Levels Of CD1a Research Articles

Comparative Evaluation of the Immunophenotypic Profile of Acute Lymphoblastic Leukaemia in Adult and Paediatric Age Groups in Iraq

Background: Immunophenotyping is a primary tool for investigating acute lymphoblastic leukemia (ALL). Acute lymphoblastic leukemia accounts for 75 - 80% of pediatric leukemia cases and 14% of adult leukemia cases. However, the prognostic factors, treatment strategies, and pathogenic pathways differ between these groups. Objectives: This study aims to compare the pattern of antigen expression between adult and pediatric ALL patients in the Iraqi population and to evaluate the frequency of aberrant myeloid antigen expression in both groups. Methods: A total of 131 patients with de-novo ALL were studied from May 2014 to May 2015. The diagnosis of ALL was confirmed by morphology and immunophenotyping of peripheral blood and bone marrow aspirate specimens using a panel of fluorochrome-labeled antibodies in 6 - colored flow cytometry. Results: The B-ALL phenotype was prevalent in 44 (68.7%) of adult patients compared to 51 (76.1%) in the pediatric group. CD10 expression was present in 49 (76.6%) of adults, mostly with B-ALL. The pediatric group had lower CD10 expression in T-ALL (P = 0.039). CD20 expression was significantly lower in adults compared to children. In T-ALL, significant differences between adult and pediatric groups were observed in CD1a and CD2 expression. Aberrant myeloid antigen expression was present in 29 (45.3%) adults and 30 (44.8%) pediatric patients, with CD33 and CD13 being the most prevalent in both groups. Conclusions: There was an increased T-cell lineage and aberrant antigen expression in both adult and pediatric ALL cases in this Iraqi sample. Adults with ALL tend to display higher levels of CD20 and lower levels of CD1a and CD2 compared to children, markers associated with poorer prognoses.

High expression of circulating exosomal PD-L1 contributes to immune escape of hepatocellular carcinoma and immune clearance of chronic hepatitis B.

To investigate the expression of programmed death ligand-1 (PD-L1) in circulating exosomes, and to define the role of exosomal PD-L1 in promoting immune escape mechanism during chronic hepatitis B infection (CHB) and related liver diseases. The levels of PD-L1 expressed in exosomes were detected by ELISA. CD8+T cells were sorted and cytotoxicity test was assessed by flow cytometry. PD-L1 protein expression in hepatocellular carcinoma (HCC) and normal adjacent tissues were detected by immunohistochemistry. Circulating exosomal PD-L1 levels were significantly higher in patients with CHB and HCC than in healthy controls (F =7.46, P=0.001). Levels of CD107a on CD8+T cells in patients with CHB receiving PD-L1 blocking antibody were significantly lower than in patients receiving isotype blocking antibody (t = 4.96, P < 0.01). Levels of TNF-α in cell culture supernatants of the PD-L1 blocking antibody group were significantly higher than in the isotype blocking antibody group (t =5.92, P < 0.01). Compared with patients receiving isotype blocking antibody, levels of CD107a on CD8+T cells significantly increased in patients with HCC receiving anti-PD-L1 antibody (t = 3.51, P<0.05). Compared with adjacent tissues, the levels of PD-L1 protein expression in HCC tissues were slightly higher; however, no significant difference between the two groups was observed. PD-L1 blockade in exosomes might promote the cytotoxic function of CD8+T cells and inhibit immune evasion during progression of HCC. Blocking PD-L1 in exosomes reduced the cytotoxic function of CD8+T cells in patients with CHB while enhancing the production of proinflammatory cytokines.

