Efficacy of different hemodialysis methods on dendritic cell marker CD40 and CD80 and platelet activation marker CD62P and P10 in patients with chronic renal failure.

BACKGROUND: Chronic renal failure (CRF) has become a major public health concern, which increases the risk of stroke and systemic thromboembolism. Therefore, therapeutic strategies are in urgent requirement. This study was conducted for investigating efficacy of hemodialysis (HD), hemodiafiltration (HDF), and hemoperfusion (HP) in patients with CRF and the correlation with the presence of complications following HD therapy. METHODS: The therapeutic effect, living quality, biochemical indicators, and dry weight were detected before and after the treatment regimens. Flow cytometry was conducted to detect expressions of dendritic cell markers (CD40 and CD80) and platelet activation markers (CD62P and P10), and the relationship between their expression and therapeutic effect as well as the association of these expressions with complications was analyzed. RESULTS: After HD therapy, patients presented with decreased serum creatinine, serum phosphorus, triglyceride, parathyroid hormone, and ß2 -MG expression; increased hemoglobin, plasma albumin expressions, and dry weight; and enhanced therapeutic effect and living quality. CD62P and P10 expressions decreased, while CD40 and CD80 expressions increased following HD therapy. The therapeutic effect improved in patients with low expressions of CD40 and CD80 and high expressions of CD62P and P10 following HP treatment and complications were lower after treatment of HDF and HP. CONCLUSION: The aforementioned results indicated that CRF patients treated with HP exhibited higher expression of CD40 and CD80 and lower expression of CD62P and P10, suggesting that HP is conferred to have better efficacy than HDF and HD. Therefore, HP may be a promising clinical regimen for treatment of CRF patients.

Interaction between tumour-infiltrating B cells and T cells controls the progression of hepatocellular carcinoma.

OBJECTIVE: The nature of the tumour-infiltrating leucocytes (TILs) is known to impact clinical outcome in carcinomas, including hepatocellular carcinoma (HCC). However, the role of tumour-infiltrating B cells (TIBs) remains controversial. Here, we investigate the impact of TIBs and their interaction with T cells on HCC patient prognosis. DESIGN: Tissue samples were obtained from 112 patients with HCC from Singapore, Hong Kong and Zurich and analysed using immunohistochemistry and immunofluorescence. RNA expression of CD19, CD8A, IFNG was analysed using quantitative PCR. The phenotype of freshly isolated TILs was analysed using flow cytometry. A mouse model depleted of mature B cells was used for functional study. RESULTS: Tumour-infiltrating T cells and B cells were observed in close contact with each other and their densities are correlated with superior survival in patients with HCC. Furthermore, the density of TIBs was correlated with an enhanced expression of granzyme B and IFN-γ, as well as with reduced tumour viability defined by low expression of Ki-67, and an enhanced expression of activated caspase-3 on tumour cells. CD27 and CD40 costimulatory molecules and TILs expressing activation marker CD38 in the tumour were also correlated with patient survival. Mice depleted of mature B cells and transplanted with murine hepatoma cells showed reduced tumour control and decreased local T cell activation, further indicating the important role of B cells. CONCLUSIONS: The close proximity of tumour-infiltrating T cells and B cells indicates a functional interaction between them that is linked to an enhanced local immune activation and contributes to better prognosis for patients with HCC.

Antibody Drug-Target Engagement Measurement in Tissue Using Quantitative Affinity Extraction Liquid Chromatography-Mass Spectrometry: Method Development and Qualification.

We demonstrate a novel strategy using affinity extraction (AE) LC-MS to directly measure drug exposure and target engagement, two critical pharmacological questions, with a single assay. The assay measures total drug and target concentration at the site of therapeutic action, as well as the amount of target bound to drug. The case study presented applies the strategy to measure drug engagement of a membrane bound receptor (CD40) that is critical to immune regulation in colon biopsies collected from monkey dosed with an anti-CD40 antibody. Unlike other techniques that measure receptor occupancy, such as flow cytometry, this technique does not rely on viable cells allowing measurement of frozen samples in a remote setting from the clinic.

