A novel humanized mouse model to study the function of human cutaneous memory T cells in vivo in human skin

AbstractHuman skin contains a population of memory T cells that support tissue homeostasis and provide protective immunity. The study of human memory T cells is often restricted to in vitro studies and to human PBMC serving as primary cell source. Because the tisse environment impacts the phenotype and function of memory T cells, it is crucial to study these cells within their tissue. Here we utilized immunodeficient NOD-scid IL2rγnull (NSG) mice that carried in vivo-generated engineered human skin (ES). ES were generated from human keratinocytes and fibroblasts and is initially devoid of skin-resident immune cells. Upon adoptive transfer of human PBMC this reductionist system allowed to study human T cell recruitment from a circulating pool of T cells into non-inflamed human skin in vivo. Circulating human memory T cells preferentially infiltrated ES and showed diverse functional profiles of T cells found in fresh human skin. The chemokine and cytokine microenvironment of ES closely resembled that of non-inflamed human skin. Upon entering the ES T cells assumed a resident memory T cell-like phenotype in the absence of infection, and a proportion of these cutaneous T cells can be locally activated upon injection of monocyte derived dendritic cells (moDCs) that presented Candida albicans. Interestingly, we found that CD69+ memory T cells produced higher levels of effector cytokines in response to Candida albicans, compared to CD69- T cells. Overall, this model has broad utility in many areas of human skin immunology research, including the study of immune-mediated skin diseases.

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IL-7 and IL-21 are superior to IL-2 and IL-15 in promoting human T cell–mediated rejection of systemic lymphoma in immunodeficient mice

Blood ◽

10.1182/blood-2009-09-241398 ◽

2010 ◽

Vol 115 (17) ◽

pp. 3508-3519 ◽

Cited By ~ 118

Author(s):

John C. Markley ◽

Michel Sadelain

Keyword(s):

T Cells ◽

T Cell ◽

Adoptive Transfer ◽

Memory T Cells ◽

Tumor Rejection ◽

Effector Memory ◽

Transfer Model ◽

Human T Cell

Abstract The γc-cytokines are critical regulators of immunity and possess both overlapping and distinctive functions. However, comparative studies of their pleiotropic effects on human T cell–mediated tumor rejection are lacking. In a xenogeneic adoptive transfer model, we have compared the therapeutic potency of CD19-specific human primary T cells that constitutively express interleukin-2 (IL-2), IL-7, IL-15, or IL-21. We demonstrate that each cytokine enhanced the eradication of systemic CD19+ B-cell malignancies in nonobese diabetic/severe combined immunodeficient (NOD/SCID)/γcnull mice with markedly different efficacies and through singularly distinct mechanisms. IL-7– and IL-21–transduced T cells were most efficacious in vivo, although their effector functions were not as enhanced as IL-2– and IL-15–transduced T cells. IL-7 best sustained in vitro T-cell accumulation in response to repeated antigenic stimulation, but did not promote long-term T-cell persistence in vivo. Both IL-15 and IL-21 overexpression supported long-term T-cell persistence in treated mice, however, the memory T cells found 100 days after adoptive transfer were phenotypically dissimilar, resembling central memory and effector memory T cells, respectively. These results support the use of γc-cytokines in cancer immunotherapy, and establish that there exists more than 1 human T-cell memory phenotype associated with long-term tumor immunity.

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Gut Homing Potential of Human Naïve and Memory Alloreactive T Cells.

Blood ◽

10.1182/blood.v104.11.2864.2864 ◽

2004 ◽

Vol 104 (11) ◽

pp. 2864-2864

Author(s):

Michael W. Boyer ◽

Kelly McQuown ◽

Lindsay Hendey ◽

Thomas G. Gross

Keyword(s):

