ACE2 Mouse
Description
Overview of ACE2 Mouse Models
ACE2 mice are designed to overcome the natural resistance of wild-type mice to SARS-CoV-2, which binds poorly to murine ACE2. By introducing human ACE2 via transgenic or knock-in strategies, researchers create platforms for studying:
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Viral tropism and replication dynamics
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Immune response mechanisms
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Therapeutic and vaccine efficacy
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Long-term sequelae of infection (e.g., long COVID)
Table 1: Comparison of ACE2 Mouse Models
Table 2: Infection Outcomes Across Models
Immunopathological Insights
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K18-hACE2: Exhibits cytokine storms (IL-6, TNF-α ↑ 10–100×) and macrophage-dominated lung infiltration .
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ACE2-GR: Generates neutralizing antibodies without severe inflammation, enabling vaccine efficacy studies .
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hACE2ki: Shows persistent tau protein accumulation in brains post-recovery, linking SARS-CoV-2 to neurodegenerative pathways .
Applications in COVID-19 Research
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Therapeutic Testing: CAG-hACE2 demonstrated efficacy of CD24-conjugated antibodies in reducing mortality .
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Vaccine Development: K18-hACE2 validated spike protein-based vaccines through survival rate improvements .
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Long COVID Modeling: hACE2ki revealed ACE2 downregulation correlates with prolonged pulmonary and neurological sequelae .
Limitations and Considerations
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Overexpression Artifacts: K18-hACE2 and CAG-hACE2 exhibit non-physiological ACE2 levels, exaggerating neurotropism .
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Age Dependency: hACE2ki mortality rates increase in aged cohorts, requiring age-stratified studies .
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Variant Specificity: Omicron shows reduced virulence in ACE2-GR compared to Delta .
Product Specs
Angiotensin-converting enzyme 2 (ACE2) is an enzyme found on the surface of cells in various organs, including the intestines, arteries, lungs, heart, and kidneys. ACE2 serves as an entry point for SARS coronaviruses, including SARS-CoV-2. The spike (S) glycoprotein of the coronavirus is a key factor in viral infection. It's a class I viral fusion antigen located on the virion's outer envelope. The S protein identifies and binds to host cell receptors, facilitating the fusion of viral and cellular membranes, thus enabling infection. The S1 domain of the coronavirus S protein has two main parts: the N-terminal domain and the C-terminal domain. Either or both of these domains can act as receptor-binding domains. SARS-CoV and MERS-CoV both use their C-domain to bind to their respective receptors. ACE2 is a transmembrane protein with its N-terminal domain, containing the catalytic site, located outside the cell and its C-terminal tail located inside the cell. ACE2 acts as a mono-carboxypeptidase, an enzyme that cleaves peptide bonds. Specifically, it degrades Angiotensin I to produce Angiotensin 1-9 and Angiotensin II to produce Angiotensin 1-7.
ACE2 Mouse, produced in Sf9 insect cells using a baculovirus expression system, is a single, glycosylated polypeptide chain. It comprises 731 amino acids (specifically, amino acids 18-740) and has a molecular weight of 84.5 kDa. The protein includes a 6-amino acid Histidine tag (His-Tag) attached to its C-terminus to facilitate purification. It is purified using proprietary chromatographic techniques.
The ACE2 solution is buffered in Phosphate-Buffered Saline (pH 7.4) and contains 10% Glycerol as a stabilizing agent.
For short-term storage (up to 2-4 weeks), the ACE2 solution can be stored at 4°C. For extended storage, it is recommended to freeze the solution at -20°C. To further enhance stability during long-term storage, adding a carrier protein such as Albumin (0.1% HSA or BSA) is advised. Repeated freezing and thawing of the solution should be avoided.
The purity of the ACE2 protein is greater than 95%, as determined by SDS-PAGE analysis.
