Glycolytic metabolism of pathogenic T cells enables early detection of GvHD by 13C-MRI

AbstractGraft-versus-host disease (GvHD) is a prominent barrier to allogeneic hematopoietic stem cell transplantation (HSCT). Definitive diagnosis of GvHD is invasive and biopsies of involved tissues pose a high risk of bleeding and infection. Our previous studies in a chronic GvHD mouse model demonstrated that alloreactive CD4+ T cells are distributed to target organs ahead of overt symptoms, meanwhile CD4+ T cell activation is tied to increased glycolysis. Thus, we hypothesized that metabolic imaging of glycolysis would allow non-invasive detection of insipient GvHD in target organs infiltrated by glycolytic effector memory CD4+ T cells. We metabolically characterized CD4+ T cell subsets on day 14 post-transplant before the onset of chronic GvHD in a pre-clinical mouse model and performed 13C hyperpolarized magnetic resonance imaging (MRI) to quantify glycolytic activity in the liver of mice over the course of the disease. Intracellular metabolic screening and ex vivo metabolic profiling of CD4+ T cell subsets at day 14 confirmed that activated CD4+ T cells were highly glycolytic. Concurrently, hyperpolarized 13C-pyruvate MRI of the liver showed high conversion of pyruvate to lactate, indicative of increased glycolytic activity, that distinguished allogeneic from syngeneic HSCT recipients prior to the development of overt chronic GvHD. Furthermore, single cell sequencing of T cells in patients undergoing allogeneic HSCT indicated that similar metabolic changes may play a role in acute GvHD, providing a rationale for testing this imaging approach in the clinical post-HSCT setting. Our imaging approach is amenable to clinical translation and may allow early, non-invasive diagnosis of GvHD.

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Proportions of T-Cell CD8+ Subsets Lacking CD28 Expression at Day 30 after Myeloablative Allogeneic Stem Cell Transplantation Are Predictive for Relapse and Chronic GVHD.

Blood ◽

10.1182/blood.v110.11.2979.2979 ◽

2007 ◽

Vol 110 (11) ◽

pp. 2979-2979

Author(s):

Ibrahim Yakoub-Agha ◽

Pasquine Saule ◽

Leonardo Magro ◽

Pascale Cracco ◽

Valerie Coiteux ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Chronic Gvhd ◽

