Creation of “Humanized” Mice to Study Human Immunity

Diabetes Division, University of Massachusetts Medical School, Worcester, Massachusetts, USA.
Current protocols in immunology / edited by John E. Coligan ... [et al.] 06/2008; Chapter 15:Unit 15.21. DOI: 10.1002/0471142735.im1521s81
Source: PubMed


"Humanized" mice are a promising translational model for studying human hematopoiesis and immunity. Their utility has been enhanced by the development of new stocks of immunodeficient hosts, most notably mouse strains such as NOD-scid IL2rgamma(null) mice that lack the IL-2 receptor common gamma chain. These stocks of mice lack adaptive immune function, display multiple defects in innate immunity, and support heightened levels of human hematolymphoid engraftment. Humanized mice can support studies in many areas of immunology, including autoimmunity, transplantation, infectious diseases, and cancer. These models are particularly valuable in experimentation where there is no appropriate small animal model of the human disease, as in the case of certain viral infections. This unit details the creation of humanized mice by engraftment of immunodeficient mice with hematopoietic stem cells or peripheral blood mononuclear cells, provides methods for evaluating engraftment, and discusses considerations for choosing the appropriate model system to meet specific goals.

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    • "2.4. The generation of humanized mice As previously reported [17] "
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    ABSTRACT: Evaluation of the immunogenicity of human mesenchymal stem cells (MSCs) in an allogeneic setting during therapy has been hampered by lack of suitable models due to technical and ethical limitations. Here, we show that allogeneic human umbilical cord blood derived-MSCs (hUCB-MSCs) maintained low immunogenicity even after immune challenge in vitro. To confirm these properties in vivo, a humanized mouse model was established by injecting isolated hUCB-derived CD34+ cells intravenously into immunocompromised NOD/SCID IL2γnull (NSG) mice. After repeated intravenous injection of human peripheral blood mononuclear cells (hPBMCs) or MRC5 cells into these mice, immunological alterations including T cell proliferation and increased IFN-γ, TNF-α, and human IgG levels, were observed. In contrast, hUCB-MSC injection did not elicit these responses. While lymphocyte infiltration in the lung and small intestine and reduced survival rates were observed after hPBMC or MRC5 transplantation, no adverse events were observed following hUCB-MSC introduction. In conclusion, our data suggest that allogeneic hUCB-MSCs have low immunogenicity in vitro and in vivo, and are therefore "immunologically safe" for use in allogeneic clinical applications.
    Biochemical and Biophysical Research Communications 03/2014; 446(4). DOI:10.1016/j.bbrc.2014.03.051 · 2.30 Impact Factor
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    • "Human PBMCs are capable of engrafting and proliferating as T cells in NOD-scid IL2rγ-/- adult mice, and these engrafted mice can be challenged with live R5-tropic HIV-1.14 Engraftment and expansion of PBMCs treated ex vivo with NPs therefore allows for the in vivo functional evaluation of HIV-1 resistance conferred by triplex-mediated gene modification. "
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    ABSTRACT: Biodegradable poly (lactic-co-glycolic acid) (PLGA) nanoparticles (NPs) encapsulating triplex-forming peptide nucleic acids (PNAs) and donor DNAs for recombination-mediated editing of the CCR5 gene were synthesized for delivery into human peripheral blood mononuclear cells (PBMCs). NPs containing the CCR5-targeting molecules efficiently entered PBMCs with low cytotoxicity. Deep sequencing revealed that a single treatment with the formulation resulted in a targeting frequency of 0.97% in the CCR5 gene and a low off-target frequency of 0.004% in the CCR2 gene, a 216-fold difference. NP-treated PBMCs efficiently engrafted immunodeficient NOD-scid IL-2rγ(-/-) mice, and the targeted CCR5 modification was detected in splenic lymphocytes 4 weeks posttransplantation. After infection with an R5-tropic strain of HIV-1, humanized mice with CCR5-NP-treated PBMCs displayed significantly higher levels of CD4(+) T cells and significantly reduced plasma viral RNA loads compared with control mice engrafted with mock-treated PBMCs. This work demonstrates the feasibility of PLGA-NP-encapsulated PNA-based gene-editing molecules for the targeted modification of CCR5 in human PBMCs as a platform for conferring HIV-1 resistance.Molecular Therapy-Nucleic Acids (2013) 2, e135; doi:10.1038/mtna.2013.59; published online 19 November 2013.
    Molecular Therapy - Nucleic Acids 11/2013; 2(11):e135. DOI:10.1038/mtna.2013.59 · 4.51 Impact Factor
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    • "These mice develop no mature lymphocytes and NK cells [7] because the γ chain is an important component of many receptors for lymphoid-related cytokines and is crucial for the signaling through these receptors [13], [14]. The advantage of these mice is the lack of an adaptive immune system in addition to the lack of NK cells and an excellent engraftment of human cells [7], [15]. For the production of humanized mice, different human sources can be used e.g., hematopoietic stem cells from fresh cord blood (FCB) or more rarely, mobilized human stem cells (mSCs) [10]. "
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    ABSTRACT: To study the function and maturation of the human hematopoietic and immune system without endangering individuals, translational human-like animal models are needed. We compare the efficiency of CD34(+) stem cells isolated from cryopreserved cord blood from a blood bank (CCB) and fresh cord blood (FCB) in generating highly engrafted humanized mice in NOD-SCID IL2Rγ(null) (NSG) rodents. Interestingly, the isolation of CD34(+) cells from CCB results in a lower yield and purity compared to FCB. The purity of CD34(+) isolation from CCB decreases with an increasing number of mononuclear cells that is not evident in FCB. Despite the lower yield and purity of CD34(+) stem cell isolation from CCB compared to FCB, the overall reconstitution with human immune cells (CD45) and the differentiation of its subpopulations e.g., B cells, T cells or monocytes is comparable between both sources. In addition, independent of the cord blood origin, human B cells are able to produce high amounts of human IgM antibodies and human T cells are able to proliferate after stimulation with anti-CD3 antibodies. Nevertheless, T cells generated from FCB showed increased response to restimulation with anti-CD3. Our study reveals that the application of CCB samples for the engraftment of humanized mice does not result in less engraftment or a loss of differentiation and function of its subpopulations. Therefore, CCB is a reasonable alternative to FCB and allows the selection of specific genotypes (or any other criteria), which allows scientists to be independent from the daily changing birth rate.
    PLoS ONE 10/2012; 7(10):e46772. DOI:10.1371/journal.pone.0046772 · 3.23 Impact Factor
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