Qiu, L. Y., Wang, H. B., Wang, Y. M., Duan, C. M., Liu, Z. Q., Zhou, J., Wang, C. Y.
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Abstract
This study attempted to use collagen-Matrigel as extracellular matrix (ECM) to supply cells with three-dimensional (3D) culture condition and employ alginate-poly-l-lysine-alginate (APA) microcapsules to control the formation of alveolus-like structure in vitro. We tested mice foetal pulmonary cells (FPCs) by immunohistochemistry after 2D culture. The alveolus-like structure was reconstructed by seeding FPCs in collagen-Matrigel mixed with APA microcapsules 1.5 ml. A self-made mould was used to keep the structure from contraction. Meanwhile, it provided static stretch to the structure. After 7, 14 and 21 days of culture, the alveolus-like structure was analysed histologically and immunohistochemically, or by scanning transmission electron microscopy (TEM). We also observed these structures under inverted phase contrast microscope. The expression of pro-surfactant protein C (SpC) was detected by reverse transcription-polymerase chain reaction (RT-PCR). We obtained fibroblasts, epithelial cells and alveolar type II (AE2) cells in FPCs. In the reconstructed structure, seeding cells surrounding the APA microcapsules constructed alveolus-like structures, the size of them ranges from 200 to 300 mum. In each reconstructed lung tissue sheet, microcapsules had integrity. Pan-cytokeratin, vimentin and SpC positive cells were observed in 7- and 14-day cultured structures. TEM showed lamellar bodies of AE2 cells in the reconstructed tissues whereas RT-PCR expressed SpC gene. Primary mice FPCs could form alveolus-like structures in collagen-Matrigel/APA microcapsules engineered scaffolds, which could maintain a differentiated state of AE2 cells.
This study attempted to use collagen-Matrigel as extracellular matrix (ECM) to supply cells with three-dimensional (3D) culture condition and employ alginate-poly-l-lysine-alginate (APA) microcapsules to control the formation of alveolus-like structure in vitro. We tested mice foetal pulmonary cells (FPCs) by immunohistochemistry after 2D culture. The alveolus-like structure was reconstructed by seeding FPCs in collagen-Matrigel mixed with APA microcapsules 1.5 ml. A self-made mould was used to keep the structure from contraction. Meanwhile, it provided static stretch to the structure. After 7, 14 and 21 days of culture, the alveolus-like structure was analysed histologically and immunohistochemically, or by scanning transmission electron microscopy (TEM). We also observed these structures under inverted phase contrast microscope. The expression of pro-surfactant protein C (SpC) was detected by reverse transcription-polymerase chain reaction (RT-PCR). We obtained fibroblasts, epithelial cells and alveolar type II (AE2) cells in FPCs. In the reconstructed structure, seeding cells surrounding the APA microcapsules constructed alveolus-like structures, the size of them ranges from 200 to 300 mum. In each reconstructed lung tissue sheet, microcapsules had integrity. Pan-cytokeratin, vimentin and SpC positive cells were observed in 7- and 14-day cultured structures. TEM showed lamellar bodies of AE2 cells in the reconstructed tissues whereas RT-PCR expressed SpC gene. Primary mice FPCs could form alveolus-like structures in collagen-Matrigel/APA microcapsules engineered scaffolds, which could maintain a differentiated state of AE2 cells.