Density and Polarization States of Tumor-Associated Macrophages in Human Cutaneous Squamous Cell Carcinomas Arising in Solid Organ Transplant Recipients.

BACKGROUND: Solid organ transplant recipients (SOTRs) are 100 times more likely to develop cutaneous squamous cell carcinoma (SCC) with greater metastatic propensity compared with the general population, likely due to chronic immunosuppression and adverse drug effects on keratinocytes. Tumor-associated macrophages (TAMs) play critical roles in malignancies, either aiding in eradication of malignant cells or promoting tumor growth. OBJECTIVE: The authors examined whether TAM density and polarization states differ between SOTRs and nontransplant individuals. MATERIALS AND METHODS: The authors obtained normal skin, SCC in situ (SCCis), and SCC from SOTRs and nontransplant patients (N = 45) and stained with macrophage marker CD68, M1 marker CD40, and M2 marker arginase-1. RESULTS: The authors report a significantly higher density of TAMs in both SCCis and SCC. The intratumoral macrophage infiltration in SCCis from SOTR was significantly decreased compared with nontransplant patients. Tumor-associated macrophages in SCCis and SCC displayed both M1 and M2 polarization, and M2 activation levels were significantly lower in SCC from SOTR. CONCLUSION: Tumor-associated macrophages are present in early carcinogenesis and may play a critical role in the transition from SCCis to SCC, before invasion of the basement membrane by tumor cells. The intratumoral macrophage density in early stages of tumor development is significantly affected in SOTR.

Impact of cardiac surgery on the expression of CD40, CD80, CD86 and HLA-DR on B cells and monocytes.

OBJECTIVE: We measured and compared changes in the percentage of cells expressing CD80, CD86, CD40, HLA-DR and the expression of these molecules on B cells and monocytes of patients who underwent either on-pump, mini on-pump or off-pump cardiac surgery. METHODS: Blood samples from patients who underwent either on-pump, mini on-pump or off-pump cardiac surgery were collected before surgery, instantly after surgery and on the 1(st), 3(rd) and 7(th) days after surgery. Surface expression of CD80, CD86, CD40 and HLA-DR molecules was determined by flow cytometry. RESULTS: Our results show that all three surgical techniques altered the expression of these molecules, as well as the percentage relative number of specific cell populations. We identified statistically significant differences when comparing different surgical techniques. On-pump surgery revealed a more pronounced impact on the phenotype of immune system cells than the other techniques. Therefore, it is likely that the function of immune cells is changed the most by on-pump surgery. We found a lower decrease in the number of CD80(+) monocytes and a lower drop in the CD40 expression on monocytes in off-pump patients in comparison with on-pump patients. CONCLUSION: All the types of cardiac surgical techniques, off-pump, on-pump and modified mini-invasive on-pump, are associated with changes in CD80, CD86, CD40 and HLA-DR expression. We found several significant differences in the expression of the selected molecules when we compared all three groups of patients.

Unconventional maturation of dendritic cells induced by particles from the laminated layer of larval Echinococcus granulosus.

