DR Expression Research Articles

Clinical Outcome of Critically Ill Patients Cannot Be Defined by Cutoff Values of Monocyte Human Leukocyte Antigen-DR Expression

Septic shock is the most common cause of death in intensive care units. During the last two decades, new strategies have focused on the diagnosis and on the immunological changes in critically ill patients. There have been conflicting reports whether monocyte human leukocyte antigen (HLA) DR expression poses a useful parameter to characterize clinical outcome of these patients. To elucidate the role of monocyte HLA-DR expression, we hypothesized that low expression of HLA-DR on circulating human monocytes in critically ill patients correlates with higher mortality and that cutoff values of HLA-DR discriminate surviving from nonsurviving patients. In this retrospective study, monocyte HLA-DR expression in 413 critically ill patients was investigated during their intensive care unit stay. Human leukocyte antigen DR was determined in a quantitative and standardized procedure by flow cytometry (anti-HLA-DR monoclonal antibodies bound per cell [mABs/cell]) at least every third day or when clinical changes in the patients conditions were observed. Healthy probands served as control group to determine the range of "normal" values. As expected, HLA-DR expression was significantly higher in the group of survivors (n = 279) than in the group of nonsurvivors (n = 134; mABs/cell: 23,038 [SD, 11,150] vs. 18,070 [SD, 8,906]; P < 0.001). When minimal HLA-DR values per patient were compared, no cutoff values could be identified between the groups of survivors and nonsurvivors (mABs/cell: 19,611 [SD, 11,129] vs. 14,944 [SD, 8,013]; P < 0.001). In conclusion, in this sizable cohort we could again show that HLA-DR expression is decreased in critically ill patients but it is not suitable as a prognostic or predictive parameter for clinical outcome.

Biomarker Identification of Vitreous Fluid for Diabetic Retinopathy

For investigating the pathogenesis of DR, 2 main strategies are applied for the analysis of vitreous protein. The first strategy is antibody-based detection of vitreous cytokines. In previous studies of vitreous cytokines in DR, conventional enzyme-linked immunosorbent assay (ELISA) were used [4,5]. Recently, simultaneous analysis of the expression profiles of multiple cytokines and chemokines in the vitreous fluid was performed using an array system of antibody-coated beads [6]. Compared with the control group, interleukin (IL)-6, IL-8, IL-10, IL-13, interferon-inducible 10-kDa protein (IP-10), monocyte chemotactic protein-1 (MCP-1), macrophage inflammatory protein-1 beta (MIP-1β), platelet-derived growth factor (PDGF), and VEGF levels in the vitreous fluid were significantly higher in the DR group. The second strategy is proteomic analysis of vitreous proteins [7-11]. Fluorescence-based difference gel electrophoresis (DIGE) combined with matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) has enabled accurate quantitative comparisons of multiple proteins [12]. MALDI-TOF MS analysis of vitreous fluid detected approximately 1300 protein spots, and 25 of these intravitreal proteins were differentially expressed between DME and control groups. Hemopexin, beta-crystallin S, clusterin, and transthyretin were found to be specifically associated with DME. In a study using the same technique, 1242 protein spots were detected containing 19 differentially expressed intravitreal proteins between proliferative DR and control groups [13]. Differences in hemopoxin, clusterin, and pigment epithelium-derived factor (PEDF) levels were also observed between proliferative DR and control groups. Therefore, hemopexin expression was upregulated in DME and proliferative DR, whereas clusterin expression was downregulated in DME and proliferative DR and PEDF expression was downregulated only in proliferative DR. Hemopexin is an acute-phase plasma glycoprotein and is expressed in multiple cell lines derived from different tissues [14]. Importantly, increasing glucose concentrations in vitro increased hemopexin expression and modulated the reactive oxygen species levels in cells; these effects were partially reversed by addition of reduced glutathione. Clusterin is associated with protection from apoptosis of retinal cells [15]. In a mouse model of DR, clusterin reduced the leakage from vessels in the diabetic retina and restored expression of tight junction

Isolation and Characterization of Chorionic Mesenchymal Stromal Cells from Human Full Term Placenta

