Abstract
BackgroundOcular abnormalities present in microcephalic infants with presumed Zika virus (ZIKV) congenital disease includes focal pigment mottling of the retina, chorioretinal atrophy, optic nerve abnormalities, and lens dislocation. Target cells in the ocular compartment for ZIKV infectivity are unknown. The cellular response of ocular cells to ZIKV infection has not been described. Mechanisms for viral dissemination in the ocular compartment of ZIKV-infected infants and adults have not been reported. Here, we identify target cells for ZIKV infectivity in both the inner and outer blood-retinal barriers (IBRB and OBRB), describe the cytokine expression profile in the IBRB after ZIKV exposure, and propose a mechanism for viral dissemination in the retina.MethodsWe expose primary cellular components of the IBRB including human retinal microvascular endothelial cells, retinal pericytes, and Müller cells as well as retinal pigmented epithelial cells of the OBRB to the PRVABC56 strain of ZIKV. Viral infectivity was analyzed by microscopy, immunofluorescence, and reverse transcription polymerase chain reaction (RT-PCR and qRT-PCR). Angiogenic and proinflammatory cytokines were measured by Luminex assays.ResultsWe find by immunofluorescent staining using the Flavivirus 4G2 monoclonal antibody that retinal endothelial cells and pericytes of the IBRB and retinal pigmented epithelial cells of the OBRB are fully permissive for ZIKV infection but not Müller cells when compared to mock-infected controls. We confirmed ZIKV infectivity in retinal endothelial cells, retinal pericytes, and retinal pigmented epithelial cells by RT-PCR and qRT-PCR using ZIKV-specific oligonucleotide primers. Expression profiles by Luminex assays in retinal endothelial cells infected with ZIKV revealed a marginal increase in levels of beta-2 microglobulin (β2-m), granulocyte macrophage colony-stimulating factor (GMCSF), intercellular adhesion molecule 1 (ICAM-1), interleukin-6 (IL-6), monocyte chemotactic protein-1 (MCP1), and vascular cell adhesion molecule 1 (VCAM-1) and higher levels of regulated upon activation, normal T cell expressed and presumably secreted (RANTES) but lower levels of interleukin-4 (IL-4) compared to controls.ConclusionsRetinal endothelial cells, retinal pericytes, and retinal pigmented epithelial cells are fully permissive for ZIKV lytic replication and are primary target cells in the retinal barriers for infection. ZIKV infection of retinal endothelial cells and retinal pericytes induces significantly higher levels of RANTES that likely contributes to ocular inflammation.
Highlights
Ocular abnormalities present in microcephalic infants with presumed Zika virus (ZIKV) congenital disease includes focal pigment mottling of the retina, chorioretinal atrophy, optic nerve abnormalities, and lens dislocation
Retinal endothelial and retinal pericytes are permissive for ZIKV infectivity but not retinal Müller cells To identify target cells for ZIKV infection in the eye, we first examined cellular components of the retinal vascular unit that represents the Inner blood-retinal barrier (IBRB) and is a gateway to the retina
We show semiquantitative RT-Polymerase chain reaction (PCR) amplification of a 364-bp DNA fragment using ZIKV-specific primers, and no amplification using complementary DNA (cDNA) from total RNA obtained from retinal pigmented epithelial cells mock-infected or retinal pigmented epithelial cells exposed to heat-killed ZIKV (Fig. 5c)
Summary
Ocular abnormalities present in microcephalic infants with presumed Zika virus (ZIKV) congenital disease includes focal pigment mottling of the retina, chorioretinal atrophy, optic nerve abnormalities, and lens dislocation. The lesions included focal pigment mottling of the retina, chorioretinal atrophy, optic nerve abnormalities, bilateral iris coloboma (congenital fissure), and lens dislocation [10] These lesions are considered vision-threatening, and children should be screened as a process of differential diagnosis to rule out other causes such as West Nile virus infection, toxoplasmosis, cytomegalovirus, rubella, herpes simplex virus, and syphilis [13, 14]. We identify target cells in both the inner and outer blood-retinal barriers (IBRB and OBRB), describe the cytokine expression profile in retinal endothelial cells after ZIKV exposure, and propose a mechanism for viral dissemination in the retina
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