Study of the T-cell receptor repertoire by CDR3 spectratyping

Fozza, Claudio; Barraqueddu, Francesca; Corda, G; Contini, Salvatore; Virdis, Patrizia; Dore, Fausto; Bonfigli, Silvana; Longinotti, M

doi:10.1016/j.jim.2016.11.001

Cited by 22 publications

(19 citation statements)

References 123 publications

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“… 8 , 41 Clonality of T-cell subsets can be studied by flow cytometry analysis of Vβ usage, CDR3 size spectratyping or deep sequencing of VDJ combinations, and CDR3 nucleotide and amino acid sequences. 17 – 20 , 47 In SAA patients, expansion of at least one Vβ family in both CD4 + and CD8 + effector CD28 dim cells by flow cytometry with polyclonal features in CD4 + and oligoclonal characteristics in CD8 + cells has been described. 1 , 8 , 15 , 41 , 44 The CDR3 size skewing by spectratyping in the CD8 + population but not in CD4 + cells confirmed clonality in CD8 + cells.…”

Section: Discussionmentioning

confidence: 99%

Deep sequencing and flow cytometric characterization of expanded effector memory CD8 ⁺ CD57 ⁺ T cells frequently reveals T-cell receptor Vβ oligoclonality and CDR3 homology in acquired aplastic anemia

et al. 2018

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Oligoclonal expansion of CD8+ CD28− lymphocytes has been considered indirect evidence for a pathogenic immune response in acquired aplastic anemia. A subset of CD8+ CD28− cells with CD57 expression, termed effector memory cells, is expanded in several immune-mediated diseases and may have a role in immune surveillance. We hypothesized that effector memory CD8+CD28−CD57+ cells may drive aberrant oligoclonal expansion in aplastic anemia. We found CD8+CD57+ cells frequently expanded in the blood of aplastic anemia patients, with oligoclonal characteristics by flow cytometric Vβ usage analysis: skewing in 1–5 Vβ families and frequencies of immunodominant clones ranging from 1.98% to 66.5%. Oligoclonal characteristics were also observed in total CD8+ cells from aplastic anemia patients with CD8+CD57+ cell expansion by T-cell receptor deep sequencing, as well as the presence of 1–3 immunodominant clones. Oligoclonality was confirmed by T-cell receptor repertoire deep sequencing of enriched CD8+CD57+ cells, which also showed decreased diversity compared to total CD4+ and CD8+ cell pools. From analysis of complementarity-determining region 3 sequences in the CD8+ cell pool, a total of 29 sequences were shared between patients and controls, but these sequences were highly expressed in aplastic anemia subjects and also present in their immunodominant clones. In summary, expansion of effector memory CD8+ T cells is frequent in aplastic anemia and mirrors Vβ oligoclonal expansion. Flow cytometric Vβ usage analysis combined with deep sequencing technologies allows high resolution characterization of the T-cell receptor repertoire, and might represent a useful tool in the diagnosis and periodic evaluation of aplastic anemia patients. (Registered at clinicaltrials.gov identifiers: 00001620, 01623167, 00001397, 00071045, 00081523, 00961064)

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Section: Discussionmentioning

confidence: 99%

Deep sequencing and flow cytometric characterization of expanded effector memory CD8 ⁺ CD57 ⁺ T cells frequently reveals T-cell receptor Vβ oligoclonality and CDR3 homology in acquired aplastic anemia

et al. 2018

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“…This work also evaluated the TCR repertoire diversity with high sensitivity by using a panel of 24 Vβ-specific mAbs that recognize approximately 70% of the Vβ repertoire of CD3 + T cells present in healthy individuals. This strategy allowed us to assess the frequency of T cells expressing individual Vβ families ( 37 ) and provides a full picture of a large fraction of the T-cell compartment ( 38 ).…”

Section: Discussionmentioning

confidence: 99%

Loss of T-Cell Multifunctionality and TCR-Vβ Repertoire Against Epstein-Barr Virus Is Associated With Worse Prognosis and Clinical Parameters in HIV+ Patients

et al. 2018

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Epstein-Barr virus (EBV) is an oncogenic virus associated with the development of aggressive and poor-prognosis B-cell lymphomas in patients infected with human immunodeficiency virus (HIV+ patients). The most important risk factors for these malignancies include immune dysfunction, chronic immune activation, and loss of T-cell receptor (TCR) repertoire. The combination of all these factors can favor the reactivation of EBV, malignant cell transformation, and clinical progression toward B-cell lymphomas. The overarching aim of this study was to evaluate the frequency, phenotype, functionality, and distribution of TCR clonotypes for EBV-specific T-cell subpopulations in HIV+ patients at different clinical stages and for HIV+ patients with B-cell lymphoma, as well as to establish their association with clinical variables of prognostic value. Factors were studied in 56 HIV+ patients at different clinical stages and in six HIV+ subjects with diagnosed B-cell lymphoma. We found a significant decrease in all subpopulations of EBV-specific CD4+ T cells from HIV+ patients at stage 3 and with B-cell lymphoma. EBV-specific effector CD8+ T cells, particularly effector memory cells, were also reduced in HIV+ patients with B-cell lymphoma. Interestingly, these cells were unable to produce IFN-γ and lacked multifunctionality in HIV+ patients. The TCR-Vβ repertoire, which is key for protection against EBV in healthy individuals, was less diverse in HIV+ patients due to a lower frequency of TCR-Vβ2+, Vβ4+, Vβ7.1+, Vβ9+, Vβ13.6+, Vβ14+, Vβ17+, Vβ22+ CD4+, Vβ14+, and Vβ17+ CD8+ T cells. HIV+ patients with positive plasma EBV loads (EBV+HIV+) had a noteworthy decrease in the levels of both TNF-α+ and multifunctional TNF-α+/IL-2+ and TNF-α+/IFN-γ+ CD8+ T cells. Altogether, our findings demonstrate that HIV+ patients have significant alterations in the immune response to EBV (poor-quality immunity) that can favor viral reactivation, escalating the risk for developing EBV-associated B-cell lymphomas.

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“…Following stimulation, spectratyping was conducted to analyze the TCR Vβ families of Tregs at the CDR3 level and screen for clonal expansion of specific T cells. CDR3 spectratyping is a well-described method for measuring oligoclonality within T cell populations ( 32 ). Normal PBMC samples of a single Vβ family display a Gaussian distribution of 6–11 peaks, each separated by three nucleotides.…”

Section: Discussionmentioning

confidence: 99%

“…Normal PBMC samples of a single Vβ family display a Gaussian distribution of 6–11 peaks, each separated by three nucleotides. Each peak corresponds to a TCR transcript with a given CDR3 length that may contain numerous sequences ( 32 ). The number of TCR transcripts with a specific CDR3 length is proportional to the area under each peak.…”

Section: Discussionmentioning

confidence: 99%

Adoptive transfer of xenoantigen‑stimulated T cell receptor Vβ‑restricted human regulatory T cells prevents porcine islet xenograft rejection in humanized mice

Jin

Gong

et al. 2018

Mol Med Report

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Polyclonal expansion of human regulatory T cells (Tregs) prevents xenogeneic rejection by suppressing effector T cell responses in vitro and in vivo. However, a major limitation to using polyclonally expanded Tregs is that they may cause pan-immunosuppressive effects. The present study was conducted to compare the ability of ex vivo expanded human xenoantigen-stimulated Tregs (Xeno-Treg) and polyclonal Tregs (Poly-Treg) to protect islet xenografts from rejection in NOD-SCID interleukin (IL)-2 receptor (IL2r)γ−/− mice. Human cluster of differentiation (CD)4+CD25+CD127lo Tregs, expanded either by stimulating with porcine peripheral blood mononuclear cells (PBMCs) or anti-CD3/CD28 beads, were characterized by immune cell phenotyping, T cell receptor (TCR) Vβ CDR3 spectratyping and performing suppressive activity assays in vitro. The efficiency of adoptively transferred ex vivo human Tregs was evaluated in vivo using neonatal porcine islet cell clusters (NICC) transplanted into NOD-SCID IL-2rγ−/− mice, which received human PBMCs with or without Xeno-Treg or Poly-Treg. Xeno-Treg, which expressed increased levels of human leukocyte antigen-DR and secreted higher levels of IL-10, demonstrated enhanced suppressive capacity in a pig-human mixed lymphocyte reaction. Spectratypes of TCR Vβ4, Vβ10, Vβ18 and Vβ20 in Xeno-Treg showed restriction and expanded clones at sizes of 205, 441, 332 and 196 respectively, compared to those of Poly-Treg. Reconstitution of mice with human PBMCs and Poly-Treg resulted in NICC xenograft rejection at 63 days. Adoptive transfer with human PBMCs and Xeno-Treg prolonged islet xenograft survival beyond 84 days, with grafts containing intact insulin-secreting cells surrounded by a small number of human CD45+ cells. This study demonstrated that adoptive transfer of ex vivo expanded human Xeno-Treg may potently prevent islet xenograft rejection in humanized NOD-SCID IL2rγ−/− mice compared with Poly-Treg. These findings suggested that adoptive Treg therapy may be used for immunomodulation in islet xenotransplantation by minimizing systemic immunosuppression.

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Study of the T-cell receptor repertoire by CDR3 spectratyping

Cited by 22 publications

References 123 publications

Deep sequencing and flow cytometric characterization of expanded effector memory CD8 ⁺ CD57 ⁺ T cells frequently reveals T-cell receptor Vβ oligoclonality and CDR3 homology in acquired aplastic anemia

Deep sequencing and flow cytometric characterization of expanded effector memory CD8 ⁺ CD57 ⁺ T cells frequently reveals T-cell receptor Vβ oligoclonality and CDR3 homology in acquired aplastic anemia

Loss of T-Cell Multifunctionality and TCR-Vβ Repertoire Against Epstein-Barr Virus Is Associated With Worse Prognosis and Clinical Parameters in HIV+ Patients

Adoptive transfer of xenoantigen‑stimulated T cell receptor Vβ‑restricted human regulatory T cells prevents porcine islet xenograft rejection in humanized mice

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Study of the T-cell receptor repertoire by CDR3 spectratyping

Cited by 22 publications

References 123 publications

Deep sequencing and flow cytometric characterization of expanded effector memory CD8 + CD57 + T cells frequently reveals T-cell receptor Vβ oligoclonality and CDR3 homology in acquired aplastic anemia

Deep sequencing and flow cytometric characterization of expanded effector memory CD8 + CD57 + T cells frequently reveals T-cell receptor Vβ oligoclonality and CDR3 homology in acquired aplastic anemia

Loss of T-Cell Multifunctionality and TCR-Vβ Repertoire Against Epstein-Barr Virus Is Associated With Worse Prognosis and Clinical Parameters in HIV+ Patients

Adoptive transfer of xenoantigen‑stimulated T cell receptor Vβ‑restricted human regulatory T cells prevents porcine islet xenograft rejection in humanized mice

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Product

Resources

About

Deep sequencing and flow cytometric characterization of expanded effector memory CD8 ⁺ CD57 ⁺ T cells frequently reveals T-cell receptor Vβ oligoclonality and CDR3 homology in acquired aplastic anemia

Deep sequencing and flow cytometric characterization of expanded effector memory CD8 ⁺ CD57 ⁺ T cells frequently reveals T-cell receptor Vβ oligoclonality and CDR3 homology in acquired aplastic anemia