NUSAP1 Promotes Immunity and Apoptosis by the SHCBP1/JAK2/STAT3 Phosphorylation Pathway to Induce Dendritic Cell Generation in Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) is the most common type of liver cancer and is associated with high morbidity and mortality rates. The aims of this study were to investigate the immune-promoting action of nucleolar and spindle-associated protein 1 (NUSAP1) and identify an immunotherapy target for HCC. The Cancer Genome Atlas (TCGA) was used to analyze interaction molecules and immune correlation. The interaction between NUSAP1 and SHC binding and spindle associated 1 (SHCBP1) was examined. The role of the SHCBP1/Janus kinase 2/signal transducer and activator of transcription 3 (SHCBP1/JAK2/STAT3) pathway in this process was explored. After co-culture with HCC cell lines, the differentiation of peripheral blood mononuclear cells (PBMCs) into dendritic cells (DC) was evaluated by measuring the expression of surface factors CD1a and CD86. Pathological tissues from 50 patients with HCC were collected to validate the results of cell experiments. The expression levels of CD1a and CD86 in tissues were also determined. The results show that NUSAP1 interacted with SHCBP1 and was positively correlated with DC. In HCC cell lines, an interaction was observed between NUSAP1 and SHCBP1. It was verified that NUSAP1 inhibited the JAK2/STAT3 phosphorylation pathway by blocking SHCBP1. After co-culture, the levels of CD1a and CD86 in PBMC were elevated. In the clinical specimens, CD1a and CD86 expression levels were significantly higher in the high-NUSAP1 group versus the low-NUSAP1 group. In Summary, NUSAP1 enhanced immunity by inhibiting the SHCBP1/JAK2/STAT3 phosphorylation pathway and promoted DC generation and HCC apoptosis. NUSAP1 may be a target of immunotherapy for HCC.

CD1a and bound lipids drive T-cell responses in human skin disease.

In addition to serving as the main physical barrier with the outside world, human skin is abundantly infiltrated with resident αβ T cells that respond differently to self, infectious, microbiome, and noxious stimuli. To study skin T cells during infection and inflammation, experimental biologists track T-cell surface phenotypes and effector functions, which are often interpreted with the untested assumption that MHC proteins and peptide antigens drive measured responses. However, a broader perspective is that CD1 proteins also activate human T cells, and in skin, Langerhans cells (LCs) are abundant antigen presenting cells that express extremely high levels of CD1a. The emergence of new experimental tools, including CD1a tetramers carrying endogenous lipids, now show that CD1a-reactive T cells comprise a large population of resident T cells in human skin. Here, we review studies showing that skin-derived αβ T cells directly recognize CD1a proteins, and certain bound lipids, such as contact dermatitis allergens, trigger T-cell responses. Other natural skin lipids inhibit CD1a-mediated T-cell responses, providing an entry point for the development of therapeutic lipids that block T-cell responses. Increasing evidence points to a distinct role of CD1a in type 2 and 22 T-cell responses, providing new insights into psoriasis, contact dermatitis, and other T-cell-mediated skin diseases.

Natural Killer Cells in SARS-CoV-2-Vaccinated Subjects with Increased Effector Cytotoxic CD56dim Cells and Memory-Like CD57+NKG2C+CD56dim Cells.

The infection and negative effects of the SARS-CoV-2 (severe acute respiratory syndrome coronavirus) virus are mitigated by vaccines. It is unknown whether vaccination has worked by eliciting robust protective innate immune responses with high affinity. Twenty healthy volunteers received three doses of Comirnaty (Pfizer Australia Pty Ltd.) and were evaluated 9 months after the second vaccination and 1 month after the booster dose. The exclusion criteria were the presence of adverse effects following the vaccination, a history of smoking, and heterologous immunization. The inclusion criteria were the absence of prior Coronavirus Disease (COVID)-19 history, the absence of adverse effects, and the absence of comorbidities. Specific phenotype and levels of CD107a and granzyme production by blood NK (natural killer) cells were analyzed after exposure to SARS-CoV-2 spike antigen (Wuhan, Alpha B.1.1.7, Delta B.1.617.2, and Omicron B1.1.529 variants), and related with anti-SARS-CoV-2 antibody production. The booster dose caused early NK CD56dim subset activation and memory-like phenotype. We report the relevance of the innate immune response, especially NK cells, to SARS-CoV-2 vaccines to guarantee efficient protection against the infection following a booster dose.