This study focused on the characterization of mesenchymal stromal cells (MSCs) from the chorion of human full term placenta from 15 donors. Chorionic MSCs revealed homologous fibroblast-like morphology and expressed CD73, CD29, CD105, and CD90. The hematopoietic stem cell markers including HLA DR, CD11b, CD34, CD79a, and CD45 were not expressed. The growth kinetics of their serial passage was steady at the later passages (passage 10). The multilineage capability of chorionic MSCs was demonstrated by successful adipogenic, osteogenic and chondrogenic differentiation and associated gene expression. Chorionic MSCs expressed genes associated with undifferentiated cells (NANOG, OCT4, REX1) and cardiogenic or neurogenic markers such as SOX2, FGF4, NES, MAP2, and NF. TERT was negative in all the samples. These findings suggest that chorionic MSCs undifferentiated stem cells and less likely to be transformed into cancer cells. A low HLA DR expression suggests that chorionic MSCs may serve as a great source of stem cells for transplantation because of their immune-privileged status and their immunosuppressive effect. Based on these unique properties, it is concluded that chorionic MSCs are pluripotent stem cells that are probably less differentiated than BM-MSCs, and they have considerable potential for use in cell-based therapies.

What’s New in Shock, December 2011?

What’s New in Shock, December 2011?

Changes Identified by Flow Cytometry and WT1 Expression in Consecutive Bone Marrow Samples in Refractory Cytopenia of Childhood and Aplastic Anemia Before Start of the Therapy

Abstract 1342 Introduction.Aplastic anemia (AA) and myelodysplastic syndrome (MDS) are rare diseases in childhood. The most common subtype of MDS is refractory cytopenia (RC). Both diseases typically exhibit with overlapping features and in both disorders dysregulation of immune system variably contributes to the degree of bone marrow (BM) failure. In the diagnostic algorithm plays role also analysis of consecutive BM samples by morphology. Patients and methods.Patients diagnosed between 2005 – 2011 with at least two BM samples analyzed by flow cytometry (FC) before treatment has started and with centrally evaluated BM biopsy according to EWOG MDS criteria were included into the study. We compared first and the last available sample before treatment (immunosupression or stem cell transplantation). By FC we analyzed following parameters: cell subsets (granulocytes, monocytes, lymphoid cells, erythroid precursors), BM precursors (CD34pos, CD117pos), T cells (CD3pos, CD3pos4pos, CD3pos8pos, CD3posHLA DRpos out of all cells, HLA DRpos out of CD3pos/CD3pos4pos/CD3pos8pos cells); B cells (CD19pos, CD19pos10pos, CD19pos45dim to neg, CD19pos34posout of all cells, CD10pos and CD20pos10neg out of CD19pos). In total 22 patients with AA (12 girls, 10 boys, mean age 11 years; 1.1–18 years) and 20 patients with RC (11 girls, 9 boys, mean age 11 years; 3.7–18) were included into the study. Median of time interval between both samples was 139 (1–1343) days in RC and 15 (1–56) days in AA. WT1 expression on mRNA level was analyzed in the sample before treatment with the highest number of CD34pos precursors to avoid blood contamination. All patients were screened by FISH for changes on chromosome 7 and 8. We asked following questions: Are there differences in the parameters in both bone marrow samples between SAA and RC? Is there any different pattern between d0 and before therapy sample between AA and RC? Are there any differences in WT1 expression between AA and RC group? Results.RC and AA significantly differ in both time points. AA patients have significantly decreased precursors (CD34, CD117); the difference is more pronounced at the later time point. More lymphocytes (both B and T) and less granulocytes are present at later time point in AA patients (p<0.05, Mann-Whitney test). Activation of CD8 cytotoxic T cells according to HLA DR expression is more distinct in AA patients at later time point. The most significant different parameter between RC and AA is a ratio CD19/CD34 also with the significant trend between two time points (Two way ANOVA, p<0.05). WT1 expression is statistically higher in RC patients; the higher expression is associated with presence of monosomy 7. Conclusion.By FC statistical differences can be identified in both samples (d0 and before treatment) between RC and AA. More pronounced differences are at later time point, which can be explained by further destruction of precursor and myeloid compartment more pronounced in AA patients compared to RC. WT1 expression is typically high in patients with RC and monosomy 7. Disclosures:No relevant conflicts of interest to declare.

