Induction of interferon-λ contributes to TLR3 and RIG-I activation-mediated inhibition of herpes simplex virus type 2 replication in human cervical epithelial cells.
OBJECTIVE
Is it possible to immunologically activate human cervical epithelial cells to produce antiviral factors that inhibit herpes simplex virus type 2 (HSV-2) replication?


RESULTS
Our results indicate that human cervical epithelial cells possess a functional TLR3/RIG-I signaling system, the activation of which can mount an Interferon-λ (IFN-λ)-mediated anti-HSV-2 response.


BACKGROUND
There is limited information about the role of cervical epithelial cells in genital innate immunity against HSV-2 infection.


METHODS
We examined the expression of toll-like receptors (TLRs) and retinoic acid-inducible I (RIG-I) in End1/E6E7 cells by real-time PCR. The IFN-λ induced by TLR3 and RIG-I activation of End1/E6E7 cells was also examined by real-time PCR and ELISA. HSV-2 infection of End1/E6E7 cells was evaluated by the real-time PCR detection of HSV-2 gD expression. The antibody to IL-10Rβ was used to determine whether IFN-λ contributes to TLR3/RIG-I mediated HSV-2 inhibition. Expression of interferon regulatory factor 3 (IRF3), IRF7, IFN-stimulated gene 56 (ISG56), 2'-5'-oligoadenylate synthetase I (OAS-1) and myxovirus resistance A (MxA) were determined by the real-time PCR and western blot. End1/E6E7 cells were transfected with shRNA to knockdown the IRF3, IRF7 or RIG-I expression. Student's t-test and post Newman-Keuls test were used to analyze stabilized differences in the immunological parameters above between TLR3/RIG-I-activated cells and control cells.


RESULTS
Human cervical epithelial cells expressed functional TLR3 and RIG-I, which could be activated by poly I:C and 5'ppp double-strand RNAs (5'ppp dsRNA), resulting in the induction of endogenous interferon lambda (IFN-λ). The induced IFN-λ contributed to TLR3/RIG-I-mediated inhibition of HSV-2 replication in human cervical epithelial cells, as an antibody to IL-10Rβ, an IFN-λ receptor subunit, could compromise TLR3/RIG-I-mediated inhibition of HSV-2. Further studies showed that TLR3/RIG-I signaling in the cervical epithelial cells by dsRNA induced the expression of the IFN-stimulated genes (ISGs), ISG56, 2'-5'-oligoadenylate synthetase I (OAS-1) and myxovirus resistance A (MxA), the key antiviral elements in the IFN signaling pathway. In addition, we observed that the topical treatment of genital mucosa with poly I:C could protect mice from genital HSV-2 infection.


CONCLUSIONS
Future prospective studies with primary cells and suitable animal models are needed in order to confirm these outcomes.


CONCLUSIONS
The findings provide direct and compelling evidence that there is intracellular expression and regulation of IFN-λ in human cervical epithelial cells, which may have a key role in the innate genital protection against viral infections.


METHODS
Not applicable.


BACKGROUND
This work was supported by the National Natural Science Foundation of China (81301428 to L.Z. and 81271334 to W.-Z.H.), the Fundamental Research Funds for the Central Universities (2042015kf0188 to L.Z.), the China Postdoctoral Science Foundation (2013M531745 to L.Z.), the Development Program of China ('973', 2012CB518900 to W.-Z.H.) from the Ministry of Science and Technology of the People's Republic of China, grants (DA12815 and DA022177 to W.-Z.H.) from the National Institute on Drug Abuse (NIDA) and the open project of Hubei Key Laboratory of Wudang Local Chinese Medicine Research (WDCM005 to M.S.). The authors declare no competing financial interests.
Animal Biosafety Level III Laboratory at the Center for Animal Experiment, Wuhan University, Wuhan 430071, China State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan 430071, China.
Li
L
Zhou
Department of Pathology and Laboratory Medicine, Temple University School of Medicine, Philadelphia, PA 19140, USA.
Jie-Liang
JL
Li
Department of Pathology and Laboratory Medicine, Temple University School of Medicine, Philadelphia, PA 19140, USA.
Yu
Y
Zhou
Animal Biosafety Level III Laboratory at the Center for Animal Experiment, Wuhan University, Wuhan 430071, China State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan 430071, China.
Jin-Biao
JB
Liu
Animal Biosafety Level III Laboratory at the Center for Animal Experiment, Wuhan University, Wuhan 430071, China State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan 430071, China.
Ke
K
Zhuang
Animal Biosafety Level III Laboratory at the Center for Animal Experiment, Wuhan University, Wuhan 430071, China.
Jian-Feng
JF
Gao
State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan 430071, China.
Shi
S
Liu
Animal Biosafety Level III Laboratory at the Center for Animal Experiment, Wuhan University, Wuhan 430071, China Present address: College of Basic Medical Sciences, Central Laboratory of the Fourth Affiliated Hospital in Xiangyang, Hubei Key Laboratory of Wudang Local Chinese Medicine Research, Hubei University of Medicine, Shiyan 44200, China.
Ming
M
Sang
State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan 430071, China.
Jian-Guo
JG
Wu
Animal Biosafety Level III Laboratory at the Center for Animal Experiment, Wuhan University, Wuhan 430071, China Department of Pathology and Laboratory Medicine, Temple University School of Medicine, Philadelphia, PA 19140, USA wenzheho@temple.edu.
Wen-Zhe
WZ
Ho
D000071457:DEAD Box Protein 58; D053487:DEAD-box RNA Helicases / Q000235:genetics / Q000378:metabolism*; D004847:Epithelial Cells / Q000378:metabolism / Q000821:virology*; D018258:Herpesvirus 2, Human / Q000235:genetics / Q000502:physiology*; D006801:Humans; D011070:Poly I-C / Q000235:genetics; D011446:Prospective Studies; D011971:Receptors, Immunologic; D015398:Signal Transduction / Q000235:genetics / Q000502:physiology; D051196:Toll-Like Receptor 3 / Q000235:genetics / Q000378:metabolism*; D014779:Virus Replication / Q000235:genetics* / Q000502:physiology
D011971:Receptors, Immunologic; C495344:TLR3 protein, human; D051196:Toll-Like Receptor 3; C452820:RIGI protein, human; D000071457:DEAD Box Protein 58; D053487:DEAD-box RNA Helicases; D011070:Poly I-C
Toll-like receptor 3; herpes simplex virus type 2; human cervical epithelial cells; interferon-stimulated genes; interferon-λ; retinoic acid-inducible gene I