Zika virus infection of cellular components of the blood-retinal barriers: implications for viral associated congenital ocular disease.
Zika virus infection of cellular components of the blood-retinal barriers: implications for viral associated congenital ocular disease.
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DOI：
10.1186/s12974-017-0824-7
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发表时间：
2017-03-03
影响因子：
9.3
通讯作者：
Alcendor DJ
中科院分区：
文献类型：
--
作者：
Roach T;Alcendor DJ
患有寨卡病毒（ZIKV）先天性疾病的小头症婴儿中存在的眼部异常包括视网膜的局灶性色素斑点、脉络膜视网膜萎缩、视神经异常和透镜脱位。ZIKV感染性的眼隔室中的靶细胞是未知的。尚未描述眼细胞对ZIKV感染的细胞应答。ZIKV感染的婴儿和成人的眼部隔室中病毒传播的机制尚未报道。在这里，我们鉴定了内部和外部血液-视网膜屏障（IBRB和OBRB）中ZIKV感染性的靶细胞，描述了ZIKV暴露后IBRB中的细胞因子表达谱，并提出了视网膜中病毒传播的机制。我们将IBRB的原代细胞组分，包括人视网膜微血管内皮细胞、视网膜周细胞和Müller细胞以及OBRB的视网膜色素上皮细胞暴露于ZIKV的PRVABC 56株。通过显微镜、免疫荧光和逆转录聚合酶链反应（RT-PCR和qRT-PCR）分析病毒感染性。通过Luminex测定法测量血管生成和促炎细胞因子。我们通过使用黄病毒4G 2单克隆抗体的免疫荧光染色发现，当与模拟感染对照相比时，IBRB的视网膜内皮细胞和周细胞以及OBRB的视网膜色素上皮细胞完全允许ZIKV感染，但不允许Müller细胞。我们使用ZIKV特异性寡核苷酸引物通过RT-PCR和qRT-PCR证实了视网膜内皮细胞、视网膜周细胞和视网膜色素上皮细胞中的ZIKV感染性。在用ZIKV感染的视网膜内皮细胞中通过Luminex测定的表达谱揭示了β-2微球蛋白（β2-m）、粒细胞巨噬细胞集落刺激因子（GMCSF）、细胞间粘附分子1（ICAM-1）、白细胞介素-6（IL-6）、单核细胞趋化蛋白-1（MCP 1）、和血管细胞粘附分子1（VCAM-1），以及较高水平的活化后调节的正常T细胞表达和可能分泌的（RANTES），但较低水平的白细胞介素4（IL-4）。视网膜内皮细胞、视网膜周细胞和视网膜色素上皮细胞完全允许ZIKV裂解性复制，并且是视网膜感染屏障中的主要靶细胞。视网膜内皮细胞和视网膜周细胞的ZIKV感染诱导显著更高水平的RANTES，这可能有助于眼部炎症。
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. 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. We 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. We 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. Retinal 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.