Immunosuppressive Treatment ◽

Cd8 T Cell ◽

T Cell Subsets ◽

Mixed Chimerism ◽

Effector Memory ◽

Myeloablative Conditioning ◽

Risk Of Relapse

Abstract The curative potential of allo-SCT for malignancies derives from the progressive reconstitution of the immune system and the development of effective anti-tumor immunity, but GVHD and disease relapse remain considerable obstacles to improvement in overall outcomes. Because in recipients target antigens are persisting, donor-derived T-cell responses may be expected to lead to the accumulation of a sizable proportion of differentiated T-cells, as happens following infection with persisting pathogens. A few cross-sectional studies have pointed to the preponderance of certain memory T-cell subsets associated with chronic GVHD (cGVHD), but the subset identified differed between studies. Inasmuch as qualitative T-cell recovery takes months to years to complete and there is substantial variability in time to development of GVHD or relapse, serial analysis might be more suitable to unveil early changes in T-cell subset composition attributable to transplantation-related events. From October 2003 on, 55 pts who underwent an allo-SCT after myeloablative conditioning were monitored prospectively in terms of clinical post-graft complications, including graft rejection, infections, GVHD and relapse. Blood samples were obtained on days 30±2, 60±3, 90±5, 180±10 and 365±15 post-transplant. Naive (CD45RA+CCR7+), central memory (TCM, CD45RAnegCCR7+), effector memory (TEM, CD45RAnegCCR7neg), and terminally differentiated effector (TTD, CD45RA+CCR7neg) were enumerated within the CD4+ and CD8+ pools, and the percentage of cells coexpressing CD28 was calculated within each eight subsets. The degree of donor-derived T-cell chimerism was assessed by real time PCR (sensitivity ≤ 1%). Median follow-up was 733 d (404–1251). Dynamics of CD4+ and CD8+ naive, TCM, TEM, and TTD were similar between the pts who developed cGVHD (n=15) and those who did not and between pts who relapsed and those who did not. However, costaining to detect CD28 demonstrated contrasting differences between cGVHD and relapse. At day 30, pts who subsequently relapsed (n=17) had elevated percentages of cells keeping CD28 expression within CD8+ T-cell subsets (TCM, p=.001; TCM, p=.021; and TTD, p=.007). Conversely, pts who subsequently developed cGVHD (n=15; only one relapsed) had diminished percentages of CD28+ cells within the two CD8+CCR7+ subsets at day 30 (p=.002 and p=.034, respectively). Loss of CD28 expression is known to be a hallmark of CMV infection but multivariate analysis ruled out, however, a confounding effect of CMV. Adjusted hazard ratios were 0.10 (95% CI, 0.01-0.76; p=.026) and 5.56 (95% CI, 1.16-25.00; p=.032) with CD28neg cells 16.7% of all CD8+ TCM at day 30 for relapse and cGVHD, respectively. Furthermore, pts with relapse had more often mixed chimerism at day 30 while those with cGVHD had more often full-donor chimerism (p=.042 and p=.023, respectively). CONCLUSION: This prospective study is the first to associate an early contrasting change in CD8+CD28neg T-cells with the risk of relapse and cGVHD after a myeloablative conditioning. Determination at day 30 of the proportions of CD8+ T-cell subsets expressing CD28 and of the level of T-cell chimerism could assist in predicting risk of relapse and cGVHD and help build an algorithm for the management of immunosuppressive treatment.

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NaïVe T Cell Depletion of PBSC Grafts Results in Very Low Rates of Chronic Gvhd and High Survival

Blood ◽

10.1182/blood.v128.22.668.668 ◽

2016 ◽

Vol 128 (22) ◽

pp. 668-668

Author(s):

Marie Bleakley ◽

Ted A. Gooley ◽

Barbara Hilzinger ◽

Stanley R Riddell ◽

Warren D Shlomchik

Keyword(s):

