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Ag receptor diversity involves the introduction of DNA double-stranded breaks during lymphocyte development. To ensure fidelity, cleavage is confined to the G0-G1 phase of the cell cycle. One established mechanism of regulation is through periodic degradation of the RAG2 recombinase protein. However, there are additional levels of protection. In this paper, we show that cyclical changes in the IL-7R signaling pathway functionally segregate pro-B cells according to cell cycle status. In consequence, the level of a downstream effector of IL-7 signaling, phospho-STAT5, is inversely correlated with cell cycle expression of Rag, a key gene involved in recombination. Higher levels of phopho-STAT5 in S-G2 correlate with decreased Rag expression and Rag relocalization to pericentromeric heterochromatin. These cyclical changes in transcription and locus repositioning are ablated upon transformation with v-Abl, which renders STAT5 constitutively active across the cell cycle. We propose that this activity of the IL-7R/STAT5 pathway plays a critical protective role in development, complementing regulation of RAG2 at the protein level, to ensure that recombination does not occur during replication. Our data, suggesting that pro-B cells are not a single homogeneous population, explain inconsistencies in the role of IL-7 signaling in regulating Igh recombination. [ABSTRACT FROM AUTHOR]

Obesity is a chronic inflammatory condition characterized by activation and infiltration of proinflammatory immune cells and a dysregulated production of proinflammatory cytokines. While known as a key regulator of immune natural killer (NK) cell function and development, we have recently demonstrated that reduced expression of the cytokine Interleukin-15 (IL- 15) is closely linked with increased body weight and adiposity in mice and humans. Previously, we and others have shown that obese individuals have lower circulating levels of IL-15 and NK cells. Lean IL-15 overexpressing (IL-15 tg) mice had an accumulation in adipose NK cells compared to wildtype and NK cell deficient obese IL-15-/- mice. Since IL-15 induces weight loss in IL-15-/- and diet induced obese mice and has effects on various lymphocytes, the aim of this paper was to determine if lymphocytes, particularly NK cells, play a role in IL-15 mediated weight loss. Acute IL-15 treatment resulted in an increased accumulation of NK, NKT, and CD3+ T cells in adipose tissue of B6 mice. Mice depleted of NK and NKT cells had similar weight loss comparable to controls treated with IL-15. Finally, IL-15 treatment induces significant weight loss in lymphocyte deficient RAG2-/-γc-/- mice independent of food intake. Fat pad cross-sections show decreased pad size with cytokine treatment is due to adipocyte shrinkage. These results clearly suggest that IL-15 mediates weight loss independent of lymphocytes. [ABSTRACT FROM AUTHOR]

Background: Non-obese diabetic (NOD) mice develop Sjögren's-like disease (SS-like) with loss of saliva flow and increased lymphocytic infiltrates in salivary glands (SGs). There are recent reports using multipotent mesenchymal stromal cells (MSCs) as a therapeutic strategy for autoimmune diseases due to their anti-inflammatory and immunomodulatory capabilities. This paper proposed a combined immuno- and cell-based therapy consisting of: A) an injection of complete Freund's adjuvant (CFA) to eradicate autoreactive T lymphocytes, and B) transplantations of MSCs to reselect lymphocytes. The objective of this was to test the effectiveness of CD45-/TER119- cells (MSCs) in re-establishing salivary function and in reducing the number of lymphocytic infiltrates (foci) in SGs. The second objective was to study if the mechanisms underlying a decrease in inflammation (focus score) was due to CFA, MSCs, or CFA+MSCs combined. Methodology/Principal Findings: Donor MSCs were isolated from bones of male transgenic eGFP mice. Eight week-old female NOD mice received one of the following treatments: insulin, CFA, MSC, or CFA+MSC (combined therapy). Mice were followed for 14 weeks post-therapy. CD45-/TER119- cells demonstrated characteristics of MSCs as they were positive for Sca-1, CD106, CD105, CD73, CD29, CD44, negative for CD45, TER119, CD11b, had high number of CFU-F, and differentiated into osteocytes, chondrocytes and adipocytes. Both MSC and MSC+CFA groups prevented loss of saliva flow and reduced lymphocytic infiltrations in SGs. Moreover, the influx of T and B cells decreased in all foci in MSC and MSC+CFA groups, while the frequency of Foxp3+ (Treg) cell was increased. MSC-therapy alone reduced inflammation (TNF-a, TGF-b), but the combination of MSC+CFA reduced inflammation and increased the regenerative potential of SGs (FGF-2, EGF). Conclusions/Significance: The combined use of MSC+CFA was effective in both preventing saliva secretion loss and reducing lymphocytic influx in salivary glands. [ABSTRACT FROM AUTHOR]

Abstract: Fetal cattle B-cell development proceeds via a pre-B cell stage that is characterized by the expression of surrogate light chain and recombination activation genes. In this paper, we identify a new member of bovine pre-B lymphocyte genes, VPREB2. Using RT-qPCR, we assess the expression of VPREB2 and three other surrogate light chain genes as well as RAG1 and RAG2 in fetal and adult cattle tissues. The absence of VPREB1, IGLL1, RAG1 and RAG2 expression in adult tissues and the lack of B-lymphoid differentiation in adult bone marrow – OP9 stromal cell co-culture, suggest a decline of B lymphopoiesis in adult cattle. The marked differences in the expression profiles of VPREB2 and VPREB3 in comparison to those of VPREB1, IGLL1 and RAGs suggest that the biological roles of VPREB2 and VPREB3 are unrelated to the pre-B cells. [Copyright &y& Elsevier]

Abstract: In the Czech Republic, the Human Biomonitoring Project (CZ-HBM) was launched in 1994 as an integral part of the nationwide Environmental Health Monitoring System (EHMS). Until now, the HBM covers two time periods: the first covered 1994–2003 and the second, 2005–2009. Altogether three population groups were included in the HBM: adults (blood donors aged 18–58 years), children aged 8–10 years, and breastfeeding primiparas. Sampling is organized on a yearly basis. Altogether three groups of biomarkers were analyzed: (a) selected heavy metals (Pb, Cd, Hg) and essential elements (Cu, Se, Zn) in blood and urine of adults and children, (b) indicator PCBs, DDT, DDE, HCB and HCHs in human milk and blood serum of adults and (c) cytogenetic changes in peripheral lymphocytes in blood of adults and children. This paper is focused on the general design of the CZ-HBM, the trends over time, and reference values. A significant downward time trend was observed for the blood lead levels in adults and children. Increased urinary cadmium levels were observed in Czech children compared to German children. The blood and urinary mercury levels were higher in women than in men. The levels of indicator PCB congeners in the Czech human milk samples were still higher than in most other European countries, because of existing hot spots. The levels of organochlorine pesticides showed a substantial continuous downward trend. [Copyright &y& Elsevier]

Stromal cell-derived factor-1 (SDF-1/CXCL12) induces intracellular signaling pathways crucial for mobilization, migration, proliferation and survival of many cell types via CXCR4, a chemokine CXC-motif receptor, member of the G protein-coupled receptor family. Despite playing a key role in such major processes as embryogenesis, cell differentiation and organ regeneration, molecular mechanisms underlying CXCR4 signaling remain elusive, even more so, as CXCR4 seems to activate both G-protein-dependent and G-protein-independent pathways. CXCR4 is expressed on multiple cell types including lymphocytes, hematopoietic stem cells, endothelial and epithelial cells, and cancer cells. In fact, overexpression of this receptor has been detected in many different types of cancer. The SDF-1/CXCR4 axis is also involved in tumor progression, angiogenesis, metastasis, and survival. This pathway is therefore a target for therapeutics that can block the SDF-1/CXCR4 interaction or inhibit downstream intracellular signaling. Clinical mobilization of hematopoietic stem cells (HSC), a nowadays popular method of collecting material for hematopoietic progenitor stem cell transplantation, is also dependent on the SDF-1/CXCR4 axis. Granulocyte colony-stimulating factor (G-CSF), administered to a transplant donor during clinical treatment, violates interactions between CXCR4 and its ligand, which results in degradation of HSC anchorage in bone marrow and the release of these cells into peripheral blood. In this paper we describe the clinical significance of CXCR4 and its ligand, as well as the role of CXCR4 and its gene polymorphisms in disease susceptibility. [ABSTRACT FROM AUTHOR]