Age-Dependent Alterations of HLA-DR Expression and Effect of Lipopolysaccharide on Cytokine Secretion of Peripheral Blood Mononuclear Cells in the Elderly Population

HLA-DR, a major histocompatibility complex, MHC class II, is involved in several autoimmune conditions, disease susceptibility and disease resistance. Here, we investigate the impact of different age individuals on HLA-DR expression on peripheral blood mononuclear cells (PBMCs) induced by lipopolysaccharide (LPS). The results indicate that HLA-DR expression on PBMCs in the population aged above 70 years significantly increased as compared with that in the lower-age groups by flow cytometry analysis (B-D; r = 0.690, P = 0.000265). In addition, followed by LPS stimulation, the levels of cytokine TNF-α, IL-6 and IL-10 secretion by allogeneic T lymphocytes from different age groups (A-D) were significantly increased (P < 0.05). Notably, levels of TNF-α and IL-6 were significantly reduced in groups B, C and D as compared with group A (P < 0.05), whereas levels of IL-10 increased (P = 0.012). However, no significant difference in PBMC-secreted TNF-α, IL-6 and IL-10 levels was observed among groups A, B, C and D (P > 0.05) before LPS stimulation. We firstly present data showing an age-dependent increase in HLA-DR expression in the population aged above 70 years and the individual with aged above 70 years has reduced the secretion of inflammatory mediators (TNF-α and IL-6) by PBMCs and increased the secretion of anti-inflammatory cytokine (IL-10) after LPS stimulation in vitro. The better understanding of HLR-DR expression on human PBMCs in aged individuals and its relation to the host cellular immunity would provide important insights into the immunity of aged individuals and particularly, in elderly patients with sepsis.

Association of an increased frequency of CD14+HLA‐DRlo/neg monocytes with decreased time to progression in chronic lymphocytic leukaemia (CLL)

Association of an increased frequency of CD14⁺HLA‐DR^lo/neg monocytes with decreased time to progression in chronic lymphocytic leukaemia (CLL)

Hyperosmolar Stress Upregulates HLA-DR Expression in Human Conjunctival Epithelium in Dry Eye Patients and In Vitro Models

To investigate the immune response of human conjunctival epithelium to hyperosmolar stress. Tear osmolarity was measured in 15 normal subjects and 25 dry eye (DE) patients; conjunctival imprint cytology samples were obtained at the nasal bulbar area. Subconfluent primary human conjunctival epithelial cells (pHCECs) and human conjunctival organ cultures (hCOCs), both cultured in iso-osmolar medium (305 mOsm/L), were exposed for 24 hours to media with progressively higher osmolarity, with or without the ion channel inhibitor ruthenium red (RuR). Human leukocyte antigen (HLA)-DR expression was evaluated by immunocytochemistry, on imprints from subjects, on primary human conjunctival epithelial cells, on formalin fixed-paraffin embedded hCOCs, and by RT-PCR. Statistical evaluation was performed by applying the unpaired Student's t test, as well as Spearman's rho and Pearson's r correlation coefficients (significance P < 0.05). HLA-DR expression increased in DE subjects with respect to control (% mean ± SD, respectively, 46.16 ± 7.2 vs. 7.48 ± 1.14, P < 0.0001) and exhibited significantly high correlations with tear osmolarity values (r = 0.614; P < 0.0001). In vitro experiments showed a progressive increase in HLA-DR expression as the osmolarity of the medium was increased from 6.75 ± 1.16 (% mean ± SD) in iso-osmolar-cultured cells to 9.96 ± 1.37 and 12.94 ± 4.04 in cells cultured in, respectively, 350 and 400 mOsm/L (P < 0.05). A stepwise progressive increase was also found in hCOCs. Results were confirmed by RT-PCR. Ruthenium red significantly reduced HLA-DR expression in hyperosmolar-cultured cells. Data from complementary techniques demonstrate that extracellular hyperosmolarity induces HLA-DR overexpression in human conjunctival epithelial cells in both DE patients and in vitro cell culture models.

