ABSTRACT
The development of "humanized" mice has become a prominent tool for translational animal studies of human diseases. Immunodeficient mice can be humanized by injections of human umbilical cord stem cells. The engraftment of these cells and their development into human lymphocytes has been made possible by the development of novel severely immunodeficient mouse strains. Proven protocols for the generation and analysis of humanized mice in the NSG mouse background are presented here. © 2023 The Authors. Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: Human umbilical stem cell engraftment of neonatal immunodeficient mice Basic Protocol 2: Human umbilical stem cell engraftment of 4-week-old immunodeficient mice Support Protocol 1: Preparation of human umbilical stem cells Support Protocol 2: Submandibular blood collection from humanized mice and analysis of peripheral blood via flow cytometry.
Subject(s)
Stem Cells , Wiskott-Aldrich Syndrome , Humans , Animals , Mice , Flow Cytometry , Umbilical Cord , UmbilicusABSTRACT
Osteolytic bone lesions and hypercalcemia are common, serious complications in adult T cell leukemia/lymphoma (ATL), an aggressive T cell malignancy associated with human T cell leukemia virus type 1 (HTLV-1) infection. The HTLV-1 viral oncogene HBZ has been implicated in ATL tumorigenesis and bone loss. In this study, we evaluated the role of HBZ on ATL-associated bone destruction using HTLV-1 infection and disease progression mouse models. Humanized mice infected with HTLV-1 developed lymphoproliferative disease and continuous, progressive osteolytic bone lesions. HTLV-1 lacking HBZ displayed only modest delays to lymphoproliferative disease but significantly decreased disease-associated bone loss compared with HTLV-1-infected mice. Gene expression array of acute ATL patient samples demonstrated increased expression of RANKL, a critical regulator of osteoclasts. We found that HBZ regulated RANKL in a c-Fos-dependent manner. Treatment of HTLV-1-infected humanized mice with denosumab, a monoclonal antibody against human RANKL, alleviated bone loss. Using patient-derived xenografts from primary human ATL cells to induce lymphoproliferative disease, we also observed profound tumor-induced bone destruction and increased c-Fos and RANKL gene expression. Together, these data show the critical role of HBZ in driving ATL-associated bone loss through RANKL and identify denosumab as a potential treatment to prevent bone complications in ATL patients.
Subject(s)
Basic-Leucine Zipper Transcription Factors/genetics , Basic-Leucine Zipper Transcription Factors/metabolism , Leukemia-Lymphoma, Adult T-Cell/genetics , Leukemia-Lymphoma, Adult T-Cell/metabolism , Retroviridae Proteins/metabolism , Adult , Animals , Bone Resorption/genetics , Bone Resorption/metabolism , Bone Resorption/pathology , Bone and Bones/pathology , Disease Models, Animal , Disease Progression , Female , Gene Expression Regulation, Neoplastic , Heterografts , Human T-lymphotropic virus 1 , Humans , Kaplan-Meier Estimate , Leukemia-Lymphoma, Adult T-Cell/pathology , Leukemia-Lymphoma, Adult T-Cell/virology , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Osteoclasts/metabolism , RANK Ligand/genetics , RANK Ligand/metabolism , Retroviridae Proteins/genetics , TranscriptomeABSTRACT
The development of humanized mice has become a prominent tool for translational animal studies of human diseases. Here we show how immune deficient mice can be "humanized" by injections of human umbilical cord stem cells. The engraftment of these cells and development into human lymphocytes has been possible because of the development of novel severely immune deficient mouse strains. Here we present proven protocols for the generation and analysis of humanized mice on the NSG mouse background.
Subject(s)
Stem Cells/cytology , Animals , Flow Cytometry , Humans , Mice , Mice, SCID , Stem Cells/physiology , Umbilical Cord/cytologyABSTRACT
Chronic infection with human T-cell leukemia virus type 1 (HTLV1) can lead to adult T-cell leukemia (ATL). In contrast, infection with HTLV2 does not lead to leukemia, potentially because of distinct virus-host interactions and an active immune response that controls virus replication and, therefore, leukemia development. We created a humanized mouse model by injecting human umbilical-cord stem cells into the livers of immunodeficient neonatal NSG mice, resulting in the development of human lymphocytes that cannot mount an adaptive immune response. We used these mice to compare the ability of molecular clones of HTLV1, HTLV2, and select recombinant viruses to induce leukemia-lymphoma in vivo. Infection with HTLV1 strongly stimulated the proliferation of CD4+ T cells, whereas HTLV2 preferentially stimulated the proliferation of CD8+ T cells; both HTLV1 and HTLV2 induced lymphoproliferative disease. Uninfected and HTLV-infected humanized mice both showed granulomatous inflammation as a background lesion. Similarly, recombinant viruses that expressed the HTLV1 envelope protein (Env) on an HTLV2 background (HTLV2-Env1) or Env2 on an HTLV1 background (HTLV1-Env2) induced lymphoproliferative disease. HTLV2-Env1 stimulated the proliferation of CD4+ T cells, whereas HTLV1-Env2 stimulated both CD4+ and CD8+ T-cell subsets. Our results show that T-cell transformation in vivo is guided by the Env protein of the virus. Furthermore, our humanized mouse model is useful for exploring the preferred T-cell tropisms of HTLV1 and HTLV2.
