key: cord-0951327-elaqvw2d
authors: Bangma, Amber; Voskuil, Michiel Dirk; Weersma, Rinse K.
title: TNFα-antagonist use and mucosal inflammation are associated with increased intestinal expression of SARS-CoV-2 host protease TMPRSS2 in patients with inflammatory bowel disease
date: 2020-06-15
journal: Gastroenterology
DOI: 10.1053/j.gastro.2020.05.091
sha: 0f2c9135e46454ff0a9db3c76c315238ea086812
doc_id: 951327
cord_uid: elaqvw2d

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Conflicts of interest R.K.W. acted as consultant for Takeda, received unrestricted research grants from Takeda, Johnson and Johnson, Tramedico and Ferring and received speaker fees from MSD, Abbvie and Janssen Pharmaceuticals. The remaining authors disclose no (potential) conflicts of interest.

angiotensin-converting enzyme 2 CD:

Crohn's disease COVID-19: coronavirus disease 2019 IBD:

inflammatory bowel disease IBD-U inflammatory bowel disease unclassified SARS-CoV-2: severe acute respiratory syndrome coronavirus 2 UC ulcerative colitis TMPRSS2:

transmembrane protease serine protease 2 TNFα:

tumor necrosis factor alpha Dear Editor, Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) viral transfection is dependent on ACE2 and TMPRSS2, and increased intestinal receptor expression might support viral replication [1, 2, 3] . Therefore, we read with great interest the work by Krzysztof et al, reporting that anatomical location, intestinal inflammation and age are key determinants of intestinal expression of angiotensinconverting enzyme 2 (ACE2) and transmembrane serine protease 2 (TMPRSS2) in patients with inflammatory bowel disease (IBD) [1] . However, several additional questions remain as the authors themselves also describe. It is important to understand the effects of immunomodulating drugs on ACE2 and TMPRSS2 expression, and the cell-type specific context in which these genes are expressed [1] . In addition, it is important to understand genetic determinants of intestinal ACE2 and TMPRSS2 expression. To address these remaining questions, we queried host genetic data and RNA sequencing data of intestinal biopsies from patients with IBD. We independently confirm that anatomical location, intestinal inflammation and age are determinants of intestinal ACE2 and TMPRSS2 expression. Furthermore, we demonstrate that immunomodulating drugs are associated with increased intestinal TMPRSS2 expression, and highlight the cell-type specific context of ACE2 and TMPRSS2 expression. We analyzed bulk RNA sequencing data of 92 ileal and 199 colonic snap frozen mucosal biopsies from 168 patients with IBD, as part of the 1000IBD project [4] . Genotypes were obtained using both imputed Global Screening Array data and whole exome sequencing. We analyzed clinical data regarding the use of immunomodulating drugs, age, sex, diagnosis (CD, UC or IBD-U) and BMI. Multivariate linear mixed regression analyses were performed to assess the effects of the abovementioned clinical factors on intestinal gene expression levels of ACE2 and TMPRSS2 (R v.3.6.0). In addition, we assessed the effects of host genetic variation on gene expression (cis-expression quantitative trait loci). All of the above-mentioned clinical factors, as well as sequencing batch, were included as covariates. We performed deconvolution analyses on bulk RNA sequencing data to assess cell-type-specific expression of ACE2 and TMPRSS2. We validated this approach using single-cell RNA sequencing data from an independent set of 18 colonic biopsies from 11 patients with ulcerative colitis (unpublished data).

First, we replicated the effects of anatomical location, intestinal inflammation and age on intestinal expression of ACE2 and TMPRSS2. Expression of ACE2 was higher, and TMPRSS2 was lower in ileum compared to colon (P< 2.2x10 -16 ). Moreover, expression of ACE2 was lower, and expression of TMPRSS2 was higher in inflamed ileum compared to non-inflamed ileum, independent of medication use, age, sex, diagnosis and BMI (P= 4.4x10 -6 and P= 8.6x10 -8 , respectively). Ileal TMPRSS2 expression was associated with increasing age (P= .04). Furthermore, we found increased TMPRSS2 expression in intestinal (ileal and colonic) biopsies of male patients (P= .02).

Second, we studied the effects of immunomodulating drugs on ACE2 and TMPRSS2 expression in intestinal biopsies. We found increased TMPRSS2 expression in ileal biopsies of patients using Tumor Necrosis Factor alpha (TNFα)-antagonists (P= 8.8x10 -6 ) independent of intestinal inflammation, age, sex, diagnosis and BMI. Because aminosalicylates were only used in the context of ulcerative colitis, we assessed the influence of aminosalicylates in colonic tissue only, and observed an increased TMPRSS2 expression (P= .02). The use of thiopurines or steroids was not associated with differential expression of ACE2 or TMPRSS2. Furthermore, host genetic variation was not associated with differential expression of ACE2 or TMPRSS2.

Third, we interrogated the cell-type-specific context of intestinal ACE2 and TMPRSS2 expression. Using bulk RNA sequencing data, we quantified cell-type proportions and observed that an enrichment of epithelial cells is associated with increased expression of ACE2 and TMPRSS2 in both ileum and colon (all P< 0.004). Using single-cell RNA sequencing data from colonic biopsies, we observed that TMPRSS2 is mainly expressed by absorptive enterocytes, and that intestinal inflammation is associated with increased TMPRSS2 expression within absorptive enterocytes (P= 3.2x10 -22 ). ACE2 was also primarily expressed by absorptive enterocytes, but expression was not affected by intestinal inflammation.

In conclusion, ACE2 and TMPRSS2 are key proteins for cellular entry of SARS-CoV-2, and are highly expressed in the intestinal mucosa. Next to intestinal inflammation, age, sex and anatomical location, also the use of TNFα-antagonists and aminosalicylates influence intestinal expression of TMPRSS2. We demonstrate that intestinal inflammation is associated with increased expression of TMPRSS2 in absorptive enterocytes, suggesting that the increased expression is not merely an effect of change in cellular composition during inflammation. Altered intestinal expression could render patients with IBD particularly susceptible to COVID-19 and absorptive enterocytes could provide targets for interventional studies. Indeed, clinical studies are needed to monitor the impact of COVID-19 on patients with IBD.

Published Online First: 12

Published Online First: 01

We thank Shixian Hu in help with statistical analyses, and Eleonora Festen, Werna Uniken Venema, Harry van Goor and Arno Bourgonje for critical discussion of the results. Furthermore, we thank all the participants of the 1000IBD cohort.