The larval stage of the cestode parasite Echinococcus granulosus causes hydatid disease in humans and livestock. This infection is characterized by the growth in internal organ parenchymae of fluid-filled structures (hydatids) that elicit surprisingly little inflammation in spite of their massive size and persistence. Hydatids are protected by a millimeter-thick layer of mucin-based extracellular matrix, termed the laminated layer (LL), which is thought to be a major factor determining the host response to the infection. Host cells can interact both with the LL surface and with materials that are shed from it to allow parasite growth. In this work, we analyzed the response of dendritic cells (DCs) to microscopic pieces of the native mucin-based gel of the LL (pLL). In vitro, this material induced an unusual activation state characterized by upregulation of CD86 without concomitant upregulation of CD40 or secretion of cytokines (interleukin 12 [IL-12], IL-10, tumor necrosis factor alpha [TNF-α], and IL-6). When added to Toll-like receptor (TLR) agonists, pLL-potentiated CD86 upregulation and IL-10 secretion while inhibiting CD40 upregulation and IL-12 secretion. In vivo, pLL also caused upregulation of CD86 and inhibited CD40 upregulation in DCs. Contrary to expectations, oxidation of the mucin glycans in pLL with periodate did not abrogate the effects on cells. Reduction of disulfide bonds, which are known to be important for LL structure, strongly diminished the impact of pLL on DCs without altering the particulate nature of the material. In summary, DCs respond to the LL mucin meshwork with a "semimature" activation phenotype, both in vitro and in vivo.

Porphyromonas gingivalis lipopolysaccharide weakly activates M1 and M2 polarized mouse macrophages but induces inflammatory cytokines.

Porphyromonas gingivalis is associated with chronic periodontitis, an inflammatory disease of the tooth's supporting tissues. Macrophages are important in chronic inflammatory conditions, infiltrating tissue and becoming polarized to an M1 or M2 phenotype. As responses to stimuli differ between these phenotypes, we investigated the effect of P. gingivalis lipopolysaccharide (LPS) on M1 and M2 macrophages. M1 and M2 polarized macrophages were produced from murine bone marrow macrophages (BMMϕ) primed with gamma interferon (IFN-γ) or interleukin-4 (IL-4), respectively, and incubated with a low or high dose of P. gingivalis LPS or control TLR2 and TLR4 ligands. In M1-Mϕ, the high dose of P. gingivalis LPS (10 µg/ml) significantly increased the expression of CD40, CD86, inducible nitric oxide synthase, and nitric oxide secretion. The low dose of P. gingivalis LPS (10 ng/ml) did not induce costimulatory or antibacterial molecules but did increase the secretion of IL-1α, IL-6, IL-12p40, IL-12p70, and tumor necrosis factor alpha (TNF-α). P. gingivalis LPS marginally increased the expression of CD206 and YM-1, but it did enhance arginase expression by M2-Mϕ. Furthermore, the secretion of the chemokines KC, RANTES, eotaxin, and MCP-1 from M1, M2, and nonpolarized Mϕ was enhanced by P. gingivalis LPS. TLR2/4 knockout macrophages combined with the TLR activation assays indicated that TLR2 is the main activating receptor for P. gingivalis LPS and whole cells. In conclusion, although P. gingivalis LPS weakly activated M1-Mϕ or M2-Mϕ compared to control TLR ligands, it induced the secretion of inflammatory cytokines, particularly TNF-α from M1-Mϕ and IL-10 from M2-Mϕ, as well as chemotactic chemokines from polarized macrophages.

Differential partial activation phenotype and production of tumour necrosis factor-α by conventional dendritic cells in response to lipopolysaccharide in HIV+ viraemic subjects and HIV+ controllers.

HIV(+) subjects are reported to have increased soluble CD14 (sCD14) in plasma, an indicator of microbial translocation. We evaluated if microbial translocation has a differential impact on the activation and function of conventional dendritic cells (cDC) from viraemic HIV(+) subjects and HIV(+) controllers (CTs). The HIV(+) subjects were classified into two groups according to their plasma viral load (pVL): CT and viraemic. Subjects without HIV were included as controls (HIV(-) ). The frequencies and phenotypes of cDC from these subjects were evaluated by multi-parameter flow cytometry. In addition, peripheral blood mononuclear cells (PBMCs) were stimulated with lipopolysaccharide (LPS) or single-stranded RNA40 (ssRNA40), the phenotype of the cDC and the intracellular production of tumour necrosis factor (TNF)-α by the cDC were evaluated by flow cytometry. We observed a partial activation phenotype for the cDC in the viraemic subjects and CTs ex vivo and after LPS activation, which showed differences in the expression of CD40 and CD86. Furthermore, in response to LPS the cDC from the viraemic subjects produced more TNF-α compared to the cDC from CTs. Interestingly, the percentage of TNF-α(+) cDC was found to be correlated positively with the pVL. The partial activation of cDC and the over-production of TNF-α in response to LPS in viraemic HIV(+) subjects might be related to the increased chronic activation observed in these subjects. In contrast, cDC from CTs seem to have a regulated response to LPS, indicating that they respond differently to chronic immune activation. These results may have implications in the development of HIV therapies and vaccines using DC.

Removal of biological response modifiers associated with platelet transfusion reactions by columns containing adsorption beads.

BACKGROUND: Biological response modifiers (BRMs), such as soluble CD40 ligand (sCD40L); regulated upon activation, normal T-cell expressed, and secreted (RANTES); and transforming growth factor-ß1 (TGF-ß1), are released from platelets (PLTs) during storage and may trigger adverse effects after PLT transfusion. Although washing PLTs is effective at reducing the level of BRMs and the incidence of transfusion reactions, the washing procedure is time-consuming and may induce PLT activation. Furthermore, some BRMs continue to accumulate during the storage of washed PLTs. A method to remove BRMs using adsorbent columns has not yet been developed. STUDY DESIGN AND METHODS: We evaluated the ability of columns packed with Selesorb and Liposorber beads, which are both clinically used, to remove BRMs from PLT concentrates (PCs) stored for 5 days. The levels of these BRMs were determined before and after adsorption. RESULTS: The adsorption columns significantly reduced the levels of RANTES and sCD40L and partially reduced TGF-ß1. There were no significant effects on PLT activation, aggregation, morphology, and plasma lactate dehydrogenase (an indicator of PLT lysis) levels, or hypotonic shock response. Adsorption, however, reduced the PLT recovery to approximately 60% of the untreated value. CONCLUSIONS: This study showed that the levels of BRMs were substantially reduced using columns of clinically available adsorption beads. PLT functions and the quality of PCs were maintained after adsorption. The use of adsorption columns may be useful in reducing the incidence of nonhemolytic transfusion reactions.

Dendritic cell, monocyte and T cell activation and response to glatiramer acetate in multiple sclerosis.

BACKGROUND: Treatment with glatiramer acetate (GA) modestly decreases disease activity in multiple sclerosis (MS). The mechanism of action is incompletely understood and differences in the response to treatment between individuals may exist. OBJECTIVE: To study the activation of CD4+ T cells, monocytes and dendritic cells (DC) in relation to disease activity in MS patients treated with GA. METHODS: Flow cytometry was used to study the activation of CD4+ T cells and T cell subsets (CD25(high) and CD26(high) cells), monocytes and DCs in a cross-sectional study of 39 untreated and 29 GA-treated MS patients, the latter followed prospectively for one year. Gd-enhanced magnetic resonance imaging (MRI) studies were conducted in all patients. Disease activity was assessed as relapses. RESULTS: The median percentage of DCs expressing CD40 was 10% in untreated MS patients and 5.9% in GA-treated patients (Bonferroni-corrected p=0.0005). The hazard ratio of relapse was 1.32 (95% confidence interval 1.05-1.64) per 1% increase in CD40+ DCs. Patients treated with GA had fewer CD4+ T cells expressing surface markers associated with T helper type 1 effector responses and more CD4+ T cells expressing surface markers associated with regulatory, naïve or central memory T cell populations, but CD4+ T cell activation was not related with relapse risk. CONCLUSIONS: MS patients treated with GA show prominent changes in circulating antigen-presenting cells and CD4+ T cells. Expression of CD40 on DCs is significantly lower and associated with relapse risk in MS patients treated with GA.

CD40 signal regulates CXCR4 mediating ovarian carcinoma cell migration: implications for extrapelvic metastastic factors.

