Ciliogenesis and Mucus Synthesis in Cultured Human Respiratory Epithelial Cells

Hanamure, Yutaka; Deguchi, Kiyotaka; Ohyama, Masaru

doi:10.1177/000348949410301111

Cited by 21 publications

(18 citation statements)

References 20 publications

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“…As shown in Figure 3, MUC5AC, 5B, and 8 mRNAs were expressed in both ALI and LCC culture of passaged HNE cells, which suggests that culture of HNE cells induced mucous cell differentiation. This is also consistent with our previous report that observed the mucin granules in TEM study (Yoo et al 2003) and with the recent reports on the differentiation process of nasal epithelial cells (Yoon et al 2003;Hanamaure, Deguchi, and Ohyama 1994;Usui et al 2000;Gray et al 1996). Moreover, mucin gene expression was more significant in ALI culture in 21 days than that of LCC in 21 days, which again confirmed the importance of retinoic acid in BEGM and ALI condition on the growth and differentiation of airway epithelial cells (Yoon et al 2000;Karrtinen et al 1993).…”

Section: Rt-pcrsupporting

confidence: 92%

Air-Liquid Interface Culture of Serially Passaged Human Nasal Epithelial Cell Monolayer forIn VitroDrug Transport Studies

et al. 2005

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The objective of this study was to establish a drug transport study using human nasal epithelial (HNE) cell monolayers cultured by the air-liquid interface (ALI) method using serum-free medium (BEGM:DME/F12, 50:50). The cells were developed and characterized in comparison to those that have been previously cultured by the liquid-covered culture (LCC) method. The epithelial cell monolayer cultured by the ALI method resulted in a significantly higher transepithelial electrical resistance value (3,453 +/- 302 ohm x cm(2)) that was maintained (>1,000 ohm x cm(2)) for up to 20 days compared with that cultured by the LCC method. Observation by scanning electron microscopy revealed mature cilia after 2 weeks in the ALI culture, while flatten unhealthy ciliated cells were observed in the LCC method. After 21 days, higher level of MUC5AC and 8 mRNA were expressed in ALI culture which confirmed the secretory differentiation of HNE monolayers in vitro. No significant difference in the permeability coefficients of a model hydrophilic marker ((14)C-mannitol) and a lipophilic drug (budesonide) was observed between the two conditions on day 7. The passage 2-3 of the HNE monolayer using ALI condition retained the morphology and differentiated features of normal epithelium. Thus it would be a suitable model for in vitro nasal drug delivery studies.

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Section: Rt-pcrsupporting

confidence: 92%

Air-Liquid Interface Culture of Serially Passaged Human Nasal Epithelial Cell Monolayer forIn VitroDrug Transport Studies

et al. 2005

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“…In contrast, the percentage of secretory cells that could be detected by anti-mucin antibody (H6C6) remained almost constant throughout the study. As our results show, mucous differentiation generally precedes ciliary differentiation in airway epithelial cell culture (13,15). However, the appearance of ciliated cells occurred : 10 days later than in the other airway epithelium (9,12,13).…”

Section: Discussionsupporting

confidence: 57%

Ciliary and Secretory Differentiation of Normal Human Middle Ear Epithelial Cells

Choi¹,

Kim²,

Lee³

et al. 2002

Acta Oto-Laryngologica

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Recent technical advances now permit the serial culture of normal human middle ear epithelial (NHMEE) cells. However, the ciliary differentiation of these cells has not been achieved. The purpose of this study was to establish a culture system in order to differentiate serially cultured NHMEE cells into ciliated cells. If ciliated cells developed, the percentages of ciliated cells and secretory cells were measured throughout the duration of culture. We also examined the levels of mucin and lysozyme secretion and their mRNAs in a time-dependent manner. Human middle ear mucosa with a normal appearance was harvested and serially cultured after enzymatic disaggregation. These cells were cultured in an air-liquid interface (ALI) culture system for 2, 7, 14, 21 and 28 days after confluence. Ciliogenesis usually began 16-18 days after confluence. The percentage of ciliated cells detected by means of immunohistochemical staining increased over time up to a maximum of 10.6% but the percentage of secretory cells remained stable at approximately 40% throughout the duration of culture. By Day 14 after confluence, the amounts of mucin and lysozyme secretion, as measured by dot-blotting analysis, had increased significantly and then remained stable. The expression levels of mucin gene 5B (MUC5B), MUC8 and lysozyme increased with the duration of culture. MUC8 in particular showed a dramatic increase on Day 28 after confluence. In contrast, the level of MUC5AC mRNA peaked on Day 14 after confluence, and then decreased. In conclusion, ciliary differentiation of NHMEE cells can be induced using an ALI culture system. Our study also suggests that secretory function develops earlier than ciliogenesis, and that the expressions of MUC5B and MUC8 mRNAs increase as a function of differentiation.

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“…Recently, Hanamure et al [8] demonstrated differentiation of human respiratory cells when cultured on acellular collagen gels, even though formation of a basement membrane formation was not shown. The cell culture features reported are confirmed by our observations that ciliated cells mostly disappeared within 3 weeks.…”

Section: Discussionmentioning

confidence: 99%

“…As reported by Jorissen et al [10], the cilia reappeared on human nasal epithelial cells when the monolayers of deciliated cells were resuspended to form aggregates. Since this is not useful for the fabrication of EMC as grafts, the floating culture system described by Hanamure et al [8], which also facilitates ciliogenesis, seems to be appropriate.…”

Section: Discussionmentioning

confidence: 99%

Living epithelial-mesenchymal compounds formed in vitro suitable for autografting

Ross

Wittmann

1997

Eur Arch Otorhinolaryngol

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Three different, human epithelial-mesenchymal compounds (EMC) were generated in vitro for prospective grafting in epithelial defects. All compounds consisted of a fibroblast-populated collagen lattice as a mesenchymal component seeded with different types of cultured epithelial cells isolated from biopsies of healthy skin, oral mucosa and respiratory mucosa. Maturation of the epithelial cells was enabled by the presence of a high calcium concentration (1.8 mM) when cultures were lifted to the air-liquid interface. Light and electron microscopy revealed moderate differentiation of the multilayered epithelium in all compounds as well as basement membrane development at the epithelial-mesenchymal junction after 2-3 weeks. A coherent, tissue-like consistency of the collagen lattice and the presence of a basement membrane preventing detachment of the epithelium permitted easy handling and even loose suturing of the compounds produced.

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Ciliogenesis and Mucus Synthesis in Cultured Human Respiratory Epithelial Cells

Cited by 21 publications

References 20 publications

Air-Liquid Interface Culture of Serially Passaged Human Nasal Epithelial Cell Monolayer forIn VitroDrug Transport Studies

Air-Liquid Interface Culture of Serially Passaged Human Nasal Epithelial Cell Monolayer forIn VitroDrug Transport Studies

Ciliary and Secretory Differentiation of Normal Human Middle Ear Epithelial Cells

Living epithelial-mesenchymal compounds formed in vitro suitable for autografting

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