Multiple myeloma (MM) is a life-threatening haematological malignancy for which standard therapy is inadequate. Autologous stem cell transplantation is a relatively effective treatment, but residual malignant sites may cause relapse. Allogeneic transplantation may result in durable responses due to antitumour immunity mediated by donor lymphocytes. However, morbidity and mortality related to graft-versus-host disease remain a challenge. Recent advances in understanding the interaction between the immune system of the patient and the malignant cells are influencing the design of clinically more efficient study protocols for MM. Cellular immunotherapy using specific antigen-presenting cells (APCs), to overcome aspects of immune incompetence in MMpatients, has received great attention, and numerous clinical trials have evaluated the potential for dendritic cell (DC) vaccines as a novel immunotherapeutic approach. This paper will summarize the data investigating aspects of immunity concerning MM, immunotherapy for patients with MM, and strategies, on the way, to target the plasma cell more selectively.We also include theMM antigens and their specific antibodies that are of potential use for MM humoral immunotherapy, because they have demonstrated the most promising preclinical results. [ABSTRACT FROM AUTHOR]

Lung cancer is the leading cause of cancer deaths worldwide and one of the most common types of cancers. The limited success of chemotherapy and radiotherapy regimes have highlighted the need to develop new therapies like antitumor immunotherapy. CD8+ T-cells represent a major arm of the cell-mediated anti-tumor response and a promising target for developing T-cell-based immunotherapies against lung cancer. Lung tumors, however, have been considered to possess poor immunogenicity; even so, lung tumor-specific CD8+ T-cell clones can be established that possess cytotoxicity against autologous tumor cells. This paper will focus on the alterations induced in CD8+ T-cells by lung cancer. Although memory CD8+ T-cells infiltrate lung tumors, in both tumorinfiltrating lymphocytes (TILs) and malignant pleural effusions, these cells are dysfunctional and the effector subset is reduced. We propose that chronic presence of lung tumors induces dysfunctions in CD8+ T-cells and sensitizes them to activation-induced cell death, which may be associated with the poor clinical responses observed in immunotherapeutic trials. Getting a deeper knowledge of the evasion mechanisms lung cancer induce in CD8+ T-cells should lead to further understanding of lung cancer biology, overcome tumor evasion mechanisms, and design improved immunotherapeutic treatments for lung cancer. [ABSTRACT FROM AUTHOR]

Inflammation is recognized as an important factor in the pathophysiology of hypertension, with the renin-angiotensin-aldosterone system (RAAS) playing a key role in the disease. Initially described because of its contribution to extracellular fluid and electrolyte homeostasis, the RAAS has been implicated in endothelial dysfunction, vascular remodeling, oxidative stress, proinflammatory cytokine production, and adhesion molecule synthesis by the vascular wall. Both angiotensin II and aldosterone are involved in these systemic effects, activating innate and adaptive immune responses. This paper highlights some aspects connecting RAAS to the hypertensive phenotype, based on experimental and clinical studies, with emphasis on new findings regarding the contribution of an increasingly studied population of T lymphocytes: the T-regulatory lymphocytes. These cells can suppress inflammation and may exert beneficial vascular effects in animal models of hypertension. [ABSTRACT FROM AUTHOR]

Presenting processed antigens to CD4+ lymphocytes during the immune response involves major histocompatibility complex class II molecules. MHC class II genes transcription is regulated by four transcription factors: CIITA, RFXANK, RFX5 and RFXAP. Defects in these factors result in major histocompatibility complex class II expression deficiency, a primary combined immunodeficiency frequent in North Africa. Autosomal recessive mutations in the RFXANK gene have been reported as being the principal defect found in North African patients with this disorder. In this paper, we describe clinical, immunological and genetic features of 11 unrelated Algerian patients whose monocytes display a total absence of MHC class II molecules. They shared mainly the same clinical picture which included protracted diarrhoea and respiratory tract recurrent infections. Genetic analysis revealed that 9 of the 11 patients had the same RFXANK founder mutation, a 26 bp deletion (named I5E6-25_I5E6+1, also known as 752delG26). Immunological and genetic findings in our series may facilitate genetic counselling implementation for Algerian consanguineous families. Further studies need to be conducted to determine 752delG26 heterozygous mutation frequency in Algerian population. [ABSTRACT FROM AUTHOR]

Background: The memory response to LCMV in mice persists for months to years with only a small decrease in the number of epitope specific CD8+T cells. This long persistence is associated with resistance to lethal LCMV disease. In contrast to studies focused on the number and surface phenotype of the memory cells, relatively little attention has been paid to the diversity of TCR usage in these cells. CD8+ T cell responses with only a few clones of identical specificity are believed to be relatively ineffective, presumably due to the relative ease of virus escape. Thus, a broad polyclonal response is associated with an effective anti-viral CD8+ T cell response. Results: In this paper we show that the primary CD8+ T cell response to the LCMV gp33-41 epitope is extremely diverse. Over time while the response remains robust in terms of the number of gp33-tetramer+ T cells, the diversity of the response becomes less so. Strikingly, by 26 months after infection the response is dominated by a small number TCRVβ sequences. In addition, it is of note the gp33 specific CD8+ T cells sorted by high and low tetramer binding populations 15 and 22 months after infection. High and low tetramer binding cells had equivalent diversity and were dominated by a small number of clones regardless of the time tested. A similar restricted distribution was seen in NP396 specific CD8+ T cells 26 months after infection. The identical TCRVβ sequences were found in both the tetramerhi and tetramerlo binding populations. Finally, we saw no evidence of public clones in the gp33-specific response. No CDR3 sequences were found in more than one mouse. Conclusions: These data show that following LCMV infection the CD8+ gp33-specific CD8 T cell response becomes highly restricted with enormous narrowing of the diversity. This narrowing of the repertoire could contribute to the progressively ineffective immune response seen in aging. [ABSTRACT FROM AUTHOR]

The immunomodulatory agent, lenalidomide, is a structural analogue of thalidomide approved by the US Food and Drug Administration for the treatment of myelodysplastic syndrome (MDS) and multiple myeloma (MM). This agent is also currently under active investigation for the treatment of chronic lymphocytic leukemia (CLL) and non-Hodgkin's lymphoma (NHL), as well as in drug combinations for some solid tumors and mantle cell lymphoma (MCL). Although treatment with lenalidomide has translated into a significant extension in overall survival in MM and MDS and has superior safety and efficacy relative to thalidomide, the mechanism of action as it relates to immune modulation remains elusive. Based on preclinical models and clinical trials, lenalidomide, as well as other structural thalidomide derivatives, enhances the proliferative and functional capacity of T-lymphocytes and amplifies costimulatory signaling pathways that activate effector responses and suppress inflammation. This paper summarizes our current understanding of T- and natural killer (NK) cell pathways that are modified by lenalidomide in hematopoietic neoplasms to inform future decisions about potential combination therapies. [ABSTRACT FROM AUTHOR]