Ovarian carcinomas are highly invasive, especially in the peritoneal cavity. SDF-1α and its receptor, CXCR4, play a crucial role in migration of cancer cells. Here, SDF-1α directed HO8910 cell migration, but not SKOV3 cells. After being educated to express CXCR4 in vivo or by treating with sCD40L, SDF-1α reexhibited the ability of directing SKOV3 cell migration, which could be antagonized by CXCR4-neutralizing antibody. Furthermore, concomitant expression of CXCR4/CD40 in ovarian carcinoma tissues had stronger correlation with pelvic metastasis than did each alone. It is suggest that SDF-1α acts through CXCR4 to induce ovarian cancer cell migration, which could be facilitated by CD40 activation. Simultaneously examining the expression of CXCR4 and CD40 will provide valuable diagnosis of pelvic metastasis for ovarian carcinomas.

Markers CD40, VEGF, AKT, PI3K, and S100 correlate with tumor stage in gastric cancer.

BACKGROUND: To better understand gastric cancer occurrence and prognosis, we explored the expression of molecules in the CD40 pathway and their correlation with gastric cancer prognosis. PATIENTS AND METHODS: We measured the expression of CD40, VEGF, AKT, PI3K, and S100 in gastric cancer tissues and adjacent normal tissues from 128 patients by immunohistochemistry. RESULTS: The expression of CD40, VEGF, AKT, and PI3K were significantly higher in tumor tissue than in normal tissue, while S100 expression in dendritic cells (DC) was lower. Expression of CD40, VEGF, AKT, and PI3K significantly increased with T stage, while S100 expression decreased with T stage. Lymph node metastasis was associated with low or negative S100 expression. PI3K expression increased with clinical stage, while negative S100 expression was associated with higher clinical stages. Multivariate analysis did not indicate significant associations between any of these markers and recurrence or mortality. CONCLUSION: The correlation between T stage of gastric cancer and the higher expression of CD40, VEGF, AKT, and PI3K, along with lower S100 expression in DC, may provide insights into future targets for more effective immunotherapy for cancer.

Infiltration of CD40-positive tumor-associated macrophages indicates a favorable prognosis in colorectal cancer patients.

BACKGROUND/AIMS: Recently the role of tumor-associated macrophages (TAMs) on immunity has been variously discussed. We studied a series of cell surface antigens in TAMs in colorectal cancer tissues and their corresponding normal tissues using flow cytometry to find out prognostic indicators of these patients. METHODOLOGY: We assessed the numbers of CD14+ macrophages positive for each of the cell surface antigens (CD80, CD86, HLA-DR, CD1a, CD40 and CD83) in cancer tissues and corresponding normal tissues among 31 patients with colorectal cancer, and performed the univariate and multivariate analysis to find out prognostic indicators for overall survival among the patients. RESULTS: The numbers of CD80+, CD86+ and HLA-DR+ TAMs in the cancer tissues were higher than those in corresponding normal tissues. Inversely CD40+ and CD83+ macrophages in cancer tissues were less than those in normal tissues. With the multivariate analysis, the number of CD40+ TAMs, as well as lymph node metastasis and distant metastasis, was shown to be an independent prognostic factor of colorectal cancer patients. CONCLUSIONS: The dense infiltration of CD40+ TAM in colorectal cancer tissues indicates a favorable prognosis, which suggests that CD40 plays an important role in the tumor immunity of colorectal cancer.

CD40 expression in pancreatic cancer.

