TY - JOUR
T1 - Generation of a human airway epithelium derived basal cell line with multipotent differentiation capacity
AU - Walters, Matthew S.
AU - Gomi, Kazunori
AU - Ashbridge, Beth
AU - Moore, Malcolm A.S.
AU - Arbelaez, Vanessa
AU - Heldrich, Jonna
AU - Ding, Bi Sen
AU - Rafii, Shahin
AU - Staudt, Michelle R.
AU - Crystal, Ronald G.
N1 - Funding Information:
We thank B-G. Harvey, R.J. Kaner, A.E. Tilley and J. Yee-Levin for coordinating and help in obtaining the large airway epithelium samples for basal cell culture; N. Hackett and G. Wang for useful discussions and N. Mohamed and D.N. McCarthy for help in preparing this manuscript. These studies were supported, in part, by R01HL107882, P50 HL084936, UL1 TR000457, UL1 RR024143 and The Starr Foundation/Starr Cancer Consortium.
PY - 2013/12/3
Y1 - 2013/12/3
N2 - Background: As the multipotent progenitor population of the airway epithelium, human airway basal cells (BC) replenish the specialized differentiated cell populations of the mucociliated airway epithelium during physiological turnover and repair. Cultured primary BC divide a limited number of times before entering a state of replicative senescence, preventing the establishment of long-term replicating cultures of airway BC that maintain their original phenotype.Methods: To generate an immortalized human airway BC cell line, primary human airway BC obtained by brushing the airway epithelium of healthy nonsmokers were infected with a retrovirus expressing human telomerase (hTERT). The resulting immortalized cell line was then characterized under non-differentiating and differentiating air-liquid interface (ALI) culture conditions using ELISA, TaqMan quantitative PCR, Western analysis, and immunofluorescent and immunohistochemical staining analysis for cell type specific markers. In addition, the ability of the cell line to respond to environmental stimuli under differentiating ALI culture was assessed.Results: We successfully generated an immortalized human airway BC cell line termed BCi-NS1 via expression of hTERT. A single cell derived clone from the parental BCi-NS1 cells, BCi-NS1.1, retains characteristics of the original primary cells for over 40 passages and demonstrates a multipotent differentiation capacity into secretory (MUC5AC, MUC5B), goblet (TFF3), Clara (CC10) and ciliated (DNAI1, FOXJ1) cells on ALI culture. The cells can respond to external stimuli such as IL-13, resulting in alteration of the normal differentiation process.Conclusion: Development of immortalized human airway BC that retain multipotent differentiation capacity over long-term culture should be useful in understanding the biology of BC, the response of BC to environmental stress, and as a target for assessment of pharmacologic agents.
AB - Background: As the multipotent progenitor population of the airway epithelium, human airway basal cells (BC) replenish the specialized differentiated cell populations of the mucociliated airway epithelium during physiological turnover and repair. Cultured primary BC divide a limited number of times before entering a state of replicative senescence, preventing the establishment of long-term replicating cultures of airway BC that maintain their original phenotype.Methods: To generate an immortalized human airway BC cell line, primary human airway BC obtained by brushing the airway epithelium of healthy nonsmokers were infected with a retrovirus expressing human telomerase (hTERT). The resulting immortalized cell line was then characterized under non-differentiating and differentiating air-liquid interface (ALI) culture conditions using ELISA, TaqMan quantitative PCR, Western analysis, and immunofluorescent and immunohistochemical staining analysis for cell type specific markers. In addition, the ability of the cell line to respond to environmental stimuli under differentiating ALI culture was assessed.Results: We successfully generated an immortalized human airway BC cell line termed BCi-NS1 via expression of hTERT. A single cell derived clone from the parental BCi-NS1 cells, BCi-NS1.1, retains characteristics of the original primary cells for over 40 passages and demonstrates a multipotent differentiation capacity into secretory (MUC5AC, MUC5B), goblet (TFF3), Clara (CC10) and ciliated (DNAI1, FOXJ1) cells on ALI culture. The cells can respond to external stimuli such as IL-13, resulting in alteration of the normal differentiation process.Conclusion: Development of immortalized human airway BC that retain multipotent differentiation capacity over long-term culture should be useful in understanding the biology of BC, the response of BC to environmental stress, and as a target for assessment of pharmacologic agents.
KW - Airway
KW - Basal cell
KW - Differentiation
KW - Immortalized
KW - Progenitor
KW - hTERT
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U2 - 10.1186/1465-9921-14-135
DO - 10.1186/1465-9921-14-135
M3 - Article
C2 - 24298994
AN - SCOPUS:84888793869
SN - 1465-9921
VL - 14
JO - Respiratory Research
JF - Respiratory Research
IS - 1
M1 - 135
ER -