T Cells ◽

T Cell ◽

Memory T Cells ◽

Human Memory ◽

Cell Manipulation ◽

Integrin Expression ◽

Donor Pool ◽

Tissue Specific ◽

Relative Contribution

Abstract T cells are either naïve, having never encountered cognate antigen, or memory, with a history of activation, proliferation, and acquistion of effector specialization including tissue specific homing properties. We hypothesized that memory T cells would contain a subpopulation of alloantigen specific cells that might have acquired tissue specific homing characteristics such as upregulation of α4β7 integrin that directs T cells to gut lymphoid and non-lymphoid tissue. In mice, α4β7 integrin dependent migration to the peyer’s patch is essential to instigate lethal GVHD in MHC mismatched BMT models. To test this hypothesis, immunomagnetic sorting for CD45RO and CD45RA was used to obtain populations either depleted or enriched for naïve T cells respectively. Using CFDA dye dilution and CD69 upregulation, alloantigen specific T cells were tracked with flow cytometry at days 3, 5, and 7 of a mixed lymphocyte reaction, with an autologous control. ModFit LT software was used to estimate precursor frequencies, which showed that approximately 1 in 200 CD4 and CD8 positive T cells are alloreactive (n=9) regardless of whether they were CD45RO or CD45RA selected, demonstrating that alloreactivity could be potentially recruited from either the naïve or memory donor pool in a GVHD reaction. Analysis of α4β7 integrin coexpression before alloantigen stimulation demonstrated that for both CD4 and CD8 positive T cells, the memory pool had two-fold higher levels of coexpression that were statistically significant compared to the naive pool, with CD8 memory T cells demonstrating the highest coexpression (24 +/− 12%). When examined at day 5 of the MLR, both CD4 and CD8 positive alloantigen specific T cells from the CD45RO selected group maintained levels of α4β7 integrin coexpression similar to baseline (14% and 25% respectively). In contrast, CD4 and CD8 positive alloantigen specific T cells from the CD45RA selected group had levels of α4β7 integrin coexpression that were four-fold higher compared to baseline, with 20% and 48% coexpression respectively (n=6). The relative contribution of α4 and β7 integrins was examined by comparing the mean flourescent intensity (MFI) of the alloantigen specific T cells to the resting T cells within the same day 5 MLR. The greatest increase in expression was seen for β7 integrin on the CD45RA selected CD4 and CD8 positive T cells with 3.1 and 3.2 times higher expression respectively (both p<.001). In contrast, the CD45RO selected CD4 and CD8 positive T cells had 1.78 (p<.01) and 1.35 (p<.05) times higher β7 expression levels respectively. With regard to α4 integrin expression, CD45RA selected CD4 and CD8 positive T cells had 2.2 and 2.0 times higher expression respectively (both p<.01). The α4 integrin expression on CD45RO selected CD4 and CD8 positive T cells was 2.0 (p<.05) and 1.43 (p= NS) times higher respectively. These data suggest that both naïve and memory CD4 and CD8 positive T cells may contribute to alloreactivity, although there are differences in the regulation of α4β7 integrin expression which could significantly affect in vivo T cell homing and therefore instigation of GVHD. Based upon murine studies, it has been demonstrated that memory T cells do not contribute to GVHD; our data suggest in contrast that human memory T cells could contribute in a significant way to GVHD and further study is necessary before development of successful T cell manipulation strategies aimed at attenuating GVHD.

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Circulating CD4+ memory T cells give rise to a CD69+ resident memory T cell population in non-inflamed human skin

10.1101/490094 ◽

2018 ◽

Cited By ~ 2

Author(s):

Maria M Klicznik ◽

Ariane Benedetti ◽

Angelika Stoecklinger ◽

Daniel J Campbell ◽

Iris K Gratz

Keyword(s):

T Cells ◽

Candida Albicans ◽

T Cell ◽

Human Skin ◽

Cell Population ◽

Memory T Cells ◽

Humanized Mouse Model ◽

Normal Human Skin ◽

Memory T Cell ◽

Resident Memory T Cells

The blood of human adults contains a pool of circulating CD4+ memory T cells and normal human skin contains a CD4+CD69+ memory T cell population that produce IL17 in response to Candida albicans. Here we studied the generation of CD4+CD69+ memory T cells in human skin from a pool of circulating CD4+ memory T cells. Using adoptive transfer of human PBMC into a skin-humanized mouse model we discovered the generation of CD4+CD69+ resident memory T cells in human skin in absence of infection or inflammation. These CD4+CD69+ resident memory T cells were activated and displayed heightened effector function in response to Candida albicans. These studies demonstrate that a CD4+CD69+ T cell population can be established in human skin from a pool of circulating CD4+ memory T cells in absence of infection/inflammation. The described process might be a novel way to spread antigen-specific immunity at large barrier sites even in absence of infection or inflammation.

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In Vivo Selection of T-Cell Receptor Junctional Region Sequences by HLA-A2 Human T-Cell Lymphotropic Virus Type 1 Tax11-19 Peptide Complexes

Journal of Virology ◽

10.1128/jvi.75.2.1065-1071.2001 ◽

2001 ◽

Vol 75 (2) ◽

pp. 1065-1071 ◽

Cited By ~ 11

Author(s):

Mineki Saito ◽

Graham P. Taylor ◽

Akiko Saito ◽

Yoshitaka Furukawa ◽

Koichiro Usuku ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

T Cell Receptor ◽

Virus Type ◽

Cell Receptor ◽

Antigen Specificity ◽

Human T Cell ◽

Cdr3 Region

ABSTRACT Using HLA-peptide tetrameric complexes, we isolated human T-cell lymphotrophic virus type 1 Tax peptide-specific CD8+ T cells ex vivo. Antigen-specific amino acid motifs were identified in the T-cell receptor Vβ CDR3 region of clonally expanded CD8+ T cells. This result directly confirms the importance of the CDR3 region in determining the antigen specificity in vivo.