Leukocyte subsets dynamics following open pulmonary lobectomy for lung cancer: a prospective, observational study

This study was planned to observe prospectively the effects of standard open pulmonary lobectomy on leukocyte subsets and their connection with oncological outcome. Leukocyte subsets from 200 patients undergoing pulmonary lobectomy were analyzed: 151 patients had non-small-cell lung cancer, and 49 had non-malignant diseases. Blood samples were taken for leukocyte flow cytometry before and five, 30 and 60 days after operation. The end points were: observation of postoperative leukocyte subsets that are dynamic in patients with lung cancer vs. patients without malignant disease; correlations between leukocyte subsets trend and disease-free interval or survival; and identification of prognostic factors related to preoperative leukocyte subsets. Lymphocyte counts significantly decreased at five days after lobectomy while monocyte counts increased, and complete recovery of the preoperative leukocyte setting was documented at 30 and 60 days. The patients with lung cancer showed a significant low percentage of human leukocyte antigens on their monocytes before surgery (P=0.0017), followed by a peculiar disarrangement of leukocytes subsets compared with patients without malignant disease at the five-day control point. There was no correlation between leukocyte subset dynamics and disease free interval or survival. This study proves reductions of T-, B and natural killer cells, and the expression of DR on T-lymphocyte after pulmonary lobectomy; oncological patients were significantly less affected by surgery than non-cancer patients.

Influence of 1,25-Dihydroxyvitamin D3 on Human Leukocyte Antigen-DR Expression, Macrophage Infiltration, and Graft Survival in Renal Allografts

Vitamin D plays an important role in the regulation of cellular growth and cell proliferation as well as exerting immunoregulatory activities. We sought to show the influence of vitamin D on renal graft survival. Among 102 patients, 40 were treated with vitamin D (group D) and the remaining 62 cases were not, forming a control group (group C). We studied human leukocyte antigen (HLA)-DR expression on tubules, interstitial cells, peritubular capillaries (PTCs), and evaluated macrophage infiltration of the interstitium and the PTCs. Compared to group C, group D patients showed fewer acute rejection episodes. Compared to group C patients group D patients showed significantly lower degrees of tubular and interstitial HLA-DR expression as well as interstitial and PTC macrophage infiltration. In addition in the PTC, HLA-DR expression was greater and therefore PTC destruction less in group D patients ( P < .001). Twenty-seven (43.5%) of 62 group C patients lost their grafts at 29.2 ± 15 months, whereas only 7/40 (17.5%) group D patients lost their grafts at 43.2 ± 13 months. A significant difference was noted between the two groups in regard to the time of graft loss ( P < .01). Testing vitamin D therapy along with several other covariates showed an independent effect on 5-year graft survival ( P < .001). These data confirmed that vitamin D therapy shows an independent positive impact on long-term graft survival, which may be explained by its immunosuppressive effects of to reduce renal HLA-DR expression and renal macrophage infiltration and in turn mitigate PTC destruction. In conclusion, these results highlighted the potential use of vitamin D in renal allograft patients.

Age-dependent differences in postoperative HLA-DR expression in cardiac surgery patients

Objective: Advanced age is a risk factor for clinical complications in cardiac surgery. Delayed recovery of human leucocyte antigen (HLA)-DR expression is also associated with complications after cardiac surgery. Little is currently known concerning the effect of age on HLA-DR expression.

Serial increase of IL-12 response and human leukocyte antigen-DR expression in severe sepsis survivors

IntroductionSepsis-induced immunosuppression may result in death. The mechanisms of immune suppression include loss of macrophage and monocyte expression of the major histocompatibility complex, increased anti-inflammatory cytokine expression and decreased expression of proinflammatory cytokines. In this study, we sought to determine the mechanisms of immune suppression in severe sepsis by repeated detection.MethodsWe designed this prospective observational study to measure monocyte human leukocyte antigen (HLA)-DR expression, plasma cytokine levels and cytokine responses on days 1 and 7 in stimulated peripheral blood mononuclear cells (PBMCs) of healthy controls and patients with severe sepsis.ResultsOf the 35 enrolled patients, 23 survived for 28 days and 12 died, 6 of whom died within 7 days. Plasma levels of IL-1β, IL-6, IL-10, IL-17, transforming growth factor (TGF)-β1 and TNF-α were higher, but plasma IL-12 level was lower in septic patients than those in controls. Day 1 plasma levels of IL-1β, IL-6, IL-10 and TGF-β1 in nonsurvivors were higher than those in survivors. Day 7 plasma IL-10 levels in nonsurvivors were higher than in survivors. IL-1β response was higher, but IL-12 and TNF-α responses were lower in septic patients than in controls. Day 1 IL-6 response was lower, but day 1 TGF-β1 response was higher in nonsurvivors than in survivors. Plasma IL-6 and IL-10 levels were decreased in survivors after 6 days. IL-6 response was decreased in survivors after 6 days, but IL-12 response was increased. Monocyte percentage was higher, but positive HLA-DR percentage in monocytes and mean fluorescence intensity (MFI) of HLA-DR were lower in septic patients than in controls. MFI of HLA-DR was increased in survivors after 6 days.ConclusionsMonocyte HLA-DR expression and IL-12 response from PBMCs are restored in patients who survive severe sepsis.

