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ACE2 contributes to the maintenance of mouse epithelial barrier function

As the COVID-19 pandemic spreads worldwide, the absence of any efficient treatment and our limited understanding of the virus will limit our ability to control the disease, if we are to defeat this pandemic further research is essential. SARS-COV-2 enters the cell via Angiotensin I converting enzyme 2 (ACE2), a zinc metalloprotease, there it replicates causing damage to the lung and other tissues. Further research into this mechanism and regulation of ACE2 may prove beneficial in the development of effective COVID-19 treatments.

ACE2 is expressed highly throughout the gut where it inhibits the renin/angiotensin system (RAS). Recent COVID-19 studies have implicated the virus in gastrointestinal diseases and the faecal microbiome of COVID-19 patients is significantly different to healthy patients. ACE2 upregulation has also been shown to worsen the effects of COVID-19 infection by increasing viral entry into cells. The use of ACE2 inhibitors has been discussed as a possible treatment for COVID-19, however the importance of the renin/angiotensin system may impair their use due to significant side effects. A few studies with ACE inhibitors have demonstrated lower mortality rates for COVID-19 patients, however inhibiting ACE2 caused intestinal dysbiosis and increased inflammation.

In this study, the authors explored the role of ACE2 in the gut and its role in the inflammatory and regenerative response using intestinal organoids and mouse IBD models.

The study used ACE2 knockout mice and demonstrated no expression of ACE2 in colon or intestinal organoids derived from these mice.

In intestinal organoids, ACE2 deletion did not alter the morphology of the organoid, however LGR5, a marker of intestinal stem cells and ki67, a proliferation marker were significantly lower in ACE2 knockout organoids compared to normal.

Consistent with these findings, ACE2 deleted organoids had slower proliferation and differentiation resulting in smaller organoids.

In an intestinal barrier function assay using FITC-dextran, ACE2 knockout organoids absorbed more FITC-dextran than normal organoids suggesting a role for ACE2 in barrier function, however ACE2 KO mice did not show any difference in plasma FITC compared to normal mice.

In a mouse model of DSS-induced IBD, ACE2 deletion exacerbated the IBD effect with more pronounced weight loss, earlier bloody faeces and higher faeces album level compared to normal mice.

More crypt loss and abnormal intestinal structures were also observed in ACE2 KO mice with DSS and increased inflammatory cell infiltration and shorter colon length was noted.

There was also significant reduction in the expression of LGR5 and MUC2.

In ACE2 KO organoids there was also a calcium overload in response to ATP and elevated reactive oxygen species (ROS) production, which inhibited the normal function of intestinal stem cells.

Recent studies have shown that patients with IBD may be susceptible to poor outcome and increased risk from COVID-19. The Sars-Cov-2 virus enters the cell via the ACE2 protein and given that a number of ACE2 inhibitors are in use for hypertension there may be a benefit in using these to combat COVID-19. In this study the authors demonstrated that deletion of ACE2 affected the stemness of intestinal stem cells and exacerbated the effect of IBD-inducing agents, resulting in perturbation of barrier function via calcium overload and increased ROS. The authors advise against the use of ACE2 inhibitors to treat COVID-19 patients with IBD but suggest a combination of ACE2 inhibitors and calcium blockers merits further investigation.

Reference

Yu et al. ACE2 contributes to the maintenance of mouse epithelial barrier function. Biochemical and Biophysical Research Communications (2020).
doi: 10.1016/j.bbrc.2020.10.002

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Epistem's Organoid, IBD and COVID-19 Services

We have recently launched our COVID‑19 testing services for both the detection of the virus as well as the detection of antibodies to the virus indicating prior infection. Analysis, from oral or nasal swabs and blood, is carried out in our accredited labs with quick turnaround times. We also perform NGS analysis to characterise COVID‑19 strains.

We have leveraged our 10X Genomics NGS services for immune repertoire sequencing of both B-cell and T-cell receptors, and can assess clonal expansion and VDJ usage for both COVID‑19 infection and response to vaccine treatment.

We also offer preclinical models that replicate aspects of IBD in humans that can be used to generate target validation and mechanism of action data to confirm the efficacy of novel therapeutic agents. In addition, we produce acute models of IBD using DSS and TNBS, as well as an adoptive T-cell transfer model of chronic IBD.

Epistem offers validated small intestinal in vitro organoid models derived from a variety of species, including human, canine and rodents. Our organoid cultures display cellular architecture that closely resembles that observed in vivo. Cultures mimic the stem cell niche allowing cell proliferation and differentiation to occur. The established organoid protocols allow MOA determination and provide in vitro to in vivo biological and PD linkages through following readouts:

  • Viability assays (visual or with MTS)
  • Histology/IHC or RNAscope to assess the effect on specific protein expression including LGR5
  • Gene expression (NGS, Microarray or qRT-PCR) to evaluate the effect of treatment on expression of signalling pathways genes
  • FACS analysis to identify specific cell populations affected
  • ELISA to identify and quantify cytokines of interest within organoid's spend media

Epistem's intestinal organoid models allow screening of multiple agents, with over 5000 screened to date including Wnt pathway regulators, cytokines, growth factor, chemotherapy drugs, steroids and lectins.

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