The biological activity of ACE2 is defined as its enzymatic ability to hydrolyze the substrate McaYVADAPK(Dnp)-OH. This ACE2 Mouse product exhibits an activity greater than 200 pmol/min/µg, meaning that 1 µg of the enzyme can hydrolyze more than 200 picomoles of the substrate per minute at a pH of 7.5 and a temperature of 25°C.
ACE2, 2010305L05Rik, Angiotensin I Converting Enzyme, Angiotensin I Converting, Enzyme (Peptidyl-Dipeptidase A), Angiotensin-Converting Enzyme Homolog, Angiotensin-Converting Enzyme, ACE-Related Carboxypeptidase, Metalloprotease MPROT15, Peptidyl-Dipeptidase A, ACEH, EC 3.4.17.23, EC 3.4.17.
QSLTEENAKT FLNNFNQEAE DLSYQSSLAS WNYNTNITEE NAQKMSEAAA KWSAFYEEQS KTAQSFSLQE IQTPIIKRQL QALQQSGSSA LSADKNKQLN TILNTMSTIY STGKVCNPKN PQECLLLEPG LDEIMATSTD YNSRLWAWEG WRAEVGKQLR PLYEEYVVLK NEMARANNYN DYGDYWRGDY EAEGADGYNY NRNQLIEDVE RTFAEIKPLY EHLHAYVRRK LMDTYPSYIS PTGCLPAHLL GDMWGRFWTN LYPLTVPFAQ KPNIDVTDAM MNQGWDAERI FQEAEKFFVS VGLPHMTQGF WANSMLTEPA DGRKVVCHPT AWDLGHGDFR IKMCTKVTMD NFLTAHHEMG HIQYDMAYAR QPFLLRNGAN EGFHEAVGEI MSLSAATPKH LKSIGLLPSD FQEDSETEIN FLLKQALTIV GTLPFTYMLE KWRWMVFRGE IPKEQWMKKW WEMKREIVGV VEPLPHDETY CDPASLFHVS NDYSFIRYYT RTIYQFQFQE ALCQAAKYNG SLHKCDISNS TEAGQKLLKM LSLGNSEPWT KALENVVGAR NMDVKPLLNY FQPLFDWLKE QNRNSFVGWN TEWSPYADQS IKVRISLKSA LGANAYEWTN NEMFLFRSSV AYAMRKYFSI IKNQTVPFLE EDVRVSDLKP RVSFYFFVTS PQNVSDVIPR SEVEDAIRMS RGRINDVFGL NDNSLEFLGI HPTLEPPYQPPVTLEHHHHH H.
Q&A
What is the fundamental limitation of using standard mice for SARS-CoV-2 research?
Wild-type mice are not susceptible to SARS-CoV-2 infection due to the low affinity of mouse ACE2 for the viral spike protein. In mouse ACE2, seven of the S-ACE2 interface residues differ from human ACE2, with most changed to uncharged polar amino acids, resulting in significantly lower binding affinity compared to human, gorilla, and monkey ACE2 . This structural incompatibility necessitates the development of specialized mouse models expressing human ACE2 to enable SARS-CoV-2 research in mice .
What are the major types of ACE2 mouse models available for SARS-CoV-2 research?
Several distinct ACE2 mouse models have been developed with different genetic modifications:
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K18-hACE2 transgenic mice: Express human ACE2 cDNA under the control of the keratin 18 (K18) promoter. These mice show severe disease with high mortality and marked neurodissemination following SARS-CoV-2 infection .
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ACE2 Gene Replacement (ACE2-GR) mice: The entire mouse Ace2 genomic locus is replaced with the human ACE2 gene locus. These mice show mild disease without CNS involvement .
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Humanized ACE2 knockin (hACE2ki) mice: Express human ACE2 in tissue and cell-specific patterns similar to endogenous mouse Ace2 .
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Double-transgenic mice: Express both human ACE2 and TMPRSS2, showing increased viral infectivity and more severe disease manifestations .
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Optimized codon hACE2 (opt-hACE2) mice: Feature codon-optimized human ACE2 for improved translation efficiency, resulting in enhanced expression .