BACKGROUND: CD40, a tumor necrosis factor receptor family member, is expressed in a variety of cell types. This widespread expression suggests that CD40 may play an important role in normal physiology and disease pathogenesis. The objective of the current study was to investigate the expression of CD40, and its association with clinicopathological features and survival in patients with pancreatic ductal adenocarcinoma. METHODOLOGY: CD40 expression was assessed in 53 pancreatic ductal adenocarcinoma surgical specimens by immunohistochemistry, and expression was correlated with patient clinicopathological parameters and outcome. RESULTS: Among 53 pancreatic cancer specimens, CD40 expression was detected in 13 specimens (24.5%), and peritumoral lymphocytes were present in 45 specimens (84.9%). Patients with CD40-positive tumors exhibited prolonged median disease-free survival (DFS) compared with patients with CD40-negative tumors (15.60 +/- 3.87 versus 10.03 +/- 1.92); however, this was not significant (p = 0.845). Patients with peritumoral lymphocytic reaction exhibited prolonged median DFS compared with patients without peritumoral lymphocytes (10.96 +/- 1.40 vs. 7.60 +/- 0.47); however, this was not significant (p = 0.624). Patients with peritumoral lymphocytic reaction exhibited higher median overall survival compared with patients without peritumoral lymphocytes (15.20 +/- 1.78 vs. 10.13 +/- 1.39); however, again this was not significant (p = 0.100). CONCLUSIONS: These results suggest that CD40 expression on pancreatic cancer cells and peritumoral lymphocytic reaction may serve as prognostic markers.

Site-specific dynamics of CD11b+ and CD103+ dendritic cell accumulations following ozone exposure.

Pulmonary dendritic cells (DCs) are among the first responders to inhaled environmental stimuli such as ozone (O(3)), which has been shown to activate these cells. O(3) reacts with epithelial lining fluid (ELF) components in an anatomically site-specific manner dictated by O(3) concentration, airway flow patterns, and ELF substrate concentration. Accordingly, the anatomical distribution of ELF reaction products and airway injury are hypothesized to produce selective DC maturation differentially within the airways. To investigate how O(3) affects regional airway DC populations, we utilized a model of O(3)-induced pulmonary inflammation, wherein C57BL/6 mice were exposed to 0.8 ppm O(3) 8 h/day for 1, 3, and 5 days. This model induced mild inflammation and no remarkable epithelial injury. Tracheal, but not more distant airway sites, and mediastinal lymph node (MLN) DC numbers were increased significantly after the third exposure day. The largest increase in each tissue was of the CD103(+) DC phenotype. After 3 days of exposure, fewer DCs expressed CD80, CD40, and CCR7, and, at this same time point, total MLN T cell numbers increased. Together, these data demonstrate that O(3) exposure induced site-specific and phenotype changes in the pulmonary and regional lymph node DC populations. Possibly contributing to ozone-mediated asthma perturbation, the phenotypic changes to DCs within pulmonary regions may alter responses to antigenic stimuli. Decreased costimulatory molecule expression within the MLN suggests induction of tolerance mechanisms; increased tracheal DC number may raise the potential for allergic sensitization and asthmatic exacerbation, thus overcoming O(3)-induced decrements in costimulatory molecule expression.

Areca nut extracts suppress the differentiation and functionality of human monocyte-derived dendritic cells.

BACKGROUND AND OBJECTIVE: Areca quid chewing, a major risk factor contributing to the occurrence of oral cancer and precancer, has been reported to be associated with the severity and high prevalence of periodontal diseases in areca quid chewers. As dendritic cells are critically involved in the regulation of innate and adaptive immunity in oral mucosa, the objective of the present study was to investigate the effect of areca nut extracts (ANE) on the differentiation and reactivity of dendritic cells derived from monocytes. MATERIAL AND METHODS: Human peripheral blood monocytes were cultured in the presence of granulocyte-monocyte colony-stimulating factor and interleukin-4 for 7 d to generate dendritic cells. To examine the effect of ANE on the generation of dendritic cells, the monocytes were exposed to ANE throughout the 7 d culture period. In addition, the effect of ANE on the maturation of monocyte-derived dendritic cells induced by lipopolysaccharide (LPS) was examined. RESULTS: Monocytes cultured in granulocyte-monocyte colony-stimulating factor and interleukin-4 exhibited a typical phenotype of dendritic cells, as evidenced by the heightened expression of human leukocyte antigen (HLA)-DR, CD11c and the co-stimulatory molecules CD40, CD80 and CD86. Exposure of the monocytes to ANE did not influence the expression of HLA-DR and CD11c, but markedly attenuated the proportion of CD40-positive cells and the mean fluorescence intensity of CD86. The expression of co-stimulatory molecules in LPS-activated dendritic cells was not affected, whereas the mRNA expression of interleukin-12 induced by LPS was markedly suppressed by ANE treatment in a concentration-dependent manner. CONCLUSION: These results suggest that ANE exposure interfered with the differentiation of dendritic cells from monocytes. Moreover, the functionality of mature monocyte-derived dendritic cells was attenuated in the presence of ANE.