Malondialdehyde (MDA) is used for the estimation of damage by reactive oxygen species. MDA is a major reactive aldehyde resulting from the peroxidation of biological membranes. The most common method used to assess MDA production is the thiobarbituric acid (TBARS) assay. However, the value of this method is curbed by low specificity and has been criticized for its use in human studies. In the present study we have used an alternative method for the estimation of MDA production i.e. reaction of MDA with a chromogenic agent 1-methyl-2-phenylindole at 45°C. The paper describes the method of preparing standards for the estimation of MDA (lipid peroxidation) after the treatment with an oxidative stress inducing agent hydrogen peroxide (H2O2). In the present study, the treatments of 1, 5, 10, 20, 50, 100, 150 and 200 μM of H2O2 induced significant increase in lipid peroxidation as compared to the untreated ones. The results suggest that the present method can be used to measure the lipid peroxidation in cultured human peripheral blood lymphocytes and is specific for MDA estimation. [ABSTRACT FROM AUTHOR]

Abstract: Boolean networks have been successfully used in modelling gene regulatory networks. However, for large networks, analysis by simulation becomes unfeasible. In this paper we propose a reduction method for Boolean networks that decreases the size of the network, while preserving important dynamical properties and topological features. As a result, the reduced network can be used to infer properties about the original network and to gain a better understanding of the role of network topology on the dynamics. In particular, we use the reduction method to study steady states of Boolean networks and apply our results to models of Th-lymphocyte differentiation and the lac operon. [Copyright &y& Elsevier]

The blood cancer T cell large granular lymphocyte (T-LGL) leukemia is a chronic disease characterized by a clonal proliferation of cytotoxic T cells. As no curative therapy is yet known for this disease, identification of potential therapeutic targets is of immense importance. In this paper, we perform a comprehensive dynamical and structural analysis of a network model of this disease. By employing a network reduction technique, we identify the stationary states (fixed points) of the system, representing normal and diseased (T-LGL) behavior, and analyze their precursor states (basins of attraction) using an asynchronous Boolean dynamic framework. This analysis identifies the T-LGL states of 54 components of the network, out of which 36 (67%) are corroborated by previous experimental evidence and the rest are novel predictions. We further test and validate one of these newly identified states experimentally. Specifically, we verify the prediction that the node SMAD is over-active in leukemic T-LGL by demonstrating the predominant phosphorylation of the SMAD family members Smad2 and Smad3. Our systematic perturbation analysis using dynamical and structural methods leads to the identification of 19 potential therapeutic targets, 68% of which are corroborated by experimental evidence. The novel therapeutic targets provide valuable guidance for wet-bench experiments. In addition, we successfully identify two new candidates for engineering long-lived T cells necessary for the delivery of virus and cancer vaccines. Overall, this study provides a bird's-eye-view of the avenues available for identification of therapeutic targets for similar diseases through perturbation of the underlying signal transduction network. [ABSTRACT FROM AUTHOR]

Abstract: Purpose: With the development of diagnostic techniques, renal cell carcinoma (RCC) is currently diagnosed in earlier stages, allowing the introduction of less invasive techniques in its management. One of the most promising new treatment methods is based on the utilization of high temperature created by radiofrequency current circulating around the needle probe introduced into the tumor. Besides the direct destruction of the cancer tissue, the treatment may induce immunologic reaction to tumor antigens released from destroyed tumor cell. This paper describes changes observed in the peripheral blood lymphocyte population after radiofrequency ablation (RFA) of RCC. Methods: Blood was tested before, and 2, 4, and 6 weeks after the RFA in 6 patients with RCC for the proportions and numbers of CD3+, CD3+HLA-DR+, CD3+CD4+, CD3+CD8+, and CD56+CD16+ cells. The blood was stained with fluorochrome-conjugated monoclonal antibodies and percentages of cells expressing various markers were determined by flow cytometry. Results: In all patients, the changes were most pronounced 2 weeks after the procedure. The proportion of CD4+ and CD8+ lymphocytes were changed. In 1 patient, an increase in both CD4+ and CD8+ cells was observed. In 5 out of 6 patients, the proportion of activated (DR+) cells was increased over the whole follow-up period with the highest values in the second week after RFA. The percentage of the CD56+CD16+ was decreased in most of the patients. Conclusions: Our study confirms that in the majority of patients, RFA of the renal tumors causes significant changes in the proportion of the peripheral immune cells. We suggest that the results presented in this article shows the necessity for further studies. [Copyright &y& Elsevier]

Immunopathology is a major cause of influenza-associated morbidity and mortality worldwide. However, the role and regulatory mechanisms of CD4 T cells in severe lung immunopathology following acute influenza infection are poorly understood. In this paper, we report that the emergence of immunopathogenic CD4 T cells is under the control of a transmembrane immunoadaptor DAP12 pathway during influenza infection. We find that the mice lacking DAP12 have unaltered viral clearance but easily succumb to influenza infection as a result of uncontrolled immunopathology. Such immunopathology is associated with markedly increased CD4 T cells displaying markedly increased cytotoxicity and Fas ligand expression. Furthermore, the immunopathogenic property of these CD4 T cells is transferrable. Thus, depletion of CD4 T cells or abrogation of Fas/Fas ligand signaling pathway improves survival and immunopathology. We further find that DAP12 expressed by dendritic cells plays an important role in controlling the immunopathogenic CD4 T cells during influenza infection. Our findings identify a novel pathway that controls the level of immune-pathogenic CD4 T cells during acute influenza infection. [ABSTRACT FROM AUTHOR]

Abstract: The comet assay is now the method of choice for measuring most kinds of DNA damage in cells. However, due to the lack of a standardised protocol inter-laboratory comparisons are of limited value. The aim of this paper is to demonstrate how small changes in comet-assay variables may significantly affect the results. We examined the effect of varying agarose concentrations, alkaline unwinding time, electrophoresis time, voltage and current, by use of two cell types, viz. human peripheral blood lymphocytes and the lymphoblastoid cell line TK-6. All these variables have marked effects on assay performance and, therefore, on the determination of DNA damage. Here we identify factors of particular importance. [Copyright &y& Elsevier]

Abstract: This paper considers how well standard calibration curve for translocations constructed for lymphocyte cultures irradiated in vitro with gamma-rays from 60Co compares with the translocations yield in lymphocytes taken from people at a long post-exposure time. Data were used from radiation accident victims overexposed to doses ranging from 0.2 to 8.5 Gy and who were cytogenetically followed-up for various times upto 50 y. Their cultured lymphocytes had been scored both by the conventional dicentric method and by FISH for all translocations involving painted chromosomes (2, 3, 8); (2, 3, 5) or (2, 4, 12). The in vivo dose response relationship was derived by fitting translocation frequencies to the contemporary individual doses obtained independently and confirmed by different biological assays and physical dosimetry. A comparison with the conventional in vitro curve indicates reductions of translocation frequencies with increasing time which would prejudice retrospective dose assessment by FISH. This has led to the possibility to amend the in vitro dose response curve for translocations to make it more suitable for use in retrospective biodosimetry. This approach for retrospective biodosimetry therefore uses a dose response relationship based on truly persisting translocations. [Copyright &y& Elsevier]

Although essential for T cell function, the identity of the T cell receptor (TCR) "inside-out" pathway for the activation of lymphocyte function-associated antigen 1 (LFA-1) is unclear. SKAP1 (SKAP-55) is the upstream regulator needed for TCR-induced RapL-Rap1 complex formation and LFA-1 activation. In this paper, we show that SKAP1 is needed for RapL binding to membranes in a manner dependent on the PH domain of SKAP1 and the PI3K pathway. A SKAP1 PH domain-inactivating mutation (i.e. R131M) markedly impaired RapL translocation to membranes for Rap1 and LFA-1 binding and the up-regulation of LFA-1-intercellular adhesion molecule 1 (ICAM-1) binding. Further, N-terminal myr-tagged SKAP1 for membrane binding facilitated constitutive RapL membrane and Rap1 binding and effectively substituted for PI3K and TCR ligation in the activation of LFA-1 in T cells. [ABSTRACT FROM AUTHOR]

