Establishment of an immortalized human subglottic epithelial cell line

Abstract

Objective: Translational research into subglottic disease is restricted by the availability of primary human tissue originating from this subsite. Primary epithelial cells are also limited by their inability to survive beyond several divisions in culture outside of the body. Specific subglottic cell lines, useful for in vitro studies, have not yet been described. We therefore demonstrate what we believe to be the first immortalized subglottic epithelial cell line.Methods: Subglottic tissue was derived from a single adult patient's neoplasia-free human subglottic brushing specimen. Cells were immortalized using a lentiviral vector expressing simian virus 40 T antigen. Karyotyping was performed on the transformed cells using single nucleotide polymorphism array comparative genomic hybridization. Transformed cells were phenotypically characterized by light microscopy, immunohistochemistry, and electrophysiology studies.Results: The immortalized subglottic cell line (SG01) was able to divide successfully beyond 20 passages. Karyotyping demonstrated no significant genomic imbalance after immortalization. The cells demonstrated normal epithelial morphology and cytokeratin expression throughout. SG01 cells were also successfully cultured at air-liquid interface (ALI). At ALI cells demonstrated cilia, mucus production, and relevant ion channel expression.Conclusion: The novel SG01 subglottic epithelial cell line has been established. This cell line provides a unique resource for researchers to investigate subglottic diseases, such as subglottic stenosis.

Novel immortalized human vocal fold epithelial cell line: In vitro tool for mucosal biology

“…18,19 Immortalization of epithelial cells from various human tissues (subglottis, posterior commissure, and ventricles) that are in close proximity to and surround the true membranous VF have also been described. 20 In each of these aforementioned immortalizations, the type of human epithelial cells immortalized were pseudostratified ciliated epithelium which differs from the stratified squamous epithelium from the free edge of the true membranous VF. Herein we describe and characterize, to the best of our knowledge, the first generation of immortalized hVFE cells obtained from the luminal surface of the true VF.…”

“…In earlier reports, multiple human epithelial subsites have been immortalized by SV40 or HPV E6/E7, including ovarian, subglottic, esophageal, and corneal. 3,20,27,28 Furthermore, previous work report that human epithelial cells from airway, nasopharyngeal, and ovarian were immortalized with combination of oncogene expression and activation of telomerase. 4,29,30 Use of both virus oncogenes, such as HPV E6/7 and SV40 T antigen and expression of hTERT has produced immortal epithelial cell lines with abrogated cell cycle control and apoptosis machinery, with spontaneous malignant transformation during prolong culture.…”

RNA Sequencing Reveals Cancer‐Associated Changes in Laryngeal Cells Exposed to Non‐Acid Pepsin

“…Epithelial cell lines were recently developed from human posterior commissure and subglottis. 30,61 TVC HPV would provide a useful comparison to these lines for study of laryngeal subsite cancer proneness. As with the posterior commissure line developed in our Laryngoscope 131: January 2021 Samuels et al: RNA Seq of Pepsin Treated Laryngeal Cells laboratory, TVC HPV cells will be made available for research through a biological repository.…”

“…While fibroblasts play a role in promoting tumorigenesis, carcinoma arises from epithelial cells, hence epithelial cell models are best suited for study of laryngeal carcinogenesis. Epithelial cell lines were recently developed from human posterior commissure and subglottis 30,61 . TVC HPV would provide a useful comparison to these lines for study of laryngeal subsite cancer proneness.…”

Establishment of Immortalized Laryngeal Epithelial Cells Transfected with Bmi1

“…The first two immortalized laryngeal epithelial cell lines were established by the exogenous introduction of a lentivirus-encoding HPV E6/E7 12 or simian virus 40 (SV40) 13 . The cells immortalized by these methods have proven to be reliable due to their stable genetic background 14 , 15 .…”