How does human ACE2 expression affect SARS-CoV-2 pathogenesis in different mouse models?
The pattern, level, and regulation of human ACE2 expression significantly impact disease manifestation:
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K18-hACE2 mice: Show aberrant expression of hACE2 in the neuroepithelium, leading to viral neuroinvasion and rapid death, which is not typical of most human COVID-19 cases .
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ACE2-GR mice: Display physiological expression patterns of human ACE2, resulting in milder disease without CNS involvement, more closely resembling asymptomatic or mild human infections .
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Double-transgenic mice (hACE2+TMPRSS2): Co-expression of both receptors increases viral infectivity both in vitro and in vivo, leading to significant weight loss, clinical symptoms, acute lung injury, and lethality .
How should researchers select the appropriate ACE2 mouse model for their specific research question?
Selection should be based on the research objectives:
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For studying severe COVID-19 pathology and testing therapeutic interventions against severe disease: K18-hACE2 or double-transgenic mice may be appropriate as they develop severe symptoms .
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For investigating mild or asymptomatic infection and immune responses: ACE2-GR mice are preferable as they develop antibody responses without severe disease .
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For long COVID/PASC research: hACE2ki mice have been specifically designed for this purpose, showing tau protein pathologies associated with Alzheimer's disease post-infection .
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For studying differential pathogenesis of SARS-CoV-2 variants: The double-transgenic model has demonstrated utility in comparing variant susceptibility and pathogenesis .
What are the optimal infection parameters for ACE2 mouse models?
Infection parameters vary by model and research question:
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Dose: For hACE2ki mice, a dose of 5 × 10^5 PFU/mouse via nasal instillation has been shown effective for studying variant effects without causing mortality in young adult mice .
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Age consideration: Age significantly affects disease severity. Young adult hACE2ki mice (6 weeks old) show no mortality, making them suitable for long-term studies .
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Infection route: Intranasal infection is standard across most models to mimic natural respiratory infection .
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Variant selection: Different SARS-CoV-2 variants (WA, Delta, Omicron) produce distinctive phenotypes in terms of viral load, weight loss, and inflammatory responses, allowing for comparative studies .
What analytical methods are essential for characterizing SARS-CoV-2 infection in ACE2 mouse models?
Comprehensive analysis should include:
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Viral load quantification: In key tissues including lung, trachea, nasal turbinate, and potentially brain (for K18-hACE2 models) .
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Immunohistochemistry: To assess tissue and cell-specific expression of human ACE2 and viral tropism .
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Cytokine profiling: Measurement of pro-inflammatory cytokines in bronchoalveolar lavage fluid to characterize immune responses .
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Immune cell phenotyping: Analysis of immune cell profiles in infected tissues to understand immunopathology .
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Weight monitoring and clinical scoring: To assess disease progression and severity .
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Antibody response measurement: Particularly important for models with mild disease to confirm infection .
How do ACE2-GR mice differ from K18-hACE2 transgenic mice in disease manifestation?
The differences are substantial and critical for research applications:
| Feature | ACE2-GR Mice | K18-hACE2 Mice |
|---|---|---|
| Disease severity | Mild disease | Severe disease |
| Mortality | Non-lethal | Rapid death |
| CNS involvement | Not detectable | Marked neurodissemination |
| Viral replication | Supported but limited | Extensive |
| Expression pattern | Physiological (under human ACE2 regulatory elements) | Non-physiological (under K18 promoter) |
| Antibody response | Reliable | May be limited by rapid mortality |
| Research application | Immune responses, long-term consequences | Severe COVID-19, therapeutic testing |
ACE2-GR mice express human ACE2 under the control of its native regulatory elements, leading to a more physiologically relevant expression pattern. In contrast, K18-hACE2 mice show aberrant expression in the neuroepithelium, contributing to the rapid death and neurodissemination not typically seen in human COVID-19 .
How does co-expression of TMPRSS2 with human ACE2 affect SARS-CoV-2 pathogenesis?
