SARS-CoV-2 receptor and entry genes are expressed by sustentacular cells in the human olfactory neuroepithelium
Open accessAbstract
Various reports indicate an association between COVID-19 and anosmia, suggesting an infection of the olfactory sensory epithelium, and thus a possible direct virus access to the brain. To test this hypothesis, we generated RNA-seq libraries from human olfactory neuroepithelia, in which we found substantial expression of the genes coding for the virus receptor angiotensin-converting enzyme-2 (ACE2), and for the virus internalization enhancer TMPRSS2. We analyzed a human olfactory single-cell RNA-seq dataset and determined that sustentacular cells, which maintain the integrity of olfactory sensory neurons, express ACE2 and TMPRSS2 . We then observed that the ACE2 protein was highly expressed in a subset of sustentacular cells in human and mouse olfactory tissues. Finally, we found ACE2 transcripts in specific brain cell types, both in mice and humans. Sustentacular cells thus represent a potential entry door for SARS-CoV-2 in a neuronal sensory system that is in direct connection with the brain.
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TL;DR
Human olfactory neuroepithelium expresses SARS-CoV-2 entry genes, with ACE2 and TMPRSS2 enriched in sustentacular cells rather than olfactory sensory neurons. ACE2 transcripts/protein were also found in specific brain cell types, suggesting potential routes for infection and anosmia.
Key findings
- Bulk RNA-seq of human nasal biopsies found mean ACE2 and TMPRSS2 expression in olfactory sensory neuroepithelium (4.6 and 76.1 TPM, respectively) and respiratory epithelium (7.1 and 63.7 TPM, respectively), indicating entry-gene expression in both tissues.
- In a public single-cell RNA-seq dataset of 28,726 cells from four adult patients, sustentacular cells and respiratory ciliated cells showed the highest coexpression of ACE2 and TMPRSS2; olfactory sensory neurons showed little or no expression. ACE2-positive sustentacular cells numbered 71 versus 3,011 ACE2-negative, and ACE2-positive respiratory ciliated cells numbered 19 versus 567 ACE2-negative.
- Immunohistochemistry confirmed ACE2 protein in a subset of sustentacular cells in mouse and human olfactory tissues; in mouse, ACE2 labeling was apical and restricted to dorsal, not ventral, olfactory epithelium. TMPRSS2 labeling was observed in many nasal epithelium constituents.
- In mouse brain datasets, Ace2 was limited mainly to pericytes/mural cells and not neurons; in human brain datasets, ACE2 was found in non-neuronal and neuronal types including astrocytes, oligodendrocytes, Purkinje neurons, and cortical layer 5 neurons, with some coexpressing TMPRSS2.
Methods
- Bulk RNA-seq libraries were generated from human olfactory and respiratory nasal biopsies collected via nasal endoscopic surgery from four adult patients, followed by differential expression analysis.
- Public single-cell RNA-seq data from human olfactory/nasal tissue (Durante et al., 2020; 28,726 cells, 26 clusters) were reanalyzed for ACE2, TMPRSS2, ERMN, and GSTA2 expression and coexpression.
- Immunohistochemistry was performed on mouse and human olfactory tissues and mouse olfactory bulb using anti-ACE2 and anti-TMPRSS2 antibodies with neuronal/sustentacular markers (TUJ1, OMP, ERMN); olfactory epithelium from two deceased SARS-CoV-2-infected individuals was also examined.
Limitations
- Stated: the olfactory sensory epithelium from two deceased SARS-CoV-2-infected individuals was highly damaged and largely detached from the basal lamina, hindering identification of viral particles in sustentacular cells; whether the damage was caused by the virus could not be determined from the few samples analyzed.
- Evident from the text: sample sizes are small (four adult patients for biopsies; two deceased infected individuals), and brain expression conclusions rely partly on public datasets with possible differences in sequencing depth, protocol, mRNA half-lives, or cell-type selection.
- Evident from the text: the study does not demonstrate functional SARS-CoV-2 infection of sustentacular cells or establish causality for anosmia; the authors note that inflammation or infection of other non-neuronal cell types cannot be excluded.
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Claims the paper makes
0 of 36 cited sources checked · 0 supported · 0 flagged
- Findingown data
In human nasal epithelium, TMPRSS2 labeling is observed in many constituents.
“Congruently with its pervasiveness in the single-cell transcriptomes, TMPRSS2 labeling was observed in many constituents of the nasal epithelium (Figures 4H, S4F, and S4G).”
Quote in the paperNot a substantial claim - Findingown data
In the Zeisel et al. (2018) mouse brain dataset, a very limited number of cell types express Ace2.
“In the first collection, published by Zeisel et al. (2018), which includes non-neuronal cells, we found a very limited number of cell types that expressed Ace2 (Figure 5A).”
Quote in the paperSubstantial claim0 of 1 sources checked- cites Zeisel et al. (2018)
Not checked in the free run.
- Findingown data
Human olfactory neuroepithelia show substantial expression of ACE2 and TMPRSS2 transcripts.
“we generated RNA-seq libraries from human olfactory neuroepithelia, in which we found substantial expression of the genes coding for the virus receptor angiotensin-converting enzyme-2 (ACE2) and for the virus internalization enhancer TMPRSS2.”
- Also stated in: § Expression of ACE2 and TMPRSS2 Human Sustentacular and Respiratory Ciliated Cells · hedged
“We found a mean of 4.6 and 76.1 TPMs in the sensory neuroepithelium samples for ACE2 and TMPRSS2, respectively (Figures 1F and 1G), indicating the presence in this tissue of cells that may express both genes, or of a mix of cells that express either TMPRSS2 or ACE2.”
Quote in the paperSubstantial claim- Rests on an assumption: The four adult patients from whom olfactory and respiratory biopsies were collected are representative of the general human population for ACE2/TMPRSS2 expression in these tissues.
