Effect of cytomegalovirus (CMV) serostatus on the incidence and virological features of active CMV infection in allogeneic stem cell transplant recipients.

Polyethyleneimine-based immunopolyplex for targeted gene transfer in human lymphoma celllines

Background Specific and efficient delivery of genes into targeted cells is a priority objective in non-viral gene therapy. Polyethyleneimine-based polyplexes have been reported to be good non-viral transfection reagents. However, polyplex-mediated DNA delivery occurs through a non-specific mechanism. This article reports the construction of an immunopolyplex, a targeted non-viral vector based on a polyplex backbone, and its application in gene transfer over human lymphoma cell lines. Methods Targeting elements (biotin-labeled antibodies), which should recognize a specific element of the target cell membrane and promote nucleic acid entry into the cell, were attached to the polyplex backbone…

Loss of a novel tumor suppressor gene locus at chromosome 8p is associated with leukemic mantle cell lymphoma

Abstract Patients with mantle cell lymphoma (MCL) may present with either nodal or leukemic disease. The molecular determinants underlying this different biologic behavior are not known. This study compared the pattern of genetic abnormalities in patients with nodal and leukemic phases of MCL using comparative genomic hybridization (CGH) and fluorescence in situ hybridization (FISH) for specific gene loci. Although both leukemic and nodal MCL showed similar genomic patterns of losses (involving 6q, 11q22-q23, 13q14, and 17p13) and gains (affecting 3q and 8q), genomic loss of chromosome 8p occurred more frequently in patients with leukemic disease (79% versus 11%,P < .001). Subsequent…

Study of the Expression of Angiogenic Factors and Dopaminergic Receptors: Correlation with ZAP70, CD38 and the Mutational State in Chronic Lymphocytic Leukaemia (CLL).

Abstract The angiogenesis is a well known process implicate in the progression of CLL. In this disease has been described an increase of angiogenic and pro-angiogenic factors in serum and angiogenic receptors in B cells, mainly in advances stages. Recently had been known that the agonist of the dopaminergic receptor D2 can inhibit the tumour growth and a possible mechanism mediator is the internalization of VEGFR-2. The aim of this work had been to analyze the expression of the dopaminergic receptors (D1, D2, D3, D4 and D5), the angiogenic receptors and its correlation with the main biological factor implicated in the illness (ZAP-70, CD38 and the mutational status of IgVH/BCL-6). We had de…

Treatment of B-CLL Cells with Bortezomib and Rituximab Reduces Cell Viability In Vitro..

Introduction : In B-CLL, somatic mutation of IgVH genes defines a group of patients with favorable prognosis, whereas the absence of IgVH mutations is correlated with a worse outcome. Our previous data suggested that BCL-6 mutations identify a subgroup of patients with high risk of progression despite the presence of mutated IgVH gene, but the clinical significance of this molecular alteration remains uncertain. New approaches are now being tested for the treatment of B-CLL. Proteasome inhibitor, Bortezomib (Btz), and monoclonal antibodies specific for surface antigens, Rituximab (Rtx), represent potential therapeutic strategy. Objetives : To study the effects of Btz and Rtx on viability of…

Performance of the QuantiFERON-cytomegalovirus (CMV) assay for detection and estimation of the magnitude and functionality of the CMV-specific gamma interferon-producing CD8(+) T-cell response in allogeneic stem cell transplant recipients.

ABSTRACTThe performance of the QuantiFERON-cytomegalovirus (CMV) assay was compared to that of a flow cytometry intracellular cytokine staining (ICS) method for the detection of CMV-specific gamma interferon (IFN-γ)-producing CD8+T-cell responses in allogeneic stem cell transplant (allo-SCT) recipients and for estimations of their magnitude and functionality. A total of 90 whole-blood specimens from 23 allo-SCT recipients was analyzed by both methods. Overall, the percentage of specimens that yielded concordant results by both methods was 68.8% (κ = 0.691; 95% confidence interval [CI], 0.548 to 0.835), and the sensitivity of the QuantiFERON-CMV assay for the detection of positive IFN-γ T-ce…

Targeted oligonucleotide delivery in human lymphoma cell lines using a polyethyleneimine based immunopolyplex.

