Research Trend   |   Jun 28, 2020

A powerful mouse model for studying Covid-19

Wuhan Virus

We have all been engulfed by the current SARS-CoV-2 pandemic with the lives of millions affected. To date, COVID-19 has resulted in almost 8 million confirmed cases with over 400,000 deaths. Considerable effort and money have been invested in trying to understand the mode of infection, pathogenicity and immune reactivity in order to develop therapeutics and vaccines.

Mice have been studied in the laboratory for over 100 years and due to their genetic similarity with humans, have proven to be the model organism of choice for understanding human development, homeostasis and disease. The lack of useful mouse models to study COVID-19 is compounded by the fact that mouse ACE2 protein, unlike ACE2 in human and bat that acts as the receptor for SARS-CoV-2 entry (1), does not facilitate cellular entry of SARS-CoV-2 in mouse (2). This is analogous to MERS-CoV, which was also shown to be incapable of infecting murine cells, leading several groups to develop mouse models that can facilitate viral entry (3,4). This included the generation of a transgenic knockin mouse that expresses the MERS-CoV receptor DPP4 and a model whereby mice are transduced with adenovirus carrying the human DPP4 gene, thus sensitizing them to MERS-CoV infection. Such models successfully recapitulated the human disease as did SARS mouse models expressing human ACE2 (hACE2) (5). However, the use of such hACE2 models to study the current SARS-CoV-2 appears restricted as demonstrated by poor pathogenesis. The development of a new mouse model that displays the hallmarks of COVID-19 upon SARS-CoV-2 infection is clearly of importance.

Adenovirus is a preferred system for gene delivery into most mammalian cells since it does not integrate into the genome and, by design, is rendered replication incompetent. In a very recent paper, Sun et al (6) modified an existing approach (7) to develop a COVID-19 mouse model whereby adenovirus expressing hACE2 under the control of a CMV promoter (Ad5-ACE2) is administered intranasally to 6-8 week old BALB/c mice followed by infection with SARS-CoV-2 isolated from COVID-19 patients. In line with previous work (8,9), hACE2 expression was observed in the alveolar epithelium and to a lesser extent the airway epithelium.  In contrast to control mice that only received empty Ad5 virus, Ad5-ACE2-transduced and SARS-CoV-2-infected BALB/c mice developed pneumonia, characterised by breathing difficulties and significant (20%) weight loss. Importantly, C57BL/6 mice, which, unlike BALB/c, are not immunocompromised, showed a similar phenotype. Lung pathology of both models revealed inflammatory infiltrates, necrosis and alveolar edema consistent with recent reports of severe gross lung damage in SARS-CoV-2 human lungs.

To better understand how SARS-CoV-2 interacts with the innate immune system, knockout (KO) mice lacking components of the interferon-1 (IFN-1) signalling pathway were infected with SARS-CoV-2 following sensitization with Ad5-ACE2. Genetic depletion of interferon-α/β receptor (IFNAR-/- mice) led to delayed viral clearance and attenuated inflammation in contrast to mice lacking type II IFN (IFN-γ), which remained asymptomatic. A more dramatic phenotype was observed in STAT1 KO mice with SARS-CoV-2 infection leading to exacerbated weight loss, enhanced lung inflammation and delayed virus clearance. In support of previous work, a protective role of IFN-1 against SARS-CoV-2 was proposed since pre-treatment of mice with the potent IFN-1 inducer resulted in a better clinical outcome.

To identify changes in gene expression resulting from SARS-CoV-2 infection, lungs from Ad5-ACE2-transduced mice were analysed by RNA-Seq 2 days post-infection. A total of 3056 genes differentially expressed compared to Ad5-Empty transduced mice were observed. Consistently, genes associated with inflammation and innate and adaptive immunity were upregulated together with genes involved in T cell migration (CD4 and CD8). Immuno-depletion of CD4+ and CD8+ T cells confirmed the requirement for both arms of the immune response. T cell epitopes were identified in the N protein and S1 region of the S protein, as demonstrated by the response of T cells harvested from infected mouse lungs to peptide pools encompassing SARS-CoV-2 structural and accessory ORFs. Additionally, a number of cytokines and chemokines shown to be over expressed in COVID-19 patients (10) were also upregulated.

Therapeutic intervention and vaccine development were evaluated with detection of neutralizing antibodies in mice sensitized to Ad5-hACE2. Using the Venezuelan equine encephalitis replicon particle (VRP) vaccine platform, SARS-CoV-2 titers were greatly reduced in BALB/c and C57BL/6 models following immunization with VRPs expressing SARS-CoV-2 spike protein (VRP-S).  In contrast, no increase in virus clearance was observed following immunization with VRPs expressing transmembrane, nucleocapsid or envelope proteins of SARS-CoV-2. The potential therapy of plasma transfer was also characterised in Ad5-hACE2 mice with plasma from 3 COVID-19 patients administered one day prior to infection. Compared to plasma from a non-infected donor, plasma from SARS-CoV-2 infected patients prevented weight loss and lung damage together with increased virus clearance. No effect was observed with plasma from patients who survived infections of SARS and MERS. Lastly, a similar regime was applied for treatment of the U.S. Food and Drug Administration approved drug, Remdesivir. Treatment with Remdesivir improved clinical outcome in infected Ad5-hACE2-transduced mice.

In summary, the work here describes a mouse model that recapitulates pneumonia seen in COVID-19 patients. Further studies are required for understanding the different contributions of STAT and IFN-1 signalling as is the need to address targeting of non-pulmonary target cells (11). However, this newly developed SARS-CoV-2 sensitization model, together with a similar model by Hassan et al (12), should prove extremely useful for deciphering the molecular mechanisms involved in viral clearance as well as therapeutic intervention and vaccine development.

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VectorBuilder offers adenovirus vectors and packaging of HAd5 and chimeric Ad5/F35 adenovirus. Our SARS-CoV-2 services include coronavirus vectors, lentivirus pseudotyped with coronavirus S protein, cell lines and animal models. Contact us to find out more, or for help designing your custom vectors. Use our award-winning online platform to design and order custom vectors specific to your research needs.

Reference

  1. Hoffman et al. Cell. 2020. 181 (2): 271-280 
  2. Zhou et el. Nature. 2020. 579: 270–273
  3. Cockrell et al. Nat Microbiol. 2016. 2: 16226
  4. Li et al. Proc Natl Acad Sci. 2017. 114:3119-e312 
  5. Bao et al. Nature. 2020. May 7
  6. Sun et al. Cell. 2020
  7. Zhao et al. Proc Natl Acad Sci U S A. 2014. 111: 4970-4975
  8. Crystal et al. Nat Genet. 1994. 8: 42-51
  9. Nabel, G.J. Nat Med. 2004. 10: 135-141
  10. Chen et al. J Clin Invect. 2019. 130: 2620-2629
  11. Meng-Yuan et al. Infect Dis Poverty. 2020. 9(1):45
  12. Hassan et al. Cell. 2020. 182: 1–10

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