Circulating microparticles are associated with plaque burden and cause eNOS uncoupling in patients with carotid atherosclerosis.

Aims: The study aimed to evaluate the correlation of different microparticle (MP) phenotypes with plaque burden and their diagnostic value and preliminarily explore the role of MPs in atherosclerosis (AS). Methods: Carotid intima-media thickness (CIMT) and maximal plaque area in 23 patients with carotid atherosclerosis (CAS) and 22 healthy subjects were measured by ultrasound. Transmission electron microscopy, nanoparticle tracking analysis and western blot were used to identify MPs. Flow cytometry assay measured absolute number of MPs, and receiver operating characteristic (ROC) analysis was used to assess the relationship between plaque burden and MPs. To study the preliminary mechanism of MPs in AS, MPs were administered to 32 male Kunming mice, which were randomly divided into control, CAS, healthy, and tetrahydrobiopterin (BH4) groups. Hematoxylin-eosin staining, immunohistochemistry staining, and Western blot were adopted to detect relevant indexes 24h after the injection. Results: The plasma levels of CD45+ leukocyte-derived microparticle (LMP), CD11a+ LMP, CD11a+/CD45+ LMP, and CD31+/CD42b+ platelet-derived microparticle (PMP) in CAS patients were significantly higher than those in healthy subjects, and were positively correlated with the maximal plaque area. Moreover, the levels of CD11a+ LMP, CD11a+/CD45+ LMP were also positively correlated with CIMT. The area under the ROC curve of the four MPs was 0.689, 0.747, 0.741, and 0.701, respectively. Compared with healthy subjects, MPs from CAS patients resulted in a significantly lower expression of endothelial nitric oxide synthase (eNOS) dimer/monomer, and BH4 could improve eNOS uncoupling. Moreover, the level of VCAM-1 in intima in the CAS group was significantly higher than in the other three groups. Conclusion: CD11a+ LMP and CD11a+/CD45+ LMP might be potential biomarkers for CAS prediction. BH4-related eNOS uncoupling occurs in CAS patients, and circulating MPs from them lead to endothelial dysfunction through eNOS uncoupling.

A Multi-Organ-on-Chip Approach to Investigate How Oral Exposure to Metals Can Cause Systemic Toxicity Leading to Langerhans Cell Activation in Skin.

Investigating systemic toxicity in vitro is still a huge challenge. Here, a multi-organ-on-chip approach is presented as a typical case of topical exposure of oral mucosa to metals, which are known to activate the immune system and in turn may result in skin inflammation. Reconstructed human gingiva (RHG) and reconstructed human skin containing MUTZ-3–derived Langerhans cells (MUTZ-LC) in the epidermis (RHS-LC) were incorporated into a HUMIMIC Chip3plus, connected by dynamic flow and cultured for a total period of 72 h. Three independent experiments were performed each with an intra-experiment replicate in order to assess the donor and technical variations. After an initial culture period of 24 h to achieve stable dynamic culture conditions, nickel sulfate was applied topically to RHG for 24 h, and LC activation (maturation and migration) was determined in RHS-LC after an additional 24 h incubation time. A stable dynamic culture of RHG and RHS-LC was achieved as indicated by the assessment of glucose uptake, lactate production, and lactate dehydrogenase release into the microfluidics compartment. Nickel exposure resulted in no major histological changes within RHG or RHS-LC, or cytokine release into the microfluidics compartment, but did result in an increased activation of LC as observed by the increased mRNA levels of CD1a, CD207, HLA-DR, and CD86 in the dermal compartment (hydrogel of RHS-LC (PCR)). This is the first study to describe systemic toxicity and immune cell activation in a multi-organ setting and can provide a framework for studying other organoids in the future.