The efficacy of antisense gene therapy depends on efficient delivery of oligonucleotides into targeted cells. Although polyethyleneimine based polyplexes have been reported as good transfection reagents, they are inefficient in lymphoid cell transfection. We report the construction of an immunopolyplex, a targeted nonviral vector based on a polyplex backbone and its application for oligonucleotide transfer on human lymphoma cell lines. The salient characteristic of immunopolyplex lies in the possibility of easily replacing the targeting element (antibody), leaving the polyplex backbone intact. Furthermore, a study was made of the influence of endocytosis inhibitors on immunopolyplex activit…

Decrease of CD4+ T Lymphocytes after myocardial infarction is related with extensive myocardial fibrosis

Purpose: Myocardial fibrosis plays a potential role in left ventricular remodeling and patients' outcome. After myocardial infarction innate immune cells infiltrate infarcted area and replace necrotic tissue by fibrotic tissue. However the role of adaptive immunity, especially T cells, has not yet been investigated in this scenario. Methods: We studied 94 patients with a first STEMI treated with percutaneous revascularization. Leucocyte subsets and a wide variety of lymphocyte subtypes were determined in peripheral blood 24 h after reperfusion by means of flow cytometry. Infarct size and cardiac fibrosis were measured by late enhancement Cardiac Magnetic Resonance (CRM) 1 week and 6 months …

Human cytomegalovirus (HCMV)-specific CD4+ T lymphocyte response in AIDS patients with no past or current HCMV disease following HAART.

Abstract Background: The incidence of Human Cytomegalovirus (HCMV) end-organ disease has dramatically decreased since the implementation of highly active antiretroviral therapies (HAARTs), but the precise immune mechanism whereby HCMV is controlled remains to be elucidated. Objectives: To investigate the effect of (HAART) on CD4 + T-cell immunity to HCMV in AIDS patients with no past or current HCMV disease. Study design: Seventeen patients were prospectively examined for CD4 + (CD45RO + and CD45 RA + ) T-cell counts (flow cytometry), HIV RNA load (Amplicor HIV test), HCMV leukoDNAemia and HCMV DNA in urine (nested PCR), lymphoproliferative response (LPR) to HCMV, phytohemagglutinin (PHA) a…

Peripheral T-cell lymphoma associated consecutively with hemophagocytic lymphohistiocytosis and hypereosinophilic syndrome

:  Both hemophagocytic lymphohistiocytosis and hypereosinophilic syndrome have been associated with hematologic neoplasms and are respectively related to an overproduction of the cytokines Thelper 1 (Th1) and Th2 by tumor cells or reactive cells. To our knowledge, this is the first time a case of a peripheral T-cell lymphoma consecutively associated with both paraneoplastic conditions has been reported. Importantly, in this case when the lymphoma exclusively involved the bone marrow, severe paraneoplastic systemic damage, a CD8+ suppressor/cytotoxic phenotype and a hypereosinophilia not related to high levels of interleukin (IL)-5 was found. Interestingly, progression of the lymphoma coinci…

Influence of different complement sources in apoptosis of B-CLL cells with rituximab

Abstract Introduction: Rituximab (Rtx) is an anti-CD20 monoclonal antibody that is clinically active in B-cell chronic lymphocytic leukemia (B-CLL). In vitro and in vivo data indicate that Rtx mediates its biologic effect through multiple mechanisms including apoptosis, complement dependent cytotoxicity (CDC), and antibody dependent cellular cytotoxicity (ADCC). Objetive: To study in vitro sensitivity to apoptosis by Rtx of isolated cells obtained from B-CLL patients Binet stage A, in the presence of complement from different sources and correlate with CD20 and CD59 molecules expression. Methods: PBMCs were isolated from peripheral blood samples by centrifugation on a Ficoll/Hypaque gradien…

Characterization of the Proteomic and Genomic Profiles of Chronic Lymphocytic Leukemia Patients with Distinct Clinical Prognosis According to the Mutational Status of the IgVH and BCL6 and Expression Level of CD38 and ZAP70.

