Establishment of a humanized ST6GAL1 mouse model for influenza research

Abstract

Background: This study aimed to construct and characterize a humanized influenza mouse model expressing hST6GAL1.

Methods: Humanized fragments, consisting of the endothelial cell-specific K18 promoter, human ST6GAL1-encoding gene, and luciferase gene, were microinjected into the fertilized eggs of mice. The manipulated embryos were transferred into the oviducts of pseudopregnant female mice. The offspring were identified using PCR. Mice exhibiting elevated expression of the hST6GAL1 gene were selectively bred for propagation, and in vivo analysis was performed for screening. Expression of the humanized gene was tested by performing immunohistochemical (IHC) analysis. Hematologic and biochemical analyses using the whole blood and serum of humanized hST6GAL1 mice were performed.

Results: Successful integration of the human ST6GAL1 gene into the mouse genome led to the overexpression of human SiaT ST6GAL1. Seven mice were identified as carrying copies of the humanized gene, and the in vivo analysis indicated that hST6GAL1 gene expression in positive mice mirrored influenza virus infection characteristics. The IHC results revealed that hST6GAL1 was expressed in the lungs of humanized mice. Moreover, the hematologic and biochemical parameters of the positive mice were within the normal range.

Conclusion: A humanized influenza mouse model expressing the hST6GAL1 gene was successfully established and characterized.

Keywords

hST6GAL1 / humanized mice / influenza animal model

References

[1] Glezen WP. Emerging infections: pandemic influenza. Epidemiol Rev. 1996;18(1):64-76.

[2] Dawood FS, Iuliano AD, Reed C, et al. Estimated global mortality associated with the first 12 months of 2009 pandemic influenza a H1N1 virus circulation: a modelling study. Lancet Infect Dis. 2012;12(9):687-695.

[3] Organization, W.H. Influenza (Seasonal). 2023. Available from https://www.who.int/news-room/fact-sheets/detail/influenza-(seasonal)

[4] Liu WJ, Wu Y, Bi Y, et al. Emerging HxNy Influenza A Viruses. Cold Spring Harb Perspect Med. 2022;12(2):a038406.

[5] Nelli RK, Kuchipudi SV, White GA, Perez BB, Dunham SP, Chang KC. Comparative distribution of human and avian type sialic acid influenza receptors in the pig. BMC Vet Res. 2010;6:4.

[6] Nicholls JM, Bourne AJ, Chen H, Guan Y, Peiris JSM. Sialic acid receptor detection in the human respiratory tract: evidence for widespread distribution of potential binding sites for human and avian influenza viruses. Respir Res. 2007;8(1):73.

[7] Abdelwhab EM, Hafez HM. An overview of the epidemic of highly pathogenic H5N1 avian influenza virus in Egypt: epidemiology and control challenges. Epidemiol Infect. 2011;139(5):647-657.

[8] Murray J, Martin DE, Hosking S, Orr-Burks N, Hogan RJ, Tripp RA. Probenecid inhibits influenza A(H5N1) and A(H7N9) viruses in vitro and in mice. Viruses. 2024;16(1):152.

[9] Lei Y, Sun Y, Wu W, et al. Influenza H7N9 virus disrupts the monolayer human brain microvascular endothelial cells barrier in vitro. Virol J. 2023;20(1):219.

[10] Wilson JL, Akin E, Zhou R, et al. The influenza B virus victoria and yamagata lineages display distinct cell tropism and infection induced host gene expression in human nasal epithelial cell cultures. Viruses. 2023;15(9):1956.

[11] Rosu ME, Lexmond P, Bestebroer TM, et al. Substitutions near the HA receptor binding site explain the origin and major antigenic change of the B/Victoria and B/Yamagata lineages. Proc Natl Acad Sci USA. 2022;119(42):e2211616119.

[12] Ibricevic A, Pekosz A, Walter MJ, et al. Influenza virus receptor specificity and cell tropism in mouse and human airway epithelial cells. J Virol. 2006;80(15):7469-7480.

[13] Lehoux S, Groux-Degroote S, Cazet A, et al. Transcriptional regulation of the human ST6GAL2 gene in cerebral cortex and neuronal cells. Glycoconj J. 2010;27(1):99-114.

