Science / Health

Recent Advances in Mouse Models for the Study of Human Coronavirus Infectious Diseases

The hosts involved in the pathogenesis of human coronaviruses (hCoVs) include camels, reptiles, birds, bats, rodents, pigs, dogs, cattle, and cats. Seven coronaviruses have been identified to date including OC43, NL-63, MERS-CoV, SARS-CoV, and 2019-nCoV. The 2019-nCoV is responsible for the COVID-19 global pandemic. SARS-CoV and MERS-CoV have previously caused global epidemics associated with severe respiratory distress syndrome (SARS), with civets and camels serving as intermediate hosts.

The hCoVs genome comprises 16 non-structural and 4 structural protein genes. These structural genes encode for structural proteins that include nucleocapsid protein (N), spike protein (S), membrane protein (M), and envelope protein (E). Interaction of surface proteins on host cells and the viral proteins is integral to viral invasion into the host cell and subsequent viral replication. The receptor-binding domain (RBD) of SARS-CoV-2, located in the S protein, interacts with the human ACE2 (hACE2); on the contrary, DPP4 serves as a host cell receptor for MERS-CoV.

The development of protective antibodies and vaccines is attributed to the RBD for S protein as therapeutic targets. Other structural, non-structural, and enzymatically active proteins can also function as therapeutic targets for drugs, vaccines, and antibodies against hCoVs.

The clinical features of infections caused by hCoVs include diffuse alveolar injury accompanied by hyaline membrane formation, cellular debris, alveolar fibrous deposition, hemorrhage, edema, interstitial inflammation, alveolar inflammation, as well as lung tissue necrosis. An accurate mouse model for the SARS-CoV-2 study must demonstrate similar pathologic changes as observed in SARS-CoV-2 patients.

What Are the Strategies and Methods for Studying the Pathogenicity of SARS-Cov-2 Infection?

Three types of infectious mouse models of SARS-CoV-2 include the following:

  1. Inbred mice directly infected with the human SARS-CoV virus
  2. The use of gene editing mouse technology for transferring host cell binding receptors and mouse-related genes into the mice
  3. A mouse model was subjected to transfer of wild-type SARS-CoV virus to acquire pathogenic mouse-adapted viruses.

1. Direct Application of Inbred Mouse Models to Study Viral Pathogenicity

These inbred mouse models are subjected to early SARS-CoV studies. For example, young mice of 4-6 weeks were nasally infected with Urbani SARS-CoV strain which demonstrated that the virus replicates in the upper or lower respiratory tract. The replication peaked on the 3rd day while it resolved on the 7th day with no pathogenic effects. This also revealed that the SARS-CoV infection elicits a humoral immune response that leads to the development of resistance to viral replication. However, owing to the lack of pathogenic effects, it is difficult to evaluate the effectiveness of therapeutic interventions in this model.

12-14 months old mice models were subjected to assess the course of disease in elderly patients suffering from SARS-CoV infection. In these mice models, histopathologic manifestations include inflammatory cell infiltration, interstitial pneumonia, and fine bronchial cell necrosis persisting up to the 9th day of infection. Increased severity of symptoms in older mice as compared to the younger mice demonstrate the association between age and genetic composition of the host with SARS-CoV pathogenicity.  

2. Application of Gene-Editing Technology to Establish a Mouse Model of Viral Infection Disease

Wild and immunodeficient mouse models are relatively less susceptible to human SARS-CoV infection and death, indicating the need for the establishment of SARS-CoV susceptible mouse models by using gene-editing technology. The immunodeficient mice demonstrated weight loss and severe lung disease at the onset of SARS-CoV infection, with the viral replication persisting for up to 22 days. These mouse models’ pathologic features are somewhat similar to human SARS-CoV infection. On the contrary, it demonstrated lower viral replication in the respiratory tract as compared to gene-edited mouse models.

The RBD present on the S protein binds to the hACE2 receptors found on human host cells. This receptor is encoded by genes on the X chromosome. Gene-edited mice that express hACE2 receptors are susceptible to SARS-CoV infection with pathologic changes in the respiratory tract. The magnitude of hACE2 expression is proportional to the severity of the disease. Brain infection was observed in these mice models, resulting in the onset of encephalitis in the mice. 

