Unlike SARS-CoV-1 and MERS-CoV, infection with SARS-CoV-2, the viral pathogen responsible for COVID-19, is often associated with neurologic symptoms that range from mild to severe, yet increasing evidence argues the virus does not exhibit extensive neuroinvasive properties.
A new study by researchers from the University of California-Irvine along with scientists from the Scripps Research Institute-California has confirmed that the SARS-CoV-2 coronavirus is able to infect and replicate in the human host neurons.
Infections with SARS-CoV-2, the viral pathogen responsible for COVID-19, are often associated with neurologic symptoms that range from mild to severe, yet increasing evidence argues the virus does not exhibit extensive neuroinvasive properties.
The study team demonstrated that the SARS-CoV-2virus can infect and replicate in human iPSC-derived neurons and that infection shows limited anti-viral and inflammatory responses but increased activation of EIF2 signaling following infection as determined by RNA sequencing.
he study findings were published on a preprint server and are currently being peer reviewed. https://www.biorxiv.org/content/10.1101/2021.11.15.468761v1
The clinical spectrum of COVID-19 is complex, and numerous risk factors and comorbidities are considered important in affecting disease severity including age, obesity, chronic respiratory disease, and cardiovascular disease . In addition, neurological symptoms are common in COVID-19 patients, suggesting the virus can potentially infect and replicate in the central nervous system (CNS).
Indeed, encephalitis and meningitis have been reported in COVID-19 patients, and viral RNA and protein have been detected within the CSF of infected patients [2-4]. Additionally, human brain organoids are susceptible to SARS-CoV-2 infection [2, 5], yet demonstration of extensive CNS penetrance by SARS-CoV-2 has remained elusive.
It is imperative to develop pre-clinical animal models of COVID-19 that capture consistent and reproducible clinical and histologic readouts of many disease-associated symptoms following experimental infection with clinical isolates of SARS-CoV-2 .
Importantly, these models should be able to reliably evaluate interventional therapies to limit viral replication and mute immune-mediated pathology, as well as evaluate effectiveness of novel vaccines, all while remaining cost-effective. To date, the most common animal models employed to evaluate COVID-19 pathogenesis include mice, non-human primates (rhesus macaques, cynomolgus macaques and African green monkeys), Syrian hamsters, ferrets, and cats .
Human ACE2 (hACE2) transgenic mouse models have provided important insights into the pathogenesis of COVID-19. Perlman and colleagues  developed the K18-hACE2 mice, initially used as a mouse model of SARS-CoV-1, which has been successfully employed as a model of COVID-19 .
Intranasal inoculation of SARS-CoV-2 in K18-hACE2 mice results in a dose-dependent increase in weight loss and mortality with the lung being the major site of viral infection, while lower amounts of virus are detected in the heart, liver, spleen, kidney, small intestine, and colon . Examination of lungs revealed distribution of viral antigen associated with alveolar damage, interstitial lesions, edema, and inflammation.
Lung infection resulted in an increase in expression of interferons as well as inflammatory cytokines and chemokines associated with neutrophil, macrophage/monocyte, and T cell infiltration. Viral RNA was also detected within the sinonasal epithelium, and viral antigen was present in sustentacular cells associated with anosmia .
This may reflect the SARS-CoV-2 isolate being studied as well as the dose of virus being administered. However, in those animals in which virus penetrates the brain, there can be extensive spread of the virus throughout different anatomic regions accompanied by cell death , and these results are consistent with early studies examining SARS-CoV-1 infection of K18-hACE2 mice .
High-level of CNS infection in K18-hACE2 is accompanied by meningeal inflammation associated with immune cell infiltration into the brain parenchyma and microglia activation . Enhanced CNS penetrance and replication of SARS-CoV-2 within the CNS of K18-hACE2 is associated with increased mortality; although the mechanisms by which this occurs remain unclear.
The present study was undertaken to i) expand on earlier studies examining SARS-CoV-2 infection of human CNS resident cells, ii) evaluate the immune response that occurs in response to SARS-CoV-2 infection of the CNS of K18-hACE2 mice and iii) assess the contributions of microglia in host defense following CNS infection by SARS-CoV-2.
Previous studies have indicated neurons are susceptible to infection by SARS-CoV-2 (2); therefore, we infected human iPSC-derived neurons with SARS-CoV-2. Similar to earlier reports, SARS-CoV-2 was able to infect and replicate within neurons as determined by staining for nucleocapsid protein (Figures 1A and B). By 48h p.i., viral nucleocapsid protein had spread from the neuron cell body and extended down dendritic and axonal projections (Figures 1C and D).
Notably, we did not detect syncytia formation in neuron cultures at any time following infection with SARS-CoV-2, suggesting that virus may not spread via fusion with neighboring cells. RNA sequencing analysis revealed that expression of both anti-viral and inflammatory responses in infected neurons was limited relative to the genes within the heatmap at both 24h and 48h post-infection (Figure 1E).
We then evaluated pathways that may progressively change between 24h and 48h post-infection comparing the Transcripts per million (TPMs) as input for Ingenuity Pathway Analysis (IPA).
Figure 1F shows the top 12 IPA canonical pathways that are overrepresented. eIF2 signaling is the predominant pathway induced upon SARS-CoV-2 infection of neurons, followed by pathways associated with oxidative phosphorylation, eIF4, MTOR signaling and mitochondrial dysfunction, the latter with no prediction of activity. Notably, the Coronavirus Pathogenesis Pathway is also overrepresented, however is inhibited in response to neuronal infection by SARS-CoV-2 over the time frame tested with the majority of the genes represented encoding ribosomal proteins (Figure 1F).