Abstract 1536: Functional characterization of EDIT-201, a multiplexed CRISPR-Cas12a gene edited healthy donor derived NK cell therapy, reveals increased granzyme B and degranulation supporting improved serial killing capacity

Abstract We have previously described EDIT-201, a healthy donor derived NK cell therapy with a CISH and TGFBR2 knockout mediated by CRISPR-Cas12a that results in resistance to TGF-β inhibition and increased effector function. EDIT-201 kills tumor cells more efficiently and for a longer duration than control NK cells in the presence of TGF-β. Based on these data, we hypothesized that EDIT-201 has increased serial killing capacity. First, we explored the transcriptional changes contributed by each gene edit with a focus on transcripts critical for NK cell effector function and metabolism using Nanostring analysis. Unedited, mock electroporated, and control edited (targeting a biologically irrelevant site) NK cells were included as controls in addition to CISH and TGFBR2 single and double gene knockout (KO) NK cells to interrogate the potential impact of electroporation and double-stranded DNA breaks on NK cell function. All samples included in the analysis were cultured for 3 days in IL-15 post-electroporation. No significant transcriptional changes were detected in all control conditions, while samples that contained CISH editing clearly upregulated transcripts relevant for NK cell effector function, including contents of cytolytic granules (GZMB, GZMA and GZMH). EDIT-201 expressed an average of 22 fold more GZMB transcript than control NK cells as measured by RT-qPCR in four unique NK cell donors. Furthermore, EDIT-201 showed significantly higher levels of CD107a, a marker of degranulation, after 14hrs of co-culture with SKOV-3 tumor cells, suggesting that EDIT-201 had an increased capacity to degranulate relative to control NK cells. To determine the presence of granzyme proteins within tumor cells following engagement with NK cells, we developed a novel GzmB reporter gene and used lentiviral vectors to introduce this reporter into tumor cell lines. Using SK-OV-3 tumor cells transduced with our reporter, we demonstrated that upon coculture of EDIT-201 with tumor cells, intratumoral GzmB activity was identified 4 hours sooner than when the target cells were cultured with control NK cells. In addition, EDIT-201 affected 80% more SK-OV-3 tumor cells with granzyme B compared to control NK cells over a 20 hour period. Most significantly, our data demonstrated that EDIT-201 not only discharged GzmB more rapidly than control NK cells, but also the amount of GzmB degranulated is greater as well, confirming that enhanced degranulation is a key mechanism by which EDIT-201 has superior functional capacity relative to control NK cells. Together, these data demonstrate one mechanism by which EDIT-201 demonstrates superior cytotoxicity during in vitro killing of SK-OV-3 tumor targets, and continues to support the advancement of EDIT-201 as a potential novel gene edited NK cell-based treatment for cancer. Citation Format: Christopher M. Borges, Shuqi Zhang, Kevin Wasko, Kelly Donahue, Lincy Prem Antony, Jared Nasser, Steven Sexton, Amanda Pfautx, Richard A. Morgan, Karrie K. Wong. Functional characterization of EDIT-201, a multiplexed CRISPR-Cas12a gene edited healthy donor derived NK cell therapy, reveals increased granzyme B and degranulation supporting improved serial killing capacity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1536.