Abstract Introduction: In recent years several molecular prognostic factors have been identified in Chronic Lymphocytic Leukemia B (B-CLL). These include mutations in the variable region of the immunoglobulin genes (IgVH), somatic mutations in the BCL6 gene (Leukemia (2004) 18, 743–746) and the expression level of CD38 and ZAP70. However its biological significance is not clear. In order to identify novel molecular markers with prognostic and therapeutic value we have analyzed the proteomic and genomic profile of 40 B-CLL patients (Binet stage A). Material and methods: 100 μg of total PBMC proteins were used for IEF followed by 2D electrophoresis. Image analysis of scanned gels was used to …

Kinetics of cytomegalovirus (CMV) pp65 and IE-1-specific IFNgamma CD8+ and CD4+ T cells during episodes of viral DNAemia in allogeneic stem cell transplant recipients: potential implications for the management of active CMV infection.

The dynamics of CMV pp65 and IE-1-specific IFNgamma-producing CD8(+) (IFNgamma CD8(+)) and CD4(+) (IFNgamma CD4(+)) T cells and CMV DNAemia were assessed in 19 pre-emptively treated episodes of active CMV infection. Peripheral counts of IFNgamma CD8(+) and IFNgamma CD4(+) T cells inversely correlated with CMV DNAemia levels (P = <0.001 and P = 0.003, respectively). A threshold value of 1.3 cells/microl predicting CMV DNAemia clearance was established for IFNgamma CD8(+) T cells (PPV, 100%; NPV, 93%) and for IFNgamma CD4(+) T cells (PPV, 100%; NPV, 75%). Undetectable T-cell responses were usually observed at the time of initiation of pre-emptive therapy. Either a rapid (within 7 days) or a d…

An investigation on the relationship between the occurrence of CMV DNAemia and the development of invasive aspergillosis in the allogeneic stem cell transplantation setting

Cytomegalovirus (CMV) infection might increase the risk of fungal superinfection in allogeneic stem cell transplant patients. The potential association between the occurrence of CMV DNAemia and the development of invasive aspergillosis in this clinical setting was investigated. The current retrospective observational study included 167 patients undergoing T cell-replete allogeneic stem cell transplantation. Virological monitoring of active CMV infection was performed by the pp65 antigenemia assay and/or by a plasma real-time PCR assay. A total of 109 out of 167 patients developed CMV DNAemia. Twenty-three patients had proven (n = 4) or probable (n = 19) invasive aspergillosis. The occurrenc…

Genomic Abnormalities Acquired in the Blastic Transformation of Splenic Marginal Zone B-cell Lymphoma

Among 20 cases of typical splenic marginal zone lymphoma (SMZL), two cases had blastic transformation. The genetic mechanisms underlying the morphologic transformation were investigated by comparing genetic changes in initial and blastic phases. A complex karyotype including trisomy of 3q and genomic gain of 17q22-q24 was seen in both cases at diagnosis. However, the extra copy of 3q was lost during the transformation process in both tumors. Additionally, the Karpas 1718 cell line, which was derived from a patient with transformed SMZL and carried a trisomy of 3q, also evidenced the spontaneous loss of the extra 3q during the culturing process. Other acquired abnormalities observed exclusiv…

Virological and immunological features of active cytomegalovirus infection in nonimmunosuppressed patients in a surgical and trauma intensive care unit.

