Research Papers:
Ex vivo 2D and 3D HSV-2 infection model using human normal vaginal epithelial cells
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Abstract
Yaqi Zhu1,2,*, Yan Yang1,2,*, Juanjuan Guo3, Ying Dai1,2, Lina Ye2, Jianbin Qiu2, Zhihong Zeng2, Xiaoting Wu2, Yanmei Xing2, Xiang Long4, Xufeng Wu5, Lin Ye6, Shubin Wang4, Hui Li1,2
1State Key Laboratory of Virology, Institute of Medical Virology, Wuhan University School of Basic Medical Sciences, Wuhan, Hubei 430071, China
2Shenzhen R and D Center of State Key Laboratory of Virology, Wuhan University Shenzhen Institute, Shenzhen, Guangdong 518057, China
3Xiangyang No.1 People’s Hospital, Xiangyang, Hubei 441000, China
4Peking University Shenzhen Hospital, Shenzhen, Guangdong 518036
5Hubei Maternal and Child Health Hospital, Wuhan, Hubei 430070, China
6Shenzhen Eye Hospital, Shenzhen, Guangdong 518040, China
*These authors contributed equally to this work
Correspondence to:
Hui Li, email: [email protected]
Keywords: three dimension (3D), ex vivo, human normal vaginal epithelial cells (HNVEC), air-liquid interface (ALI) culture, HSV-2 infection model
Received: November 14, 2016 Accepted: January 13, 2017 Published: January 27, 2017
ABSTRACT
Herpes simplex virus type 2 (HSV-2) infects human genital mucosa and establishes life-long latent infection. It is unmet need to establish a human cell-based microphysiological system for virus biology and anti-viral drug discovery. One of barriers is lacking of culture system of normal epithelial cells in vitro over decades. In this study, we established human normal vaginal epithelial cell (HNVEC) culture using co-culture system. HNVEC cells were then propagated rapidly and stably in a defined culture condition. HNVEC cells exhibited a normal diploid karyotype and formed the well-defined and polarized spheres in matrigel three-dimension (3D) culture, while malignant cells (HeLa) formed disorganized and nonpolar solid spheres. HNVEC cells had a normal cellular response to DNA damage and had no transforming property using soft agar assays. HNVEC expressed epithelial marker cytokeratin 14 (CK14) and p63, but not cytokeratin 18 (CK18). Next, we reconstructed HNVEC-derived 3D vaginal epithelium using air-liquid interface (ALI) culture. This 3D vaginal epithelium has the basal and apical layers with expression of epithelial markers as its originated human vaginal tissue. Finally, we established an HSV-2 infection model based on the reconstructed 3D vaginal epithelium. After inoculation of HSV-2 (G strain) at apical layer of the reconstructed 3D vaginal epithelium, we observed obvious pathological effects gradually spreading from the apical layer to basal layer with expression of a viral protein. Thus, we established an ex vivo 2D and 3D HSV-2 infection model that can be used for HSV-2 virology and anti-viral drug discovery.
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