Higher CD1a Levels Correlate with PD-L1 Expression and Predict Worse Overall Survival in Triple-Negative Breast Carcinoma

Objective: The aim of this study was to measure the expression of PD-L1, CD1a (a marker for immature dendritic cells), and CD83 (a marker for mature dendritic cells) and further examine the associations of PD-L1, CD83, and CD1a with overall survival (OS) in triple-negative breast carcinoma patients. Methods: PD-L1, CD1a, and CD83 expression in breast carcinoma tissues and CD83 expression in lymph node tissues were examined by immunohistochemistry and tissue microarray in 159 patients. Patients were classified into the low, medium, and high PD-L1, CD1a, and CD83 levels. Pearson χ² test was used to analyze the correlations between PD-L1, CD1a, and CD83. The Kaplan-Meier method was used to calculate the OS. Multivariate analysis was used to identify determinants of 3- and 5-year OS. Results: 25.1, 25.8, and 49.1% of the patients had low, medium, and high PD-L1 levels, respectively. PD-L1 levels significantly correlated with CD1a (r = 0.30409, p < 0.001) and CD83 levels (r = 0.6146, p < 0.001) in breast carcinoma tissue, as well as CD83 levels (r = 0.17508, p = 0.027) in lymph node. The median OS was 83 months (range 12–106), and the 3- and 5-year OS rates were 94.97% (95% CI 91.57–98.37) and 86.79% (95% CI 81.53–92.06), respectively. Moreover, patients with high median CD1a levels had a significantly lower 5-year OS rate (75.6%) than those with low median CD1a levels (93.5%, p = 0.038). Conclusion: PD-L1, CD1a, and CD83 are variably expressed in triple-negative breast carcinoma tissues, and PD-L1 expression correlates with CD1a and CD83. Higher CD1a levels correlate with PD-L1 expression and predict worse OS in triple-negative breast carcinoma.

Dynamics of cytokines in patients with enteral insufficiency in advanced peritonitis

In this work, changes in the cytokine profile in patients with common forms of peritonitis with enteral insufficiency were studied. The obtained results were compared in patients who survived after surgery and programmed peritoneal sanation and patients who died in the early postoperative period. The aim of the study was to investigate the dynamics of cytokine profile of patients with common forms of peritonitis complicated by enteral insufficiency in order to determine their prognostic value. Material and methods. The object of the study was the blood of 23 patients with common forms of peritonitis and 17 healthy volunteers. To assess the condition of patients performed a dynamic determination in a comprehensive examination of laboratory markers of endogenous toxemia and systemic inflammatory response syndrome: immunological changes in patients by examining blood cells expressing clusters of CD11a+, CD162+, CD95+, CD16+, and molecules of the main histocompatibility complex HLA-DR+ and the level of interleukins IL-2, IL-4, IL-6. Results. During the study in patients with widespread peritonitis complicated by enteral insufficiency, at the time of hospitalization, there was a six-fold decrease in blood levels of CD11a+ cells and the content of CD162+ and CD16+ cells, respectively, 2,8 and 2 times, compared with healthy people. It has been shown that a single remediation programmed relaparotomy within two weeks effectively corrects these changes except for expression on CD16+ cells, which indicates a low effect of this type of surgery on the elimination of immune complexes. It was found that the concentration in plasma IL-2 increased 5 times and was 27,1% higher than that in almost healthy individuals. At the same time, the level of IL-4 in the blood decreased sharply – 4 times. Plasma IL-6 content also reduced. One week after relaparotomy, the expression of HLA-DR + molecules on immunocompetent cells increased again and was 1,8 times higher than the control parameters. In addition, the content of cytokines in the blood plasma increased sharply: the level of IL-2 exceeded the control by 2,4 times, IL-4 by 4 times, and IL-6 by 2 times. Conclusions. 1. Progressive and noticeable decrease in IL-2 content occurs in patients with widespread peritonitis complicated by enteral insufficiency in the complete absence of IL-4 in the blood on the background of a permanent and significant increase in plasma concentrations of IL-6. 2. Under conditions of suppression of the immune response, due to intoxication of the patient's body, the mechanism of endogenous inhibition of the immune response "IL-6 – corticoliberin – corticotropin – corticosteroids" is additionally realized.

Programmed cell death protein-1/programmed death-ligand 1 blockade enhances the antitumor efficacy of adoptive cell therapy against non-small cell lung cancer.