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Generation of primary antigen-specific human cytotoxic T lymphocytes in human/mouse radiation chimera

Blood ◽

10.1182/blood.v88.2.721.bloodjournal882721 ◽

1996 ◽

Vol 88 (2) ◽

pp. 721-730 ◽

Cited By ~ 30

Author(s):

H Segall ◽

I Lubin ◽

H Marcus ◽

A Canaan ◽

Y Reisner

Keyword(s):

T Cells ◽

T Cell ◽

Human Lymphocytes ◽

Scid Mice ◽

Human Antibody ◽

Adoptive Therapy ◽

Activation Markers ◽

Human T Cell ◽

Human T Cells

Severe combined immunodeficient (SCID) mice are increasingly used as hosts for the adoptive transfer of human lymphocytes. Human antibody responses can be obtained in these xenogeneic chimeras, but information about the functionality of the human T cells in SCID mice is limited and controversial. Studies using human peripheral blood lymphocytes (PBL) injected intraperitoneally (IP) into SCID mice (hu-PBL-SCID mice) have shown that human T cells from these chimeras are anergic and have a defective signaling via the T-cell receptor. In addition, their antigenic repertoire is limited to xenoreactive clones. In the present study, we tested the functionality of human T cell in a recently described chimeric model. In this system, BALB/c mice are conditioned by irradiation and then transplanted with SCID bone marrow, followed by IP injection of human PBL. Our experiments demonstrated that human T cells, recovered from these hu-PBL-BALB mice within 1 month posttransplant, proliferated and expressed activation markers upon stimulation with anti-CD3 monoclonal antibody. A vigorous antiallogeneic human cytotoxic T-lymphocyte (CTL) response could be generated in these mice by immunizing them with irradiated allogeneic cells. Moreover, anti-human immunodeficiency virus type 1 (HIV-1) Net- specific human CTLs could be generated in vivo from naive lymphocytes by immunization of mouse-human chimeras with a recombinant vaccinia-nef virus. This model may be used to evaluate potential immunomodulatory drugs or cytokines, and could provide a relevant model for testing HIV vaccines, for production of antiviral T-cell clones for adoptive therapy, and for studying human T-cell responses in vivo.

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AAV-Mediated In Vivo CAR Gene Therapy for Targeting Human T Cell Leukemia

10.1101/2021.02.15.431201 ◽

2021 ◽

Author(s):

Waqas Nawaz ◽

Bilian Huang ◽

Shijie Xu ◽

Yanlei Li ◽

Linjing Zhu ◽

...

Keyword(s):

Gene Therapy ◽

T Cells ◽

T Cell ◽

T Cell Leukemia ◽

Cell Leukemia ◽

Car T Cells ◽

Car T Cell ◽

Human T Cell ◽

Car T

AbstractChimeric antigen receptor (CAR) T cell therapy is the most active field in immuno-oncology and brings substantial benefit to patients with B cell malignancies. However, the complex procedure for CAR T cell generation hampers its widespread applications. Here, we describe a novel approach in which human CAR T cells can be generated within the host upon injecting an Adeno-associated virus (AAV)vector carrying the CAR gene, which we call AAV delivering CAR gene therapy (ACG). Upon single infusion into a humanized NCG tumor mouse model of human T cell leukemia, AAV generates sufficient numbers of potent in vivo CAR cells, resulting in tumor regression; these in vivo generated CAR cells produce antitumor immunological characteristics. This instantaneous generation of in vivo CAR T cells may bypass the need for patient lymphodepletion, as well as the ex vivo processes of traditional CAR T cell production, which may make CAR therapy simpler and less expensive. It may allow the development of intricate, individualized treatments in the form of on-demand and diverse therapies.Significance StatementAAV can generate enough CAR cells within the host. That act as a living drug, distributed throughout the body, and persist for weeks, with the ability to recognize and destroy tumor cells.

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Tax and Overlapping Rex Sequences Do Not Confer the Distinct Transformation Tropisms of Human T-Cell Leukemia Virus Types 1 and 2

Journal of Virology ◽

10.1128/jvi.77.14.7728-7735.2003 ◽

2003 ◽

Vol 77 (14) ◽

pp. 7728-7735 ◽

Cited By ~ 19

Author(s):

Jianxin Ye ◽

Li Xie ◽

Patrick L. Green

Keyword(s):