Cytomegalovirus (CMV) reactivation occurs frequently in critically ill patients. The natural course of CMV infection and the interaction between CMV and the adaptive immune system in this setting remain poorly defined. Fifty-three CMV-seropositive patients in a surgical and trauma intensive care unit were included in this study. The CMV DNA load in tracheal aspirates (TA) and plasma (PL) was monitored by qPCR. CMV-specific T-cell immunity was assessed by intracellular cytokine staining. Plasma TNF-alpha levels were determined by ELISA. CMV reactivation occurred in 39.7% of patients (23% had CMV DNA detected only in TA). The analysis of TA allowed an earlier diagnosis in 28% of patients. Cle…

An Assessment of the Effect of Human Herpesvirus-6 Replication on Active Cytomegalovirus Infection after Allogeneic Stem Cell Transplantation

Human herpesvirus-6 (HHV-6) may enhance cytomegalovirus (CMV) replication in allogeneic stem cell transplant (allo-SCT) recipients either through direct or indirect mechanisms. Definitive evidence supporting this hypothesis are lacking. We investigated the effect of HHV-6 replication on active CMV infection in 68 allo-SCT recipients. Analysis of plasma HHV-6 and CMV DNAemia was performed by real-time PCR. Enumeration of pp65 and IE-1 CMV-specific IFNgamma CD8(+) and CD4(+)T cells was performed by intracellular cytokine staining. HHV-6 DNAemia occurred in 39.8% of patients, and was significantly associated with subsequent CMV DNAemia in univariate (P=.01), but not in multivariate analysis (P…

Immunological insights into the pathogenesis of active CMV infection in non-immunosuppressed critically ill patients.

Dissociation of cytomegalovirus (CMV) DNA loads between the lower respiratory tract and blood, with high levels in the former compartment and low or undetectable levels in the latter, commonly occurs during active CMV infection in critically ill patients despite the presence of high frequencies of CMV-specific IFN-γ-producing CD8+ and CD4+ T cells in blood. Data presented in this case report suggest that inter-compartmental differences in interleukin-10 (IL-10) levels may, in part, explain the pathobiology of this phenomenon. In the absence of ganciclovir treatment, a significant correlation was observed between IL-10 levels and CMV DNA loads in lower respiratory tract specimens (P = 0.016)…

Reconstitution of lymphocyte populations and cytomegalovirus viremia or disease after allogeneic peripheral blood stem cell transplantation

Early reconstitution of lymphoid populations was monitored by immunophenotyping in 57 allogeneic peripheral blood stem cell (allo-PBSC) transplant patients either with or without cytomegalovirus (CMV) viremia or disease. Cell counts for total lymphocytes and CD4(+) T cells above the percentile 60th at day 14 postransplant were associated significantly with CMV viremia-free survival within 120 days after transplant. Recovery of total lymphocyte, CD3(+), and CD8(+) T-cell counts proceeded at a more rapid rate in CMV viremic patients than in nonviremic patients, irrespective of whether preemptive treatment with ganciclovir had been prescribed. Significant expansion of CD8(+) and CD8(+) CD57(+)…

Dynamics of cytomegalovirus (CMV) plasma DNAemia in initial and recurrent episodes of active CMV infection in the allogeneic stem cell transplantation setting: implications for designing preemptive antiviral therapy strategies.

Preemptive antiviral therapy strategies for active cytomegalovirus (CMV) infection occurring in allogeneic stem cell transplant recipients should be optimized to avoid overtreatment. The current study was aimed at determining whether the analysis of the kinetics of CMV DNA load in plasma may provide useful information for the therapeutic management of active CMV infection in this setting. A total of 59 consecutive patients were included in the study, of which 40 (67.8%) developed 1 (n = 21) or more (n = 19) episodes of CMV DNAemia. The need for antiviral therapy for initial or secondary episodes of CMV DNAemia could not be predicted on the basis of the CMV DNA load value in the first plasma…