Purpose: Over the past five years the Center for Minimally Invasive Radiation Biodosimetry at Columbia University has developed the Rapid Automated Biodosimetry Tool (RABiT), a completely automated, ultra-high throughput biodosimetry workstation. This paper describes recent upgrades and reliability testing of the RABiT. Materials and methods: The RABiT analyses fingerstick-derived blood samples to estimate past radiation exposure or to identify individuals exposed above or below a cut-off dose. Through automated robotics, lymphocytes are extracted from fingerstick blood samples into filter-bottomed multi-well plates. Depending on the time since exposure, the RABiT scores either micronuclei or phosphorylation of the histone H2AX, in an automated robotic system, using filter-bottomed multi-well plates. Following lymphocyte culturing, fixation and staining, the filter bottoms are removed from the multi-well plates and sealed prior to automated high-speed imaging. Image analysis is performed online using dedicated image processing hardware. Both the sealed filters and the images are archived. Results: We have developed a new robotic system for lymphocyte processing, making use of an upgraded laser power and parallel processing of four capillaries at once. This system has allowed acceleration of lymphocyte isolation, the main bottleneck of the RABiT operation, from 12 to 2 sec/sample. Reliability tests have been performed on all robotic subsystems. Conclusions: Parallel handling of multiple samples through the use of dedicated, purpose-built, robotics and high speed imaging allows analysis of up to 30,000 samples per day. [ABSTRACT FROM AUTHOR]

Abstract: DOCK10 is a member of the dedicator of cytokinesis (DOCK) family of Rho GTPase activators preferentially expressed in lymphocytes. In this paper, we analyzed DOCK10 mRNA diversity produced because of alternative splicing. Alternative first coding exon usage led to 2 main protein-coding transcripts, DOCK10.1 and DOCK10.2. Full-length cDNA clones of both isoforms were obtained from both normal human peripheral blood mononuclear cells and mouse spleen for the first time for human DOCK10.1, mouse DOCK10.1, and mouse DOCK10.2. Human and mouse DOCK10.1 clones corresponded to the protein coding assemblies provided by the National Center for Biotechnology Information as Reference Sequences for DOCK10. Our analysis especially focused on human cDNA clones, of which 63% were alternatively spliced forms involving diverse exons and introns. DOCK10.1 expression was enriched in normal T cells, and DOCK10.2 expression was enriched in normal B cells and chronic lymphocytic leukemia (CLL) B cells. Both isoforms were upregulated in response to interleukin-4 in B cells, both normal and CLL, but not in T cells. Our data suggest that cell-specific mechanisms regulate expression of the alternative first exon variants of DOCK10 in vertebrates. [Copyright &y& Elsevier]

In this paper we show the dose response curves in vitro obtained at first time in Cuba, with several biological indicators for radiation damage: rings in premature chromosome condensation (PCC-R); PCC index and γ-H2AX foci, all of these in peripheral human lymphocytes. These curves cover an interval of doses between 0.1 Gy to 25 Gy, several radiation qualities and times post-exposition, strengthening the capabilities of the Cuban laboratory to give answer to the radiological emergencies and in the same way increase its contribution to elucidate any radiological abnormal event take place at the region where the main indicator at the present time is the dicentric assay in lymphocytes of peripheral blood. [ABSTRACT FROM AUTHOR]

Diabetes can be associated with a reduction in functional β cell mass, which must be restored if the disease is to be cured or progress is to be arrested. To study the cell count, it is also necessary to determine the number of nuclei within the insulin stained area. It can take a single experimentalist several months to complete a single study of this kind, results of which may still be quite subjective. In this paper, we propose a framework based on a novel measure of local symmetry for detection of cells. The local isotropic phase symmetry measure (LIPSyM) is designed to give high values at or near the cell centers. We demonstrate the effectiveness of our algorithm for detection of two types of specific cells in histology images, cells in mouse pancreatic sections and lymphocytes in human breast tissue. Experimental results for these two problems show that our algorithm performs better than human experts for the former problem, and outperforms the best reported results for the latter. [ABSTRACT FROM AUTHOR]

Abstract: ASP3258 is a novel, orally active, selective phosphodiesterase (PDE) 4 inhibitor which has an improved therapeutic window over second generation compounds such as roflumilast and cilomilast. Here, we investigated the effect of ASP3258 on cigarette smoke exposure-induced lung injury in guinea pigs, a well-defined model for chronic obstructive pulmonary disease (COPD). COPD-like lung injury was induced by repeated cigarette smoke exposure (10 cigarettes/day, 5days/week, for 4weeks). Orally administered ASP3258 (0.3, 1, and 3mg/kg) dose-dependently suppressed pulmonary accumulation of mononuclear cells and neutrophils, and the inhibitory effect of ASP3258 (1mg/kg) was almost the same as that of roflumilast (1mg/kg). In contrast, a glucocorticoid prednisolone (10mg/kg, p.o.) did not show any effect. Histological examination revealed that ASP3258 treatment significantly inhibited infiltration of neutrophils and macrophages into either or both alveolar or peribronchiolar areas, as well as hyperplastic and squamous metaplastic changes of epithelium in the bronchi. Decreasing trends in histological scores for accumulation of lymphocytes in the alveoli and alveolar wall thickening were also observed in ASP3258-treated animals. Further, ASP3258 attenuated augmentation of matrix metalloproteinase-9 activity in the bronchoalveolar lavage fluid. These findings suggest that ASP3258 has therapeutic potential for treating COPD not only through inhibition of pulmonary cellular accumulation but also by preventing lung structural alterations initiated by repeated cigarette smoke exposure. To our knowledge, this is the first paper demonstrating that PDE4 inhibitors exert significant inhibitory effects on subchronic cigarette smoke exposure-induced lung injury in guinea pigs. [Copyright &y& Elsevier]

Abstract: Somatostatin (SS) and growth hormone-releasing hormone (GHRH) are synthesized and secreted by the hypothalamus, which mediates the synthesis and secretion of the growth hormone (GH) from the hypophysis, as well as regulates the GH concentrations in animals and humans. In this paper, we describe the regulation of animal growth using plasmid DNA-encoding GHRH gene. In this study, we introduce a gene transfer technique mediated by PLGA microspheres. The eukaryon expression vectors based on the RNA replicon, namely, pCMV-Rep-GHRH, were constructed; further, poly(d, l-lactide-co-glycolide) (PLGA) microsphere-encapsulating plasmids were prepared. Gilts were intramuscularly injected at day 85 of gestation with 1 or 5mg of pCMV-Rep-GHRH microspheres. Piglets were weighed following birth and at days 20 and 50 postbirth. Blood samples were collected at day 50 to evaluate serum IGF-1 and GHRH concentrations. The result showed that the birth weight of the piglets at 20d and 50d were significantly higher (by 20.32–110.76%; P< 0.01) than that in the group injected with saline-MP. After 50days, the concentrations of serum IGF-I in the treatment groups injected with plasmid were observed to be significantly higher (2.5 times; P< 0.01) than that in the group injected with saline-MP; the concentration of serum GHRH was higher than 21% in 1 case (P< 0.01). The lymphocyte stimulation indices detected confirmed that GHRH gene transfer could improve immunity in first-generation progeny pigs. The results of this study confirmed that PLGA microsphere-mediated GHRH expression in pregnant pig skeletal muscle enhanced the body weight of piglets prior to and following birth. [Copyright &y& Elsevier]

