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Age and gender-related variability of main lymphocyte subsets (T, B and NK cell absolute counts and percentages from Ly; T4, T8 and DN cell absolute counts and percentages from lymphocytes and from T cells; T4:T8 and T:B ratios) was studied in a large cohort of pediatric patients (2 days - 17 years) at yearly intervals. 4128 6-color TBNK tests performed on BD FACSCanto II flow cytometer were assessed; patients with immune deficiencies and tumors were not included. The study revealed significant age and gender-related changes in all subsets. Absolute counts of T, B, T4 cells dropped from neonates to adolescents, decrease of T8 and NK cells was milder; relative count of T cells increased with age and that of B cells decreased; T4:T8 ratio went down and T:B ratio grew. Total T, T4 cells and T4:T8 ratio were significantly higher in girls, while T8, NK and DN cells - in boys; significantly higher relative and absolute B cell counts in boys appeared in adolescence. We compared our results with reference values for healthy children (Tosato et al., 2015), there was a good concordance, except for DN cells. Advantages of using patient cohort instead of healthy children as reference, possibilities for adjusting age and gender-specific reference ranges and potential international data pooling are discussed. This article is protected by copyright. All rights reserved.
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BACKGROUND Hodgkin lymphoma (HL) transformation into diffuse large B-cell lymphoma (DLBCL) is uncommon, and scant information has been published on transformed high-grade lymphomas. Therefore, it is important to present and discuss cases of lymphoma transformation to make new information on disease progression, diagnosis, and treatment more readily available. In this paper, we present a case of HL transformation into DLBCL with atypical dissemination. CASE REPORT A 39-year-old woman presented with severe hip pain. A computed tomography (CT) scan was performed, which showed massive pathological retroperitoneal and pelvic lymphadenopathy. The lymph nodes were biopsied and revealed HL. The patient then underwent 7 cycles of ABVD therapy; however, clinical concern was raised for persistent disease due to the poor response to therapy. A vertebral body biopsy was performed to clarify the diagnosis, and histological analysis revealed DLBCL. Therefore, specific chemotherapy with the R-CHOP scheme was begun; the patient received 8 cycles of rituximab and residual lymphoma tissue irradiation. Two months later, magnetic resonance imaging later demonstrated radiological disease progression with multiple widespread metastases in the spinal vertebrae as well as prevertebral, epidural, intradural, and intramedullary metastatic spread. The patient underwent intrathecal chemotherapy and radiation therapy, after which, full metabolic remission was observed on PET/CT. CONCLUSIONS Vigilance should be maintained for patients with poor response to HL treatment owing to the possible transformation into DLBCL. However, even in such cases, full metabolic remission can be achieved with appropriate treatment.
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Doença de Hodgkin , Linfoma Difuso de Grandes Células B , Adulto , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Bleomicina/uso terapêutico , Dacarbazina/uso terapêutico , Doxorrubicina/uso terapêutico , Feminino , Doença de Hodgkin/tratamento farmacológico , Humanos , Linfoma Difuso de Grandes Células B/patologia , Tomografia por Emissão de Pósitrons combinada à Tomografia Computadorizada , Rituximab/uso terapêutico , Vimblastina/uso terapêuticoRESUMO
Monitoring of minimal residual disease (MRD) by flow cytometry (FCM) is a powerful prognostic tool for predicting outcomes in acute lymphoblastic leukemia (ALL). To apply FCM-MRD in large, collaborative trials, dedicated laboratory staff must be educated to concordantly high levels of expertise and their performance quality should be continuously monitored. We sought to install a unique and comprehensive training and quality control (QC) program involving a large number of reference laboratories within the international Berlin-Frankfurt-Münster (I-BFM) consortium, in order to complement the standardization of the methodology with an educational component and persistent quality control measures. Our QC and quality assurance (QA) program is based on four major cornerstones: (i) a twinning maturation program, (ii) obligatory participation in external QA programs (spiked sample send around, United Kingdom National External Quality Assessment Service (UK NEQAS)), (iii) regular participation in list-mode-data (LMD) file ring trials (FCM data file send arounds), and (iv) surveys of independent data derived from trial results. We demonstrate that the training of laboratories using experienced twinning partners, along with continuous educational feedback significantly improves the performance of laboratories in detecting and quantifying MRD in pediatric ALL patients. Overall, our extensive education and quality control program improved inter-laboratory concordance rates of FCM-MRD assessments and ultimately led to a very high conformity of risk estimates in independent patient cohorts.