Antigen-presenting cell marker expression and phagocytotic activity in periodontal ligament cells.

BACKGROUND: Periodontal ligament (PDL) cells are the main cellular constituents of the periodontium, maintain the integrity of the connective tissue, and impact pathology in periodontitis. The aim of this study was to analyze whether PDL cells recognize foreign particles and participate in the immune response to periodontal pathogens. METHODS: Expression of surface proteins characteristic of antigen-presenting cells (APCs) (major histocompatibility complex [MHC] class II, CD40, CD80, CD86) was analyzed in PDL cells after challenge with the cytokines interleukin (IL)-1ß, IL-17A, and interferon-gamma (IFN-γ) or with heat-killed Aggregatibacter actinomycetemcomitans using real-time PCR and flow cytometry. Confocal laser scanning microscopy, transmitted light microscopy, flow cytometry, and time-lapse microscopy were applied to analyze their phagocytotic capacity of collagen (carboxylate-modified microspheres), non-periodontal (Escherichia coli) and periodontal (Aggregatibacter actinomycetemcomitans) pathogens. Furthermore, it was examined whether cytokine activation of PDL cells affects the phagocytosis of collagen or bacteria. RESULTS: PDL cells upregulated MHC class II after cytokine stimulation on transcriptional level, whereas co-stimulatory molecules characteristic of professional APCs were not induced. Analyses on protein level revealed that MHC class II was not constitutively expressed in all PDL cell lines used. PDL cells phagocytosed both collagen and bacteria via acidic vesicles, suggesting the formation of phagosomes. Phagocytosis could be partially inhibited by inhibitors of phagocytosis, i.e., dynasore and wortmannin. Pre-incubation with cytokines did not further enhance the phagocytosis rate of collagen or bacteria. CONCLUSIONS: These results suggest that PDL cells do not only represent bystanders in periodontal infections, but display non-professional APC characteristics, suggesting possible participation in immune reactions of the oral cavity.

Impaired dendritic cell maturation and IL-10 production following H. pylori stimulation in gastric cancer patients.

The current study was to investigate the interaction between Helicobacter pylori and human dendritic cells (DCs). Whether impaired DC function can influence the outcome of H. pylori infections. Human monocyte-derived DCs (MDDCs) from five gastric cancer patients and nine healthy controls were stimulated with H. pylori. Maturation markers of MDDC were examined by flow cytometry. IL-10 and TNF-α released by MDDCs and IL-17 produced by T cells were measured by ELISA. Regulatory signaling pathways of IL-10 were examined by ELISA, western blotting, and chromatin immunoprecipitation assay. The results showed that as compared with healthy individuals, the maturation marker CD40 in MDDCs, IL-17A expression from T cells, and IL-10 expression from MDDCs were significantly lower in gastric cancer patients. Blocking DC-SIGN, TLR2, and TLR4 could reverse H. pylori-associated IL-10 production. Activation of the p38 MAPK and NF-kB signaling pathways concomitant with decreased tri-methylated H3K9 and increased acetylated H3 accounted for the effect of H. pylori on IL-10 expression. Furthermore, upregulated IL-10 expression was significantly suppressed in H. pylori-pulsed MDDCs by histone acetyltransferase and methyltransferase inhibitors. Taken together, impaired DC function contributes to the less effective innate and adaptive immune responses against H. pylori seen in gastric cancer patients. H. pylori can regulate IL-10 production through Toll-like and DC-SIGN receptors, activates p-p38 MAPK signaling and the transcription factors NF-kB, and modulates histone modification.

