From Bench to Cageside: Immunology of Humanizing...
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From Bench to Cageside:Immunology of Humanizing Mice
Zachary T Freeman
Goals• Understand relevant immunology related to
humanized mice• Understand three main components that contribute to
formation of humanized mice• Clinical management of severely immunodeficient
mice
Goals• Understand relevant immunology related to
humanized mice• Understand three main components that contribute to
formation of humanized mice• Clinical management of severely immunodeficient
mice
Dranoff, G Nat. Rev. Cancer, 2004
Innate Lymphoid Cells (ILCs) Are a Novel Cell Type for Immune Responses
Shortman K, et al Nat Rev Imm2002
Innate Lymphoid Cells (ILCs) Are a Novel Cell Type for Immune Responses
Spits H, Nat Rev Imm, 2013
Humoral Immunity Cellular Immunity
Kumar et al. Robbins Basic Pathology, 8 edn. 2007, pp. 109, Copyright Elsevier.
B Cells Produced Antibodies as the Main Mediator of Humoral Immunity
Cooper MD, Nat Rev Imm, 2015
Zou W, Nat Rev Imm, 2010
CD4 T Cells Coordinate and Modulate Cellular Immune Responses
Restifo NP, Nat Rev Imm, 2012
CD8 T Cells Mediate Effector Portion of Cellular Immunity
Goals• Understand relevant immunology related to
humanized mice• Understand three main components that contribute to
formation of humanized mice• Clinical management of severely immunodeficient
mice
Crotty S, Nat Rev Imm, 2015
T and B cell Interactions are Important for Protective Immunity
https://www.jax.org/jax-mice-and-services/in-vivo-pharmacology/humanized-mice/cd34
What is an NSG mouse?
https://www.jax.org/strain/005557
NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ
Non Obese Diabetic Mice Have Altered Innate and Adaptive Immunophenotype
Calis JJA, Trends in Imm, 2014
• T cell mediated islet destruction/diabetes• Defective NK cell function• Defective macrophage cytokine production • Altered immunoregulatory functions of antigen
presenting cells
What is an NSG mouse?
https://www.jax.org/strain/005557
NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ
Antigen Specificity Defines B and T cells
Calis JJA, Trends in Imm, 2014
B and T cell Receptors Share Common Features
Calis JJA, Trends in Imm, 2014
Lymphoid Specific General
Villartay, JP, Nat Rev Imm, 2003
Deletion of Prkdc Leads to Defect in DNA Repair Mechanisms in All Cells
What is an NSG mouse?
https://www.jax.org/strain/005557
NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ
IL-2 Receptor Gamma is Essential B, T, NK and DC Function
Rochman Y, Nat Rev Imm, 2009
https://www.jax.org/jax-mice-and-services/in-vivo-pharmacology/humanized-mice/cd34
Ionizing Radiation Has Variable Effects on Immune Cells
Finkelstein SE, et al Frontiers in Oncology, 2012
https://www.jax.org/jax-mice-and-services/in-vivo-pharmacology/humanized-mice/cd34
Origins of “Humanizing” Component
• Peripheral blood mononuclear cells (PBMC/PBL)• CD34+ hematopoietic stem cells (HSCs)• Bone marrow, liver, thymus (BLT mice)
CD34+ Cells Serve as Total Precursor to Immune System
Sharpless NE et al, Nat Rev Mol Cell Bio, 2007
Shultz LD, Nat Rev Imm, 2012
Unknown Cross talk between Mouse and Human Cytokines
• S: Human Stem Cell Factor (SCF) gene• maintains normal basal hematopoiesis.