- Also stated in: § Expression of ACE2 and TMPRSS2 Human Sustentacular and Respiratory Ciliated Cells · hedged
- Findingown data
ACE2 protein is strongly expressed in a subset of sustentacular cells in the mouse olfactory epithelium.
“In the olfactory cavity, a strong and defined labeling was observed in the respiratory epithelium in the nasal cavity and nasopharyngeal duct (Figures S3A–S3H), and in a subset of sustentacular cells lining the nasal cavity (Figures 3C–3G and S3B).”
Quote in the paperSubstantial claim - Findingown data
Olfactory sensory neurons show little or no expression of TMPRSS2 and ACE2.
“Olfactory sensory neurons showed little or no expression of TMPRSS2 and ACE2, respectively (Figure 2C).”
Quote in the paperSubstantial claim
- Findingown data
In a human olfactory single-cell RNA-seq dataset, sustentacular cells express ACE2 and TMPRSS2.
“We analyzed a human olfactory single-cell RNA-seq dataset and determined that sustentacular cells, which maintain the integrity of olfactory sensory neurons, express ACE2 and TMPRSS2.”
- Also stated in: § Expression of ACE2 and TMPRSS2 Human Sustentacular and Respiratory Ciliated Cells · plain
“Sustentacular cells and ciliated respiratory cells exhibited the highest levels of expression of both genes.”
Quote in the paperSubstantial claim0 of 1 sources checked- cites Durante et al. (2020)
Not checked in the free run.
- Rests on an assumption: The single-cell RNA-seq dataset from Durante et al. (2020) accurately reflects the cell types and gene expression of the human olfactory neuroepithelium.
- Also stated in: § Expression of ACE2 and TMPRSS2 Human Sustentacular and Respiratory Ciliated Cells · plain
- Findingown data
ACE2 protein is strongly labeled in capillary-associated cells throughout the layers of the mouse olfactory bulb.
“A strong labeling of capillary-associated cells was observed through the different layers of the olfactory bulb (Figures 5D–5H), in agreement with the single-cell RNA-seq observations.”
Quote in the paperNot a substantial claim - Findingown data
In the mouse brain datasets, none of the Ace2-expressing cell populations coexpress Tmprss2.
“None of these cell populations coexpressed Tmprss2.”
Quote in the paperSubstantial claim - Findingcitation
In the Lake et al. (2018) human brain single-nucleus RNA-seq dataset, neuronal and glial cell types, in particular Purkinje neurons and cerebellar astrocytes, express ACE2 or TMPRSS2.
“In the first dataset (Lake et al. (2018)), we found neuronal and glial cell types, in particular Purkinje neurons and cerebellar astrocytes, that expressed ACE2 or TMPRSS2 (Figures S6A–S6E).”
Quote in the paperSubstantial claim0 of 1 sources checked- cites Lake et al. (2018)
Not checked in the free run.
- Rests on an assumption: The human brain single-nucleus RNA-seq datasets are representative of the human brain.
- Findingown data
The paper's data showed substantial expression of ACE2 in the mouse brain.
“Although our data showed substantial expression of ACE2 in the mouse brain, they did not replicate the widespread expression of ACE2 in the central nervous system previously reported by others”
Quote in the paperNot a substantial claim - Findingown data
The paper's data did not replicate the widespread ACE2 expression in the mouse central nervous system previously reported.
“Although our data showed substantial expression of ACE2 in the mouse brain, they did not replicate the widespread expression of ACE2 in the central nervous system previously reported by others, which found ACE2 in the motor cortex, raphe, and nuclei involved in the regulation of cardiovascular function (Doobay et al., 2007).”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites Doobay et al., 2007
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- Findingown data
Populations containing olfactory horizontal cells, microvillar cells, and Bowman's glands also show transcription of both ACE2 and TMPRSS2, although at lower levels.
“Populations containing olfactory horizontal cells, microvillar cells, and Bowman's glands also showed transcription of both genes, although at lower levels.”
Quote in the paperNot a substantial claim - Findingcitation
In the Saunders et al. (2018) mouse brain dataset, mural cells are the main cell type expressing Ace2.
“These observations were confirmed by the second mouse dataset (Saunders et al., 2018), in which mural cells were the main cell type that expressed Ace2 (Figure 5C).”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites Saunders et al., 2018
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- Findingown data
ACE2 protein is strongly expressed in some sustentacular cells of the human olfactory sensory epithelium.
“In accordance with the single-cell data, a strong labeling of ACE2 was observed in some sustentacular cells of the sensory epithelium (Figures 4C–4G).”
Quote in the paperSubstantial claim- Rests on an assumption: The four adult patients from whom olfactory and respiratory biopsies were collected are representative of the general human population for ACE2/TMPRSS2 expression in these tissues.
- Findingown data
In the human olfactory neuroepithelium, ACE2 staining in sustentacular cells is mainly present in the somata.
“In the human olfactory neuroepithelium, a similar labeling of ACE2 was observed in sustentacular cells, although the staining was mainly present in the somata.”
Quote in the paperNot a substantial claim - Findingown data
Mean ACE2 expression in sensory neuroepithelium samples was 4.6 TPM.
“We found a mean of 4.6 and 76.1 TPMs in the sensory neuroepithelium samples for ACE2 and TMPRSS2, respectively (Figures 1F and 1G), indicating the presence in this tissue of cells that may express both genes, or of a mix of cells that express either TMPRSS2 or ACE2.”
Quote in the paperSubstantial claim - Findingown data
Mean ACE2 expression in respiratory epithelium samples was 7.1 TPM.
“We observed a mean of 7.1 and 63.7 TPMs in the respiratory epithelium samples for ACE2 and TMPRSS2, respectively (Figures 1F and 1G), reflecting the presence of ciliated cells, which represent targets of SARS-CoV-2.”
Quote in the paperNot a substantial claim - Findingown data
A subset of olfactory sustentacular cells in the human olfactory neuroepithelium expresses ACE2.