Cytokine-induced killer (CIK) cells and natural killer (NK) cells are employed by two different approaches to adoptive cell immunotherapy for cancer. It has been reported that adoptive cell immunotherapy could prolong the overall survival (OS) of advanced cancer patients. The introduction of agents that induce immune checkpoint blockades has improved the efficacy of immune-mediated therapy for metastatic cancers. However, the effects of combining a checkpoint inhibitor with CIK cells or NK cells to target non-small cell lung cancer (NSCLC)remain unknown. The present study investigated the effects of combining CIK cells with a programmed cell death protein-1 (PD-1) inhibitor (an anti-PD-1 monoclonal antibody). During the expansion cultivation, the addition of the PD-1 antibody promoted CIK-mediated cytotoxicity in H1975 lung adenocarcinoma cells. Co-cultivation of CIK cells with the PD-1 antibody for 6 days induced CD3+CD56+ T cell expansion, with increases in the levels of CD107a and interferon γ (IFN-γ). When NK cells were co-cultured with 5 µg/mL of an anti-programmed death-ligand 1 (PD-L1) mAb for 24 hours at an effector cell: target ratio of 10:1, it led to more potent cytotoxicity compared to other time points and concentrations. However, combining NK cells with the anti-PD-L1 mAb showed no significant advantages over treatment with NK cells alone. Our results suggest that combining CIK cells with PD-1 blockade before transfusion might improve the efficiency of CIK therapy for NSCLC patients. This effect does not seem to occur for NK cell therapy.

Effects of disodium cantharidinate on dendritic cells of patients with bladder carcinoma

The present study explored the effects of disodium cantharidinate (DC) on the peripheral blood-derived dendritic cells of patients with bladder carcinoma. The peripheral blood mononuclear cells from the 15 cases of urinary bladder carcinoma of middle and advanced stage were separated, and dendritic cells were prepared. The morphological changes of dendritic cells were observed. Flow cytometry was used to detect the expression levels of CD1a and CD83 on dendritic cell surface. MTT assay was utilized to measure the proliferation ability of allogeneic lymphocyte stimulated by DC. Annexin V-FITC/propidium iodide (PI) double staining flow cytometry method was carried out to detect cell apoptosis after treatment with DC. The changes in caspase-3 and PARP expression levels were investigated by western blot method. The high-dose DC resulted in a significant increase in the expressions of dendritic cell phenotyptic molecules CDla and CD83 as compared to control group. In addition, the proliferation index of allogenic lymphocyte stimulated by DC was significantly higher than that of control group. Moreover, MTT assay showed significant inhibition of the growth of BIU-87 cells. After 24 h of DC treatment, double staining flow cytometry confirmed the ability of DC to induce cell apoptosis. Further, western blot method showed a significant elevation of caspase-3 and PARP protein expression after DC treatment. In conclusion, DC treatment could induce dendritic cell maturation of patient with carcinoma of urinary bladder and promote its functional changes. Furthermore, DC was able to inhibit the proliferation of cell BIU-87 and also has the ability to induce cell apoptosis.

Variation and significance of secretory immunoglobulin A, interleukin 6 and dendritic cells in oral cancer.