TMPRSS2 (transmembrane serine protease 2) plays a critical role in viral entry by cleaving the SARS-CoV-2 spike protein. In double-transgenic mice:
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Co-expression significantly increases viral infectivity both in vitro and in vivo compared to single-transgenic models .
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Double-transgenic mice show more pronounced clinical disease symptoms, including significant weight loss and acute lung injury .
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These mice demonstrate higher lethality following SARS-CoV-2 infection .
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The model is highly responsive to TMPRSS2 inhibitors like nafamostat, which effectively reduces virus-induced weight loss, viral replication, and mortality, making it valuable for antiviral drug testing .
What are the key differences between various human ACE2 expression strategies in mice?
Different genetic strategies for introducing human ACE2 yield distinct outcomes:
| Strategy | Key Characteristics | Expression Pattern | Research Value |
|---|---|---|---|
| K18 promoter-driven | hACE2 cDNA under K18 promoter | Aberrant expression, including in neuroepithelium | Severe disease model |
| Gene replacement | Entire human ACE2 locus replaces mouse Ace2 | Physiological, matches human tissue specificity | Mild disease, immune studies |
| Opt-hACE2 | Codon-optimized hACE2 | Enhanced expression in multiple tissues | More active immune responses, severe phenotypes |
| β-globin insertion | hACE2 with β-globin enhancer elements | Variable depending on element placement | Enhances transcription when placed downstream |
| Dual receptor expression | hACE2 + TMPRSS2 | Complementary expression enabling efficient viral entry | Highly susceptible model for therapeutic testing |
The positioning of regulatory elements like β-globin within the hACE2 cassette significantly influences expression levels, with downstream placement enhancing transcription. Additionally, codon optimization of hACE2 improves translation efficiency across multiple tissues .
How can ACE2 mouse models be utilized to study long-term consequences of SARS-CoV-2 infection?
Humanized ACE2 mouse models, particularly hACE2ki mice, provide valuable platforms for studying long COVID/PASC (Post-Acute Sequelae of SARS-CoV-2):
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These models enable the investigation of tau protein pathologies linked to Alzheimer's disease in the brains of mice post-infection .
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Researchers can study the accumulation and longitudinal propagation of tau protein, a key marker of neurodegeneration .
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The non-lethal nature of infection in certain models (like ACE2-GR and hACE2ki) allows for long-term follow-up studies that would be impossible in more severe models .
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These models facilitate examination of persistent immune dysregulation and organ damage that may contribute to long COVID manifestations .
How can ACE2 mouse models be used to compare pathogenesis of different SARS-CoV-2 variants?
These models are invaluable for comparative variant studies:
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Different variants (WA, Delta, Omicron) have been shown to produce distinctive phenotypes in hACE2ki mice, with variations in:
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Double-transgenic mice have demonstrated differential susceptibility and pathogenesis profiles between variants, making them useful for assessing emerging variant threats .
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The models allow for standardized comparison of variant virulence, immune evasion, and tissue tropism under controlled conditions .
What insights have ACE2 mouse models provided about the mechanisms of SARS-CoV-2 neuroinvasion?
These models have revealed important aspects of SARS-CoV-2 neuroinvasion:
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K18-hACE2 mice show marked neurodissemination of the virus, which correlates with aberrant expression of the hACE2 transgene in the neuroepithelium .
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In contrast, ACE2-GR mice show no detectable CNS involvement despite supporting viral replication in respiratory tissues, suggesting that physiological ACE2 expression patterns may limit neuroinvasion .
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hACE2ki mice exhibit tau protein pathologies in the brain following infection, even without direct evidence of extensive viral neuroinvasion, suggesting potential indirect mechanisms of neurological damage .
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These contrasting findings help differentiate between artifacts of mouse models and genuine pathogenic mechanisms relevant to human COVID-19 neurological complications .
What are the key limitations of current ACE2 mouse models for COVID-19 research?