“we demonstrate here that a subset of olfactory sustentacular cells in the olfactory neuroepithelium, but not olfactory sensory neurons, expresses ACE2”
- Also stated in: § Discussion · boosted
“We first determined, using transcriptomic analyses of whole tissue and single cells from human olfactory epithelia, that a subset of sustentacular cells expresses ACE2.”
Quote in the paperSubstantial claim - Also stated in: § Discussion · boosted
- Findingown data
Mean TMPRSS2 expression in respiratory epithelium samples was 63.7 TPM.
“We observed a mean of 7.1 and 63.7 TPMs in the respiratory epithelium samples for ACE2 and TMPRSS2, respectively (Figures 1F and 1G), reflecting the presence of ciliated cells, which represent targets of SARS-CoV-2.”
Quote in the paperNot a substantial claim - Findingown data
Human ACE2-expressing sustentacular cells are found isolated or in small clusters, intermingled with ACE2-negative sustentacular cells or respiratory epithelium.
“These human ACE2-expressing sustentacular cells were found isolated or in small clusters, intermingled with sustentacular cells negative for ACE2, or even sometimes intermingled with respiratory epithelium”
Quote in the paperSubstantial claim - Findingown data
Olfactory sensory neurons in the human olfactory neuroepithelium do not express ACE2.
“we demonstrate here that a subset of olfactory sustentacular cells in the olfactory neuroepithelium, but not olfactory sensory neurons, expresses ACE2”
Quote in the paperSubstantial claim - Findingown data
ACE2 and TMPRSS2 are coexpressed in cells of the human brain.
“Coexpression of ACE2 and TMPRSS2 in the different subclasses (n = 19 subclasses).”
- Also stated in: § SARS-CoV-2 Receptor Expression in the Brain · plain
“ACE2 and TMPRSS2 coexpressing cells, highlighted in black on the t-SNE plot.”
Quote in the paperSubstantial claim - Also stated in: § SARS-CoV-2 Receptor Expression in the Brain · plain
- Findingown data
Olfactory-specific genes including olfactory receptor genes, CNGA2, ANO2, OMP, and ERMN were significantly enriched in olfactory samples compared to respiratory samples.
“As expected, olfactory-specific genes, including olfactory receptor genes, CNGA2 and ANO2, OMP, and ERMN (encoding critical elements of the olfactory transduction cascade, a specific marker of mature olfactory sensory neurons and a marker of sustentacular cells, respectively), were significantly enriched in olfactory samples (Figures 1C–1E).”
Quote in the paperNot a substantial claim - Findingown data
ACE2 is expressed in specific cell types of the human brain, including neuronal and non-neuronal cell types.
“We also found ACE2 expression in specific cell types of the brain, including neuronal and non-neuronal cell types.”
- Also stated in: § Discussion · plain
“We identified various cell types in the brain that express ACE2, although at relatively low levels in humans.”
Quote in the paperSubstantial claim - Also stated in: § Discussion · plain
- Findingown data
In the mouse, ACE2 expression in brain is detected in vascular endothelial cells and pericytes, but not in neurons.
“In the mouse, we detected ACE2 expression in brain vascular endothelial cells and pericytes, but not in neurons.”
Quote in the paperNot a substantial claim - Findingown data
In the mouse, ACE2 is expressed in the apical border of sustentacular cells.
“we observed (similarly to humans) a clear expression of ACE2 in the apical border of sustentacular cells”
Quote in the paperNot a substantial claim - Findingown data
In the human olfactory neuroepithelium, some sustentacular cells are positive and others negative for ACE2.
“Again, as observed in the mouse, some of these cells were positive and others were negative for ACE2.”
Quote in the paperNot a substantial claim - Findingown data
ACE2-expressing sustentacular cells coexpress TMPRSS2.
“These cells were found to coexpress TMPRSS2, a serine protease known to facilitate viral entry.”
Quote in the paperSubstantial claim- Rests on an assumption: TMPRSS2 is a serine protease that facilitates viral entry.
- Findingown data
Mean TMPRSS2 expression in sensory neuroepithelium samples was 76.1 TPM.
“We found a mean of 4.6 and 76.1 TPMs in the sensory neuroepithelium samples for ACE2 and TMPRSS2, respectively (Figures 1F and 1G), indicating the presence in this tissue of cells that may express both genes, or of a mix of cells that express either TMPRSS2 or ACE2.”
Quote in the paperSubstantial claim - Findingown data
The amounts of ACE2 transcripts and the intensity of immunohistological staining in sustentacular cells are in the same range as those observed in respiratory ciliated cells.
“the amounts of ACE2 transcripts as well as the intensity of the immunohistological staining we observed in sustentacular cells are in the same range as the ones observed in respiratory ciliated cells”
Quote in the paperSubstantial claim - Findingown data
In the mouse, ACE2 expression in sustentacular cells is non-homogeneous, present in the most dorsally located cells and absent from more ventrally located ones.
“This distribution was, however, non-homogenous, since ACE2 was present in the most dorsally located sustentacular cells, but was entirely absent from more ventrally located ones.”
Quote in the paperNot a substantial claim - Premisecitation
SARS-CoV has been observed in human brains.
“SARS-CoV (i.e., the virus responsible for the previous SARS epidemic in 2002), the genome of which is closely related to the one of SARS-CoV-2 and whose effects on human tissues appear similar to those observed with SARS-CoV-2, has been observed in human brains (Ding et al., 2004; Gu et al., 2005; Xu et al., 2005).”
Quote in the paperNot a substantial claim0 of 3 sources checked- cites Ding et al., 2004
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- cites Gu et al., 2005
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- cites Xu et al., 2005
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- Interpretationown data
Whether the damage to the olfactory sensory epithelium in deceased SARS-CoV-2-infected individuals was caused by the virus cannot be determined based on the few samples analyzed.
