Graft-versus-host disease is a severe complication of allogeneic stem cell transplantation. The major role is played by alloreactive donor T-cell clones leading to host tissue damage. Selective depletion is a strategy to eliminate host-reactive donor T cells from haematopoietic stem cell allografts to prevent graft-versus-host disease while conserving useful donor immune functions. We have used irradiated peripheral blood mononuclear cells from cancer patients and healthy donor cells as responder cells in primary mixed leukocyte reaction. To prepare graft-versus leukaemia/myeloma-specific T cells, alloreactive T cells in primary mixed leukocyte reaction were depleted with anti-CD25 immunotoxin. The remaining T cells had insignificant alloreactivity in secondary mixed leukocyte reaction. Then, allodepleted donor T cells were repeatedly stimulated using purified leukaemia/tumour cells from the same cancer patient. Leukaemia/tumour-reactive donor T cells were purified using cell sorter on the basis of CD4 and CD8 activation. Their specificity was tested in nonradioactive cytotoxicity test. We performed 22 reactions (15 samples with leukemic and 7 samples with multiple myeloma cells). Selective depletion of alloreactive donor T cells with anti-CD25 immunotoxin led to significant depletion (99.2-100 %, median 99.7%). The effect of donor T cells was well preserved, while the graft-versus-host reactivation of donor cells was negligible, even after repeated stimulation with patient's non-tumour cells. Thus, it is possible to selectively deplete donor alloreactive T cells with anti-CD25 immunotoxin. In the cases of leukaemia patients, a strong graft-versus-leukaemia reactivity was noticed in allodepleted donor T cells; in myeloma patients, graft-versus-myeloma reactivity was less significant.

• MeSH
• antigeny CD4 metabolismus   MeSH
• cytotoxicita imunologická MeSH
• dárci tkání * MeSH
• homologní transplantace MeSH
• leukemie imunologie   patologie   MeSH
• lidé MeSH
• lymfocytární deplece * MeSH
• mnohočetný myelom imunologie   patologie   MeSH
• nemoc štěpu proti hostiteli imunologie   patologie   MeSH
• proliferace buněk MeSH
• receptor interleukinu-2 - alfa-podjednotka metabolismus   MeSH
• T-lymfocyty imunologie   MeSH
• test smíšené lymfocytární kultury MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Regulatory T cells (Tregs) are critical regulators of autoimmune diseases, including type 1 diabetes mellitus. It is hypothesised that Tregs' function can be influenced by changes in the expression of specific microRNAs (miRNAs). Thus, we performed miRNAs profiling in a population of Tregs separated from peripheral blood of five type 1 diabetic patients and six healthy donors. For more detailed molecular characterisation of Tregs, we additionally compared miRNAs expression profiles of Tregs and conventional T cells. Tregs were isolated according to CD3+, CD4+, CD25(hi)+ and CD127- by flow cytometry, and miRNA expression profiling was performed using TaqMan Array Human MicroRNA Panel-1 (384-well low density array). In Tregs of diabetic patients we found significantly increased expression of miRNA-510 (p=0.05) and decreased expression of both miRNA-342 (p<0.0001) and miRNA-191 (p=0.0079). When comparing Tregs and T cells, we revealed that Tregs had significant higher expression of miRNA-146a and lower expression of eight specific miRNAs (20b, 31, 99a, 100, 125b, 151, 335, and 365). To our knowledge, this is the first study demonstrating changes in miRNA expression profiles occurring in Tregs of T1D patients and a miRNAs signature of adult Tregs.

• MeSH
• diabetes mellitus 1. typu genetika   krev   MeSH
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mikro RNA genetika   MeSH
• mladý dospělý MeSH
• regulační T-lymfocyty metabolismus   MeSH
• sekvenční analýza hybridizací s uspořádaným souborem oligonukleotidů metody   MeSH
• shluková analýza MeSH
• stanovení celkové genové exprese MeSH
• T-lymfocyty metabolismus   MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladý dospělý MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• práce podpořená grantem MeSH

Despite novel treatment strategies, multiple myeloma (MM) remains an incurable disease with low immunogenicity and multiple immune defects. We developed an ex vivo strategy for inducing myeloma-specific cytotoxic T lymphocytes (CTLs) and demonstrate the possibility of identification and long-term in vivo monitoring of individual myeloma-specific T-cell clones using the most sensitive clonotypic assay that is able to detect low frequencies of T-cell clones (1 clonotypic cell in 10(6) cells). Ten patients with MM were examined for the presence of tumour-reactive T cells using dendritic cells loaded with autologous tumour cells. All patients had detectable myeloma-reactive T cells in vitro. Expanded myeloma-reactive T cells demonstrated specific cytotoxic effects against autologous tumour cells in vitro (median 39.6% at an effector:target ratio of 40:1). The clonality of myeloma-specific T cells was studied with a clonotypic assay, which demonstrated both oligoclonal and monoclonal populations of myeloma-specific T cells. CD8(+) CTLs were the most immunodominant myeloma-specific T-cell clones and clinical responses were closely associated with the in vivo expansion and long-term persistence of individual CD8(+) T-cell clones, usually at very low frequencies (10(-3)-10(-6)). We conclude that the clonotypic assay is the most sensitive tool for immunomonitoring of low-frequency T cells.

• MeSH
• aktivace lymfocytů imunologie   MeSH
• antigeny nádorové imunologie   MeSH
• buněčná diferenciace imunologie   MeSH
• buněčné klony imunologie   MeSH
• cytotoxicita imunologická imunologie   MeSH
• cytotoxické T-lymfocyty imunologie   MeSH
• dendritické buňky imunologie   MeSH
• hypervariabilní oblasti genetika   imunologie   MeSH
• imunodominantní epitopy imunologie   MeSH
• imunomagnetická separace metody   MeSH
• lidé středního věku MeSH
• lidé MeSH
• mnohočetný myelom imunologie   terapie   MeSH
• molekulární sekvence - údaje MeSH
• monitorování imunologické metody   MeSH
• nádorové buňky kultivované MeSH
• následné studie MeSH
• prezentace antigenu imunologie   MeSH
• sekvence aminokyselin MeSH
• senioři MeSH
• Check Tag
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH