Background: Humanized mouse models are essential for studying the human immune response and antibody development. However, conventional models show limited B cell maturation and antigen-specific humoral responses. To overcome these limitations, we used the NOG-EXL mice expressing human interleukin 3 (IL-3) and granulocyte-macrophage colony-stimulating factor (GM-CSF) to enhance myeloid and B-cell lineage differentiation.
Methods: Human CD34+ hematopoietic stem cells (HSC) were transplanted into NOG-EXL mice to produce humanized immune systems. After immune cell reconstitution was confirmed across 12 weeks, the mice were immunized twice with inactivated severe fever with thrombocytopenia syndrome virus (SFTSV) antigens. Peripheral blood mononuclear cells and splenocytes were analyzed using multicolor flow cytometry to assess human immune cell subsets. Antigen-specific immunoglobulin G (IgG) production was quantified using enzyme-linked immunosorbent assay (ELISA), and virus-specific B cells were isolated using antigen-labeled recombinant protein probes.
Results: Twelve weeks after transplantation of HSCs into NOG-EXL mice, they exhibited robust engraftment of human leukocytes, including T, B, and dendritic cells, compared to NOG mice. Unlike NOG mice, humanized NOG-EXL mice exhibited an increase in human IgG levels, indicating the production of human antibody responses to antigens. Humanized NOG-EXL mice were immunized twice every 2 weeks with inactivated SFTSV, and antigen-specific human antibodies against the virus were detected in the mouse sera by ELISA. Sera from SFTSV-immunized humanized mice demonstrated neutralizing activity against SFTSV, confirming the induction of functional virus-specific neutralizing antibodies. Antigen-binding IgG-positive human B cells were isolated from mouse splenocytes using recombinant protein probes.
Conclusion: This model provides a valuable platform for evaluating humoral immunity and isolating B cells using high-affinity human monoclonal antibodies without genetic engineering.
| [1] |
Winiger RR, Perez L. Therapeutic antibodies and alternative formats against SARS-CoV-2. Antivir Res. 2024; 223:105820.
|
| [2] |
Jorgensen SCJ. Nirsevimab: review of pharmacology, antiviral activity and emerging clinical experience for respiratory syncytial virus infection in infants. J Antimicrob Chemother. 2023; 78(5): 1143-1149.
|
| [3] |
Passariello M, Ferrucci V, Sasso E, et al. A novel human neutralizing mAb Recognizes Delta, gamma and omicron variants of SARS-CoV-2 and can Be used in combination with Sotrovimab. Int J Mol Sci. 2022; 23(10):5556.
|
| [4] |
Shultz LD, Ishikawa F, Greiner DL. Humanized mice in translational biomedical research. Nat Rev Immunol. 2007; 7(2): 118-130.
|
| [5] |
Rongvaux A, Willinger T, Martinek J, et al. Development and function of human innate immune cells in a humanized mouse model. Nat Biotechnol. 2014; 32(4): 364-372.
|
| [6] |
Mian SA, Anjos-Afonso F, Bonnet D. Advances in human immune system mouse models for studying human hematopoiesis and cancer immunotherapy. Front Immunol. 2020; 11:619236.
|
| [7] |
Ito R, Takahashi T, Ito M. Humanized mouse models: application to human diseases. J Cell Physiol. 2018; 233(5): 3723-3728.
|
| [8] |
Becker PD, Legrand N, van Geelen CM, et al. Generation of human antigen-specific monoclonal IgM antibodies using vaccinated “human immune system” mice. PLoS One. 2010; 5(10):e13137.
|
| [9] |
Wege AK, Schmidt M, Ueberham E, et al. Co-transplantation of human hematopoietic stem cells and human breast cancer cells in NSG mice: a novel approach to generate tumor cell specific human antibodies. MAbs. 2014; 6(4): 968-977.
|
| [10] |
Yu H, Borsotti C, Schickel JN, et al. A novel humanized mouse model with significant improvement of class-switched, antigen-specific antibody production. Blood. 2017; 129(8): 959-969.
|
| [11] |
Moldenhauer LM, Keenihan SN, Hayball JD, Robertson SA. GM-CSF is an essential regulator of T cell activation competence in uterine dendritic cells during early pregnancy in mice. J Immunol. 2010; 185(11): 7085-7096.
|
| [12] |
Renner K, Metz S, Metzger AM, et al. Expression of IL-3 receptors and impact of IL-3 on human T and B cells. Cell Immunol. 2018; 334: 49-60.
|
| [13] |
Tadmori W, Feingersh D, Clark SC, Choi YS. Human recombinant IL-3 stimulates B cell differentiation. J Immunol. 1989; 142(6): 1950-1955.
|
| [14] |
Xia X, Li L, Choi YS. Human recombinant IL-3 is a growth factor for normal B cells. J Immunol. 1992; 148(2): 491-497.
|
| [15] |
Ito R, Takahashi T, Katano I, et al. Establishment of a human allergy model using human IL-3/GM-CSF-transgenic NOG mice. J Immunol. 2013; 191(6): 2890-2899.
|
| [16] |
Zhang YZ, Zhou DJ, Qin XC, et al. The ecology, genetic diversity, and phylogeny of Huaiyangshan virus in China. J Virol. 2012; 86(5): 2864-2868.
|
| [17] |
Cui H, Shen S, Chen L, et al. Global epidemiology of severe fever with thrombocytopenia syndrome virus in human and animals: a systematic review and meta-analysis. Lancet Reg Health West Pac. 2024; 48:101133.
|
| [18] |
Yun SM, Park SJ, Park SW, et al. Molecular genomic characterization of tick- and human-derived severe fever with thrombocytopenia syndrome virus isolates from South Korea. PLoS Negl Trop Dis. 2017; 11(9):e0005893.
|
| [19] |
Kim KH, Kim J, Ko M, et al. An anti-Gn glycoprotein antibody from a convalescent patient potently inhibits the infection of severe fever with thrombocytopenia syndrome virus. PLoS Pathog. 2019; 15(2):e1007375.
|
| [20] |
Sokullu E, Gauthier MS, Coulombe B. Discovery of antivirals using phage display. Viruses. 2021; 13(6):1120.
|
| [21] |
Nogales-Gadea G, Saxena A, Hoffmann C, et al. Generation of recombinant human IgG monoclonal antibodies from immortalized sorted B cells. J Vis Exp. 2015; 100:e52830.
|
| [22] |
Liu H, Saxena A, Sidhu SS, Wu D. Fc engineering for developing therapeutic bispecific antibodies and novel scaffolds. Front Immunol. 2017; 8:38.
|
| [23] |
Saxena A, Wu D. Advances in therapeutic fc engineering - modulation of IgG-associated effector functions and serum half-life. Front Immunol. 2016; 7:580.
|
| [24] |
Lebman DA, Coffman RL. Interleukin 4 causes isotype switching to IgE in T cell-stimulated clonal B cell cultures. J Exp Med. 1988; 168(3): 853-862.
|
| [25] |
Michée-Cospolite M, Boudigou M, Grasseau A, et al. Molecular mechanisms driving IL-10- producing B cells functions: STAT3 and c-MAF as underestimated central key regulators? Front Immunol. 2022; 13:818814.
|
RIGHTS & PERMISSIONS
2026 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.
PDF
(2358KB)