The nonhomologous end-joining (NHEJ) pathway is essential for radioresistance and lymphocyte-specific V(D)J (variable [diversity] joining) recombination. Defects in NHEJ also impair hematopoietic stem cell (HSC) activity with age but do not affect the initial establishment of HSC reserves. In this paper, we report that, in contrast to deoxyribonucleic acid (DNA)-dependent protein kinase catalytic subunit (DNA-PKcs)-null mice, knockin mice with the DNA-PKcs3A/3A allele, which codes for three alanine substitutions at the mouse Thr2605 phosphorylation cluster, die prematurely because of congenital bone marrow failure. Impaired proliferation of DNA-PKcs3A/3A HSCs is caused by excessive DNA damage and p53-dependent apoptosis. In addition, increased apoptosis in the intestinal crypt and epidermal hyperpigmentation indicate the presence of elevated genotoxic stress and p53 activation. Analysis of embryonic fibroblasts further reveals that DNA-PKcs3A/3A cells are hypersensitive to DNA cross-linking agents and are defective in both homologous recombination and the Fanconi anemia DNA damage response pathways. We conclude that phosphorylation of DNA-PKcs is essential for the normal activation of multiple DNA repair pathways, which in turn is critical for the maintenance of diverse populations of tissue stem cells in mice. [ABSTRACT FROM AUTHOR]

The classical neurotransmitter, serotonin (5-HT), plays an important role outside of the central nervous system in immune signaling. Here I review recent studies describing 5-HT uptake in dendritic cells and B lymphocytes, 5-HT synthesis in T lymphocytes, and the role of specific 5-HT receptor subtypes in innate and adaptive immune cells. Furthermore, a recent paper describing the immune phenotype of 5-HT deficient mice is discussed. [ABSTRACT FROM AUTHOR]

These are personal reflections on the development of methods to use micronuclei as a measure of genetic damage and their use in research and in toxicology by four people who have been intimately involved with this work, a personal rather than a comprehensive history. About 6000 papers have been published using such methods in many tissues in vivo or in cultured cells of many organisms from plants to humans, but the majority of the work has been on mammalian erythrocytes and human lymphocytes, the areas in which we have worked primarily. Although this is by no means a complete history, those working in the field may be interested in some of the personal events that lie behind the development and acceptance of methods that are now standard. [ABSTRACT FROM AUTHOR]

Automated image analysis scoring of micronuclei (MN) in cells can facilitate the objective and rapid measurement of genetic damage in mammalian and human cells. This approach was repeatedly developed and tested over the past two decades but none of the systems were sufficiently robust for routine analysis of MN until recently. New methodological, hardware and software developments have now allowed more advanced systems to become available. This mini-review presents the current stage of development and validation of the Metasystems Metafer MNScore system for automated image analysis scoring of MN in cytokinesis-blocked binucleated lymphocytes, which is the best-established method for studying MN formation in humans. The results and experience of users of this system from 2004 until today are reviewed in this paper. Significant achievements in the application of this method in research related to mutagen sensitivity phenotype in cancer risk, radiation biodosimetry and biomonitoring studies of air pollution (enriched by new data) are described. Advantages as well as limitations of automated image analysis in comparison with traditional visual analysis are discussed. The current increased use of the Metasystems Metafer MNScore system in various studies and the growing number of publications based on automated image analysis scoring of MN is promising for the ongoing and future application of this approach. [ABSTRACT FROM AUTHOR]

The International Human Micronucleus (HUMN) Project (www.humn.org) was founded in 1997 to coordinate worldwide research efforts aimed at using micronucleus (MN) assays to study DNA damage in human populations. The central aims were to (i) collect databases on baseline MN frequencies and associated methodological, demographic, genetic and exposure variables, (ii) determine those variables that affect MN frequency, (iii) establish standardised protocols for performing assays so that data comparisons can be made more reliably across laboratories and countries and (iv) evaluate the association of MN frequency with disease outcomes both cross-sectionally and prospectively. In the first 10 years of the HUMN project, all of these objectives were achieved successfully for the MN assay using the cytokinesis-block micronucleus (CBMN) assay in human peripheral blood lymphocytes and the findings were published in a series of papers that are among the most highly cited in the field. The CBMN protocol and scoring criteria are now standardised; the effect of age, gender and smoking status have been defined, and it was shown prospectively using a database of almost 7000 subjects that an increased MN frequency in lymphocytes predicts cancer risk. More recently in 2007, the HUMN coordinating group decided to launch an equivalent project focussed on the human MN assay in buccal epithelial cells because it provides a complementary method for measuring MN in a tissue that is easily accessible and does not require tissue culture. This new international project is now known as the human MN assay in exfoliated cells (HUMNxL). At present, a database for >5000 subjects worldwide has been established for the HUMNxL project. The inter-laboratory slide-scoring exercise for the HUMNxL project is at an advanced stage of planning and the analyses of data for methodological, demographic, genetic, lifestyle and exposure variables are at a final stage of completion. Future activities will be aimed at (i) defining the genetic variables that affect MN frequencies, (ii) validation of the various automated scoring systems based on image analysis, flow cytometry and laser scanning cytometry, (iii) standardisation of protocols for scoring micronuclei (MNi) in cells from other tissues, e.g. erythrocyte and nasal cells and (iv) prospective association studies with pregnancy complications, developmental defects, childhood cancers, cardiovascular disease and neurodegenerative diseases. [ABSTRACT FROM AUTHOR]

Interleukin-6 is currently attracting significant interest as a potential therapeutic target in systemic sclerosis (SSc). In this paper, the biology of interleukin-6 is reviewed, and the evidence for interleukin-6 dysregulation in SSc is explored. The role of inteleukin-6 classical and trans signalling pathways in SSc relevant phenomena such as chronic inflammation, autoimmunity, endothelial cell dysfunction, and fibrogenesis is discussed. The existing evidence that interventions designed to block interleukin-6 signalling are of therapeutic relevance in SSc is evaluated. [ABSTRACT FROM AUTHOR]

Hepatitis C virus (HCV) has been recognized as a major cause of chronic liver diseases worldwide. It has been suggested that HCV infects not only hepatocytes but also mononuclear lymphocytes including B cells that express the CD81 molecule, a putative HCV receptor. HCV infection of B cells is the likely cause of B-cell dysregulation disorders such as mixed cryoglobulinemia, rheumatoid factor production, and B-cell lymphoproliferative disorders that may evolve into non-Hodgkin's lymphoma (NHL). Epidemiological data indicate an association between HCV chronic infection and the occurrence of B-cell NHL, suggesting that chronic HCV infection is associated at least in part with B-cell lymphomagenesis. In this paper, we aim to provide an overview of recent literature, including our own, to elucidate a possible role of HCV chronic infection in B-cell lymphomagenesis. [ABSTRACT FROM AUTHOR]

