Relevance of SARS-CoV-2 related factors ACE2 and TMPRSS2 expressions in gastrointestinal tissue with pathogenesis of digestive symptoms, diabetes-associated mortality, and disease recurrence in COVID-19 patients

  1. Ashutosh Kumar
  2. Muneeb A. Faiq
  3. Vikas Pareek
  4. Khursheed Raza
  5. Ravi K. Narayan
  6. Pranav Prasoon
  7. Pavan Kumar
  8. Maheswari Kulandhasamy
  9. Chiman Kumari
  10. Kamla Kant
  11. Himanshu N. Singh
  12. Rizwana Qadri
  13. Sada N. Pandey
  14. Santosh Kumar

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Abstract

Introduction

COVID-19 is caused by a new strain of coronavirus called SARS-coronavirus-2 (SARS-CoV-2), which is a positive sense single strand RNA virus. In humans, it binds to angiotensin converting enzyme 2 (ACE2) with the help a structural protein on its surface called the S-spike. Further, cleavage of the viral spike protein (S) by the proteases like transmembrane serine protease 2 (TMPRSS2) or Cathepsin L (CTSL) is essential to effectuate host cell membrane fusion and virus infectivity. COVID-19 poses intriguing issues with imperative relevance to clinicians. The pathogenesis of GI symptoms, diabetes-associated mortality, and disease recurrence in COVID-19 are of particular relevance because they cannot be sufficiently explained from the existing knowledge of the viral diseases. Tissue specific variations of SARS-CoV-2 cell entry related receptors expression in healthy individuals can help in understanding the pathophysiological basis the aforementioned collection of symptoms.

Materials and Methods

The data were downloaded from the Human Protein Atlas available at ( https://www.proteinatlas.org/humanproteome/sars-cov-2 ) and the tissue specific expressions (both mRNA and protein) of ACE2 and TMPRSS2 as yielded from the studies with RNA sequencing and immunohistochemistry (IHC) were analyzed as a function of the various components of the digestive tract. A digestive system specific functional enrichment map of ACE2 gene was created using g:profiler ( https://biit.cs.ut.ee/gprofiler/gost ) utility and the data were visualized using Cytoscape software, version 3.7.2 ( https://cytoscape.org/ ).

Results

The correlated expression (transcriptomic and proteomic) of ACE2 (to which SARS-CoV-2 binds through the S-spike) was found to be enriched in the lower gastrointestinal tract (GIT) (highest in small intestine, followed by colon and rectum), and was undetectable in the upper GIT components: mouth cavity (tongue, oral mucosa, and salivary glands), esophagus, and stomach. High expression of ACE2 was noted in the glandular cells as well as in the enterocytes in the lining epithelium (including brush border epithelium). Among other digestive system organs, Gall bladder (GB) showed high expression of ACE2 in glandular cells, while any protein expression was undetectable in liver and pancreas. TMPRSS2 was found enhanced in GIT and exocrine glands of pancreas, and co-localized with ACE2 in enterocytes.

Conclusions

Based on the findings of this study and supportive evidence from the literature we propose that a SARS-CoV-2 binding with ACE2 mediates dysregulation of the sodium dependent nutrient transporters and hence may be a plausible basis for the digestive symptoms in COVID-19 patients. ACE2 mediated dysregulation of sodium dependent glucose transporter (SGLT1 or SLC5A1) in the intestinal epithelium also links it to the pathogenesis of diabetes mellitus which can be a possible reason for the associated mortality in COVID-19 patients with diabetes. High expression of ACE2 in mucosal cells of the intestine and GB make these organs potential sites for the virus entry and replication. Continued replication of the virus at these ACE2 enriched sites may be a basis for the disease recurrence reported in some, thought to be cured, patients.

Graphical Abstract

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  1. ScreenIT

    SciScore for 10.1101/2020.04.14.040204: (What is this?)

    Please note, not all rigor criteria are appropriate for all manuscripts.

    Table 1: Rigor

    Institutional Review Board StatementIACUC: Since no direct subject or patient data were used in this study, clearance from the Institutional Ethics Committee was precluded.
    IRB: IHC: As described by the source labs, specimens containing normal tissue were collected and sampled from anonymized paraffin embedded material of surgical specimens, in accordance with approval from the local ethics committee.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variablenot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    Antibodies against human ACE2 (HPA000288, CAB026174) and TMPRSS2 (HPA035787) were labeled with DAB (3, 3’-diaminobenzidine) stain.
    human ACE2
    suggested: (Sigma-Aldrich Cat# HPA000288, RRID:AB_1078160)
    CAB026174
    suggested: None
    TMPRSS2
    suggested: (Sigma-Aldrich Cat# HPA035787, RRID:AB_2674782)
    HPA035787
    suggested: None
    Software and Algorithms
    SentencesResources
    A digestive system specific functional enrichment map of ACE2 gene was constructed using g:profiler (https://biit.cs.ut.ee/gprofiler/gost) utility and viewed with Cytoscape software, version 3.7.2 (https://cytoscape.org/).
    g:profiler
    suggested: (G:Profiler, RRID:SCR_006809)
    Cytoscape
    suggested: (Cytoscape, RRID:SCR_003032)
    https://cytoscape.org/
    suggested: (CluePedia Cytoscape plugin, RRID:SCR_015784)
    The extracted RNA samples were analyzed using either an Experion automated electrophoresis system (Bio-Rad Laboratories, Hercules, CA, USA) with the standard-sensitivity RNA chip or an Agilent 2100 Bioanalyzer system (Agilent Biotechnologies, Palo Alto, USA) with the RNA 6000 Nano Labchip Kit.
    Bio-Rad Laboratories
    suggested: (Bio-Rad Laboratories, RRID:SCR_008426)
    Agilent Biotechnologies
    suggested: None
    Transcript abundance estimation was performed using Kallisto v0.43.1 (https://pachterlab.github.io/kallisto/about).
    Kallisto
    suggested: (kallisto, RRID:SCR_016582)

    Results from OddPub: Thank you for sharing your data.

    Results from LimitationRecognizer: We detected the following sentences addressing limitations in the study:
    Limitations: All the aspects of the plausible SARS-CoV-2 binding receptor ACE2 mediated pathology in the digestive system which we have discussed above are based on the distribution of the virus cell entry related factors in the normal tissue. Hence, this study presents indirect evidence which needs to be validated in actual patients before reaching any conclusion. Future directions: Further studies are advisable to understand the molecular mechanisms involved in the SARS-CoV-2 binding receptor ACE2 mediated dysregulation of the intestinal nutrient transporters and finding out COVID-19 specific drug targets. Inter-individual variations in frequency of the digestive symptoms, diabetes associated mortality, and recurrences may depend upon the genotype specific variations in ACE2 expression and other patient specific characteristics (like age, sex, and comorbidity). A study of these variables in the disease pathogenesis may help in deciding personalized therapeutic management for the COVID-19 cases.

    Results from TrialIdentifier: No clinical trial numbers were referenced.

    Results from Barzooka: We did not find any issues relating to the usage of bar graphs.

    Results from JetFighter: We did not find any issues relating to colormaps.

    Results from rtransparent:
    • Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
    • Thank you for including a funding statement. Authors are encouraged to include this statement when submitting to a journal.
    • No protocol registration statement was detected.

    About SciScore

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  2. Version published to 10.1101/2020.04.14.040204 on bioRxiv