Biomol Ther.  2017 May;25(3):296-307. 10.4062/biomolther.2017.043.

Establishment of Immortalized Primary Human Foreskin Keratinocytes and Their Application to Toxicity Assessment and Three Dimensional Skin Culture Construction

• Choi M
• Park M
• Lee S
• Lee JW
• Cho MC
• Noh M
• Lee C

• KMID: 2378808
• DOI: http://doi.org/10.4062/biomolther.2017.043

Abstract

In spite of frequent usage of primary human foreskin keratinocytes (HFKs) in the study of skin biology, senescence-induced blockage of in vitro proliferation has been a big hurdle for their effective utilization. In order to overcome this passage limitation, we first isolated ten HFK lines from circumcision patients and successfully immortalized four of them via a retroviral transduction of high-risk human papillomavirus (HPV) E6 and E7 oncogenes. We confirmed expression of a keratinocyte marker protein, keratin 14 and two viral oncoproteins in these immortalized HFKs. We also observed their robust responsiveness to various exogenous stimuli, which was evidenced by increased mRNA expression of epithelial differentiation markers and pro-inflammatory genes in response to three reactive chemicals. In addition, their applicability to cytotoxicity assessment turned out to be comparable to that of HaCaT cells. Finally, we confirmed their differentiation capacity by construction of well-stratified three dimensional skin cultures. These newly established immortalized HFKs will be valuable tools not only for generation of in vitro skin disease models but also for prediction of potential toxicities of various cosmetic chemicals.

Keyword

Human foreskin keratinocyte; Immortalization; Toxicity assessment; Three-dimensional skin culture

MeSH Terms

Antigens, Differentiation
Biology
Foreskin*
Humans*
In Vitro Techniques
Keratin-14
Keratinocytes*
Oncogene Proteins
Oncogenes
RNA, Messenger
Skin Diseases
Skin*
Zidovudine
Antigens, Differentiation
Keratin-14
Oncogene Proteins
RNA, Messenger
Zidovudine