{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,5,14]],"date-time":"2026-05-14T05:16:09Z","timestamp":1778735769137,"version":"3.51.4"},"reference-count":25,"publisher":"Wiley","issue":"4","license":[{"start":{"date-parts":[[2017,6,6]],"date-time":"2017-06-06T00:00:00Z","timestamp":1496707200000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/onlinelibrary.wiley.com\/termsAndConditions#vor"}],"funder":[{"name":"Science and Technology Department of Henan (China)","award":["161100311100"],"award-info":[{"award-number":["161100311100"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Journal Cellular Physiology"],"published-print":{"date-parts":[[2018,4]]},"abstract":"<jats:sec><jats:label\/><jats:p>Normal human cervical epitheliums infected with HPVs gene in vitro are underlying molecular models to investigate physiological mechanisms of cervical epithelia and cervical disease. The current study aimed to establish a modified culture method for cervical epithelium and explore the feasibility of transfection with HPV\u201016 E6 gene mediated by lentivirus in primary cervical cells. The cells were dissociated enzymatically using Dispase II combined with 0.25% Trypsin\u20130.01% ethylenediamine tetracetic acid (EDTA) or Collagenase I. The detached effectiveness of Dispase II at different times was compared. Isolated cells were cultured and subcultured in modified keratinocyte serum\u2010free medium (K\u2010SFM) supplemented with 5% fetal bovine serum (FBS) or K\u2010SFM alone. Cytokeratin was used as the identification of cervical epitheliums. Proliferative capacity and growth curve of cervical epitheliums were evaluated by cell counting kit\u20108 (CCK\u20108). The cells at passages 3 were used to infect with HPV\u201016 E6 gene by lentivirus. The expression of green fluorescent protein (GFP) presented in the infected cells was observed via fluorescence microscopy and the levels of E6 mRNA were detected by quantitative real\u2010time PCR (qRT\u2010PCR). The results indicate that cervical epithelial cells can be isolated successfully by Dispase II combined with 0.25% Trypsin\u20130.01% EDTA method for 20\u2009hr and maintained for five or six passages in K\u2010SFM medium with 5% FBS. The present study proposed a brief and high\u2010yield protocol for isolation and culture of human cervical epitheliums. Moreover, an infected cell model with HPV\u201016 E6 gene mediated by lentivirus was established which can do duty for studies in vitro on the carcinogenic mechanism of HR\u2010HPVs.<\/jats:p><\/jats:sec>","DOI":"10.1002\/jcp.25978","type":"journal-article","created":{"date-parts":[[2017,5,2]],"date-time":"2017-05-02T16:14:02Z","timestamp":1493741642000},"page":"2773-2780","source":"Crossref","is-referenced-by-count":11,"title":["An improved method for primary culture of normal cervical epithelial cells and establishment of cell model in vitro with HPV\u201016 E6 gene by lentivirus"],"prefix":"10.1002","volume":"233","author":[{"given":"Tingting","family":"Fan","sequence":"first","affiliation":[{"name":"Department of Cytopathology The Third Affiliated Hospital of Zhengzhou University Zhengzhou China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Xiaofu","family":"Li","sequence":"additional","affiliation":[{"name":"Department of Cytopathology The Third Affiliated 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