The present study aimed to determine changes in the concentration of secretory immunoglobulin A (SIgA) and interleukin 6 (IL-6) in the saliva of patients with oral cancer, to evaluate the abnormal expression of cluster of differentiation (CD) 1a, CD83, CD80 and CD86 on dendritic cells (DCs) of oral cancer tissues and to discuss the interaction between SIgA, IL-6 and DCs in oral cancer. A total of 40 patients between 27 and 70 years of age, median age 52 years, with primary oral cancer were enrolled in the present study, and a group of 20 healthy male and female volunteers was used as the control group. The concentration of SIgA and IL-6 in the saliva of the preoperative patients was determined by ELISA. The expression levels of CD1a, CD83, CD80 and CD86 were detected by immunohistochemistry and flow cytometry, which was performed on histopathological sections from paraffin-embedded tumor and corresponding adjacent control tissues. The specimens were assessed using the semi-quantitative immunoreactive score (IRS). The concentration of SIgA in the saliva from patients with oral cancer decreased, whereas the IL-6 level significantly increased compared with the control subjects (P<0.05). In addition, the decrease of SIgA level and increase of IL-6 level exhibited a negative correlation (r=−0.543, P<0.05). According to the IRS score, the expression levels of CD1a, CD83, CD80 and CD86 in the cancer tissue were lower than the expression levels of the control group (P<0.05). Furthermore, the expression of CD80 and CD86 exhibited no correlation with histological grade or pathological type (P>0.05), but exhibited a negative correlation with clinical stage and lymph node metastasis (P<0.05). The concentration of SIgA and IL-6 in saliva may be used as an auxiliary diagnostic indicator for oral cancer. The detection of CD80 and CD86 expressed on DCs in oral cancer tissue may be useful for the diagnosis and evaluation of the prognosis of tumors. The present study hypothesized that the use of SIgA vaccines or IL-6 inhibitors may be useful for reversing the immune deficiency associated with DCs in oral cancer.

Psoriatic T cells recognize neolipid antigens generated by mast cell phospholipase delivered by exosomes and presented by CD1a.

Psoriasis is a chronic inflammatory skin disease associated with a T helper 17 response. Yet, it has proved challenging to identify relevant peptide-based T cell antigens. Antigen-presenting Langerhans cells show a differential migration phenotype in psoriatic lesions and express constitutively high levels of CD1a, which presents lipid antigens to T cells. In addition, phospholipase A2 (PLA2) is highly expressed in psoriatic lesions and is known to generate neolipid skin antigens for recognition by CD1a-reactive T cells. In this study, we observed expression of a cytoplasmic PLA2 (PLA2G4D) in psoriatic mast cells but, unexpectedly, also found PLA2G4D activity to be extracellular. This was explained by IFN-α-induced mast cell release of exosomes, which transferred cytoplasmic PLA2 activity to neighboring CD1a-expressing cells. This led to the generation of neolipid antigens and subsequent recognition by lipid-specific CD1a-reactive T cells inducing production of IL-22 and IL-17A. Circulating and skin-derived T cells from patients with psoriasis showed elevated PLA2G4D responsiveness compared with healthy controls. Overall, these data present an alternative model of psoriasis pathogenesis in which lipid-specific CD1a-reactive T cells contribute to psoriatic inflammation. The findings suggest that PLA2 inhibition or CD1a blockade may have therapeutic potential for psoriasis.

Abstract # 1731 Perioperative COX-2 inhibition and beta-adrenoceptor blockade in breast cancer patients improve markers of immunity and reduce inflammation and primary tumor epithelial-to-mesenchymal transition (EMT)

Preclinical research has implicated excess perioperative release of catecholamines and prostaglandins in mediating the metastasis-promoting effects of stress and surgery. This study reports the first clinical test of combined blockade of these pathways using the beta-adrenergic blocker, propranolol, and the COX-2 inhibitor, etodolac. In a double-blind placebo-controlled clinical trial, 36 breast cancer patients were randomized to an 11-day oral drug treatment beginning five days before surgery. Blood samples were taken before treatment (T1), on the mornings before and after surgery (T2&T3), and after treatment cessation (T4), and excised tumors were collected. No adverse effects of drug treatment were reported. Among other findings, in the placebo group, (i) serum IL-6 and CRP levels increased before surgery (T2), and (ii) stimulated IL-12 and IFN γ levels were reduced by stress and surgery (T2&T3). Both effects were blocked and further reversed by the drug treatment. Transcriptome analyses of excised tumor tissue identified 368 genes showing >1.20-fold differential expression as a function of drug treatment. Bioinformatics analyses based on a priory hypotheses indicated decreased tumor expression of several pro-metastatic gene programs, including inflammatory and epithelial-to-mesenchymal transition (EMT). Flow cytometry showed that drug treatment also abolished a postoperative (T3) circulating influx of immature-proinflammatory monocytes (CD14++CD16-), and enhanced perioperative (T2&T3) expression levels of CD11a on NK cells. These findings suggest pro-immune, anti-inflammatory, and potential anti-metastatic effects of this novel and safe therapy.