Despite their utility, these models have several limitations:
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Species differences: Even humanized mice retain many murine-specific immune and physiological characteristics that may not perfectly recapitulate human responses to SARS-CoV-2 .
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Expression pattern artifacts: Some models (particularly K18-hACE2) show non-physiological expression patterns leading to atypically severe infections and aberrant sites of viral replication .
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Background strain effects: The genetic background (typically C57BL/6) may influence infection outcomes and immune responses .
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Age and sex differences: These variables significantly affect COVID-19 outcomes in humans but are not always adequately addressed in mouse studies .
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Limited chronic disease modeling: Most models either cause rapid death or mild disease, with few accurately capturing the spectrum of moderate disease severity seen in many human patients .
How can researchers verify proper human ACE2 expression in their mouse models?
Proper validation requires multi-level analysis:
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mRNA expression analysis: RT-PCR to confirm expression of human ACE2 transcripts in relevant tissues, including alternative spliced forms .
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Protein expression verification: Immunohistochemistry to assess tissue- and cell-specific expression patterns of human ACE2 protein .
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Functional validation: Confirmation of susceptibility to SARS-CoV-2 infection and viral replication .
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Comparative expression analysis: Comparison with endogenous mouse Ace2 expression patterns in wild-type mice when possible .
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Transcript variant analysis: Assessment of all relevant transcripts, including the long noncoding RNA transcript ACE2-DT, which was detected in ACE2-GR mice .
What strategies can enhance the translational value of findings from ACE2 mouse models?
To maximize translational relevance:
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Model selection: Choose models with physiological expression patterns (like ACE2-GR) for mechanistic studies, while more severe models may be appropriate for therapeutic testing .
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Multimodal analysis: Combine viral load quantification with histopathology, immunophenotyping, and functional assessments .
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Cross-model validation: Confirm key findings across different ACE2 mouse models to distinguish model-specific artifacts from genuine biological insights .
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Age and sex considerations: Include both male and female mice of different ages to better reflect human demographic variability .
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Variant comparison: Test multiple SARS-CoV-2 variants to assess the generalizability of findings .
Product Science Overview
Angiotensin Converting Enzyme 2 (ACE2) is a crucial enzyme in the renin-angiotensin system (RAS), which plays a significant role in regulating blood pressure, fluid balance, and systemic vascular resistance. ACE2 is a monocarboxypeptidase that converts angiotensin II (a potent vasoconstrictor) into angiotensin-(1-7), which has vasodilatory and anti-inflammatory properties .
ACE2 is a type I transmembrane protein composed of 805 amino acids. It has a single catalytic domain and is expressed in various tissues, including the lungs, heart, kidneys, and intestines . The enzyme’s primary function is to counterbalance the effects of angiotensin II by producing angiotensin-(1-7), thereby promoting vasodilation and reducing inflammation .
ACE2 gained significant attention during the COVID-19 pandemic as the functional receptor for the SARS-CoV-2 virus. The spike (S) glycoprotein of SARS-CoV-2 binds to the receptor-binding domain (RBD) of ACE2, facilitating viral entry into host cells . This interaction is crucial for the virus’s ability to infect and replicate within human cells, making ACE2 a potential target for therapeutic interventions .
Recombinant ACE2 (Mouse) is a laboratory-produced version of the mouse ACE2 enzyme. It is often used in research to study the enzyme’s structure, function, and role in various physiological and pathological processes. Recombinant ACE2 can be produced using various expression systems, including bacterial, yeast, and mammalian cells .
Recombinant ACE2 has shown promise as a therapeutic agent in several studies. It has been demonstrated to protect against acute lung injury induced by SARS-CoV and avian influenza . Additionally, recombinant ACE2 can inhibit SARS-CoV-2 proliferation in vitro, suggesting its potential use in treating COVID-19 . By restoring the balance between the RAS and the ACE2/angiotensin-(1-7)/MAS axis, recombinant ACE2 may help mitigate organ injuries associated with viral infections .