The suppression of immune activation during enfuvirtide-based salvage therapy is associated with reduced CCR5 expression and decreased concentrations of circulating interleukin-12 and IP-10 during 48 weeks of longitudinal follow-up

It has been suggested that patients who initiate highly active antiretroviral therapy (HAART) late in their course of infection may have suboptimal CD4 T-cell gains, persistent alterations in T-cell subsets and residual inflammation. To address this issue, we carried out a comprehensive 48-week immunological study in HIV-infected patients who had experienced failures of prior therapies, had low CD4 cell counts, and were receiving enfuvirtide-based salvage therapy. Immunological monitoring of peripheral lymphocytes from enfuvirtide-responder patients was performed over a 48-week period. A detailed assessment of immune cell subsets, their activation state [CD38 and human leucocyte antigen (HLA)-DR expression] and homeostasis [activation-induced cell death (AICD) and Ki67 expression], and the expression of co-receptors was performed by flow cytometry. Cytokine and chemokine signatures were assessed using multianalyte profiling technology. Enfuvirtide-based salvage therapy induced a progressive restoration of naïve and central memory CD4 T cells, associated with a decrease in their activation state, suppression of premature priming for AICD and increased expression of Ki67. In addition, a significant decrease in C-C chemokine receptor 5 (CCR5) expression was detected on CD4 T cells, which was strongly correlated with the suppression of immune activation. Changes in circulating proinflammatory molecules occurred; i.e. there were decreases in the concentrations of interleukin (IL)-12, macrophage inflammatory protein MIP-1α, MIP-1β, monokine induced by IFNγ (MIG) and interferon-γ-inducible protein-10 (IP-10). The decline in circulating IL-12 and IP-10 was correlated with both the reduction in the viral load and CD4 T-cell restoration. This study shows that suppression of HIV-1 replication with enfuvirtide-based salvage therapy in patients with low CD4 cell counts may result in an immunological benefit, characterized by the restoration of CD4 T-cell subsets associated with decreased immune activation and suppression of inflammation.

Patterns of HLA-DR Surface Expression on CD14+ Monocytes During Adverse Events After Hematopoietic Stem Cell Transplantation.

AbstractAbstract 1499The human leukocyte antigen DR (HLA-DR) surface expression on CD14+ monocytes reflects the degree of activation of these cells. In line with the central role of monocytes and macrophages in the immune system, a decreased HLA DR expression on CD14+ monocytes has been shown to be a hallmark of an altered immune status during the systemic inflammatory response syndrome (SIRS). We therefore hypothesized that HLA-DR expression might likewise be altered after hematopoietic stem cell transplantation (HSCT).HLA-DR surface expression of CD14+ monocytes was assessed by flow cytometry in 30 pediatric patients (below 15 years, n=23) and young adults (15 years and above, n=7) up to one year after HSCT (7 autologous and 23 allogeneic stem cell transplants). Normal values were derived from a control group of healthy children (n=18) and young adults (n=22).During the conditioning period, a significant increased HLA-DR expression in pediatric patients and young adults was observed. This increase could not be attributed to the administration of anti-thymocyte globulin (ATG). After HSCT, HLA-DR expression was not altered in general. However, HLA-DR expression decreased significantly up to 5 days before (p<0.05) and during (p<0.01) bacterial infections or sepsis. In contrast, HLA-DR expression levels were increased 7–10 days before and at the time of diagnosis of viral infections. HLA-DR expression was also elevated during acute graft-versus-host disease (GVHD). At the time of hepatic veno-occlusive disease (VOD), HLA-DR expression on CD14+ monocytes was reduced. Neither the administration of granulocyte colony-stimulating factor (G-CSF) nor the occurrence of a relapse was associated with a change of HLA-DR expression.In conclusion, adverse events after HSCT were associated with altered HLA-DR expression levels. Therefore, HLA-DR expression on CD14+ monocytes appears as a promising parameter which might allow identifying patients at risk after HSCT. Disclosures:No relevant conflicts of interest to declare.