T Cells ◽

T Cell ◽

Acute Leukemia ◽

Chronic Gvhd ◽

Hematopoietic Cell ◽

T Cell Subsets ◽

Cell Depletion ◽

Effector Memory ◽

Phase Ii Trials ◽

T Cell Depletion

Abstract Background Graft-versus-host disease (GVHD) frequently causes morbidity and mortality after allogeneic hematopoietic cell transplantation (HCT) as a result of organ damage and infections. In HLA-identical HCT, GVHD results from recognition by donor T cells of minor histocompatibility (H) antigens on recipient tissues. Complete T cell depletion (TCD) of donor hematopoietic cell products is more effective than pharmacologic immunosuppression for preventing GVHD, but is complicated by delayed immune reconstitution and consequent life-threatening infections.Approaches to HCT which preferentially deplete the T cells that primarily cause GVHD and preserve pathogen-specific T cells may improve HCT outcomes. Mature CD3+ CD8+ and CD3+ CD4+ T cells can be classified into CD45RA+ CD62L+ naïve (TN) and CD45RO+ memory (TM) subsets, the latter of which includes effector memory (TEM) and central memory (TCM) cells. Murine studies in which allogeneic TCD bone marrow (BM) is transplanted with purified T cells from individual T cell subsets to irradiated minor H antigen disparate recipients have demonstrated that the most severe GVHD results from transplanting T cells of the TN subset. Purified TCM causes mild GVHD and TEM do not cause detectable GVHD and can transfer immunity to pathogens.In vitro studies have similarly demonstrated that human donor CD8+ T cells specific for recipient minor H antigens are found predominantly within the TN cell subset, suggesting selective TN cell depletion may alter the GVHD incidence and/or severity in human HCT. Methods and results We developed an effective process for engineering human peripheral blood stem cell (PBSC) grafts that depletes CD45RA+ TN cells and retains CD34+ stem cells and functional CD45RO+ TM cells specific for a broad range of opportunistic pathogens (Bleakley BBMT 2014). We are conducting clinical trials to evaluate the selective depletion of TN cells from HLA-matched allogeneic PBSC grafts for the prevention of GVHD in patients with acute leukemia, the first of which has been published (Bleakley JCI 2015, N=35). Seventy patients have now been treated on three consecutive phase II trials. The median age was 34 years (1-56 years), 56% of patients had a diagnosis of ALL, 46% had previously relapsed or had detectable disease (MRD or relapse) at the time of HCT, and 23% had unrelated donor (URD) grafts. Intensive myeloablative, TBI-containing (13.2Gy) conditioning was used for 63 patients, whilst 7 patients received a medium intensity 'midi' preparative regimen, including 4Gy of TBI. The TN-depletion procedure was successfully performed on URD PBSC products shipped overnight from donor centers throughout the US, as well as on MRD PBSC collected at our centers. Reliable engraftment with high-level donor chimerism was observed in recipients of 'midi' as well as intensive myeloablative conditioning. The 2-year estimates of overall survival, disease-free survival, survival free of relapse and chronic GVHD (CRFS) and survival free of relapse, grade II-IV acute GVHD, and chronic GVHD (GRFS) are 79%, 73%, 69% and 63% respectively. Median follow-up among survivors is 26 months. The frequency and severity of chronic GVHD is remarkably low (5%) compared to historical rates of 40-60% chronic GVHD in HLA-matched PBSC transplantation with conventional calcineurin inhibitor-based immunosuppression. Relapse and non-relapse mortality (NRM) are acceptably low at 19% and 8%, respectively. No NRM occurred in patients <40 years. Updated results will be presented. Conclusions The outcomes of recipients of TN-depleted PBSC grafts compare very favorably to published results of HCT for patients with acute leukemia. For example, the 69% incidence of CRFS at 2 years in TN-depleted recipients compares with reported 2-year GRFS rates of 37% and 17% in recipients of allogeneic PBSC from HLA-matched related donors with or without ATG (Kroger et al. NEJM 2016). Our results suggest that TN-depletion of PBSC grafts may reduce the risk of chronic GVHD without negatively impacting other important HCT outcomes. Disclosures Riddell: Juno Therapeutics: Equity Ownership, Patents & Royalties, Research Funding; Cell Medica: Consultancy, Honoraria; Adaptive Biotechnologies: Consultancy, Honoraria.

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Acute Gvhd Induce the Critical Depletion of Naïve Pool in Regulatory T Cells: Implication for Linked Pathogenesis into Chronic Gvhd

Blood ◽

10.1182/blood.v126.23.923.923 ◽

2015 ◽

Vol 126 (23) ◽

pp. 923-923

Author(s):

Takanori Yoshioka ◽

Yusuke Meguri ◽

Takeru Asano ◽

Yuriko Kishi ◽

Miki Iwamoto ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Regulatory T Cells ◽

