SARS-CoV-2 infection in K18-ACE2 transgenic mice replicates human pulmonary disease in COVID-19

Over the last several months, a global pandemic has developed due to the emergence of the novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which is capable of infecting

humans. The lack of preexisting immunity to SARS-CoV-2 in humans has prompted the scientific community to strive to understand the mechanisms underlying SARS-CoV-2 pathogenesis and to test

new therapeutic agents and vaccines for the clinical management of coronavirus disease 2019 (COVID-19). In these efforts, the development of appropriate animal models of the disease is one

of the most valuable strategies.

Unfortunately, although several animal models of COVID-19 have been investigated, only a few (hamsters, ferrets, minks, cats, and nonhuman primates) have been found to be susceptible to the

disease.1 In all cases, the critical issue appears to be the specificity of the viral tropism for human angiotensin-converting enzyme 2 (hACE2), the key factor in the entry of the virus into

human cells. Mice are commonly used to study emerging viruses but do not provide a suitable model for SARS-CoV-2 infection because mouse ACE2 (mACE2) is not an appropriate receptor for

SARS-CoV-2 and does not facilitate the entry of the virus into the cells. Therefore, the use of mouse models for COVID-19 is limited to genetically modified mice (or, alternatively,

engineered viruses that effectively bind mACE2).

To date, several genetically modified mouse models have been developed, in which hACE2 is permanently expressed under the regulation of either global or specific promoters.2,3,4 Winkler et

al.5 recently reported the use of transgenic mice expressing hACE2 driven by the cytokeratin-18 (K18) gene promoter (K18-hACE2), a model that was originally developed for the severe acute

respiratory syndrome (SARS),3 to investigate the effects of SARS-CoV-2 infection (Fig. 1). The study showed the presence of high levels of viral infection in the lungs following intranasal

inoculation with 1 × 102 plaque-forming units (p.f.u.) in both male and female mice. These lung infections shared many features with severe COVID-19 in humans, including the presence of an

inflammatory response characterized by high levels of proinflammatory cytokines and chemokines; a cellular infiltrate composed mainly of monocytes, neutrophils, and T cells; and perivascular

inflammation. Since mild pulmonary disease is the characteristic result of SARS-CoV-2 infection in naturally susceptible animals, the severity of disease observed following SARS-CoV-2

infection in K18-hACE2 mice provides a useful model in which to investigate pulmonary disease and test antiviral countermeasures in this species.

Overexpression of ACE2 in the lung epithelium facilitates SARS-CoV-2 infection in mice

However, there are also some caveats to this model that are mainly related to the fact that, despite the fact that viral spread to other organs such as heart, spleen, and kidney has been

detected, extrapulmonary manifestations are usually mild in these animals and, therefore, do not completely mimic the severe clinical symptoms observed in humans. Thus, evidence of

thrombosis and vasculitis is present in K18-hACE2 mice with severe pneumonia, but there is no clear proof of widespread thrombosis with microangiopathy and coagulopathy, as have been

observed in many severe cases in humans.6 Similarly, although some mice develop anosmia soon after infection, neurological involvement is less severe in these mice than in humans.7 These

differences between human and mouse findings likely reflect the fact that the expression of hACE2 is driven by a nonnative promotor, resulting in a nonphysiological tissue distribution

pattern of ACE2 expression. In fact, while hACE2 expression is mainly limited to lung epithelia in K18-hACE2 mice, the protein is expressed in the vascular endothelia, renal and

cardiovascular tissue, and epithelia of the small intestine, testes, and lung in humans.8 Therefore, the presence of high levels of hACE in the pulmonary epithelia of K18-hACE2 mice may

account for the observed severity of pulmonary manifestations, but the lack of expression in other tissues may prevent a direct effect of the virus that may be present in humans with

COVID-19.

Second, unlike ACE, which converts angiotensin I (Ang I) to angiotensin II (Ang II), ACE2 promotes the formation of Ang-1-9.9 In contrast to Ang I, which induces vasoconstriction and

promotes fibrosis and inflammation, Ang-1-9 has vasodilatory, antifibrotic, and anti-inflammatory effects. Interestingly, ACE2 activity is reduced by either downregulation or shedding upon

SARS-CoV-2 infection,10 which likely contributes to the severity of the disease in humans. As there is no evidence of the existence of reduced mACE2 expression in K18-hACE2 mice, it is

tempting to speculate that the level of Ang-1-9, and therefore its protective effects, are maintained, thus avoiding the development of some severe symptoms that may be present in humans.

Finally, as already acknowledged by Wilken et al., some of the more relevant risk factors for developing severe COVID-19 in humans, such as female sex, old age, and comorbid cardiovascular

disease or diabetes, are not present in K18-hACE2 mice, likely preventing some of the adverse clinical outcomes observed in humans.

In summary, the animal model proposed for COVID-19 mimics many of the clinical features of the disease, making it one of the best models available. A future comprehensive model will include

additional aspects, such as the vasculopathy and coagulopathy associated with SARS-CoV-2 infection in humans or the effects of different chronic diseases, advanced age, or male sex, which

put patients at greater risk of adverse outcomes after viral infection.

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