Allogeneic stem cell transplantation (alloSCT) is used for treating patients with T-prolymphocytic leukemia (T-PLL). However, direct evidence of GvL activity in T-PLL is lacking. We correlated minimal residual disease (MRD) kinetics with immune interventions and T-cell receptor (TCR) repertoire diversity alterations in patients after alloSCT for T-PLL. Longitudinal quantitative MRD monitoring was performed by clone-specific real-time PCR of TCR rearrangements (n=7), and TCR repertoire diversity assessment by next-generation sequencing (NGS; n=3) Although post-transplant immunomodulation (immunosuppression tapering or donor lymphocyte infusions) resulted in significant reduction (>1 log) of MRD levels in 7 of 10 occasions, durable MRD clearance was observed in only two patients. In all three patients analyzed by TCR-NGS, MRD responses were reproducibly associated with a shift from a clonal, T-PLL-driven profile to a polyclonal signature. Novel clonotypes that could explain a clonal GvL effect did not emerge. In conclusion, TCR-based MRD quantification appears to be a suitable tool for monitoring and guiding treatment interventions in T-PLL. The MRD responses to immune modulation observed here provide first molecular evidence for GvL activity in T-PLL which, however, may be often only transient and reliant on a poly-/oligoclonal rather than a monoclonal T-cell response.

• MeSH
• buněčné klony imunologie   MeSH
• dospělí MeSH
• genová přestavba T-lymfocytů genetika   MeSH
• homologní transplantace MeSH
• imunomodulace * MeSH
• kinetika MeSH
• kvantitativní polymerázová řetězová reakce MeSH
• lidé středního věku MeSH
• lidé MeSH
• reakce štěpu proti leukémii * MeSH
• receptory antigenů T-buněk analýza   genetika   MeSH
• reziduální nádor diagnóza   genetika   MeSH
• senioři MeSH
• T-buněčná prolymfocytární leukemie diagnóza   terapie   MeSH
• transplantace kmenových buněk metody   MeSH
• vysoce účinné nukleotidové sekvenování MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• senioři MeSH
• Publikační typ
• časopisecké články MeSH

The TCR signal transduction is initiated by the activation of Src-family kinases (SFK) which phosphorylate Immunoreceptor tyrosine-based activation motifs (ITAM) present in the intracellular parts of the T-cell receptor (TCR) signaling subunits. Numerous data suggest that after stimulation TCR interacts with membrane rafts and thus it gains access to SFK and other important molecules involved in signal transduction. However, the precise mechanism of this process is unclear. One of the key questions is how SFK access TCR and what is the importance of non-raft and membrane raft-associated SFK for the initiation and maintenance of the TCR signaling. To answer this question we targeted a negative regulator of SFK, C-terminal Src kinase (Csk) to membrane rafts, recently described "heavy rafts" or non-raft membrane. Our data show that only Csk targeted into "classical" raft but not to "heavy raft" or non-raft membrane effectively inhibits TCR signaling, demonstrating the critical role of membrane raft-associated SFK in this process.

• MeSH
• buněčná membrána metabolismus   MeSH
• fosforylace MeSH
• imunoblotting MeSH
• kultivované buňky MeSH
• lidé MeSH
• membránové mikrodomény MeSH
• protoonkogenní proteiny metabolismus   MeSH
• receptory antigenů T-buněk metabolismus   MeSH
• signální transdukce MeSH
• tyrosinkinasy metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

To confirm a diagnosis of malignant lymphomas it is imperative to distinguish between reactive and neoplastic proliferation. The PCR (polymerase chain reaction) is a method that can be used for detection of clonal rearrangements of the immunoglobulin heavy chain (IgH) and T-cell receptor (TCR) genes. This study summarizes the outcomes of PCR analysis of IgH and TCR gene rearrangements in 91 bioptic cases of lymphoproliferative disorders. In the class of B lymphomas we detected clonal IgH rearrangement in nearly 83% of cases and in class of T lymphomas in 81% of cases. We can affirm that PCR analysis of B and T cell clonality on DNA extracted from the whole section of formalin-fixed, paraffin-embedded tissue is very suitable for routinely elaborate this. Its influence on the diagnostics of morphological unclear cases in particular, is crucial and is useful in establishing a diagnosis of lymphoid neoplasias in specimens in which histological and immunophenotypic studies are inconclusive.