Tumor necrosis factor–α and interleukin-10 contribute to immunoparalysis in patients with acute pancreatitis

Immunoparalysis, defined as downregulation of human leukocyte antigen–DR (HLA-DR) expression on monocytes, is strongly associated with septic complications of acute pancreatitis. However, the possible causes of this immunoparalysis have been largely unknown. A prospective case control study was performed in 54 patients with acute pancreatitis and 24 normal volunteers. HLA-DR expression on monocytes and serum cytokine levels were measured. In addition, monocytes from normal volunteers treated with tumor necrosis factor (TNF)–α in vitro were evaluated for HLA-DR expression and cytokine release. HLA-DR expression was significantly lower in patients with severe pancreatitis than in those with mild acute pancreatitis and healthy volunteers (42.28% ± 11.49% vs. 86.85% ± 14.56% vs. 93.92% ± 7.40%, p < 0.0001). Pearson correlation analysis showed that serum TNF-α and serum interleukin-10 levels were both correlated with HLA-DR expression. In addition, exogenous TNF-α could enhance IL-10 secretion from normal monocytes in a dose–response manner. In addition, TNF-α could downregulate the HLA-DR expression on monocytes even in the presence of anti-IL-10 antibodies. Therefore, both TNF-α and IL-10 contributed to the development of immunoparalysis in patients with acute pancreatitis.

Increased HLA‐DR Expression on Tissue Eosinophils in Eosinophilic Esophagitis

The aim of the study was to investigate whether eosinophils have increased human leukocyte antigen (HLA)-DR expression in subjects with eosinophilic esophagitis (EoE) compared with controls. Patients who were undergoing an upper endoscopy with biopsies for suspected gastroesophageal reflux disease (GERD) or EoE at Lucile Packard Children's Hospital were enrolled. In total, the blood and tissue samples of 10 healthy controls (HC), 11 subjects with GERD, and 10 with EoE were studied. Multiple tissue staining to identify eosinophils (via eosinophil cationic protein-clone EG2) and major histocompatibility complex class II cell surface receptors (via HLA-DR) was performed via immunohistochemistry. The peripheral blood was analyzed using flow cytometry to detect eosinophil HLA-DR expression among these subjects. In the tissue, a greater proportion of eosinophils expressed HLA-DR among the subjects with EoE (mean 0.83 +/- 0.14, n = 9) relative to those with GERD (mean 0.18 +/- 0.19, n = 8, P < 0.01) and HC (mean 0.18 +/- 0.13, n = 6, P < 0.01). In total, 6 participants (4 HC subjects and 2 subjects with GERD) did not have any eosinophils identified on tissue staining and were unable to be included in the present statistical analysis. In the blood, there was no statistically significant difference in eosinophil HLA-DR expression among HC subjects (mean 415 +/- 217, n = 6), subjects with GERD (mean 507 +/- 429, n = 2), and those with EoE (mean 334 +/- 181, n = 6). These data demonstrate that the eosinophils from the esophagus of subjects with EoE have increased HLA-DR expression within this tissue.

Quantitative Evaluation of Ocular Surface Inflammation in Patients with Different Grade of Conjunctivochalasis

Purpose: To evaluate and compare HLA-DR expression of conjunctival epithelial cells in different grade of conjunctivochalasis (CCh).Patients and Methods: Thirty patients and seven healthy subjects underwent clinical examination, grading of CCh by lid-parallel conjunctival fold (LIPCOF) test, and impression cytology of the bulbar conjunctiva. HLA-DR expression was analyzed by flow cytometry. Schirmer test, tear break-up time, and vital staining were also performed.Results: Patients who presented with mild and moderate CCh showed a similar expression of HLA-DR to normal subjects. A significant increase (p < 0.005) of HLA-DR was found in patients with severe CCh. Positive correlation was found between fluorescein staining and HLA–DR expression (r = 0.36, p = 0.009) and between lissamine–green staining and HLA-DR expression (r = 0.30, p = 0.027). Neither correlation was found between Schirmer test nor break up time and HLA-DR expression.Conclusion: Inflammation plays a pivotal role on the ocular surface of patients with severe CCh. Since mild and moderate CCh HLA-DR expression was similar to normal controls, we can speculate that the source of inflammation could be the presence of conjunctival folds.

