Supplementary Material for: Systematic Investigation of SARS-CoV-2 Receptor Protein Distribution along Viral Entry Routes in Humans

<b><i>Background:</i></b> The novel beta-coronavirus, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), enters the human body via mucosal surfaces of the upper and/or lower respiratory tract. Viral entry into epithelial cells is mediated via angiotensin-converting enzyme 2 (ACE2) and auxiliary molecules, but the precise anatomic site of infection still remains unclear. <b><i>Methods:</i></b> Here, we systematically investigated the main SARS-CoV-2 receptor proteins ACE2 and transmembrane serine protease 2 (TMPRSS2), as well as 2 molecules potentially involved in viral entry, furin and CD147, in formalin-fixed, paraffin-embedded human tissues. Tissue microarrays incorporating a total of 879 tissue cores from conjunctival (<i>n</i> = 84), sinonasal (<i>n</i> = 95), and lung (bronchiolar/alveolar; <i>n</i> = 96) specimens were investigated for protein expression by immunohistochemistry. <b><i>Results:</i></b> ACE2 and TMPRSS2 were expressed in ciliated epithelial cells of the conjunctivae and sinonasal tissues, with highest expression levels observed in the apical cilia. In contrast, in the lung, the expression of those molecules in bronchiolar and alveolar epithelial cells was much rarer and only very focal when present. Furin and CD147 were more uniformly expressed in all tissues analyzed, including the lung. Interestingly, alveolar macrophages consistently expressed high levels of all 4 molecules investigated. <b><i>Conclusions:</i></b> Our study confirms and extends previous findings and contributes to a better understanding of potential SARS-CoV-2 infection sites along the human respiratory tract.

**背景**：新型β冠状病毒——严重急性呼吸综合征冠状病毒2型（SARS-CoV-2）可通过上、下呼吸道的黏膜表面侵入人体。病毒侵入上皮细胞的过程由血管紧张素转换酶2（ACE2）及辅助分子介导，但目前感染的确切解剖位点仍不明确。
**方法**：本研究针对福尔马林固定石蜡包埋的人体组织，系统探究了SARS-CoV-2的主要受体蛋白血管紧张素转换酶2（ACE2）与跨膜丝氨酸蛋白酶2（TMPRSS2），以及另外两种可能参与病毒侵入过程的分子——弗林蛋白酶（furin）与CD147。研究采用组织微阵列技术，对共计879份组织芯进行免疫组化检测，样本涵盖结膜组织（n=84）、鼻窦鼻腔组织（n=95）以及肺组织（细支气管/肺泡组织，n=96）。
**结果**：ACE2与TMPRSS2在结膜及鼻窦鼻腔组织的纤毛上皮细胞中表达，且在顶纤毛处的表达水平最高。与之相反，在肺组织中，上述分子在细支气管及肺泡上皮细胞中的表达极为罕见，仅在极少数情况下呈局灶性表达。弗林蛋白酶与CD147在所有受检组织中均呈更为均匀的表达，肺组织亦不例外。值得注意的是，肺泡巨噬细胞始终高表达本次研究涉及的全部4种分子。
**结论**：本研究验证并拓展了既往研究结果，有助于进一步阐明SARS-CoV-2在人体呼吸道中的潜在感染位点。