T Cells ◽

T Cell ◽

Leukemia Virus ◽

T Cell Leukemia ◽

Cell Leukemia ◽

Cell Leukemia Virus Type ◽

Human T Cell ◽

T Cell Leukemia Virus

ABSTRACT Human T-cell leukemia virus type 1 (HTLV-1) and HTLV-2 are distinct oncogenic retroviruses that infect several cell types but display their biological and pathogenic activity only in T cells. Previous studies have indicated that in vivo HTLV-1 has a preferential tropism for CD4+ T cells, whereas HTLV-2 in vivo tropism is less clear but appears to favor CD8+ T cells. Both CD4+ and CD8+ T cells are susceptible to HTLV-1 and HTLV-2 infection in vitro, and HTLV-1 has a preferential immortalization and transformation tropism of CD4+ T cells, whereas HTLV-2 immortalizes and transforms primarily CD8+ T cells. The molecular mechanism that determines this tropism of HTLV-1 and HTLV-2 has not been determined. HTLV-1 and HTLV-2 carry the tax and rex transregulatory genes in separate but partially overlapping reading frames. Since Tax has been shown to be critical for cellular transformation in vitro and interacts with numerous cellular processes, we hypothesized that the viral determinant of transformation tropism is encoded by tax. Using molecular clones of HTLV-1 (Ach) and HTLV-2 (pH6neo), we constructed recombinants in which tax and overlapping rex genes of the two viruses were exchanged. p19 Gag expression from proviral clones transfected into 293T cells indicated that both recombinants contained functional Tax and Rex but with significantly altered activity compared to the wild-type clones. Stable transfectants expressing recombinant viruses were established, irradiated, and cocultured with peripheral blood mononuclear cells. Both recombinants were competent to transform T lymphocytes with an efficiency similar to that of the parental viruses. Flow cytometry analysis indicated that HTLV-1 and HTLV-1/TR2 had a preferential tropism for CD4+ T cells and that HTLV-2 and HTLV-2/TR1 had a preferential tropism for CD8+ T cells. Our results indicate that tax/rex in different genetic backgrounds display altered functional activity but ultimately do not contribute to the different in vitro transformation tropisms. This first study with recombinants between HTLV-1 and HTLV-2 is the initial step in elucidating the different pathobiologies of HTLV-1 and HTLV-2.

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Essential Role of Human T Cell Leukemia Virus Type 1orf-Iin Lethal Proliferation of CD4+Cells in Humanized Mice

Journal of Virology ◽

10.1128/jvi.00565-19 ◽

2019 ◽

Vol 93 (19) ◽

Cited By ~ 7

Author(s):

Veronica Galli ◽

Christopher C. Nixon ◽

Natasa Strbo ◽

Maria Artesi ◽

Maria F. de Castro-Amarante ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Leukemia Virus ◽

Humanized Mice ◽

T Cell Leukemia ◽

Cell Leukemia ◽

Cell Leukemia Virus Type ◽

Human T Cell ◽

T Cell Leukemia Virus

ABSTRACTHuman T cell leukemia virus type 1 (HTLV-1) is the ethological agent of adult T cell leukemia/lymphoma (ATLL) and a number of lymphocyte-mediated inflammatory conditions, including HTLV-1-associated myelopathy/tropical spastic paraparesis. HTLV-1orf-Iencodes two proteins, p8 and p12, whose functions in humans are to counteract innate and adaptive responses and to support viral transmission. However, thein vivorequirements fororf-Iexpression vary in different animal models. In macaques, the ablation oforf-Iexpression by mutation of its ATG initiation codon abolishes the infectivity of the molecular clone HTLV-1p12KO. In rabbits, HTLV-1p12KOis infective and persists efficiently. We used humanized mouse models to assess the infectivity of both wild-type HTLV-1 (HTLV-1WT) and HTLV-1p12KO. We found that NOD/SCID/γC−/−c-kit+mice engrafted with human tissues 1 day after birth (designated NSG-1d mice) were highly susceptible to infection by HTLV-1WT, with a syndrome characterized by the rapid polyclonal proliferation and infiltration of CD4+CD25+T cells into vital organs, weight loss, and death. HTLV-1 clonality studies revealed the presence of multiple clones of low abundance, confirming the polyclonal expansion of HTLV-1-infected cellsin vivo. HTLV-1p12KOinfection in a bone marrow-liver-thymus (BLT) mouse model prone to graft-versus-host disease occurred only following reversion of theorf-Iinitiation codon mutation within weeks after exposure and was associated with high levels of HTLV-1 DNA in blood and the expansion of CD4+CD25+T cells. Thus, the incomplete reconstitution of the human immune system in BLT mice may provide a window of opportunity for HTLV-1 replication and the selection of viral variants with greater fitness.IMPORTANCEHumanized mice constitute a useful model for studying the HTLV-1-associated polyclonal proliferation of CD4+T cells and viral integration sites in the human genome. The rapid death of infected animals, however, appears to preclude the clonal selection typically observed in human ATLL, which normally develops in 2 to 5% of individuals infected with HTLV-1. Nevertheless, the expansion of multiple clones of low abundance in these humanized mice mirrors the early phase of HTLV-1 infection in humans, providing a useful model to investigate approaches to inhibit virus-induced CD4+T cell proliferation.

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