Immune Tolerance ◽

Cd4 T Cells ◽

Acute Gvhd ◽

Chronic Gvhd ◽

T Cell Subsets ◽

Effector Memory ◽

Ki 67

Abstract CD4+Foxp3+ regulatory T cells (Treg) play a central role in establishing immune tolerance after allogeneic hematopoietic stem cell transplantation (HSCT). We previously reported that the long-term severe lymphopenia could result in the collapse of Treg homeostasis leading to the onset of chronic GVHD (Matsuoka et al. JCI 2010). We recently found that, not only in the chronic phase but also in the acute phase, the homeostasis of Treg is more susceptible to the post-transplant environment as compared to other lymphocyte subsets (Yoshioka et al. ASH 2014). However, the impact of acute GVHD on Treg homeostasis and the pathogenesis of following chronic GVHD has not been well studied. In this study, we examined Treg reconstitution in the early phase after transplant in patients with or without acute GVHD. For the purpose, we obtained peripheral blood samples at 2, 4, 8 and 12 weeks after transplant from 52 patients who received allogeneic HSCT, and then analyzed CD4+CD25med-highCD127lowFoxp3+ Treg comparing with CD4+CD25neg-lowCD127highFoxp3- conventional T cell (Tcon) and CD8+ T cells. CD4 T cell subsets are further divided into subpopulations by the expression of CD45RA and CD31. The expressions of Helios, Ki-67 and Bcl-2 on these subsets were also examined. After transplant, total lymphocyte counts in examined patients were significantly lower than the counts before the start of conditioning (median lymphocytes 95/ul at 2 weeks and 302/ul at 4 weeks vs 600/ul before conditioning, P<0.01 and P<0.01, respectively). As we reported before, Treg showed the active proliferation without diminishing Bcl-2 levels in the severe lymphopenia, resulted in the increased %Treg of CD4 T cells at 4 weeks after transplant (%Treg of CD4 T cells; 12.2% at 4 weeks, 4.6% in healthy controls, P<0.005). 18 patients who developed acute GVHD were studied Treg homeostasis before and after the onset of GVHD more in detail. Before the onset of acute GVHD, % Ki-67+ cells in Treg and Tcon were in the equivalent levels in these patients. After the onset of acute GVHD, % Ki-67+ cells in Treg was dramatically increased from 19.1% to 61.2% (median) and this was significantly higher than % Ki-67+ cells in Tcon after acute GVHD (P<0.05). %Treg of total CD4 T cells were significantly increased after GVHD (% Treg; Median 7.2% vs 12.2%, P<0.004). Expanded Treg after acute GVHD showed a predominant Helios+CD45RA-CD31- effector/memory phenotype with the lower level of Bcl-2 expression as compared to CD45RA+ naïve Treg. As a consequence, naïve Treg pool including CD45RA+CD31+ recent thymic emigrant Treg (RTE-Treg) were critically decreased during acute GVHD (%CD45RA+ cells; 12.7% into 6.5%, P<0.004: CD45RA+CD31+ cells; 3.6% into 2.1%, P<0.003). In contrast, Tcon still retained a relatively higher level of naïve pool (%CD45RA+ cells; 20.5%, % CD45RA+CD31+ cells; 10.9%) after acute GVHD. These data indicated that Treg proliferation was rapidly promoted in face with the inflammatory condition during acute GVHD and this appears to contribute the amelioration of developing GVHD. However, the prompt reaction resulted in the depletion of naïve Treg pool which is important to maintain stable Treg homeostasis in the long period. In conclusion, our findings suggest that acute GVHD drives aggressive Treg proliferation resulting in the increased percentage of this subset but this also induce the severe alteration of Treg homeostasis by depleting naïve Treg, which may provide the linked pathogenesis of the subsequent onset of chronic GVHD. The careful monitoring of Treg from the point of view might provide important information to promote immune tolerance. Disclosures No relevant conflicts of interest to declare.

