Pawan Kumar Singh1, Sneha Singh1, Dustin Farr2, Ashok Kumar3. 1. Department of Ophthalmology, Visual and Anatomical Sciences, Wayne State University, Detroit, MI, USA. 2. Department of Biochemistry, Microbiology and Immunology, Wayne State University, Detroit, MI, USA. 3. Department of Ophthalmology, Visual and Anatomical Sciences, Wayne State University, Detroit, MI, USA; Department of Biochemistry, Microbiology and Immunology, Wayne State University, Detroit, MI, USA. Electronic address: akuma@med.wayne.edu.
Abstract
PURPOSE: Zika virus (ZIKV) has emerged as an important human pathogen causing ocular complications. There have been reports of the shedding of ZIKV in human as well as animal tears. In this study, we investigated the infectivity of ZIKV in corneal epithelial cells and their antiviral immune response. METHODS: Primary human corneal epithelial cells (Pr. HCECs) and an immortalized cell line (HUCL) were infected with two different strains of ZIKV (PRVABC59 & BeH823339) or dengue virus (DENV, serotypes 1-4). Viral infectivity was assessed by immunostaining of viral antigen and plaque assay. qRT-PCR and immunoblot analyses were used to assess the expression of innate inflammatory and antiviral genes. Supplementation of recombinant ISG15 (rISG15) and gene silencing approaches were used to elucidate the role of ISG15 in corneal antiviral defense. RESULTS: Pr. HCECs, but not the HUCL cells, were permissive to both ZIKV strains and specifically to DENV3 infection. ZIKV induced the expression of viral recognition receptors (TLR3, RIG-I, &MDA5), and genes involved in inflammatory (CXCL10 & CCL5) and antiviral (IFNs, MX1, OAS2, ISG15) responses in Pr. HCECs. Furthermore, ZIKV infection caused Pr. HCECs cell death, as evidenced by TUNEL staining. Silencing of ISG15 increased ZIKV infectivity while supplementation with rISG15 reduced ZIKV infection by direct inactivation of ZIKV and inhibiting its entry. CONCLUSIONS: Our study demonstrates for the first time, that ZIKV can readily infect and replicate in Pr. HCECs. Therefore, ZIKV may persist in the cornea and pose the potential risk of transmission via corneal transplantation.
PURPOSE:Zika virus (ZIKV) has emerged as an important human pathogen causing ocular complications. There have been reports of the shedding of ZIKV in human as well as animal tears. In this study, we investigated the infectivity of ZIKV in corneal epithelial cells and their antiviral immune response. METHODS: Primary human corneal epithelial cells (Pr. HCECs) and an immortalized cell line (HUCL) were infected with two different strains of ZIKV (PRVABC59 & BeH823339) or dengue virus (DENV, serotypes 1-4). Viral infectivity was assessed by immunostaining of viral antigen and plaque assay. qRT-PCR and immunoblot analyses were used to assess the expression of innate inflammatory and antiviral genes. Supplementation of recombinant ISG15 (rISG15) and gene silencing approaches were used to elucidate the role of ISG15 in corneal antiviral defense. RESULTS: Pr. HCECs, but not the HUCL cells, were permissive to both ZIKV strains and specifically to DENV3 infection. ZIKV induced the expression of viral recognition receptors (TLR3, RIG-I, &MDA5), and genes involved in inflammatory (CXCL10 & CCL5) and antiviral (IFNs, MX1, OAS2, ISG15) responses in Pr. HCECs. Furthermore, ZIKV infection caused Pr. HCECs cell death, as evidenced by TUNEL staining. Silencing of ISG15 increased ZIKV infectivity while supplementation with rISG15 reduced ZIKV infection by direct inactivation of ZIKV and inhibiting its entry. CONCLUSIONS: Our study demonstrates for the first time, that ZIKV can readily infect and replicate in Pr. HCECs. Therefore, ZIKV may persist in the cornea and pose the potential risk of transmission via corneal transplantation.
Authors: Pawan Kumar Singh; John-Michael Guest; Mamta Kanwar; Joseph Boss; Nan Gao; Mark S Juzych; Gary W Abrams; Fu-Shin Yu; Ashok Kumar Journal: JCI Insight Date: 2017-02-23
Authors: C Gustavo De Moraes; Michele Pettito; Juan B Yepez; Anavaj Sakuntabhai; Etienne Simon-Loriere; Mussaret B Zaidi; Matthieu Prot; Claude Ruffie; Susan S Kim; Rando Allikmets; Joseph D Terwilliger; Joseph H Lee; Gladys E Maestre Journal: JMM Case Rep Date: 2018-03-14
Authors: Branden R Nelson; Justin A Roby; William B Dobyns; Lakshmi Rajagopal; Michael Gale; Kristina M Adams Waldorf Journal: Viral Immunol Date: 2019-11-05 Impact factor: 2.257