“Whether this damage was caused by the virus cannot be determined based on the few samples analyzed.”
Quote in the paperNot a substantial claim - Interpretationown data
ACE2 and TMPRSS2 are expressed in a number of cell types in the human brain.
“ACE2 and TMPRSS2 are therefore expressed in a number of cell types in the human brain.”
Quote in the paperNot a substantial claim- Rests on an assumption: The human brain single-nucleus RNA-seq datasets are representative of the human brain.
- Interpretationown data
Sustentacular cells represent a potential entry door for SARS-CoV-2 in a neuronal sensory system directly connected with the brain.
“Sustentacular cells thus represent a potential entry door for SARS-CoV-2 in a neuronal sensory system that is in direct connection with the brain.”
Quote in the paperSubstantial claim - Premisecitation
Quantification of gene transcripts at single cell level often leads to inaccurate predictions, up to complete discordance between transcript and protein levels.
“Quantification of gene transcripts at the single cell level, a commonly used proxy of protein abundance, often leads to very inaccurate predictions, up to a complete discordance between real transcript and protein levels (Bauernfeind and Babbitt, 2017; Liu et al., 2016).”
Quote in the paperSubstantial claim0 of 2 sources checked- cites Bauernfeind and Babbitt, 2017
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- cites Liu et al., 2016
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- Interpretationown data
Sustentacular cells play in the same league as respiratory ciliated cells in terms of SARS-CoV-2 receptors.
“suggesting at least that sustentacular cells play in the same league in terms of SARS-CoV-2 receptors.”
Quote in the paperSubstantial claim - Study design and conductown data
The ACE2 antibodies were validated by labeling colon and kidney tissues known to contain ACE2-expressing cell types.
“These antibodies were first evaluated to label colon and kidney tissues, which contain previously described ACE2-expressing cell types (Figures S1A–S1J).”
Quote in the paperSubstantial claim - Premisecitation
SARS-CoV-2 virions have been found in the olfactory bulb.
“since SARS-CoV-2 virions have been found in the olfactory bulb (Nampoothiri et al., 2020).”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites Nampoothiri et al., 2020
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- Premisecitation
Brain lesions were observed in a transgenic mouse model expressing human ACE2 in the nose and infected intranasally with SARS-CoV.
“brain lesions were observed in a transgenic mouse model expressing the human ACE2 in the nose and infected intranasally with SARS-CoV (Netland et al., 2008).”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites Netland et al., 2008
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- Study design and conductown data
The study could not determine whether the SARS-CoV-2 virus is present in sustentacular cells of infected patients.
“we could not determine whether the SARS-CoV-2 virus is indeed present in sustentacular cells of infected patients.”
Quote in the paperNot a substantial claim - Premisecitation
SARS-CoV-2 virions have been observed in the human olfactory bulb.
“SARS-CoV-2 virions have in fact been observed in the human olfactory bulb (Nampoothiri et al., 2020).”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites Nampoothiri et al., 2020
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- Interpretationreasoning
The link between the viral molecular entry tools expressed by olfactory sustentacular cells and SARS-CoV-2-induced chemosensory alteration appears quite credible.
“the link between the viral molecular entry tools expressed by olfactory sustentacular cells and the SARS-CoV-2-induced chemosensory alteration appears quite credible.”
Quote in the paperNot a substantial claim- Rests on an assumption: ACE2 is the main player in the binding and entry of SARS-CoV-2 into human cells.
- Rests on an assumption: TMPRSS2 is a serine protease that facilitates viral entry.
- Interpretationown data
The data point to potential targets for SARS-CoV-2 infection in brain cells.
“these data point to potential targets for SARS-CoV-2 infection in brain cells.”
Quote in the paperSubstantial claim - Premisecitation
SARS-CoV-2 infection of golden hamsters via the nasal route leads to massive damage of the olfactory neuroepithelium.
“SARS-CoV-2 infection of golden hamsters via the nasal route leads to massive damage of the olfactory neuroepithelium (Bryche et al., 2020).”
Quote in the paperSubstantial claim0 of 1 sources checked- cites Bryche et al., 2020
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- Interpretationreasoning
The hijack of ACE2 by SARS-CoV-2 may by itself perturb the functioning of the olfactory sensor.
“the hijack of this receptor by SARS-CoV-2 may by itself perturb the functioning of the olfactory sensor.”
Quote in the paperSubstantial claim- Rests on an assumption: ACE2 is the main player in the binding and entry of SARS-CoV-2 into human cells.
- Interpretationreasoning
Perturbation of the horizontal basal cell stem cell pool is unlikely to represent an immediate disturbance to the olfactory system.
“a perturbation of the stem cell pool constituted by the horizontal basal cells appears unlikely to represent an immediate disturbance to the system.”
Quote in the paperSubstantial claim- Rests on an assumption: Neuronal renewal in the main olfactory epithelium is slow (a few months).
- Rests on an assumption: SARS-CoV-2-triggered anosmia develops rapidly.
- Study design and conductown data
Human olfactory neuroepithelium can be discriminated from respiratory epithelium by TUJ1-positive olfactory sensory neurons, ERMN-positive sustentacular cells, and absence of Alcian blue-labeled goblet cells.
“Similarly to what is observed in the mouse, the human neuroepithelium can be discriminated from the respiratory epithelium by the presence of TUJ1-positive olfactory sensory neurons (Figure 4A), ERMN-positive sustentacular cells along the apical border of the tissue (Figures 4A and S4C–S4E), as well as the absence of Alcian blue-labeled goblet cells (Figure 4B).”
Quote in the paperSubstantial claim - Premisecitation
Contact of olfactory mucosa with various drugs to which sustentacular cells are very sensitive leads to transient anosmia.
“Indeed, contact of the olfactory mucosa with various drugs (such as 3-methylindole [Miller and O'Bryan, 2003], the anti-thyroid drug methimazole [Bergstrom et al., 2003], or nickel sulfate [Jia et al., 2010]) to which sustentacular cells are very sensitive leads to transient anosmia.”