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Functionally specialized human CD4+ T-cell subsets express physicochemically distinct TCRs

eLife ◽

10.7554/elife.57063 ◽

2020 ◽

Vol 9 ◽

Author(s):

Sofya A Kasatskaya ◽

Kristin Ladell ◽

Evgeniy S Egorov ◽

Kelly L Miners ◽

Alexey N Davydov ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

T Cell Receptors ◽

Adaptive Immune System ◽

T Cell Subsets ◽

Effector Memory ◽

Intrinsic Properties ◽

Antigen Specificity ◽

Adaptive Immune ◽

Transition Pathways

The organizational integrity of the adaptive immune system is determined by functionally discrete subsets of CD4+ T cells, but it has remained unclear to what extent lineage choice is influenced by clonotypically expressed T-cell receptors (TCRs). To address this issue, we used a high-throughput approach to profile the αβ TCR repertoires of human naive and effector/memory CD4+ T-cell subsets, irrespective of antigen specificity. Highly conserved physicochemical and recombinatorial features were encoded on a subset-specific basis in the effector/memory compartment. Clonal tracking further identified forbidden and permitted transition pathways, mapping effector/memory subsets related by interconversion or ontogeny. Public sequences were largely confined to particular effector/memory subsets, including regulatory T cells (Tregs), which also displayed hardwired repertoire features in the naive compartment. Accordingly, these cumulative repertoire portraits establish a link between clonotype fate decisions in the complex world of CD4+ T cells and the intrinsic properties of somatically rearranged TCRs.

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Mitochondrial Dysfunction Associates With Acute T Lymphocytopenia and Impaired Functionality in COVID-19 Patients

Frontiers in Immunology ◽

10.3389/fimmu.2021.799896 ◽

2022 ◽

Vol 12 ◽

Author(s):

Yufei Mo ◽

Kelvin Kai-Wang To ◽

Runhong Zhou ◽

Li Liu ◽

Tianyu Cao ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Mitochondrial Dysfunction ◽

Cd8 T Cells ◽

Functional Impairment ◽

Cell Loss ◽

Cd8 T Cell ◽

Underlying Mechanism ◽

T Cell Subsets ◽

Effector Memory

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection results in rapid T lymphocytopenia and functional impairment of T cells. The underlying mechanism, however, remains incompletely understood. In this study, we focused on characterizing the phenotype and kinetics of T-cell subsets with mitochondrial dysfunction (MD) by multicolor flow cytometry and investigating the association between MD and T-cell functionality. While 73.9% of study subjects displayed clinical lymphocytopenia upon hospital admission, a significant reduction of CD4 or CD8 T-cell frequency was found in all asymptomatic, symptomatic, and convalescent cases. CD4 and CD8 T cells with increased MD were found in both asymptomatic and symptomatic patients within the first week of symptom onset. Lower proportion of memory CD8 T cell with MD was found in severe patients than in mild ones at the stage of disease progression. Critically, the frequency of T cells with MD in symptomatic patients was preferentially associated with CD4 T-cell loss and CD8 T-cell hyperactivation, respectively. Patients bearing effector memory CD4 and CD8 T cells with the phenotype of high MD exhibited poorer T-cell responses upon either phorbol 12-myristate-13-acetate (PMA)/ionomycin or SARS-CoV-2 peptide stimulation than those with low MD. Our findings demonstrated an MD-associated mechanism underlying SARS-CoV-2-induced T lymphocytopenia and functional impairment during the acute phase of infection.

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BCG vaccination induces enhanced frequencies of memory T cells and altered plasma levels of common γc cytokines in elderly individuals

PLoS ONE ◽

10.1371/journal.pone.0258743 ◽

2021 ◽

Vol 16 (11) ◽

pp. e0258743

Author(s):

Nathella Pavan Kumar ◽

Chandrasekaran Padmapriyadarsini ◽

Anuradha Rajamanickam ◽

Perumal Kannabiran Bhavani ◽

Arul Nancy ◽

...