Impact of immunotherapy on blood dendritic cells in patients with Hymenoptera venom allergy.

BACKGROUND: Modulation of T-cell differentiation, which is controlled by dendritic cells (DCs), plays a crucial role in specific immunotherapy (SIT). However, the number and the characteristics of blood DCs before and during immunotherapy are unknown. OBJECTIVE: To analyze the number and the characteristics of blood DC subsets in patients with Hymenoptera venom allergy before and after initiation of SIT. METHODS: In this clinical trial (NCT00947908), blood myeloid and plasmacytoid DCs were analyzed in 20 patients with Hymenoptera venom allergy (bee or wasp venom) by using 4-color flow cytometry at 3 time points: directly before SIT, and 52 hours and 12 months after initiation of SIT. In addition, 20 age-matched and sex-matched controls were examined. RESULTS: In patients with Hymenoptera venom allergy, the number of plasmacytoid DCs before SIT was comparable to that of controls. Plasmacytoid DCs decreased markedly 52 hours after initiation of SIT and returned to control levels after 12 months of treatment. Myeloid DCs were elevated in patients with Hymenoptera venom allergy before, during, and after the first 12 months of SIT. In addition, there were changes in the expression of function-associated surface molecules on myeloid DCs (such as Fc γ receptor 2 and Toll-like receptor 2) during SIT. CONCLUSION: Numbers of blood myeloid DCs are elevated in patients with Hymenoptera venom allergy, and there are specific changes in the expression of function-associated surface molecules on these cells during SIT. Numbers of plasmacytoid DCs in blood are profoundly but are only transiently decreased after initiation of SIT.

Analysis of CD40 expression in breast cancer and its relation to clinicopathological characteristics.

Breast cancer is currently the most common cancer in women worldwide. For this reason, new biomarkers for better predicting response to treatment are needed. CD40, described as expressed in haematological and epithelial tumors, is linked to apoptosis and offers promise as a new predictive/ prognostic marker. We evaluated CD40 expression in formalin-fixed, paraffin-embedded samples from 181 breast carcinomas using immunohistochemical staining with CD40 antibody. Samples were divided according to hormone (oestrogen receptor /ER/, progesterone receptor /PR/) and her-2/neu status into groups: 1.Luminal A (ER+PR+her-2/neu-), 2. Luminal B (ER+PR+her-2/neu+), 3.Triple-negative (ER-PR-her-2/neu-) and 4. Her-2/neu (ER-PR-her-2/neu+). The results of CD40 staining were correlated with clinicopathological data. CD40 was found to be expressed in membrane, cytoplasm and nucleus. Normal ducts expressed cytoplasmic CD40 in 30% of cases, in breast tumor ducts in 53% of cases. CD40 was evaluated as an independent marker and significant positive correlation was found with Bcl-2 (p =0.002), early stage (p =0.016) and preoperative chemotherapy (p =0.043). There was higher overall survival for patients with cytoplasmic CD40 expression (0.05). Differences in expression of cytoplasmic CD40 between groups with different hormonal and her-2/neu status were statistically highly significant (p=0.00003). In groups with different hormonal status, a positive statistical correlation was found for the luminal A group with relapse (p=0.024) and stage (p=0.006). No correlation was found with age, disease onset, family history of cancer/ breast cancer, patient history, hormonal replacement therapy, menopausal status at onset of disease, adjuvant chemotherapeutic treatment or disease free survival. Nuclear expression of CD40 was found to be unrelated to any clinicopathological data. However, there was higher ratio of positive cases in cancer cases (83%) than in normal tissue (30%). In conclusion, cytoplasmic expression of CD40 is related to factors connected to better prognosis and suggest that CD40 may have potential as a new prognostic factor in breast cancer.