The human brain vasculature shows a distinct expression pattern of SARS-CoV-2 entry factors

Moheb Ghobrial, Jason Charish, Shigeki Takada, Taufik Valiante, Philippe P. Monnier, Ivan Radovanovic, Gary D. Bader, Thomas Wälchli

Secret key to the brain vasculature: Can Cathepsin B open the door for SARS-CoV-2?

Selected by Kristina Kuhbandner


During the ongoing coronavirus disease 2019 (COVID-19) pandemic, so far more than 39 million people have been infected with the respiratory virus SARS-CoV-21. Next to the well characterized pulmonary symptoms, many patients suffer from neurological complications ranging from loss of smell and headache to encephalopathy and ischemic stroke2. Notably, similar manifestations were also described in neonates born to SARS-CoV-2 infected mothers 3.

Increasing evidence points towards an involvement of the brain and peripheral vasculature as SARS-CoV-2 can infect human blood vessels and cause endotheliitis in multiple organs, which might be responsible for multi-organ failure4. However, the underlying molecular mechanisms remain to be elucidated.

SARS-CoV-2 uses its spike protein on the surface to dock to angiotensin-converting enzyme 2 (ACE2), which is expressed in different organs such as lung, kidney, liver and the intestine; in the brain ACE2 is predominantly present in the vascular wall. To be able to fuse membranes and enter their host cells, the virus also requires further help of human proteases such as transmembrane protease serine 2 (TMPRSS2), cathepsin B and L (CTSB/L) and furin5. Interestingly, compared to the periphery, ACE2 and TMPRSS2 only show weak expression in the brain, suggesting the existence of alternative entry mechanisms6. In this preprint, Ghobrial et al. provide a detailed analysis of the expression pattern of SARS-CoV-2 entry-associated genes in human adult and fetal as well as mouse tissue with focus on CTSB and CTSL in the vasculature.


The authors analyzed a total of 42 human adult and fetal tissue types using several publicly available single-cell RNA sequencing (scRNA-seq) datasets from healthy individuals. The data were visualized with t-distributed stochastic neighbor embedding (t-SNE).

Key findings

1) Expression pattern of entry-associated genes is distinct in the human vasculature

ScRNA-Seq analysis revealed that CTSL was mainly present in the peripheral vasculature, while CTSB showed a higher expression in the brain vasculature. These results were corroborated by data from the Human Protein Atlas and by immunofluorescence staining of multiple tissues. Furthermore, a complementary expression pattern was observed in the adult and fetal mouse central nervous system.

2) SARS-CoV-19 entry factors are expressed in cells of the neurovascular unit

The neurovascular unit is composed of endothelial and perivascular cells, including pericytes, astrocytes, neurons and immune cells. In these cells, CTSB was one of the highest expressed SARS-CoV-2 entry-associated genes with most abundant presence in microglia and macrophages followed by endothelial cells.

3) CTSB and CTSL expression is conserved during brain vasculature development

The complex architecture of the brain vasculature is established during fetal and postnatal development, and neonatal COVID-19 was associated with neurological impairment. Comparison of adult and fetal brain endothelial cells showed a similar gene expression profile, suggesting the presence of a developmentally established gene expression pattern of SARS-CoV2-entry genes.

4) Endothelial CTSB expression correlates with multiple disease-related genes and pathways

Among the genes with highest association with CTSB were those linked to immune function and inflammation, angiogenesis and cell-extracellular matrix interactions, as well as glucose and fatty acid metabolism. Analogically, pathway analysis revealed inflammation, angiogenesis, viral-host interaction, vascular metabolism and blood brain barrier permeability as top regulated pathways in endothelial cells. Again, the results were comparable in the adult and fetal situation. Overall, the identified pathways downstream of CTSB might help to explain some of the neurovascular symptoms related to COVID-19.

Why I like this preprint

Neurological complications associated with COVID-19 are frequently reported, but existing data on the interaction of SARS-CoV-2 with the central nervous system are still often preliminary and divergent2. Given the central role of the (neuro-)vasculature in COVID-19, it is critical to create a better understanding of the underlying mechanisms which might predispose human brain endothelial cells for viral infection. In their preprint, Ghobrial et al. propose an alternative vasculature-specific entry route into the brain involving CTSB and provide a comprehensive single-cell atlas of SARS-CoV-2 related entry factors and interaction partners in the brain vasculature. Thus, this work also highlights the impressive capabilities of integrative bioinformatical analysis of multiple independent datasets. Although the results remain to be confirmed in functional assays, the established tool can be utilized in future studies in clinical COVID-19 samples and may contribute to the development of treatment strategies targeting (neuro-)vascular symptoms in COVID-19 patients.

Questions to the authors

  1. CTSB/L are known to be important for Ebola virus entry into host cells7. Are there any existing data on how factors associated with a higher risk for COVID-19 (e.g. diabetes, age…) influence viral entry via CTSB/L in Ebola patients?
  2. Given the overall low expression of ACE2 in human tissue, do you think it might be the limiting factor for viral entry?
  3. Microglia show the highest expression of CTSB in the neurovascular unit. However, there is evidence from experiments in hiPSC-derived microglia that these cells themselves are not infected8. Could you comment on this a little further?



  1. WHO Coronavirus Disease (COVID-19) Dashboard. Accessed October 18, 2020.
  2. Iadecola C, Anrather J, Kamel H. Effects of COVID-19 on the Nervous System. Cell. 2020;183(1):16-27.e1. doi:10.1016/j.cell.2020.08.028
  3. Vivanti AJ, Vauloup-Fellous C, Prevot S, et al. Transplacental transmission of SARS-CoV-2 infection. Nature Communications. 2020;11(1):3572. doi:10.1038/s41467-020-17436-6
  4. Varga Z, Flammer AJ, Steiger P, et al. Endothelial cell infection and endotheliitis in COVID-19. The Lancet. 2020;395(10234):1417-1418. doi:10.1016/S0140-6736(20)30937-5
  5. Shang J, Wan Y, Luo C, et al. Cell entry mechanisms of SARS-CoV-2. PNAS. 2020;117(21):11727-11734. doi:10.1073/pnas.2003138117
  6. Sungnak W, Huang N, Bécavin C, et al. SARS-CoV-2 entry factors are highly expressed in nasal epithelial cells together with innate immune genes. Nature Medicine. 2020;26(5):681-687. doi:10.1038/s41591-020-0868-6
  7. Chandran K, Sullivan NJ, Felbor U, Whelan SP, Cunningham JM. Endosomal Proteolysis of the Ebola Virus Glycoprotein Is Necessary for Infection. Science. 2005;308(5728):1643-1645. doi:10.1126/science.1110656
  8. Jacob F, Pather SR, Huang W-K, et al. Human Pluripotent Stem Cell-Derived Neural Cells and Brain Organoids Reveal SARS-CoV-2 Neurotropism Predominates in Choroid Plexus Epithelium. Cell Stem Cell. Published online September 21, 2020. doi:10.1016/j.stem.2020.09.016



Tags: brain vasculature, cathepsin b, covid-19, neurological complications

Posted on: 21 October 2020


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