Immune responses in humanized mice are generally inefficient without co-transplantation of human thymus or HLA transgenes. Previously, we generated humanized mice via the intra-bone marrow injection of CD133+ cord blood cells into irradiated adult immunodeficient mice (IBMI-huNSG mice), which could mount functional immune responses against HTLV-1, although the underlying mechanisms were still unknown. Here, we investigated thymocyte development in IBMI-huNSG mice, focusing on the roles of human and mouse MHC restriction. IBMI-huNSG mice had normal developmental profiles but aberrant thymic structures. Surprisingly, the thymic medulla-like regions expanded after immunization due to enhanced thymocyte expansion in association with the increase in HLA-DR+ cells, including CD205⁺ dendritic cells (DCs). The organ culture of thymus from immunized IBMI-huNSG mice with a neutralizing antibody to HLA-DR showed the HLA-DR-dependent expansion of CD4 single positive thymocytes. Mature peripheral T-cells exhibited alloreactive proliferation when co-cultured with human peripheral blood mononuclear cells. Live imaging of the thymus from immunized IBMI-huNSG mice revealed dynamic adhesive contacts of human-derived thymocytes and DCs accompanied by Rap1 activation. These findings demonstrate that an increase in HLA-DR+ cells by immunization promotes HLA-restricted thymocyte expansion in humanized mice, offering a unique opportunity to generate humanized mice with ease.

Human T-cell leukemia virus type 1 (HTLV-1) establishes lifelong infection and is associated with severe diseases such as adult T-cell leukemia/lymphoma (ATL) and HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). Cytotoxic T lymphocytes (CTLs), especially those specific for the viral protein Tax, play a pivotal role in controlling HTLV-1 infection. However, conventional humanized mouse models fail to fully reconstitute human immune responses, limiting their utility for evaluating CTL-mediated immunity. This study aimed to establish a physiologically relevant in vivo model to investigate human CTL responses against HTLV-1. To achieve this, we utilized NOG-HLA-A02 transgenic (Tg) mice expressing human HLA-A02:01 on thymic epithelial cells, enabling proper development of HLA-restricted human T cells. Compared to conventional humanized NOG mice, HTLV-1-infected humanized NOG-HLA-A02 Tg mice exhibited significantly reduced HTLV-1 proviral load (PVL), decreased expansion of infected CD4⁺ T cells, a trend toward increased frequencies of Tax-specific CD8⁺ T cells, and prolonged survival. These results demonstrate that the expression of HLA-A02:01 facilitates robust CTL-mediated immune control of HTLV-1. This model provides a powerful platform for dissecting HTLV-1 immunopathogenesis and evaluating CTL-targeted therapeutic strategies, including vaccines and immune checkpoint inhibitors.