Quote in the paperNot a substantial claim0 of 3 sources checked- cites Miller and O'Bryan, 2003
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- cites Bergstrom et al., 2003
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- cites Jia et al., 2010
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- Premisecitation
SARS-CoV-2 infection is associated with olfactory dysfunction.
“the link between SARS-CoV-2 infection and olfactory dysfunction is today clearly established (Heidari et al., 2020; Hopkins et al., 2020; Luers et al., 2020; Moein et al., 2020; Spinato et al., 2020; Xydakis et al., 2020).”
Quote in the paperSubstantial claim0 of 3 sources checked- cites Heidari et al., 2020
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- cites Hopkins et al., 2020
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- cites Luers et al., 2020
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- May combine two claims. 15
- Classified as probably not a substantial claim. 12
May combine two claims.
Findingown dataIn the Allen Brain Map human brain dataset, ACE2 is expressed at relatively limited levels in various non-neuronal and neuronal types, in particular astrocytes, oligodendrocytes, and cortical layer 5 neurons, a portion of which coexpress TMPRSS2.
“In the second dataset, publicly available in the Allen Brain Map cell types database, we found expression of ACE2 (although at relatively limited levels) in various non-neuronal and neuronal types, in particular astrocytes, oligodendrocytes, and neurons from the cortical layer 5 (Figures 6A–6G), a portion of which coexpressed TMPRSS2 (Figures 6C–6E).”
Quote in the paperNot a substantial claim- Rests on an assumption: The human brain single-nucleus RNA-seq datasets are representative of the human brain.
May combine two claims.
Findingown dataThe olfactory sensory epithelium of two deceased SARS-CoV-2-infected individuals was highly damaged and largely detached from the basal lamina.
“The olfactory sensory epithelium of two deceased SARS-CoV-2-infected individuals was analyzed. It was found to be highly damaged and largely detached from the basal lamina (Figure S5), hindering the potential identification of viral particles in sustentacular cells.”
- Also stated in: § Limitations of the Study · plain
“the epithelium was found to be highly damaged and largely detached from the basal lamina”
Quote in the paperSubstantial claim- Also stated in: § Limitations of the Study · plain
May combine two claims.
Findingown dataThe mouse brain cells expressing Ace2 are related to pericytes and coexpress Rgs5.
“These cells are related to pericytes and coexpress the mural cell marker Rgs5 (Figure 5B).”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites Zeisel et al. (2018)
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Findingown dataIn the Zeisel et al. (2018) mouse brain dataset, Tmprss2 is expressed by even fewer cell types than Ace2, with Purkinje cells showing the highest expression.
“Tmprss2, whose transcripts were barely detected, was expressed by even fewer cell types (Figure 5A), with Purkinje cells displaying the highest expression level.”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites Zeisel et al. (2018)
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Findingown dataIn mouse olfactory epithelium, sustentacular cells are positive for TMPRSS2, and several other cell types show weak expression.
“In the mouse main olfactory epithelium, sustentacular cells were positive for TMPRSS2, and several other cell types showed weak expression (Figures 3N, S2A, S2B, S2E, and S2F).”
Quote in the paperNot a substantial claimMay combine two claims.
Findingown dataDifferent types of respiratory cells and olfactory sustentacular cells express ACE2, and many of these cells coexpress TMPRSS2.
“In contrast, different types of respiratory cells, and also olfactory sustentacular cells, expressed ACE2. Many of these cells also coexpressed TMPRSS2.”
Quote in the paperSubstantial claimMay combine two claims.
Findingown dataIn humans, ACE2 transcripts are found in brain non-neuronal cell types such as astrocytes and microglia, and in neurons, particularly Purkinje and cortical layer 5 neurons.
“In humans, we found ACE2 transcripts in non-neuronal cell types such as astrocytes and microglia, and also in neurons, in particular Purkinje and cortical layer 5 neurons.”
Quote in the paperSubstantial claimMay combine two claims.
Findingown dataACE2-positive sustentacular cells in mouse olfactory epithelium are almost exclusively located in the dorsal part, with a relatively abrupt transition area.
“These were almost exclusively located in the dorsal part of the epithelium, with a relatively abrupt transition area (Figures 3H and 3I).”
Quote in the paperNot a substantial claimMay combine two claims.
Findingown dataAll three anti-ACE2 antibodies labeled the apical portion of mouse sustentacular cells, except for slight labeling of their somata with Ab2.
“All antibodies showed a labeling of the apical portion of sustentacular cells, except for a slight labeling of their somata with Ab2 (Figure 3D).”
Quote in the paperNot a substantial claimMay combine two claims.
Findingown dataIn human olfactory neuroepithelium, ACE2 is enriched in the apical half of sustentacular cells, unlike the very apical localization in mouse.
“However, the very apical localization of ACE2 observed in the mouse neuroepithelium was not recapitulated in humans, but ACE2 was rather enriched in the apical half of the cells.”
Quote in the paperNot a substantial claimClassified as probably not a substantial claim.
PremisecitationEntry of SARS-CoV-2 into target cells depends on the Spike protein present on the virus capsid.
“Entry of SARS-CoV-2 into target cells depends on the Spike protein (S), which is present on the virus capsid (Letko et al., 2020; Walls et al., 2020).”
Quote in the paperSubstantial claim0 of 2 sources checked- cites Letko et al., 2020
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- cites Walls et al., 2020
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PremisecitationIn SARS-CoV-2-infected Syrian hamsters, sustentacular cells are infected and the olfactory epithelium shows massive desquamation.
“However, a study of SARS-CoV-2-infected Syrian hamsters showed infection of sustentacular cells and a massive desquamation of the olfactory epithelium (Bryche et al., 2020).”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites Bryche et al., 2020
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Interpretationown dataRespiratory cells are not the only cells in contact with the outside world that exhibit the molecular keys involved in SARS-CoV-2 entry in the nose; sustentacular cells share the same characteristics.