Purpose: Ascorbic acid is an essential micronutrient and is considered to have an antioxidant function in living systems. For the past several decades, ascorbic acid has been the subject of considerable interest as an anticancer agent. Several studies have shown that ascorbic acid is cytotoxic to a variety of cancer cells, whereas normal cells are relatively resistant to such cytotoxic action. In this study, we propose a putative molecular mechanism that accounts for the preferential cytotoxicity of ascorbic acid against cancer cells. Methods: Standard and lysed version of alkaline single-cell gel electrophoresis (Comet assay); ferrous oxidation-xylenol orange (FOX) assay. Results: We show that ascorbic acid acts as a prooxidant and leads to oxidative DNA breakage in lymphocytes and lymphocyte nuclei. Scavengers of reactive oxygen species were able to inhibit ascorbic acid-induced DNA breakage, suggesting the involvement of reactive oxygen species in this reaction. We further show that such DNA breakage is inhibited by both iron and copper chelators in cells, whereas in nuclei, similar inhibition was achieved only by copper chelators, indicating an important role of chromatin-bound copper in the prooxidant cellular DNA breakage by ascorbic acid. Conclusion: We propose that the copper-dependent cellular redox status is an important element in the cytotoxic action of ascorbic acid against cancer cells. It is well established that cellular copper levels are considerably elevated in various malignancies. Therefore, cancer cells may be more subject to electron transfer between copper and ascorbate to generate reactive oxygen species. In light of these observations and those in literature, in this paper we explain that the preferential cytotoxicity of ascorbic acid against cancer cells is the result of elevated copper levels in such cells. Further, this study identifies nuclear copper as a novel molecular target for cytotoxic action of ascorbic acid, which has implications for its chemotherapeutic properties against cancer. [ABSTRACT FROM AUTHOR]

AbstractObjective:The aim of this study was to evaluate the effectiveness of Er:YAG laser and conventional periodontal therapy in the treatment of acute streptococcal gingivitis both clinically and microbiologically. Background Data:This case report describes a 2-year follow-up of a 30-year-old, female, chronic periodontitis patient, presenting severe gingival inflammation, sensitivity, pain, and acute gingival lesions that were treated with Er:YAG laser and conventional hand and ultrasonic instruments. Materials and Methods:Before the initial periodontal treatment, microbiological samples were taken from the lesion sites with sterile paper points from the sulcuses bilaterally, and excisional biopsies were obtained from the lesions. Following diagnostic tests, the lesions were identified as acute streptococcal gingivitis. Following the measurement of clinical indices, initial periodontal therapy was performed with Er:YAG laser on the right side and conventional hand and ultrasonic instruments on the left side, which were performed as two sessions at weekly intervals. As an adjunct to mechanical periodontal therapy, 500 mg amoxicillin was prescribed t.i.d. for a week. Results:Microbiological samples grew mostly Streptococcus sp.and black pigmented obligate anaerobic bacteria. The histopathological examination revealed acanthosis, papillomatosis, and spongiotic lesions in the keratinized stratified squamous epithelium; infiltration of polymorphonuclear neutrophils, lymphocytes, and macrophages and plasma cells in the connective tissue; infiltration and accumulation of polymorphonuclear neutrophils in the epithelium, especially in the spongiotic lesions; and formation of microabscess-like clusters. After the initial periodontal treatment, clinical and microbiological measurements were repeated and reductions in clinical indices and the number of microorganisms were observed. Both treatment modalities gave similar results, and no recurrences were observed during the 2-year follow-up. Conclusion:Er:YAG laser seems to be promising and as effective as conventional periodontal therapy in the treatment of acute streptococcal gingivitis. [ABSTRACT FROM AUTHOR]

There is a wealth of information regarding the differentiation of T-helper cells. Nevertheless, there is no general agreement on the topology and dynamical properties of the molecular network controlling the differentiation of these cells. This paper presents a continuous dynamical system to model the signaling network that controls the differentiation process of T-helper cells. The model is able to represent the differentiation from the precursor Th0 cell to any of the four effectors types (Th1, Th2, Th17, and Treg), as well as the phenotype of single null mutants. We present the first sensitivity analysis of the equations defining the Th model, showing that the qualitative dynamical behavior of the model is very robust against changes in three out of four tested parameters. The robustness of the model is in agreement with our claim that the qualitative behavior of the system is to a large extent independent of the methodological framework used for modeling. [ABSTRACT FROM AUTHOR]

Abstract: Uracil may arise in DNA as a result of spontaneous cytosine deamination and/or misincorporation of dUMP during DNA replication. In this paper we will review: (i) sources of the origin of uracil in DNA; (ii) some properties of the enzymes responsible for the excision of uracil and their role in the Ig diversification process, which comprises somatic hypermutation and class switch recombination; and (iii) consequences of cytosine deamination in other than the Ig loci, in cell types different than B lymphocytes. Furthermore, the issue concerning the basal level of uracil in DNA and consequences of the presence of U:A pairs for DNA stability and cell functions will be discussed. Finally, we will discuss the clinical significance of aberrant uracil incorporation into DNA and possible involvement of aberrantly expressed AID and the enzyme-induced presence of uracil, in carcinogenesis. Based on the literature data we conclude/hypothesize that the non-canonical base uracil may be present and well tolerated in DNA mostly as U:A pairs, likely in quantities of 104 per genome. Although a role of uracil in DNA is not fully defined, it is possible that an ancestral system which once used uracil in primordial genetic material (uracil-DNA), may have evolved to use this molecule in regulatory processes such as: (i) meiotic cell division to facilitate chromatid exchange during crossing-over (in spermatocytes); (ii) it is possible that uracil present in DNA may be a signaling molecule during metamorphosis of Drosophila melanogaster; and (iii) during transcription since some regulatory proteins (Escherichia coli lac repressor) and GCN4 can recognize uracil versus thymine in specific DNA regulatory sequences. Moreover, recent data suggest that in transcriptionally active chromatin the dUTP/dTTP pool may be significantly increased, which in turn may lead to massive uracil incorporation into DNA. [ABSTRACT FROM AUTHOR]

Please cite this paper as: Absence of modulation of CD4+CD25high regulatory T cells in CTCL patients treated with bexarotene. Experimental Dermatology 2010; 19: e95-e102. Abstract: Cutaneous T-cell lymphoma (CTCL) are a heterogeneous group of lymphoproliferative disorders, characterized by the infiltration of the epidermis by mature and activated malignant CD4+ T-lymphocytes. Retinoids such as retinoic acid and synthetic analogues have long been used alone or in combination with other therapies for CTCL. Bexarotene, the first synthetic highly selective RXR retinoid, was approved for the treatment of all stages of CTCL in patients refractory to at least one systemic therapy. Recently, six cases in which the initiation of bexarotene therapy for CTCL was associated with the progression of internal disease despite improvement of cutaneous signs and symptoms were reported. Moreover, it has been established that retinoids promote the generation of CD4+ Foxp3+ regulatory T cells, raising the question of an induction of regulatory T-cells by bexarotene. The aim of this work was to determine if bexarotene induces an increase of functional regulatory T cells which could play a role in the development of secondary extra-cutaneous lymphomas. Regulatory T cells were studied both in cutaneous biopsy specimens using an immunohistochemical analysis of CD4, CD25 and Foxp3 and in blood where proportion and functionality of circulating CD4+CD25high T-cells were determined. The study was performed in 10 patients [five patients with Sézary syndrome (SS) and five mycosis fungoïdes (MF)], treated for 6 months with bexarotene. Four healthy donors were used as controls for phenotypic and functional analysis on PBL. We found that the frequency of CD4+CD25high Treg cells was not significantly different before starting bexarotene and after 6 months of treatment in CTCL patients. However, we observed that the frequency of CD4+CD25high Treg cells before the beginning of the treatment was significantly increased compared to healthy donors. In addition, functional assays demonstrated that Foxp3 expressing CD4+CD25high T-cells were capable of suppressing autologous CD4 + CD25− T-cell proliferation. In the present work, we detected the presence of functional circulating CD4+CD25high Foxp3+ regulatory T-cells in CTCL patients, with an increased frequency compared to healthy donors. The treatment with bexarotene does not seem to affect the regulatory T-cell compartment. [ABSTRACT FROM AUTHOR]