• GM: Human Granulocyte/Macrophage-colony stimulating factor 2 (GM-CSF)• Innate cell numbers and function
• 3: Human interleukin-3 (IL-3) • multipotent hematopoietic stem cells
differentiate into myeloid progenitor cells• myeloid lineage to proliferate
Human Cytokines Enhance Innate Cell Engraftment in NSG-SGM3 Mice
Shultz LD, Nat Rev Imm, 2012
Human Cytokines Enhance Innate Cell Engraftment in Humanized Mice
Goals• Understand relevant immunology related to
humanized mice• Understand three main components that contribute to
formation of humanized mice• Clinical management of severely immunodeficient
mice
Immunology Problems Underlying Clinical Disease in Humanized Mice
• Increased susceptibility to commensal/opportunisitcorganisms• Cornybacterium bovis• Klebsiella oxytoca• Pasturella pneumonia• Pseudomonas sp• Staphlococcus sp• Pneumocystis murina
• Graft Versus Host Disease• Limited resources for the immune system• Risk of anthropozoonosis
Human Immune Cells Do No Recognize Murine Immune Cells
Kobayashi KS, Nature Reviews Immunology, 2012
Human Immune Cells Do No Recognize Murine Immune Cells
Bleakley M, Nature Reviews Cancer, 2004
Limited Host Resources Alter Immune Cell Expansion and Homeostasis
Badovinac VP, Immunity 2007
Unknown Risk of Anthropozoonosis?
• Human immune cells host to many infectious pathogens• HIV, EBV, CMV, HTLV
• Unknown risk of infection for severely immunodeficient mouse
• Unclear if murine immune system responds to these organisms
Management of Humanized Mice• Barrier housing
• Air locks/showers• Designated clothing• PPE• PAPRs
• Individually ventilated cages• HEPA filtered air• Automated water
Management of Humanized Mice• Vary levels of “sterilization”
• Cages• Food• Bedding• Laminar flow hoods
• Food/water practices• Acidification of water• Prophylactic antibiotics not recommended
Management of Humanized Mice• Infectious Agent Surveillance
• Sentinel selection• Age?• Immune status?• Direct vs. indirect
• Agent exclusion• Testing methodology/frequency
• Limited information about efficacy of treatment• Rederivation to clean up infections
Management of Humanized Mice
https://www.jax.org/news-and-insights/jax-blog/2015/april/top-5-tips-to-take-care-of-your-nsg-mice
Basic immunology of humanizing mice is important for clinical management and use of the model.
Questions?
Acknowledgements
• Jessica Xu DVM• ULAM faculty and residents
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CD8(+) T cell response to infection. Immunity 26, 827-841 (2007).• 2.Bleakley, M. & Riddell, S.R. Molecules and mechanisms of the graft-versus-leukaemia effect. Nat Rev Cancer 4, 371-380 (2004).• 3.Calis, J.J. & Rosenberg, B.R. Characterizing immune repertoires by high throughput sequencing: strategies and applications. Trends
Immunol 35, 581-590 (2014).• 4.Cooper, M.D. The early history of B cells. Nat Rev Immunol 15, 191-197 (2015).• 5.Crotty, S. A brief history of T cell help to B cells. Nat Rev Immunol 15, 185-189 (2015).• 6.de Villartay, J.P., Fischer, A. & Durandy, A. The mechanisms of immune diversification and their disorders. Nat Rev Immunol 3, 962-
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Immunol 9, 480-490 (2009).• 12. Sharpless, N.E. & DePinho, R.A. How stem cells age and why this makes us grow old. Nat Rev Mol Cell Biol 8, 703-713 (2007).• 13. Shortman, K. & Liu, Y.J. Mouse and human dendritic cell subtypes. Nat Rev Immunol 2, 151-161 (2002).• 14. Spits, H. et al. Innate lymphoid cells--a proposal for uniform nomenclature. Nat Rev Immunol 13, 145-149 (2013).• 15. Walsh, N.C. et al. Humanized Mouse Models of Clinical Disease. Annu Rev Pathol 12, 187-215 (2017).• 16. Zou, W. & Restifo, N.P. T(H)17 cells in tumour immunity and immunotherapy. Nat Rev Immunol 10, 248-256 (2010).• 17. Kumar et al. Robbins Basic Pathology, 8 edn. 2007, pp. 109, Copyright Elsevier.• https://www.jax.org/strain/005557• https://www.jax.org/news-and-insights/jax-blog/2015/april/top-5-tips-to-take-care-of-your-nsg-mice• https://www.jax.org/jax-mice-and-services/in-vivo-pharmacology/humanized-mice/cd34