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BACKGROUND AND OBJECTIVES: Adipose-derived mesenchymal stem cells (ADSCs) are promising candidates in regenerative medicine. The need for in vitro propagation to obtain therapeutic quantities of the cells imposes a risk of impaired functionality due to cellular senescence. The aim of the study was to analyze in vitro senescence of previously cryopreserved human ADSCs subjected to serial passages in cell culture. METHODS AND RESULTS: ADSC cultures from 8 donors were cultivated until proliferation arrest was reached. A gradual decline of ADSC fitness was observed by altered cell morphology, loss of proliferative, clonogenic and differentiation abilities and increased ß-galactosidase expression all of which occurred in a donor-specific manner. Relative telomere length (RTL) analysis revealed that only three tested cultures encountered replicative senescence. The presence of two ADSC subsets with significantly different RTL and cell size was discovered. The heterogeneity of ADSC cultures was supported by the intermittent nature of aging seen in tested samples. CONCLUSIONS: We conclude that the onset of in vitro senescence of ADSCs is a strongly donor-specific process which is complicated by the intricate dynamics of cell subsets present in ADSC population. This complexity needs to be carefully considered when elaborating protocols for personalized cellular therapy.
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The aim of this study was to evaluate human adipose-derived stem cells (ASCs) from passage 2 (P2) to P8 cultured in medium containing 5% autologous serum (AS) after a long-term cryopreservation with regards to their surface marker expression, differentiation potential, and immunosuppressive effect in vitro. 8-color flow cytometry and real time PCR were used to determine mesenchymal stem cell (MSC) surface marker expression on ASCs from various passages. In vitro differentiation ability and immunomodulatory properties of ASCs were also tested. Flow cytometry showed that all ASCs express typical MSC markers CD29, CD44, CD73, CD90, CD105 simultaneously, but do not express such markers as HLA-DR, CD34, CD14, CD19, and CD45. Furthermore, median fluorescence intensity of positive cell surface markers increased with each subsequent passage indicating the accumulation of protein expression. The multilineage differentiation demonstrated the ability of ASCs from P6 to efficiently differentiate into adipocytes and chondrocytes, but their potential of osteogenic differentiation was diminished. Data from co-culture of ASCs and autologous peripheral blood mononuclear cells (PBMNCs) indicated that ASCs from P3, P6, and P9 significantly reduce the proliferation of PBMNCs at ASCs:PBMNCs ratio 1:1 and this suppression is dose dependent. This study demonstrated that ASCs from P2 to P8, cultured in the presence of AS, represent a highly homogeneous cell population with a peak accumulation of MSC surface proteins at P5 possessing multilineage differentiation ability and significant immunosuppressive properties after double freezing and more than 4 years of cryopreservation.
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Tecido Adiposo/citologia , Proliferação de Células , Criopreservação , Células-Tronco Mesenquimais/metabolismo , Medicina Regenerativa/métodos , Soro/metabolismo , Adipócitos/metabolismo , Adulto , Biomarcadores/metabolismo , Comunicação Celular , Diferenciação Celular , Linhagem da Célula , Separação Celular , Forma Celular , Células Cultivadas , Condrócitos/metabolismo , Técnicas de Cocultura , Citometria de Fluxo , Humanos , Tolerância Imunológica , Leucócitos Mononucleares/imunologia , Leucócitos Mononucleares/metabolismo , Masculino , Células-Tronco Mesenquimais/imunologia , Osteoblastos/metabolismo , Fenótipo , Reação em Cadeia da Polimerase em Tempo Real , Fatores de TempoRESUMO
BACKGROUND: The aim of this study was to evaluate impact of transplantation of bone marrow mesenchymal stromal cells (BM MMSCs) on recovery after polytrauma and bone fracture repair. METHODS: A total 27 Wistar-Kyoto rats were divided into three groups (n = 9): normal control (A), polytrauma (B), and polytrauma treated with BM MMSC transplantation (C). The experimental polytrauma model was made on male rats by causing multiple fractures and hemorrhagic shock. At 36 hours 9 days after surgery, nine rats received allogeneic BM MMSCs (1 × 10(6) cells per kilogram) intravenously. The day before operation and at Days 3 and 10 after surgery as well as at the end of the experiment, blood analysis was carried out. At 10, 20, and 30 days after surgery the rats' locomotor activity was assessed in an open-field test. At Day 30, rats were euthanized, and macroscopic and histologic observations of rats' lower extremities was performed. RESULTS: The treated animals gained weight faster regained their physical activity earlier. These outcomes were associated with locomotor activity test results, blood glucose and lactate ratios, as well as less marked muscle atrophy.Rat treatment with BM MMSC transplantation stimulated bone fracture healing-bone edge consolidation and enhanced callus formation, as well as the size and maturity of newly formed trabeculae.Red blood cell analysis results showed delayed recovery after hemorrhage in the rats receiving allogeneic BM MMSCs: restoration of red blood cell counts, hematocrit level, and hemoglobin level was slower in the untreated animals. CONCLUSION: Allogeneic BM MMSC transplantation improved rats rehabilitation scores after experimental polytrauma.