• MeSH
• financování organizované MeSH
• fixativa MeSH
• formaldehyd MeSH
• genová přestavba T-lymfocytů MeSH
• lidé MeSH
• lymfoproliferativní nemoci diagnóza   genetika   MeSH
• polymerázová řetězová reakce MeSH
• přestavba genů pro těžké řetězce B-lymfocytů MeSH
• zalévání tkání do parafínu MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• hodnotící studie MeSH

TCR gene rearrangement generates diversity of T lymphocytes by V(D)J recombination. Ig genes are rearranged in B cells using the same enzyme machinery. TCRD (TCR delta) genes are frequently incompletely rearranged in B precursor leukemias and recently were found in a significant portion of physiological B lymphocytes. Incomplete TCRD rearrangements (V-D) thus serve as natural indicators of previous V(D)J recombinase activity. Functional V(D)J recombinase has recently been found in murine NK precursors. We tested whether physiological NK cells and other leukocyte subpopulations contained TCR rearrangements in humans. This would provide evidence that V(D)J recombinase was active in the ancestry cells and suggest common pathways among the positive cell types. TCRD were rearranged in 3.2-36% of NK cells but not in nonlymphoid leukocytes. The previously known phenomenon of TCRD transcription in NK cells is a possible mechanism that maintains the chromatin open at the TCRD locus. In comparison, TCRG rearrangements were frequent in T cells, low to negative in B and NK cells, and negative in nonlymphoid cells, suggesting a tighter control of TCRG. Levels of TCRD rearrangements were similar among the B lymphocyte subsets (B1-B2, naive-memory). In conclusion, human NK cells pass through a differentiation step with active V(D)J recombinase similar to T and B lymphocytes and unlike nonlymphoid leukocytes. This contradicts recent challenges to the concept of separate lymphoid and myeloid differentiation.

• MeSH
• buněčná diferenciace genetika   imunologie   MeSH
• buňky HT-29 MeSH
• buňky NK MeSH
• finanční podpora výzkumu jako téma MeSH
• genetické markery MeSH
• genová přestavba - delta řetězec receptoru antigenů T-buněk fyziologie   MeSH
• HeLa buňky MeSH
• lidé MeSH
• nádorové buněčné linie MeSH
• podskupiny B-lymfocytů MeSH
• podskupiny lymfocytů cytologie   imunologie   metabolismus   MeSH
• VDJ-rekombinasy metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• srovnávací studie MeSH

A novel immature human T-ALL cell line, UHKT-42, was established from a 12 year old male patient with acute undifferentiated leukemia. The cell line expressed surface CD7, CD5 and cytoplasmic CD3 antigens. All other T-lymphocytic antigens were undetectable on the surface or in the cytoplasm of cultured cells. Expression of the T-cell receptor (TCR) beta, TCR delta, CD3 delta and CD3 epsilon genes was detected by Northern blotting in total cellular RNA extracts, however, the expression of TCR alpha and TCR gamma was undetectable. After stimulation by TPA for 3 days, only the appearance of CD25 (Tac antigen) was detected by immunofluorescence and flow cytometry. Secretion of interleukin-2 (IL-2) into the culture media was also detected after stimulation by PHA or TPA, but not in unstimulated cells. These results suggest that UHKT-42 cells are early precursors of T cells, with TCR beta/delta expression.

• MeSH
• akutní lymfatická leukemie imunologie   patologie   MeSH
• antigeny CD5 MeSH
• antigeny CD7 MeSH
• CD antigeny metabolismus   MeSH
• diferenciační antigeny T-lymfocytů metabolismus   MeSH
• dítě MeSH
• lidé MeSH
• nádorové buňky kultivované MeSH
• receptory antigenů T-buněk alfa-beta genetika   metabolismus   MeSH
• T-lymfocyty * metabolismus   patologie   MeSH
• Check Tag
• dítě MeSH
• lidé MeSH
• mužské pohlaví MeSH
• Publikační typ
• kazuistiky MeSH

γδ T cells are considered to be innate-like lymphocytes that respond rapidly to stress without clonal selection and differentiation. Here we use next-generation sequencing to probe how this paradigm relates to human Vδ2neg T cells, implicated in responses to viral infection and cancer. The prevalent Vδ1 T cell receptor (TCR) repertoire is private and initially unfocused in cord blood, typically becoming strongly focused on a few high-frequency clonotypes by adulthood. Clonal expansions have differentiated from a naive to effector phenotype associated with CD27 downregulation, retaining proliferative capacity and TCR sensitivity, displaying increased cytotoxic markers and altered homing capabilities, and remaining relatively stable over time. Contrastingly, Vδ2+ T cells express semi-invariant TCRs, which are present at birth and shared between individuals. Human Vδ1+ T cells have therefore evolved a distinct biology from the Vδ2+ subset, involving a central, personalized role for the γδ TCR in directing a highly adaptive yet unconventional form of immune surveillance.