Clinical Significance of Cloned Expansion and CD5 Down‐Regulation in Epstein‐Barr Virus (EBV)–Infected CD8+T Lymphocytes in EBV‐Associated Hemophagocytic Lymphohistiocytosis

Epstein-Barr virus (EBV) is the pathogen that most commonly triggers infection-associated hemophagocytic lymphohistiocytosis (HLH) and ectopically infects CD8(+) T cells in EBV-associated HLH (EBV-HLH). We recently described an EBV-HLH patient who had a clonally expanded population of EBV-infected CD8(+) T cells with CD5 down-regulation. To determine whether this finding could serve as a useful marker for EBV-HLH, we investigated 5 additional patients. We found a significant increase in the subpopulation of CD8(+) T cells with CD5 down-regulation and bright human leukocyte antigen (HLA)-DR expression in all patients with EBV-HLH but not in patients with infectious mononucleosis or in control subjects. Such T cells were frequently found to be larger cells that stained positive for a specific T cell receptor VB. We also demonstrated that those cells were the major cellular target of EBV, and their numbers progressively declined in parallel with the serum ferritin levels. All together, our findings reveal the immunophenotypic characteristics of EBV-infected CD8(+) T cells and may provide a valuable tool for the diagnosis of EBV-HLH.

Enhanced Indoleamine 2,3‐Dioxygenase Activity in Patients with Severe Sepsis and Septic Shock

Severe sepsis results in a sustained deleterious immune dysregulation. Indoleamine 2,3-dioxygenase (IDO), the rate-limiting enzyme of tryptophan catabolism, plays a pivotal role in immune tolerance and is induced during various inflammatory conditions. Plasma samples obtained from patients with septic shock (n = 38), severe sepsis (n = 35), or sepsis (n = 10) and from healthy donors (n = 26) were analyzed for IDO activity by high-performance liquid chromatography. Lymphocyte, monocyte, and regulatory T cell counts as well as monocytic human leukocyte antigen DR (HLA-DR) expression were quantified by flow cytometry. Peripheral blood mononuclear cells and purified CD14(+) and CD14(-) fractions were assayed in vitro for spontaneous and inducible IDO expression and activity. IDO activity gradually increased according to sepsis severity, and septic patients who died had higher IDO activity on admission than did survivors (P = .013). Monocytes were a major source of active IDO in normal peripheral blood. The percentage and absolute number of circulating CD14(+) cells were increased in septic patients, and their monocytes remained fully able to produce functional IDO after NF-kappaB-independent interferon gamma stimulation but not through NF-kappaB-dependent Toll-like receptor engagement. IDO activity is increased during severe sepsis and septic shock and is associated with mortality. IDO production could be used to better characterize monocyte reprogramming in sepsis.

Monocyte dysfunction in Sydenham's chorea patients

Until now, there are no conclusive data about the mechanisms involved in motor symptoms of Sydenham's chorea (SC). Taking into account the autoreactive antibody-mediated hypothesis of SC pathogenesis, the SC may be associated with uncontrolled immune mechanisms. Besides the antibody hypothesis, the innate immune system has been underappreciated. Hence, we evaluated the activation state of monocytes, cells that are precursors of macrophages, to characterize the inflammation profile of patients. We assessed the surface molecules CD80, CD86, and human leukocyte antigen DR expression in patients with SC by flow cytometry analysis. Our results showed a decreased CD14 + (monocyte) frequency, with concomitant increased CD14 − frequency inside monocyte population. Although monocyte population showed a decreased human leukocyte antigen DR and CD86 frequencies, the CD14 − population showed an increased frequency of CD80 + monocyte from SC compared with controls. These data suggest that monocytes showed a reduced costimulatory potential in SC.