“Respiratory cells are thus not the only cells in contact with the outside world that exhibit the molecular keys involved in SARS-CoV-2 entry in the nose. Sustentacular cells, which lie at the interface between the central nervous system and the olfactory cavity, share the same characteristics.”
Quote in the paperSubstantial claimClassified as probably not a substantial claim. May combine two claims.
PremisecitationTMPRSS2 primes Spike in association with ACE2, leading to fusion between cellular and viral membranes.
“One reported mechanism facilitating virus entry consists, in association with ACE2, in the priming of S by the cellular serine protease TMPRSS2, also attached to the cellular membrane, which eventually leads to the fusion between the cellular and the viral membranes (Hoffmann et al., 2020; Zhou et al., 2020).”
Quote in the paperSubstantial claim0 of 2 sources checked- cites Hoffmann et al., 2020
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- cites Zhou et al., 2020
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Probably not a substantial claim: may not pass S3 (load-bearing).
PremisecitationA retrospective case study on 214 patients with COVID-19 reported neurological manifestations possibly correlated with the severity of the disease.
“a retrospective case study on 214 patients with COVID-19 reported neurological manifestations possibly correlated with the severity of the disease (Mao et al., 2020)”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites Mao et al., 2020
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Probably not a substantial claim: may not pass S3 (load-bearing).
Study design and conductown dataWhether the histological damage was caused by the virus cannot be determined based on the few samples analyzed.
“Whether the histological damage was caused by the virus cannot be determined based on the few samples analyzed.”
Quote in the paperNot a substantial claimProbably not a substantial claim: may not pass S3 (load-bearing).
InterpretationreasoningThe potential double effect on two chemosensory systems may suggest a more central alteration, involving direct infection of the brain by SARS-CoV-2.
“this potential double effect on two chemosensory systems that share nothing at the periphery may suggest a more central alteration, involving, for example, a direct infection of the brain by SARS-CoV-2.”
Quote in the paperNot a substantial claimProbably not a substantial claim: may not pass S3 (load-bearing).
Study design and conductown dataTMPRSS2 antibodies were first evaluated on kidney tissue.
“We tested two different antibodies, which were first evaluated on kidney tissue (Figure S2).”
Quote in the paperNot a substantial claimProbably not a substantial claim: may not pass S3 (load-bearing).
Study design and conductown dataA similar somatic staining was also observed, with the same antibody, in proximal tubule cells of the mouse kidney.
“A similar somatic staining was also observed, with the same antibody, in proximal tubule cells of the mouse kidney (Figure S1D).”
Quote in the paperNot a substantial claimProbably not a substantial claim: may not pass S3 (load-bearing).
PremisecitationRespiratory cells that line the respiratory airways coexpress ACE2 and TMPRSS2.
“Expectedly, the main targets of SARS-CoV-2, respiratory cells that line the respiratory airways, coexpress ACE2 and TMPRSS2 (Bertram et al., 2012).”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites Bertram et al., 2012
Not checked in the free run.
Probably not a substantial claim: may not pass S3 (load-bearing).
PremisecitationThe olfactory nerve route is used by various viruses to reach the brain.
“the olfactory nerve route is indeed used by various viruses to reach the brain, including HSV1, poliovirus, MHV (a coronavirus), paramyxoviruses, Hendra virus, Nipah virus, influenza virus, adenoviruses, bunyaviruses, and VSV (Durrant et al., 2016).”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites Durrant et al., 2016
Not checked in the free run.
May combine two claims.
PremisecitationTissue specificity of orthologs is highly conserved between tetrapod species, and in particular between mouse and humans.
“Given that ortholog tissue specificity is highly conserved between tetrapod species (Kryuchkova-Mostacci and Robinson-Rechavi, 2016), and in particular between mouse and humans (Liao and Zhang, 2006), we analyzed mouse tissues in parallel to human ones.”
Quote in the paperSubstantial claim0 of 2 sources checked- cites Kryuchkova-Mostacci and Robinson-Rechavi, 2016
Not checked in the free run.
- cites Liao and Zhang, 2006
Not checked in the free run.
May combine two claims.
Interpretationown dataThe presence of ACE2 and TMPRSS2 transcripts in sensory neuroepithelium indicates the presence of cells that may express both genes, or a mix of cells expressing either TMPRSS2 or ACE2.
“indicating the presence in this tissue of cells that may express both genes, or of a mix of cells that express either TMPRSS2 or ACE2.”
Quote in the paperSubstantial claimClassified as probably not a substantial claim.
Premisenone givenViral attachment involves an interaction between Spike and ACE2 on the target cell surface.
“Viral attachment involves an interaction between S and the angiotensin-converting enzyme-2 (ACE2), located on the surface of the target cell.”
Quote in the paperSubstantial claimProbably not a substantial claim: may not pass S3 (load-bearing).
PremisecitationACE2 is found in the motor cortex, raphe, and nuclei involved in the regulation of cardiovascular function according to a previous report.
“which found ACE2 in the motor cortex, raphe, and nuclei involved in the regulation of cardiovascular function (Doobay et al., 2007).”
Quote in the paperNot a substantial claim0 of 1 sources checked- cites Doobay et al., 2007
Not checked in the free run.
Probably not a substantial claim: may not pass S3 (load-bearing).
PremisecitationSARS-CoV particles have been found in the human brain.
“SARS-CoV particles have also been found in the human brain (Ding et al., 2004; Gu et al., 2005; Xu et al., 2005).”
Quote in the paperNot a substantial claim0 of 3 sources checked- cites Ding et al., 2004
Not checked in the free run.
- cites Gu et al., 2005
Not checked in the free run.
- cites Xu et al., 2005
Not checked in the free run.
The paper relies on this without support.
The four adult patients from whom olfactory and respiratory biopsies were collected are representative of the general human population for ACE2/TMPRSS2 expression in these tissues.
“We collected biopsies via nasal endoscopic surgery from four adult patients and explored the potential expression of ACE2 and TMPRSS2.”
The paper relies on this without support.
The single-cell RNA-seq dataset from Durante et al. (2020) accurately reflects the cell types and gene expression of the human olfactory neuroepithelium.
“To identify putative viral targets transcribing both ACE2 and TMPRSS2 in the neuroepithelium, we took advantage of a very recently published dataset reported by Durante et al. (2020).”
The paper relies on this without support.
The human brain single-nucleus RNA-seq datasets are representative of the human brain.
“Finally, we explored two single-nucleus RNA-seq datasets to evaluate ACE2 expression in the human brain.”
The paper relies on this without support.
ACE2 is the main player in the binding and entry of SARS-CoV-2 into human cells.
“ACE2, the main player in the binding and entry of the SARS-CoV-2 into human cells”
The paper relies on this without support.
TMPRSS2 is a serine protease that facilitates viral entry.
“TMPRSS2, a serine protease known to facilitate viral entry”
The paper relies on this without support.
Neuronal renewal in the main olfactory epithelium is slow (a few months).
“given the slow rate of neuronal renewal in the main olfactory epithelium (a few months)”
The paper relies on this without support.
SARS-CoV-2-triggered anosmia develops rapidly.
“the apparent rapid development of SARS-CoV-2-triggered anosmia”
The paper relies on this without support.
Sustentacular cells are in direct contact with the olfactory cavity and interact with everything that enters the nose.
“sustentacular cells are in direct contact with the olfactory cavity and thus interact with everything that enters the nose.”
The paper relies on this without support.
Sustentacular cells play a critical role in the maintenance of the olfactory neuroepithelium integrity; their loss or suffering would cause disaggregation or malfunctioning of the neuroepithelium, leading to anosmia.
“sustentacular cells play a critical role in the maintenance of the olfactory neuroepithelium integrity: their loss or suffering would cause disaggregation or malfunctioning of the neuroepithelium, respectively, leading to anosmia.”
The paper relies on this without support.
The human olfactory neuroepithelium is relatively poorly organized, particularly in aged individuals.
“the relatively poor organization of the human olfactory neuroepithelium, in particular in aged individuals.”
Claims other papers make about this one
Each is checked against this paper’s own text: does it say what the citing paper says it does?
ACE2 expression has been reported for many tissues/organs including the lung, brain, kidney, small intestine, colon, and heart.
“ACE2 expression has been reported for many tissues/organs including the lung (ie, lung type II alveolar cells/AT2, bronchial epithelial cells), brain, kidney, small intestine, colon, and heart.”
From COVID-19, Myocarditis and Pericarditis (2023) · cites this paper as [15]
The Results section on receptor expression in the brain reports ACE2 in human brain cell types. The Results section on protein expression also describes ACE2-expressing cells in mouse colon and kidney, but not human colon or kidney. The passages do not establish expression in the lung—including the specified alveolar and bronchial cells—small intestine, or heart; the respiratory cells studied are nasal. (LLM, conf=0.87)
Experimental — accuracy not yet measuredHuman olfactory sustentacular cells express both ACE2 and TMPRSS2 virus receptors.
“Studies reported that human olfactory sustentacular cells express both ACE2 and TMPRSS2 virus receptors [33].”
From Hyperactivation of P2X7 receptors as a culprit of COVID-19 neuropathology (2021) · cites this paper as [33]
Human olfactory sensory neurons do not express ACE2 and TMPRSS2 but express CD147 that could allow SARS-CoV-2 neuroinvasion.
“Although current literature data reports that human olfactory sensory neurons do not express these proteins, they express CD147 that could allow SARS-CoV-2 neuroinvasion (Fig. 1) [33, 34].”
From Hyperactivation of P2X7 receptors as a culprit of COVID-19 neuropathology (2021) · cites this paper as [33]
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Miura, T. A., Travanty, E. A., Oko, L. M., et al. (2007) · Journal of Virology · doi:10.1128/jvi.01851-07 · Cited by 63 · see citing works
Jia, C., Roman, C., & Hegg, C. C. (2010) · Toxicological Sciences · doi:10.1093/toxsci/kfq071 · Cited by 54 · see citing works
RESPIRATORY VERSUS GASTRO-INTESTINAL INFECTION IN POLIOMYELITIS
Flexner, S. (1936) · The Journal of Experimental Medicine · doi:10.1084/jem.63.2.209 · Cited by 35 · see citing works
Olfactory neural pathway in mouse hepatitis virus nasoencephalitis
Barthold, S. (2004) · Acta Neuropathologica · doi:10.1007/bf00686390 · Cited by 30 · see citing works
Olfactory receptor neuronal dendrites become mostly intra‐sustentacularly enwrapped upon maturity
Liang, F. (2018) · Journal of Anatomy · doi:10.1111/joa.12777 · Cited by 28 · see citing works
Miller, M. A., & O'Bryan, D. (2003) · Ultrastructural Pathology · doi:10.1080/01913120309944 · Cited by 18 · see citing works