• MeSH
• antigeny CD27 metabolismus   MeSH
• biologické markery metabolismus   MeSH
• buněčná diferenciace MeSH
• buněčné klony cytologie   MeSH
• CX3C chemokinový receptor 1 metabolismus   MeSH
• cytotoxicita imunologická MeSH
• dárci tkání MeSH
• dospělí MeSH
• fenotyp MeSH
• genetická variace MeSH
• hypervariabilní oblasti genetika   MeSH
• imunitní dozor * MeSH
• interleukin-15 farmakologie   MeSH
• lidé MeSH
• proliferace buněk MeSH
• receptory antigenů T-buněk gama-delta metabolismus   MeSH
• Check Tag
• dospělí MeSH
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

In spite of a comprehensive understanding of the schematics of T cell receptor (TCR) signaling, the mechanisms regulating compartmentalization of signaling molecules, their transient interactions, and rearrangement of membrane structures initiated upon TCR engagement remain an outstanding problem. These gaps in our knowledge are exemplified by recent data demonstrating that TCR triggering is largely dependent on a preactivated pool of Lck concentrated in T cells in a specific type of membrane microdomains. Our current model posits that in resting T cells all critical components of TCR triggering machinery including TCR/CD3, Lck, Fyn, CD45, PAG, and LAT are associated with distinct types of lipid-based microdomains which represent the smallest structural and functional units of membrane confinement able to negatively control enzymatic activities and substrate availability that is required for the initiation of TCR signaling. In addition, the microdomains based segregation spatially limits the interaction of components of TCR triggering machinery prior to the onset of TCR signaling and allows their rapid communication and signal amplification after TCR engagement, via the process of their coalescence. Microdomains mediated compartmentalization thus represents an essential membrane organizing principle in resting T cells. The integration of these structural and functional aspects of signaling into a unified model of TCR triggering will require a deeper understanding of membrane biology, novel interdisciplinary approaches and the generation of specific reagents. We believe that the fully integrated model of TCR signaling must be based on membrane structural network which provides a proper environment for regulatory processes controlling TCR triggering.

• Publikační typ
• časopisecké články MeSH

BACKGROUND: The Immunoglobulins (IG) and the T cell receptors (TR) play the key role in antigen recognition during the adaptive immune response. Recent progress in next-generation sequencing technologies has provided an opportunity for the deep T cell receptor repertoire profiling. However, a specialised software is required for the rational analysis of massive data generated by next-generation sequencing. RESULTS: Here we introduce tcR, a new R package, representing a platform for the advanced analysis of T cell receptor repertoires, which includes diversity measures, shared T cell receptor sequences identification, gene usage statistics computation and other widely used methods. The tool has proven its utility in recent research studies. CONCLUSIONS: tcR is an R package for the advanced analysis of T cell receptor repertoires after primary TR sequences extraction from raw sequencing reads. The stable version can be directly installed from The Comprehensive R Archive Network ( http://cran.r-project.org/mirrors.html ). The source code and development version are available at tcR GitHub ( http://imminfo.github.io/tcr/ ) along with the full documentation and typical usage examples.

• MeSH
• imunoglobuliny genetika   MeSH
• lidé MeSH
• programovací jazyk MeSH
• receptory antigenů T-buněk genetika   imunologie   MeSH
• sekvenční analýza DNA metody   MeSH
• software * MeSH
• vysoce účinné nukleotidové sekvenování metody   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

• MeSH
• finanční podpora výzkumu jako téma MeSH
• genetické markery imunologie   MeSH
• genová přestavba - delta řetězec receptoru antigenů T-buněk MeSH
• HeLa buňky MeSH
• klonování DNA MeSH
• lidé MeSH
• podskupiny B-lymfocytů imunologie   metabolismus   MeSH
• polymerázová řetězová reakce MeSH
• receptory antigenů T-buněk gama-delta analýza   genetika   krevní zásobení   MeSH
• Check Tag
• lidé MeSH

Membrane rafts and signaling molecules associated with them are thought to play important roles in immunoreceptor signaling. Rafts differ in their lipid and protein compositions from the rest of the membrane and are relatively resistant to solubilization by Triton X-100 or similar detergents, producing buoyant, detergent-resistant membranes (DRMs) that can be isolated by density gradient ultracentrifugation. One of the key signaling molecules present in T cell DRMs is the transmembrane adaptor protein LAT (linker for activation of T cells). In contrast to previous results, a recent study demonstrated that a LAT construct not present in the buoyant DRMs is fully able to support TCR signaling and development of T cells in vivo. This finding caused doubts about the real physiological role of rafts in TCR signaling. In this study, we demonstrate that these results can be explained by the existence of a novel type of membrane raft-like microdomains, producing upon detergent solubilization "heavy DRMs" containing a number of membrane molecules. At a moderate level of expression, LAT supported TCR signaling more efficiently than constructs targeted to the microdomains producing heavy DRMs or to nonraft membrane. We suggest that different types of membrane microdomains provide environments regulating the functional efficiencies of signaling molecules present therein.

• MeSH
• adaptorové proteiny signální transdukční imunologie   metabolismus   MeSH
• aktivace lymfocytů imunologie   MeSH
• Jurkat buňky MeSH
• lidé MeSH
• membránové mikrodomény chemie   imunologie   MeSH
• membránové proteiny chemie   imunologie   izolace a purifikace   metabolismus   MeSH
• receptory antigenů T-buněk imunologie   metabolismus   MeSH
• signální transdukce imunologie   MeSH
• T-lymfocyty chemie   imunologie   metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH