Ex vivo 2D and 3D HSV-2 infection model using human normal vaginal epithelial cells.
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.
State Key Laboratory of Virology, Institute of Medical Virology, Wuhan University School of Basic Medical Sciences, Wuhan, Hubei 430071, China.|Shenzhen R and D Center of State Key Laboratory of Virology, Wuhan University Shenzhen Institute, Shenzhen, Guangdong 518057, China.
Yaqi
Y
Zhu
State Key Laboratory of Virology, Institute of Medical Virology, Wuhan University School of Basic Medical Sciences, Wuhan, Hubei 430071, China.|Shenzhen R and D Center of State Key Laboratory of Virology, Wuhan University Shenzhen Institute, Shenzhen, Guangdong 518057, China.
Yan
Y
Yang
Xiangyang No.1 People's Hospital, Xiangyang, Hubei 441000, China.
Juanjuan
J
Guo
State Key Laboratory of Virology, Institute of Medical Virology, Wuhan University School of Basic Medical Sciences, Wuhan, Hubei 430071, China.|Shenzhen R and D Center of State Key Laboratory of Virology, Wuhan University Shenzhen Institute, Shenzhen, Guangdong 518057, China.
Ying
Y
Dai
Shenzhen R and D Center of State Key Laboratory of Virology, Wuhan University Shenzhen Institute, Shenzhen, Guangdong 518057, China.
Lina
L
Ye
Shenzhen R and D Center of State Key Laboratory of Virology, Wuhan University Shenzhen Institute, Shenzhen, Guangdong 518057, China.
Jianbin
J
Qiu
Shenzhen R and D Center of State Key Laboratory of Virology, Wuhan University Shenzhen Institute, Shenzhen, Guangdong 518057, China.
Zhihong
Z
Zeng
Shenzhen R and D Center of State Key Laboratory of Virology, Wuhan University Shenzhen Institute, Shenzhen, Guangdong 518057, China.
Xiaoting
X
Wu
Shenzhen R and D Center of State Key Laboratory of Virology, Wuhan University Shenzhen Institute, Shenzhen, Guangdong 518057, China.
Yanmei
Y
Xing
Peking University Shenzhen Hospital, Shenzhen, Guangdong 518036, China.
Xiang
X
Long
Hubei Maternal and Child Health Hospital, Wuhan, Hubei 430070, China.
Xufeng
X
Wu
Shenzhen Eye Hospital, Shenzhen, Guangdong 518040, China.
Lin
L
Ye
Peking University Shenzhen Hospital, Shenzhen, Guangdong 518036, China.
Shubin
S
Wang
State Key Laboratory of Virology, Institute of Medical Virology, Wuhan University School of Basic Medical Sciences, Wuhan, Hubei 430071, China.|Shenzhen R and D Center of State Key Laboratory of Virology, Wuhan University Shenzhen Institute, Shenzhen, Guangdong 518057, China.
Hui
H
Li
D015153:Blotting, Western; D018929:Cell Culture Techniques / Q000379:methods*; D018920:Coculture Techniques; D003094:Collagen; D004338:Drug Combinations; D004847:Epithelial Cells / Q000378:metabolism / Q000821:virology*; D004848:Epithelium / Q000378:metabolism / Q000821:virology; D005260:Female; D006367:HeLa Cells; D006558:Herpes Genitalis / Q000378:metabolism / Q000821:virology; D018258:Herpesvirus 2, Human / Q000502:physiology*; D054884:Host-Pathogen Interactions; D006801:Humans; D053547:Keratin-14 / Q000378:metabolism; D007797:Laminin; D018895:Microsatellite Repeats / Q000235:genetics; D008856:Microscopy, Fluorescence; D008954:Models, Biological; D016133:Polymerase Chain Reaction; D011509:Proteoglycans; D014157:Transcription Factors / Q000378:metabolism; D025521:Tumor Suppressor Proteins / Q000378:metabolism; D014621:Vagina / Q000166:cytology*
D004338:Drug Combinations; D053547:Keratin-14; D007797:Laminin; D011509:Proteoglycans; C115051:TP63 protein, human; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins; C059485:matrigel; D003094:Collagen
HSV-2 infection model; air-liquid interface (ALI) culture; ex vivo; human normal vaginal epithelial cells (HNVEC); three dimension (3D)