Tang, J., Tang, J., Ling, E. A., et al. (2009) · Neuroscience · doi:10.1016/j.neuroscience.2009.03.051 · Cited by 14 · see citing works
Tuerdi, A., Kikuta, S., Kinoshita, M., Kamogashira, T., Kondo, K., Iwasaki, S., & Yamasoba, T. (2018) · Scientific Reports · doi:10.1038/s41598-018-35607-w · Cited by 13 · see citing works
Jacobs, S., Zeippen, C., Wavreil, F., Gillet, L., & Michiels, T. (2019) · Viruses · doi:10.3390/v11080757 · Cited by 13 · see citing works
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Brann, D. H., Tsukahara, T., Weinreb, C., Lipovsek, M., Van den Berge, K., Gong, B., Chance, R., Macaulay, I. C., Chou, H.-J., Fletcher, R. B., et al. (2020) · Science Advances · doi:10.1126/sciadv.abc5801 · Cited by 1,198 · see citing works
IgA dominates the early neutralizing antibody response to SARS-CoV-2
Sterlin, D., Mathian, A., Miyara, M., Mohr, A., Anna, F., Claër, L., Quentric, P., Fadlallah, J., Devilliers, H., Ghillani, P., et al. (2021) · Science Translational Medicine · doi:10.1126/scitranslmed.abd2223 · Cited by 1,037 · see citing works
Zubair, A. S., McAlpine, L. S., Gardin, T., Farhadian, S., Kuruvilla, D. E., & Spudich, S. (2020) · JAMA Neurology · doi:10.1001/jamaneurol.2020.2065 · Cited by 923 · see citing works
Lifting the mask on neurological manifestations of COVID-19
Pezzini, A., & Padovani, A. (2020) · Nature Reviews Neurology · doi:10.1038/s41582-020-0398-3 · Cited by 447 · see citing works
Neurological Involvement in COVID-19 and Potential Mechanisms: A Review
Aghagoli, G., Gallo Marin, B., Katchur, N. J., Chaves-Sell, F., Asaad, W. F., & Murphy, S. A. (2021) · Neurocritical Care · doi:10.1007/s12028-020-01049-4 · Cited by 357 · see citing works
Anosmia in COVID-19: Underlying Mechanisms and Assessment of an Olfactory Route to Brain Infection
Butowt, R., & von Bartheld, C. S. (2021) · The Neuroscientist · doi:10.1177/1073858420956905 · Cited by 345 · see citing works
Cheemarla, N. R., Watkins, T. A., Mihaylova, V. T., Wang, B., Zhao, D., Wang, G., Landry, M. L., & Foxman, E. F. (2021) · Journal of Experimental Medicine · doi:10.1084/jem.20210583 · Cited by 213 · see citing works
COVID-19, Myocarditis and Pericarditis
Fairweather, D., Beetler, D. J., Di Florio, D. N., Musigk, N., Heidecker, B., & Cooper, L. T. (2023) · Circulation Research · doi:10.1161/circresaha.123.321878 · Cited by 181 · see citing works
Hyperactivation of P2X7 receptors as a culprit of COVID-19 neuropathology
Ribeiro, D. E., Oliveira-Giacomelli, Á., Glaser, T., Arnaud-Sampaio, V. F., Andrejew, R., Dieckmann, L., Baranova, J., Lameu, C., Ratajczak, M. Z., & Ulrich, H. (2021) · Molecular Psychiatry · doi:10.1038/s41380-020-00965-3 · Cited by 145 · see citing works
SARS-CoV-2 Dissemination Through Peripheral Nerves Explains Multiple Organ Injury
Fenrich, M., Mrdenovic, S., Balog, M., Tomic, S., Zjalic, M., Roncevic, A., Mandic, D., Debeljak, Z., & Heffer, M. (2020) · Frontiers in Cellular Neuroscience · doi:10.3389/fncel.2020.00229 · Cited by 95 · see citing works
- Authors
- Fodoulian, L., Tuberosa, J., Rossier, D., Boillat, M., Kan, C., Pauli, V., Egervari, K., Lobrinus, J. A., Landis, B. N., Carleton, A., & Rodriguez, I.
- Affiliations
- University of Geneva (CH)
- University Hospital of Geneva (CH)
- Corresponding author
- Carleton, Alan
- ORCID
- Venue
- iScience
- Publisher
- openRxiv
- Published
- 2020-04-02
- Volume
- 23
- Issue
- 12
- Pages
- 101839-101839
- ISSN
- 2589-0042 (ISSN-L)
- PMC
- PMC7685946
- OpenAlex
- W3107452553
- Cited by
- 286
- References
- 76
- Citations by year
- 2026: 8 · 2025: 7 · 2024: 29 · 2023: 46 · 2022: 68 · 2021: 68 · 2020: 60
- FWCI
- 21.23
- Type
- journal article
- Language
- en
- Open access
- gold
- doi.org
- Licence
- cc-by-nc-nd
- www.biorxiv.org/about/FAQ#license (vor)
- Locations
- iScience (publishedVersion, cc-by-nc-nd)
- bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion)
- PubMed Central (publishedVersion, cc-by-nc-nd)
- PubMed (publishedVersion)
- Archive ouverte UNIGE (University of Geneva) (publishedVersion, other-oa)
- DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
- Archive ouverte UNIGE (University of Geneva) (submittedVersion, cc-by-nd)
- Funders
- National Science Foundation
- Schweizerischer Nationalfonds zur Förderung der Wissenschaftlichen Forschung (172878, 189153)
- Université de Genève (31003A_172878, 310030_189153)
- Institute of Genetics and Genomics of Geneva
- Topics
- Life Sciences › Neuroscience › Sensory Systems › Olfactory and Sensory Function Studies
- Health Sciences › Medicine › Infectious Diseases › SARS-CoV-2 and COVID-19 Research
- Health Sciences › Medicine › Infectious Diseases › COVID-19 Clinical Research Studies
- Keywords
- SARS-CoV-2, COVID-19 pandemic, olfactory epithelium, ACE2, TMPRSS2, sustentacular cells, olfactory sensory neurons, anosmia, olfactory bulb, viral entry, single-cell RNA sequencing
About these checks
Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.
Claims read by deepseek-v4.1-flash under definition version 3 (extract-v3) on 2026-10-09.
Classified by jev-1.13-20260917 (gold-guideline-v0.2) on 2026-10-09.
Statistics and arithmetic are recomputed, not judged.
Confidence bands are shown only for fields calibrated on our annotated test set. None is calibrated yet, so no band is shown.
What counts as a claim ›How we measure confidence ›How we check this ›
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