caa a22 4500 605505837 CHVBK 20210128100624.0 cr unu---uuuuu 210128e20151101xx s 000 0 eng 10.1007/s00253-015-6794-8 doi (NATIONALLICENCE)springer-10.1007/s00253-015-6794-8 Establishment and characterization of a telomerase-immortalized canine bronchiolar epithelial cell line [Elektronische Daten] [Xing Xie, Maoda Pang, Shan Liang, Lei Yu, Yanbing Zhao, Ke Ma, Dildar Kalhoro, Chengping Lu, Yongjie Liu] Dogs are susceptible to infectious diseases that occur primarily in the respiratory tract. The airway epithelium acts as a first line of defense and is constantly exposed to microorganisms present in the environment. Respiratory epithelial cells have recently gained wide use as a cell model for studying the pathogenesis of human, murine or swine respiratory pathogen infections. However, studies of the pathogenic mechanisms of canine pathogens have been hindered by the lack of reliable respiratory cell lines. Here, we cultured primary canine bronchiolar epithelial cells (CBECs), whose characteristics were confirmed by their expression of the epithelial cell-specific marker cytokeratin 18, and have provided protocols for their isolation and ex vivo expansion. Further, we established immortalized CBECs containing the human telomerase reverse transcriptase (hTERT) gene via transfection of primary CBECs with the recombinant plasmid pEGFP-hTERT. Immortalized bronchiolar epithelial cells (hTERT-CBECs) retain the morphological and functional features of primary CBECs, as indicated by reverse transcriptase polymerase chain reaction, proliferation assays, karyotype analysis, telomerase activity assay, and Western blotting, which demonstrate that hTERT-CBECs have higher telomerase activity, an extended proliferative lifespan, and a diploid complement of chromosomes, even after Passage 50. Moreover, this cell line is not transformed, as evaluated using soft agar assays and tumorigenicity analysis in nude mice, and can therefore be safely used in future studies. The isolation and establishment of stable hTERT-CBECs is of great importance for use as an in vitro model for mechanistic studies of canine pathogenic infections. Springer-Verlag Berlin Heidelberg, 2015 Canine bronchiolar epithelial cells (CBECs) nationallicence hTERT-CBECs nationallicence Pathogenesis nationallicence Cell model nationallicence Xie Xing College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, China aut Pang Maoda College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, China aut Liang Shan College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, China aut Yu Lei College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, China aut Zhao Yanbing College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, China aut Ma Ke College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, China aut Kalhoro Dildar College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, China aut Lu Chengping College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, China aut Liu Yongjie College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, China aut Applied Microbiology and Biotechnology Springer Berlin Heidelberg 99/21(2015-11-01), 9135-9146 0175-7598 99:21<9135 2015 99 253 https://doi.org/10.1007/s00253-015-6794-8 text/html Onlinezugriff via DOI BK010053 XK010053 XK010000 Metadata rights reserved Springer special CC-BY-NC licence nationallicence 1 research-article jats NATIONALLICENCE NATIONALLICENCE NL-springer NATIONALLICENCE 856 40 https://doi.org/10.1007/s00253-015-6794-8 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Xie Xing College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, China aut NATIONALLICENCE 700 1- Pang Maoda College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, China aut NATIONALLICENCE 700 1- Liang Shan College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, China aut NATIONALLICENCE 700 1- Yu Lei College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, China aut NATIONALLICENCE 700 1- Zhao Yanbing College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, China aut NATIONALLICENCE 700 1- Ma Ke College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, China aut NATIONALLICENCE 700 1- Kalhoro Dildar College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, China aut NATIONALLICENCE 700 1- Lu Chengping College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, China aut NATIONALLICENCE 700 1- Liu Yongjie College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, China aut NATIONALLICENCE 773 0- Applied Microbiology and Biotechnology Springer Berlin Heidelberg 99/21(2015-11-01), 9135-9146 0175-7598 99:21<9135 2015 99 253