Severe Acute Respiratory Syndrome Coronavirus 2 Neurotropism
Members of the coronavirus family share several features including a pleomorphic to a spherical shape, bilipid envelope, and large spike glycoproteins that inspired the Latin corona- (or crown) name.[2,4,5] Molecular analysis of SARS-CoV-2 revealed that the spike glycoproteins are essential for viral entry via the angiotensin-converting enzyme-2 (ACE2) receptor with associated proteases, including transmembrane serine protease 2 (TMPRSS2), for spike protein priming.[3,6] Cells which co-express ACE2 receptors and TMPRSS2 or cathepsin L are seen throughout the central nervous system (CNS), including neurons, glia, and endothelium.[6–8] A regional variability appears to be present with the highest level of ACE2 expression detected within the medulla.
Transneuronal and hematogenous routes are postulated mechanisms for SARS-CoV-2 neurotropism, based primarily on evidence from other human coronavirus infections[9,10] (Table 2). A highly debated route of neurotropism for SARS-CoV-2 is a direct infection of the olfactory receptor neurons.[9,10] This would hypothetically allow the virus to pass through the olfactory mucosa to reach the olfactory bulb and the CNS via retrograde axonal transport. Although there is evidence supporting this mechanism of infection for human coronavirus-OC43 intranasal-infected mice, conflicting data has arisen in the SARS-CoV-2 literature.[9–11] ACE2-TMPRSS2 expression has been reported in olfactory epithelial support cells, stem cells, and perivascular cells, however, single-cell RNA sequencing (which eliminates RNA contamination that can occur in bulk sequencing) did not identify SARS-CoV-2 RN in olfactory neurons in two separate studies.[11,13] Another study of 33 human autopsy specimens identified SARS-CoV-2 by RT-PCR in regionally defined tissue samples from olfactory mucosa, the olfactory bulb/tubercle and the medulla. However, in situ hybridization RNA studies, which are more cell-specific and less prone to contamination by neighboring cells, only identified SARS-CoV-2 in olfactory mucosa, not nervous system tissue. The authors did co-localize SARS-CoV-2 spike protein and neuronal markers in the olfactory mucosa using immunofluorescent confocal microscopy, however, widespread detection of SARS-CoV-2 in neuronal and glial cells of the brain parenchyma was not found. To date, there is limited evidence to support retrograde axonal olfactory transport necessary for direct CNS invasion.
One preclinical study suggested that a hematogenous spread of SARS-CoV-2 to neuronal tissue may be more likely than via an olfactory route. In this study, the authors found that 50% of the S1 spike protein of SARS-CoV-2 protein can cross the blood–brain barrier in a mouse model. Although an intranasal S1 injection was found to have brain uptake, it was at much lower levels than after intravenous administration and there was no evidence for retrograde axonal transport. The spike protein may cross the blood–brain barrier, but it remains unclear if cell entry of the spike protein is sufficient for virulence. Furthermore, the ability of the spike protein to cross the blood–brain barrier does not necessarily mean the entire viral particle has this capability. In a subsequent experiment, the authors were unable to replicate spike protein crossing the blood–brain barrier of endothelialized human pluripotent stem cells, suggesting that the findings in a murine model may not translate to humans.
Variable neuropathological data from the autopsy series of COVID-19 patients has demonstrated that identifying SARS-CoV-2 in neuronal tissue is uncommon, and in some cases may represent contamination. An autopsy series of 18 COVID-19 patients found low levels of RNA by RT-PCR and no evidence of SARS-CoV-2 in multiple neuronal sites analyzed by immunohistochemistry. The authors concluded that these low RNA counts represented either blood contamination or in situ virions. Another autopsy study of 43 COVID-19 nonsurvivors detected SARS-CoV-2 by single-cell RT-PCR analysis in the frontal lobe and/or medulla of 13 patients. However, the median RNA count was low (4700 copies/ml). The authors did detect SARS-CoV-2 spike and nucleocapsid protein using immunohistochemistry in the medulla and cranial nerves IX and X in 16 cases, but the presence of the SARS-CoV-2 did not co-localize with regions of neuropathological injury. Furthermore, the pattern of microglial activation and cytotoxic T cells infiltration noted within the brainstem, cerebellum, and olfactory bulb, has also been observed in non-COVID, critically ill patients with sepsis. A study of 18 COVID-19 brains that focused on microvascular injury found perivascular activated microglia, macrophage infiltrates and focal breakdown of the blood–brain barrier, but did not detect SARS-CoV-2 by RNA PCR, in situ hybridization or immunostaining in any tissue specimens. It is possible that the presence of SARS-CoV-2 detected in some studies may be explained by translocation across a disrupted blood–brain barrier. Because SARS-CoV-2 has not been co-localized with neuropathological injury across multiple studies, it is possible that SARS-CoV-2 may enter the brain as an innocent bystander following some other insult, rather than as a neurovirulent pathogen (Table 1).[14–29]
There are also no convincing studies identifying SARS-CoV-2 in human brain tissues using electron microscopy. An aforementioned study that found evidence of SARS-CoV-2 by RT-PCR and immunohistochemistry in neuronal tissue, did not identify the virus in any studied brain region using electron microscopy. Although one case report using electron microscopy found SARS-CoV-2 in neural and capillary endothelium of the frontal lobe of a COVID-19 patient with Parkinson's disease, these findings likely represented rough endoplasmic reticulum, rather than actual virion.
Data is also inconsistent with cerebrospinal fluid (CSF) analysis for SARS-CoV-2. A systematic review of 242 papers with 430 COVID-19 patients with neurologic events that underwent CSF analysis found that only 5% (16/303) of CSF SARS-CoV-2 PCR was reported positive.[30,31] The majority of studies that reported positive CSF PCR findings provided limited information regarding cycle threshold or failed to have concomitant naso/oropharyngeal or serum PCR or antibody evidence of SARS-CoV-2 infection. Studies that did report cycle thresholds had notably high thresholds, suggestive of contamination.[32,33] Although many studies have found SARS-CoV-2 antibodies in the CSF, only one peer-reviewed paper reports evidence of intrathecal antibody synthesis with an elevated immunoglobulin G index. Though some have postulated that SARS-CoV-2 infection may trigger autoimmune encephalitis, a study of eight patients was unable to detect autoantibodies in the CSF.
Overall, the discrepancies in the neuropathologic and CSF data call into question the neuroinvasive virulence of SARS-CoV-2. Indirect mechanisms of injury from systemic insults, such as hypoxemia, thrombosis or autoimmune responses may be responsible for the majority of neurological complications observed in COVID-19 patients.
Curr Opin Infect Dis. 2021;34(3):217-227. © 2021 Lippincott Williams & Wilkins