[14] Li N, Qi Y, Zhang FY, et al. Overexpression of alpha-2, 6 sialyltransferase stimulates propagation of human influenza viruses in Vero cells. Acta Virol. 2011;55(2):147-153.

[15] Wu D, Huang W, Wang Y, et al. Gene silencing of β-galactosamide α-2, 6-sialyltransferase 1 inhibits human influenza virus infection of airway epithelial cells. BMC Microbiol. 2014;14:78.

[16] Sambrook J, Fritsch EF, Maniatis T. Molecular cloning: a laboratory manual. Cold spring harbor laboratory press; 1989.

[17] Ausubel FM et al. Short protocols in molecular biology. N Y. 1992;275:28764-28773.

[18] Chow YH, O’Brodovich H, Plumb J, et al. Development of an epithelium-specific expression cassette with human DNA regulatory elements for transgene expression in lung airways. Proc Natl Acad Sci USA. 1997;94(26):14695-14700.

[19] Rall GF, Mucke L, Oldstone MB. Consequences of cytotoxic T lymphocyte interaction with major histocompatibility complex class I-expressing neurons in vivo. J Exp Med. 1995;182(5):1201-1212.

[20] Hogan B, Constantini F, Lacy E. Manipulating the Mouse Genome. Cold Spring Harbor Laboratory Press; 1986.

[21] Mucke L, Oldstone MB, Morris JC, Nerenberg MI. Rapid activation of astrocyte-specific expression of GFAP-lacZ transgene by focal injury. New Biol. 1991;3(5):465-474.

[22] Oldstone MB, Nerenberg M, Southern P, Price J, Lewicki H. Virus infection triggers insulin-dependent diabetes mellitus in a transgenic model: role of anti-self (virus) immune response. Cell. 1991;65(2):319-331.

[23] Chen M, Li S, Liu S, et al. Infection of SARS-CoV-2 causes severe pathological changes in mouse testis. J Genet Genomics. 2023;50(2):99-107.

[24] Boehm O, Zur B, Koch A, et al. Clinical chemistry reference database for Wistar rats and C57/BL6 mice. Biol Chem. 2007;388(5):547-554.

[25] Palani S, Uddin MB, McKelvey M, Shao S, Sun K. Immune predisposition drives susceptibility to pneumococcal pneumonia after mild influenza a virus infection in mice. Front Immunol. 2023;14:1272920.

[26] Jayaraman A, Chandrasekaran A, Viswanathan K, Raman R, Fox JG, Sasisekharan R. Decoding the distribution of glycan receptors for human-adapted influenza a viruses in ferret respiratory tract. PLoS One. 2012;7(2):e27517.

[27] Belser JA, Eckert AM, Tumpey TM, Maines TR. Complexities in ferret influenza virus pathogenesis and transmission models. Microbiol Mol Biol Rev. 2016;80(3):733-744.

[28] Canning BJ, Chou Y. Using Guinea pigs in studies relevant to asthma and COPD. Pulm Pharmacol Ther. 2008;21(5):702-720.

[29] Lai F, Wee CYY, Chen Q. Establishment of humanized mice for the study of HBV. Front Immunol. 2021;12:638447.

[30] Yu CI, Gallegos M, Marches F, et al. Broad influenza-specific CD8+T-cell responses in humanized mice vaccinated with influenza virus vaccines. Blood. 2008;112(9):3671-3678.

[31] Wada Y, Nithichanon A, Nobusawa E, et al. A humanized mouse model identifies key amino acids for low immunogenicity of H7N9 vaccines. Sci Rep. 2017;7(1):1283.

[32] Wang M, Hou S, Lu X, Li J, Li R, Yan X. Interleukin-37 inhibits inflammation activation and disease severity of PM2.5-induced airway hyperresponsiveness. Ecotoxicol Environ Saf. 2021;227:112890.

[33] Perl AK et al. Conditional recombination reveals distinct subsets of epithelial cells in trachea, bronchi, and alveoli. Am J Respir Cell Mol Biol. 2005;33(5):455-462.

RIGHTS & PERMISSIONS

2024 The Author(s). Animal Models and Experimental Medicine published by John Wiley & Sons Australia, Ltd on behalf of The Chinese Association for Laboratory Animal Sciences.