Functional profile of cytomegalovirus (CMV)-specific CD8+ T cells and kinetics of NKG2C+ NK Cells associated with the resolution of CMV DNAemia in allogeneic stem cell transplant recipients

Immune mechanisms involved in control of cytomegalovirus (CMV) infection in the allogeneic stem cell transplantation setting have not been fully disclosed. CMV pp65 and IE-1-specific CD8(+) T cells expressing IFN-γ, TNF-α, and CD107a, alone or in combination, and NKG2C(+) NK cells were prospectively enumerated during 13 episodes of CMV DNAemia. The expansion of monofunctional and polyfunctional CD8(+) T cells was associated with CMV DNAemia clearance. The size and functional diversity of the expanding CD8(+) T-cell population was greater in self-resolved episodes than in episodes treated with antivirals. These differences were related to the magnitude of expansion of cognate antigen IFN-γ C…

Variant Three-Way Translocation of Inversion 16 in AML-M4Eo Confirmed by Fluorescence In Situ Hybridization Analysis

The inv(16) and t(16;16) characterize a subgroup of acute myelomonocytic leukemia (AML) with distinct morphological features and a favorable prognosis. Both cytogenetic abnormalities result in a fusion of CBF beta at 16q22 and MYH11 gene at 16p13, whose detection by PCR and fluorescence in situ hybridization (FISH) is useful for diagnosis and monitoring of the disease. Variant translocations of inv(16)/t(16;16) are very rare and whether they are also associated with a favorable prognosis is unknown. We report a patient presenting with typical AML-M4Eo and a three-way translocation of inv(16) involving 16p13, 16q22, and 3q22. FISH studies on bone marrow (BM) chromosomes using CBFB and MYH11 …

Uncontrolled immune response in acute myocardial infarction

Recently, the theory that hyperinflammation is the body's primary response to potent stimulus has been challenged. Indeed, a deregulation of the immune system could be the cause of multiple organ failure. So far, clinicians have focused on the last steps of the inflammatory cascade. However, little attention has been paid to lymphocytes, which play an important role as strategists of the inflammatory response. Experimental evidence suggests a crucial role of T lymphocytes in the pathophysiology of atherosclerosis and acute myocardial infarction (AMI). In summary, from the bottom of an imaginary inverted pyramid, a few regulatory T-cells control the upper parts represented by the wide spectr…

Identification and Characterization of Single Nucleotide Polymorphisms (SNPs) Associated with Genetic Predisposition to Develop Therapy-Related Acute Myeloid Leukemia (t-AML).

Introduction: Polymorphisms in certain genes involved on chemo/radio therapy response, as genes involved on DNA synthesis and repair, oxidative stress and drug detoxification, could be related to increased risk of develop t-AML. Objectives: To identified SNPs on genes that could be involved on putative risk of developing t-AML. To analyse the influence of relevant polymorphisms between groups of patients depending of the intensity and the type of treatment received. Material and Methods: Twelve polymorphisms located on genes from drug detoxification pathways (NOQ1, GSTP1), DNA repair (XPC[3], XRCC1[2], NBS1, ERCC5 and XRCC3) and DNA synthesis (MTHFR[2]) were studied. The analysis was carrie…

Analysis of chronic lymphotic leukemia transcriptomic profile: differences between molecular subgroups

B cell chronic lymphocytic leukemia (CLL) is a lymphoproliferative disorder with a variable clinical course. Patients with unmutated IgV(H) gene show a shorter progression-free and overall survival than patients with immunoglobulin heavy chain variable regions (IgV(H)) gene mutated. In addition, BCL6 mutations identify a subgroup of patients with high risk of progression. Gene expression was analysed in 36 early-stage patients using high-density microarrays. Around 150 genes differentially expressed were found according to IgV(H) mutations, whereas no difference was found according to BCL6 mutations. Functional profiling methods allowed us to distinguish KEGG and gene ontology terms showing…