3. Application of an Adaptive Experimental Evolutionary Approach to Obtain SARS-CoV Mutant Strong Strains

The wild mice were inoculated with SARS-CoV strains 15 times to assess the adaptive evolution of the SARS-CoV virus. This adapted virus, also called the MA15 virus strain, was capable of causing 100% lethality in all age groups of mice and demonstrated mutated amino acids. This virus strain leads to the onset of rapid weight loss, neutrophilia, lymphocytopenia, and pneumonia.

4. Establishment of Genetically Diverse Mouse Strains Using Colaboratory Cross (CC) Technology

The CC mouse strains can determine the genetic basis of host immune response to the SARS-CoV virus in the course of SARS-CoV pathogenesis. This analysis’s primary purpose is to determine the genes related to phenotypic characteristics rapidly. In addition, the CC mouse technology has led to the identification of CC mouse strains with opposite viral susceptibility along with gene localization analysis to identify the disease phenotype.

What Are the Strategies and Methods for Applying Mouse Models to Study MERS-CoV Pathogenicity? 

The wild mouse strains that used to be susceptible to the SARS-CoV virus were not susceptible MERS-CoV virus, unlike non-human primate animals that are susceptible to MERS-CoV infection. Similar to wild mouse strains, immunodeficient mice are not susceptible to the MERS-CoV virus, indicating that human response limitation and viral infection may not have a direct relationship with each other. However, the DPP4 receptor on host cells does not bind to the S protein of the MERS-CoV virus. This is the underlying mechanism of reduced susceptibility of the mice models to MERS-CoV virus infection.

1. Gene Editing DPP4 Humanized Mouse Model

The DPP4 humanized mouse model with human DPP4 receptors, using gene-editing techniques, is susceptible to MERS-CoV infection and replication. The clinical features of MERS-CoV-2 infection are relatively more significant in immunodeficient and older mice. Such mice demonstrated a high viral load in the lungs and other organs, including the brain, with severe symptoms and even death. While replacing the mouse DPP4 genome with the human one, the 5′ endogenous promoter region was retained. In these mice, moderate signs and symptoms of the disease were demonstrated with no brain damage and minimum extra-pulmonary viral invasion. DPP4 mutation exhibit a high viral load with no clinical disease symptoms. To mimic the pathogenic features of human MERS-CoV virus infection, the mice models must express full human DPP4 receptors and associated genes.

2. Adaptive Evolution of MERS-CoV Mice to Obtain a Strong Virulent Strain of Virus

Infection of mice with mutant DPP4 with adaptive MERS-CoV virus strain leads to loss of function of the respiratory tract, severe acute respiratory distress syndrome, and even death. Vaccines targeting MERS-CoV S protein can prevent respiratory dysfunction and death in these mice models.

3. CC Mouse Model of MERS-CoV Infection

The ideal mouse model to study the MERS-CoV virus infection is the one that involves infection of the mice models with strains that are not genetically modified and cause respiratory symptoms similar to the ones that occur in humans. This demonstrates that the CC mice models are not susceptible to MERS-CoV virus infection.

How to Apply the Human Coronavirus Mouse Model to Evaluate the Effect of Antiviral Therapy?

The primary goal of a mouse model is to identify the pathogenesis and efficacy of prevention and therapeutic strategies for the disease. At present, there are no specific FDA-approved vaccines and drugs for the treatment and prevention of SARS-CoV and MERS-CoV. The broad-spectrum antiviral medications are also ineffective against these viruses and may even aggravate the severity of the disease. Raltegravir is an anti-hCoV drug that alleviates the disease symptoms if administered 48 hours post-infection. 

How to Apply a Mouse Model to Study SARS-CoV-2 Infection in Neocoronavirus Pneumonia?

SARS-CoV-2 has characteristics similar to that of the SARS-CoV virus, indicating that the mouse models established for both viral infections are also similar. The severity of the disease and death is attributed to brain and lung damage in COVID-19 patients.

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