The presence of lymphocytic infiltration (LI) has been correlated with nodal metastasis and tumor recurrence in HER2+ breast cancer (BC). The ability to automatically detect and quantify extent of LI on histopathology imagery could potentially result in the development of an image based prognostic tool for human epidermal growth factor receptor-2 (HER2+) BC patients. Lymphocyte segmentation in hematoxylin and eosin (H&E) stained BC histopathology images is complicated by the similarity in appearance between lymphocyte nuclei and other structures (e.g., cancer nuclei) in the image. Additional challenges include biological variability, histological artifacts, and high prevalence of overlapping objects. Although active contours are widely employed in image segmentation, they are limited in their ability to segment overlapping objects and are sensitive to initialization. In this paper, we present a new segmentation scheme, expectation-maximization (EM) driven geodesic active contour with overlap resolution (EMaGACOR), which we apply to automatically detecting and segmenting lymphocytes on HER2+ BC histopathology images. EMaGACOR utilizes the expectation-maximization algorithm for automatically initializing a geodesic active contour (GAC) and includes a novel scheme based on heuristic splitting of contours via identification of high concavity points for resolving overlapping structures. EMaGACOR was evaluated on a total of 100 HER2+ breast biopsy histology images and was found to have a detection sensitivity of over 86% and a positive predictive value of over 64%. By comparison, the EMaGAC model (without overlap resolution) and GAC model yielded corresponding detection sensitivities of 42% and 19%, respectively. Furthermore, EMaGACOR was able to correctly resolve over 90% of overlaps between intersecting lymphocytes. Hausdorff distance (HD) and mean absolute distance (MAD) for EMaGACOR were found to be 2.1 and 0.9 pixels, respectively, and significantly better compared to the corresponding performance of the EMaGAC and GAC models. EMaGACOR is an efficient, robust, reproducible, and accurate segmentation technique that could potentially be applied to other biomedical image analysis problems. [ABSTRACT FROM AUTHOR]

Progress in microelectrode-based technologies has facilitated the development of sophisticated methods for manipulating and separating cells, bacteria, and other bioparticles. For many of these various applications, the theoretical modeling of the electrical response of compartmentalized particles to an external field is important. In this paper we address the analysis of the interaction between cells immersed in rf fields. We use an integral formulation of the problem derived from a consideration of the charge densities induced at the interfaces of the particle compartments. The numerical solution by a boundary element technique allows characterization of their dielectric properties. Experimental validation of this theoretical model is obtained by investigating two effects: (1) The influence that dipolar “pearl chaining” has on the dielectrophoretic behavior of human T lymphocytes and (2) the frequency variation of the spin and orbital torques of approaching insulinoma β-cells in a rotating field. [ABSTRACT FROM AUTHOR]

Abstract: Dendrobium nobile is commonly used as a Yin tonic for reinforcing body fluid and nourishing blood in Southeast Asia. In this paper, a pectic polysaccharide DNP-W5 was obtained from the stems of D. nobile. It contained mannose, glucose, galactose, xylose, rhamnose and galacturonic acid in molar ratios of 3.1:8.1:8.2:0.6:4.2:3.9. About 21% of the galacturonic acid existed as methyl ester and O-acetyl groups were approximately 6.9%. The structural analysis indicated that DNP-W5 possessed a backbone of a disaccharide of [→4)-α-GalAp-(1→2)-α-Rhap-(1→] with branches at O-4 of the Rhap and O-3 of the GalpA. The side chains consisted of galactosyl, mannosyl, glucosyl, and xylosyl residues. In vitro bioactive tests, DNP-W5 displayed remarkable immunoenhancing activities on T- and B-lymphocytes and its branches and acetyl groups are very important for the expression of the immunological activity. [Copyright &y& Elsevier]

Please cite this paper as: Modulating T cell functions does not alleviate chronic inflammatory skin lesions in K5.TGFβ1 transgenic mice. Experimental Dermatology 2009; 19: 406–415. To use mice with chronic hyperproliferative skin inflammation as psoriasis models, their thorough phenotypic and functional characterization is indispensable. Mice with keratin 5 promoter-controlled overexpression of latent human Transforming Growth Factor (TGF)β1 within the basal epidermis (K5.TGFβ1 mice) show a psoriasiform phenotype, but the underlying pathogenic mechanisms are not entirely clear. To elucidate the contribution of T lymphocytes to the pathogenesis in K5.TGFβ1 mice, we used three complementary approaches: first, peripheral T cells were eradicated via systemic treatment with CD3- or CD4-depleting antibodies. However, this elimination did not alleviate the chronic inflammatory disorder. Second, bone marrow transplantation from transgenic mice into wildtype recipients and vice versa resulted in the expected reconstitution of both adaptive and innate immune system but had little effect on the cutaneous phenotype both in wildtype and transgenic chimeras. Third, based on the hypothesis that the disease course could be modulated by regulatory T cells (Tregs), we expanded Tregs in vivo using a superagonistic anti-CD28 antibody. While this treatment achieved a threefold increase in Foxp3-expressing Tregs, there was little, if any, effect on the chronic skin inflammation. We conclude from our findings that T cells play little, if any, role in the skin lesions of K5.TGFβ1 mice. [ABSTRACT FROM AUTHOR]

Please cite this paper as: Para-phenylenediamine-specific lymphocyte activation test: a sensitive in vitro assay to detect para-phenylenediamine sensitization in patients with severe allergic reactions. Experimental Dermatology 2010; 19: 435–441. Patients sensitized to para-phenylenediamine (PPD) by semi-permanent tattoos increasingly develop threatening allergic reactions in response to black hair dye. The gold standard to diagnose allergic contact dermatitis is to perform epicutaneous patch tests, however, iatrogenic sensitizations and severe patch test reactions to PPD have been described, the latter especially in patients with severe allergic reactions. We examined nine patients with severe allergic reactions in response to permanent hair dyes. Patch tests using the standard concentration of 1% or 0.5% PPD resulted in severe and sometimes even bullous reactions in all patients responsive to PPD. Titration revealed that at 1% of the standard concentration (0.01% PPD), patch test sensitivity decreased and only 50% of patients responded. Consequently, we established an in vitro assay to diagnose PPD allergy. Freshly isolated peripheral blood mononuclear cells (PBMC) were cultured with titrated concentrations of PPD with or without IL-2 supplementation, and cell proliferation was determined by [3H]-thymidine incorporation. Lymphocyte activation test (LAT ) detected PBMC cell proliferation specific to PPD, with at least 3.5-fold increase in [3H]-thymidine uptake in all PPD allergic patients. Most importantly, PPD –LAT without IL-2 supplementation remained negative in three out of eight PPD allergic patients. Thus, PPD-LAT with IL-2 supplementation demonstrated a sensitivity of 100%, remained unresponsive in controls not sensitized to PPD, and in one patient sensitive to other p-amino compounds. These data demonstrate that LAT with PPD can be used to detect PPD sensitization as a possible alternative to patch testing at least in patients with severe allergic reactions to PPD. [ABSTRACT FROM AUTHOR]

In this paper, we numerically show that the dynamics of the HIV system is sensitive to both the initial condition and the system parameters. These phenomena imply that the system is chaotic and exhibits a bifurcation behavior. To control the system, we propose to initiate an HIV therapy based on both the concentration of the HIV-1 viral load and the ratio of the CD4 lymphocyte population to the CD8 lymphocyte population. If the concentration of the HIV-1 viral load is higher than a threshold, then the first type of therapy will be applied. If the concentration of the HIV-1 viral load is lower than or equal to the threshold and the ratio of the CD4 lymphocyte population to the CD8 lymphocyte population is greater than another threshold, then the second type of therapy will be applied. Otherwise, no therapy will be applied. The advantages of the proposed control strategy are that the therapy can be stopped under certain conditions, while the state variables of the overall system is asymptotically stable with fast convergent rate, the concentration of the controlled HIV-1 viral load is monotonic decreasing, as well as the positivity constraint of the system states and that of the dose concentration is guaranteed to be satisfied. Computer numerical simulation results are presented for an illustration. [ABSTRACT FROM AUTHOR]

