Purpose

SARS-CoV-2 RNA has been detected in ocular tissues, but their susceptibility to SARS-CoV-2 infection is unclear. Here, we tested whether SARS-CoV-2 can infect human conjunctival epithelial cells (hCECs) and induce innate immune response.

Methods

Conjunctival tissue from COVID-19 donors was used to detect SARS-CoV-2 spike and envelope proteins. Primary hCECs isolated from cadaver eyes were infected with the parental SARS-CoV-2 and its beta variant of concern (VOC). Viral genome copy number, expression of viral entry receptors, TLRs, interferons, innate immune response genes was determined by qPCR. Viral receptors were examined in hCECs and tissue sections by immunostaining. Spike protein was detected in the cell culture supernatant by dot blot.

Results

Spike and envelope proteins were found in conjunctiva from COVID-19 patients. SARS-CoV-2 infected hCECs showed high viral copy numbers at 24–72h post-infection; spike protein levels were the highest at 24hpi. Viral entry receptors ACE2, TMPRSS2, CD147, Axl, and NRP1 were detected in conjunctival tissue and hCECs. SARS-CoV-2 infection-induced receptor expression at early time points post-infection, but gene expression of most TLRs peaked at 48 or 72hpi. SARS-CoV-2 infected hCECs showed higher expression of genes regulating antiviral response, RIG-I, interferons (α, β, & λ), ISG15 & OAS2, cytokines (IL-6, IL1β, TNFα), and chemokines (CXCL10, CCL5). Compared to the parental strain, beta VOC induced increased viral copy number and innate response in hCECs.

Conclusions

Conjunctival epithelial cells are susceptible to SARS-CoV-2 infection. Beta VOC is more infectious than the parental strain and evokes a higher antiviral and inflammatory response.

中文翻译：

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SARS-CoV-2 及其变体感染人类结膜上皮细胞并诱导差异性抗病毒先天免疫反应

目的

眼组织中已检测到 SARS-CoV-2 RNA，但眼组织对 SARS-CoV-2 感染的易感性尚不清楚。在这里，我们测试了 SARS-CoV-2 是否可以感染人结膜上皮细胞 (hCEC) 并诱导先天免疫反应。

方法

来自 COVID-19 捐赠者的结膜组织用于检测 SARS-CoV-2 刺突蛋白和包膜蛋白。从尸体眼睛中分离出的原代 hCEC 感染了亲本 SARS-CoV-2 及其相关 β 变体 (VOC)。通过 qPCR 测定病毒基因组拷贝数、病毒进入受体、TLR、干扰素、先天免疫应答基因的表达。通过免疫染色检查 hCEC 和组织切片中的病毒受体。通过斑点印迹检测细胞培养上清液中的刺突蛋白。

结果

在 COVID-19 患者的结膜中发现了刺突蛋白和包膜蛋白。感染 SARS-CoV-2 的 hCEC 在感染后 24-72 小时显示出高病毒拷贝数；刺突蛋白水平在 24 hpi 时最高。在结膜组织和 hCEC 中检测到病毒进入受体 ACE2、TMPRSS2、CD147、Axl 和 NRP1。SARS-CoV-2 感染在感染后的早期时间点诱导受体表达，但大多数 TLR 的基因表达在感染后 48 或 72 小时达到峰值。SARS-CoV-2 感染的 hCEC 显示调节抗病毒反应、RIG-I、干扰素（α、β 和 λ）、ISG15和OAS2、细胞因子（IL-6、IL1β、TNFα）和趋化因子（CXCL10、覆铜板5）。与亲本毒株相比，β VOC 诱导 hCEC 中病毒拷贝数和先天反应增加。

结论

结膜上皮细胞易受 SARS-CoV-2 感染。Beta VOC 比亲本菌株更具传染性，并能引起更高的抗病毒和炎症反应。