Keyword(s):

T Cells ◽

Stem Cell ◽

T Cell ◽

Cd8 T Cells ◽

Plasma Levels ◽

T Cell Subsets ◽

Effector Memory ◽

Elderly Individuals ◽

Bcg Vaccination ◽

Adaptive Immune

BCG vaccination is known to induce innate immune memory, which confers protection against heterologous infections. However, the effect of BCG vaccination on the conventional adaptive immune cells subsets is not well characterized. We investigated the impact of BCG vaccination on the frequencies of T cell subsets and common gamma c (γc) cytokines in a group of healthy elderly individuals (age 60–80 years) at one month post vaccination as part of our clinical study to examine the effect of BCG on COVID-19. Our results demonstrate that BCG vaccination induced enhanced frequencies of central (p<0.0001) and effector memory (p<0.0001) CD4+ T cells and diminished frequencies of naïve (p<0.0001), transitional memory (p<0.0001), stem cell memory (p = 0.0001) CD4+ T cells and regulatory T cells. In addition, BCG vaccination induced enhanced frequencies of central (p = 0.0008), effector (p<0.0001) and terminal effector memory (p<0.0001) CD8+ T cells and diminished frequencies of naïve (p<0.0001), transitional memory (p<0.0001) and stem cell memory (p = 0.0034) CD8+T cells. BCG vaccination also induced enhanced plasma levels of IL-7 (p<0.0001) and IL-15 (p = 0.0020) but diminished levels of IL-2 (p = 0.0033) and IL-21 (p = 0.0020). Thus, BCG vaccination was associated with enhanced memory T cell subsets as well as memory enhancing γc cytokines in elderly individuals, suggesting its ability to induce non-specific adaptive immune responses.

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CD4+ T cells are essential for the development of destructive thyroiditis induced by anti–PD-1 antibody in thyroglobulin-immunized mice

Science Translational Medicine ◽

10.1126/scitranslmed.abb7495 ◽

2021 ◽

Vol 13 (593) ◽

pp. eabb7495

Author(s):

Yoshinori Yasuda ◽

Shintaro Iwama ◽

Daisuke Sugiyama ◽

Takayuki Okuji ◽

Tomoko Kobayashi ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Lymph Nodes ◽

Mononuclear Cells ◽

Critical Role ◽

T Cell Subsets ◽

Effector Memory ◽

Histopathological Analysis ◽

Activated T Cells ◽

Cervical Lymph Nodes

Immune-related adverse events induced by anti–programmed cell death–1 antibodies (PD-1-Ab), including destructive thyroiditis (thyroid-irAE), are thought to be caused by activated T cells. However, the T cell subsets that are directly responsible for damaging self-organs remain unclear. To clarify which T cell subsets are involved in the development of thyroid-irAE, a mouse model of thyroid-irAE was analyzed. PD-1-Ab administration 2.5 months after immunization with thyroglobulin caused destructive thyroiditis. Thyroiditis was completely prevented by previous depletion of CD4+ T cells and partially prevented by depleting CD8+ T cells. The frequencies of central and effector memory CD4+ T cell subsets and the secretion of interferon-γ after stimulation with thyroglobulin were increased in the cervical lymph nodes of mice with thyroid-irAE compared with controls. Histopathological analysis revealed infiltration of CD4+ T cells expressing granzyme B in thyroid glands and major histocompatibility complex class II expression on thyrocytes in mice with thyroid-irAE. Adoptive transfer of CD4+ T cells from cervical lymph nodes in mice with thyroid-irAE caused destruction of thyroid follicular architecture in the irradiated recipient mice. Flow cytometric analyses showed that the frequencies of central and effector memory CD4+ T cells expressing the cytotoxic marker CD27 were higher in peripheral blood mononuclear cells collected from patients with thyroid-irAE induced by PD-1-Ab versus those without. These data suggest a critical role for cytotoxic memory CD4+ T cells activated by PD-1-Ab in the pathogenesis of thyroid-irAE.