The currently accepted paradigm for the primary T cell response is that effector T cells commit to autonomous developmental programs. This concept is based on several experiments that have demonstrated that the dynamics of a T cell response is largely determined shortly after antigen exposure and that T cell dynamics do not depend on the level and duration of antigen stimulation. Another experimental study has also shown that T cell responses are robust to variations in antigen-specific precursor frequency. Various mathematical models have corroborated the first result that programmed T cell responses are insensitive to the level of antigen stimulation. However, this paper proposes that programmed responses do not entirely explain the robustness of T cell dynamics to variations in precursor frequency. This work studies the hypothesis that the dynamics of a T cell response may also be governed by a feedback loop involving adaptive regulatory cells rather than by intrinsic developmental programs. We formulate two mathematical models based on T cell developmental programs. In one model, effector cells undergo a fixed number of divisions before dying. In the second model, effector cells live for a fixed time during which they may divide. The study of these models suggests that developmental programs are not sufficiently robust as they produce an immune response that directly scales with precursor frequencies. Consequently, we derive a third model based on the principle that adaptive regulatory T cells develop in the course of an immune response and suppress effector cells. Our simulations show that this feedback mechanism responds robustly over a range of at least four orders of magnitude of precursor frequencies. We conclude that the proliferation program paradigm does not entirely capture the observed robustness of T cell responses to variations in precursor frequency. We propose an alternative mechanism by which the primary T cell response is governed by an emergent group dynamic and not by individual T cell programs. [ABSTRACT FROM AUTHOR]

Please cite this paper as: T-lymphocyte-induced, fas-mediated apoptosis is associated with early keratinocyte differentiation. Experimental Dermatology 2009. The development of eczematous lesions is thought to be due in part to a breakdown in skin barrier function as a result of T lymphocytes (T cells) invading the skin causing epidermal keratinocyte apoptosis. In this study, we investigated the interaction of T cells and keratinocytes on apoptosis and terminal differentiation using an in vitro co-culture system. Experiments were performed using the HaCaT keratinocyte cell line or normal human epidermal keratinocytes. Activated human peripheral blood-derived T cells were found to induce Fas-dependent keratinocyte apoptosis by up to sixfold. Increased Fas was associated with increased IFN-γ. The T-cell apoptotic signal was found to target preferentially keratinocytes in the very early stages of terminal differentiation, such as those with low levels of α6-integrin expression, and result in subsequent increased caspase 3 activity. This observation was accompanied by a marked increase in keratinocyte ICAM-1 expression and its ligand LFA-1 on T cells. Our data suggest that T cells may initiate the onset of keratinocyte terminal differentiation making them more susceptible to Fas-dependent cell death signals delivered by the T cells. [ABSTRACT FROM AUTHOR]

Abstract: In human biomonitoring, factors such as age, gender, smoking habit and alcohol consumption are usually considered and identified as agents that exert an impact on genotoxicity biomarkers. However, as a rule, factors like micronutrient status are not considered in biomonitoring studies. This paper reports on genotoxic damage in Pb-exposed workers using data obtained from the cytokinesis-block micronucleus (CBMN) cytome assay and alkaline comet assay with silver staining, considering folate and vitamin B12 nutritional status. Analysis of the results showed that the Pb-exposed group presented a 24-fold higher Pb content in the blood compared to controls. The Pb-exposed workers presented significantly greater micronuclei (MNi) counts (Z =7.9583; p <0.0001) and DNA damage, assessed by the comet assay (DF: Z =7.7056; p <0.0001 and DI: Z =7.4749; p <0.0001), but no correlation with Pb blood concentrations were detected. These two groups did not differ significantly concerning folate and vitamin B12 levels. Regarding folate, it is possible that this similarity is associated with folic acid flour-enrichment, introduced in Brazil in 2004 to prevent neural tube defects. The mean folate level obtained in the Pb-exposed group was 6.18ng/ml. The subgroup of individuals with serum folate levels equal to or higher than the mean presented significantly higher MNi (Z =2.3776; p =0.017) and nucleoplasmic bridge (NPB) frequencies (Z =1.9850; p =0.047) in peripheral blood lymphocytes. A significant positive correlation was observed between the age of Pb-exposed workers and MNi frequencies (Γ =0.3328; p <0.001), NPBs (Γ =0.1832; p <0.042) and DNA damage assessed by the comet assay (DF: Γ =0.1764; p =0.035 and DI: Γ =0.11852; p =0.028). These findings suggest that high folate levels alone do not guarantee protection against genotoxic damage. Moreover, folic acid supplementation should be studied using more efficient approaches to determine safe amounts and potential deleterious effects. [Copyright &y& Elsevier]

The ability of B cells to negatively regulate cellular immune responses and inflammation has only recently been described. Hallmark papers from a number of distinguished laboratories have identified phenotypically diverse B-cell subsets with regulatory functions during distinct autoimmune diseases, including IL-10-producing B cells, CD5+ B-1a cells, CD1d+ marginal zone B cells, and transitional-2-marginal zone precursor B cells. Most recently, a numerically rare and phenotypically unique CD1dhiCD5+CD19hi subset of regulatory B cells has been identified in the spleens of both normal and autoimmune mice. CD1dhiCD5+ B cells with the capacity to produce IL-10 have been named B10 cells as they produce IL-10 exclusively and are the predominant B-cell source of IL-10. Remarkably, B10 cells are potent negative regulators of inflammation and autoimmunity in mouse models of disease in vivo. Herein, our current understanding of B10-cell development and function is reviewed in the context of previous studies that have identified and characterized regulatory B cells, emerging evidence for B10-cell regulation of tumor immunity, and the likelihood that B10 cells exist in humans. [ABSTRACT FROM AUTHOR]

Emerging evidence suggests a strong interaction between the gut microbiota and health and disease. The interactions of the gut microbiota and the liver have only recently been investigated in detail. Receiving approximately 70% of its blood supply from the intestinal venous outflow, the liver represents the first line of defense against gut-derived antigens and is equipped with a broad array of immune cells (i.e., macrophages, lymphocytes, natural killer cells, and dendritic cells) to accomplish this function. In the setting of tissue injury, whereby the liver is otherwise damaged (e.g., viral infection, toxin exposure, ischemic tissue damage, etc.), these same immune cell populations and their interactions with the infiltrating gut bacteria likely contribute to and promote these pathologies. The following paper will highlight recent studies investigating the relationship between the gut microbiota, liver biology, and pathobiology. Defining these connections will likely provide new targets for therapy or prevention of a wide variety of acute and chronic liver pathologies. [ABSTRACT FROM AUTHOR]

Ovarian cancer is one of the leading causes of cancer-related death among women. Resistance to the disease occurs in more than 70% of the cases even after treated with chemotherapy agents such as paclitaxel- and platinum-based agents. The immune system is increasingly becoming a target for intense research in order to study the host's immune response against ovarian cancer. T cell populations, including NK T cells and Tregs, and cytokines have been associated with disease outcome, indicating their increasing clinical significance, having been associated with prognosis and as markers of disease progress, respectively. Harnessing the immune system capacity in order to induce antitumor response remains a major challenge. This paper examines the recent developments in our understanding of the mechanisms of development of the immune response in ovarian cancer as well as its prognostic significance and the existing experience in clinical studies. [ABSTRACT FROM AUTHOR]