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EP.WE.805From bench to bedside; A rationale for Immunotherapy in the adjuvant setting for Oesophageal cancer

British Journal of Surgery ◽

10.1093/bjs/znab308.094 ◽

2021 ◽

Vol 108 (Supplement_7) ◽

Author(s):

Noel Donlon ◽

Maria Davern ◽

Andrew Sheppard ◽

John Reynolds ◽

Joanne Lysaght

Keyword(s):

T Cells ◽

T Cell ◽

T Cell Activation ◽

Immune Checkpoint ◽

Cell Activation ◽

Single Agent ◽

T Cell Subsets ◽

Effector Memory ◽

Post Surgery ◽

Post Operative Period

Abstract Background Immunotherapy is being intensively investigated for its utilisation in the curative setting as a single agent and in the multimodal setting, however, the most appropriate time to incorporate ICIs remains unknown. Our study profiles systemic anti-tumour immunity perioperatively to provide a rationale for adjuvant immunotherapy. Methods Systemic immunity was immunophenotyped pre and post-oesophagectomy on days 0, 1, 3, 7 and week 6 by flow cytometry (n = 14). The frequency of circulating lymphocytes, T cells, cytotoxic and helper T lymphocytes was profiled longitudinally including the proportion of T cell subsets in circulation. This study also profiled immune checkpoint expression on circulating T cells including: PD-1, CTLA-4, TIGIT, TIM-3, LAG-3, PD-L1 and PD-L2. Markers of immunogenicity (calreticulin, HMGB1 and MIC-A/B) were also assessed. Results The frequency of circulating CD27 + T cells increases sequentially in the immediate post-operative period peaking on day 7 in OAC patients. (p < 0.01) There is a sequential decrease in the percentage of effector memory and central memory T cells in circulation and an increase in the percentage of naïve T cells in peripheral circulation of OAC patients in the immediate post-operative period. The expression of CTLA-4 on the surface of circulating CD4 + T cells decreases 6 weeks post-operatively in OAC patients. Conclusions We observed increased T cell activation and immune checkpoints immediately post-surgery with returns to baseline by week 6. These results suggest that immune checkpoint inhibitors such as anti-PD-1 may be beneficial immediately post-surgery to maintain T cell activation and prevent exhaustion of this increased population of activated T cells observed immediately post-surgery.

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Functional Analysis of Effector and Regulatory T Cells in a Parasitic Nematode Infection

Infection and Immunity ◽

10.1128/iai.01233-07 ◽

2008 ◽

Vol 76 (5) ◽

pp. 1908-1919 ◽

Cited By ~ 87

Author(s):

Sebastian Rausch ◽

Jochen Huehn ◽

Dennis Kirchhoff ◽

Justyna Rzepecka ◽

Corinna Schnoeller ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Adoptive Transfer ◽

Chronic Phase ◽

T Cell Subsets ◽

Interleukin 4 ◽

Effector Memory ◽

Parasitic Nematodes ◽

Cell Reactivity

ABSTRACT Parasitic nematodes typically modulate T-cell reactivity, primarily during the chronic phase of infection. We analyzed the role of CD4-positive (CD4+) T effector (Teff) cells and regulatory T (Treg) cells derived from mice chronically infected with the intestinal nematode Heligmosomoides polygyrus. Different CD4+ T-cell subsets were transferred into naïve recipients that were subsequently infected with H. polygyrus. Adoptive transfer of conventional Teff cells conferred protection and led to a significant decrease in the worm burdens of H. polygyrus-infected recipients. Roughly 0.2% of the CD4+ T cells were H. polygyrus specific based on expression of CD154, and cells producing interleukin 4 (IL-4) and IL-13 were highly enriched within the CD154+ population. In contrast, adoptive transfer of Treg cells, characterized by the markers CD25 and CD103 and the transcription factor Foxp3, had no effect on the worm burdens of recipients. Further analysis showed that soon after infection, the number of Foxp3+ Treg cells temporarily increased in the inflamed tissue while effector/memory-like CD103+ Foxp+ Treg cells systemically increased in the draining lymph nodes and spleen. In addition, Treg cells represented a potential source of IL-10 and reduced the expression of IL-4. Finally, under in vitro conditions, Treg cells from infected mice were more potent suppressors than cells derived from naïve mice. In conclusion, our data indicate that small numbers of Teff cells have the ability to promote host protective immune responses, even in the presence of Treg cells.

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Ex Vivo Fludarabine Selectively Depletes Human Naive T-Cell Subsets: A Step Toward Modifying Donor Lymphocyte Infusion.

Blood ◽

10.1182/blood.v106.11.1071.1071 ◽

2005 ◽

Vol 106 (11) ◽

pp. 1071-1071

Author(s):

Melody M. Smith ◽

Cynthia R. Giver ◽

Edmund K. Waller ◽

Christopher R. Flowers

Keyword(s):

T Cells ◽

T Cell ◽

Cd8 T Cells ◽

Cd4 T Cells ◽

Memory T Cells ◽

Ex Vivo ◽

T Cell Subsets ◽

Effector Memory ◽

Naïve T Cell ◽

Naive T Cell

Abstract Ex vivo modification of donor lymphocytes with purine analogs (mDL) may help to minimize graft versus host disease (GvHD) while providing beneficial graft versus leukemia (GvL) effects. In a murine model system, we have shown that allogeneic donor splenocytes, treated with fludarabine ex vivo have significantly reduced GvHD activity when transferred to irradiated recipient mice, and retain anti-viral and GvL activities (Giver, 2003). This effect appears to be mediated by relative depletion of donor CD4 CD44low, “naive” T-cells. As a first step toward developing mDL for use in patients, we sought to evaluate the effects of ex vivo fludarabine exposure on human T-cell subsets, and to determine the minimum dose of fludarabine required to achieve this effect. Methods: Peripheral blood mononuclear cell samples from 6 healthy volunteers were evaluated at 0, 24, 48, and 72 hour time points after ex vivo incubation in varying dosages of fludarabine: 2, 5, and 10(n=3) mcg/ml. Fludarabine incubated samples were compared to samples that received no fludarabine (untreated). The total viable cell number was determined and the fractions and absolute numbers of viable CD4 and CD8 naïve and memory T-cells were determined using flow cytometry after incubation with 7-AAD (dead cell stain), CD4, CD8, CD45RA, CD62L, and CCR7 antibodies, and measuring the total viable cells/ml. Results: The numbers of viable CD4 and CD8 T-cells remained relatively stable in control cultures. Without fludarabine, the average viability at 72 hr of naive and memory T-cells were 92% and 77% for CD4 and 86% and 63% for CD 8 (Fig. 1A). Naive CD4 T-cells were more sensitive to fludarabine-induced death than memory CD4 cells. At 72 hr, the average viability of fludarabine-treated naive CD4 T-cells was 33% at 2 mcg/ml (8.2X the reduction observed in untreated cells) and 30% at 5 mcg/ml, while memory CD4 T-cells averaged 47% viability at 2 mcg/ml (2.3X the reduction observed in untreated cells) (Fig. 1B) and 38% at 5 mcg/ml. The average viability of naive CD8 T-cells at 72 hr was 27% at 2 mcg/ml and 20% at 5 mcg/ml, while memory CD8 T-cell viability was 22% at 2 mcg/ml and 17% at 5 mcg/ml. Analyses on central memory, effector memory, and Temra T-cells, and B-cell and dendritic cell subsets are ongoing. The 5 and 10 mcg/ml doses also yielded similar results in 3 initial subjects, suggesting that 2 mcg/ml or a lower dose of fludarabine is sufficient to achieve relative depletion of the naive T-cell subset. Conclusions: Future work will determine the minimal dose of fludarabine to achieve this effect, test the feasibility of using ex vivo nucleoside analog incubation to reduce alloreactivity in samples from patient/donor pairs, and determine the maximum tolerated dose of mDL in a phase 1 clinical trial with patients at high risk for relapse and infectious complications following allogeneic transplantation. Figure Figure

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