414 Identification of sebocytes as a novel source for adipokines within human skin

412 Characterization of the immunomodulatory effects of sebocytes on macrophages M Lovaszi, M Mattii, K Eyerich, D Kovacs, C Zouboulis, M Stahle, S Eyerich and D Torocsik 1 Department of Dermatology, Faculty of Medicine, University of Debrecen, Debrecen, Hungary, 2 ZAUM e Center for Allergy and Environment, Technische Universitat and Helmholtz Center Munich, Munich, Germany, 3 Department of Dermatology and Allergy, Technische Universitat Munich, Munich, Germany, 4 Departments of Dermatology, Venereology, Allergology and Immunology, Dessau Medical Center, Dessau, Germany and 5 Unit of Dermatology and Venereology, Department of Medicine, Karolinska lnstitutet, Karolinska University Hospital, Stockholm, Sweden Macrophages surround the sebaceous glands both in the healthy and the inflamed pilosebaceous unit; however, the reason for this phenomenon and the possible role of sebocytes in it has not yet been investigated. By performing immunohistochemistry on healthy biopsies, we could demonstrate that macrophages in the close proximity of sebaceous glands are exclusively alternatively activated (CD163/FXIII-A/CD206/CD209). To investigate if sebocytes contribute to the differentiation, polarization and function of macrophages, human peripheral blood monocytes were differentiated and activated in the presence of either supernatant from the human SZ95 sebocyte cell line, or major sebum lipid components such as oleic-, linoleic-, palmitic-, stearicacids and squalene. Our results showed that with the secretion of CXCL8, sebocytes could exert a chemoattractant effect towards monocytes, while sebocytes derived lipids promoted monocyte differentiation into alternatively activated macrophages as marked by the up-regulation of CD206, CD209 and FXIII-A. Moreover, detection of the produced IL-1b, IL-6 and TNFa protein levels by Propionibacterium acnes activated macrophages revealed a selective inflammatory effect for the various sebum lipids. Our results altogether suggest a role for sebaceous glands in initiating and modulating innate immune responses via their proteins and lipids that are of possible pathologic and therapeutic relevance. 413 Epithelial resident and infiltrating dendritic cells amplify active and resolved psoriasis inflammation E Martini, M Wiken, S Cheuk, A Smed Sorensen, M Stahle and L Eidsmo Dept. of Medicine, Solna, Karolinska Institutet, Stockholm, Sweden In psoriasis, scaly skin lesions are caused by interactions between activated keratinocytes, T cells and dendritic cells present in the skin. Here we investigate the inflammatory profiles of skin resident Langerhans cells (LCs) and infiltrating epitdermal dendritic cells (eDCs) to determine if they have the potential to steer epithelial pathology in psoriasis. A population of LangerinCD11cCD1c eDCs with variable levels of CD1a and CD14 was strictly confined to epidermis in active psoriasis lesions and could not be detected in non lesional or resolved psoriasis or healthy skin. LCs and eDCs sorted from active psoriasis displayed shared expression of genes associated with T cell activation and neutrophil attraction. Although LCs from psoriatic lesions stimulated with Toll-like receptor (TLR) ligands produced the pro-inflammatory cytokines IL-1 beta and IL-23, eDCs were the major producer of these cytokines in the epidermis, and displayed comparable cytokine production to dDCs. In resolved psoriasis eDCs are absent but LCs promptly responded to TLR stimulation with increased IL-23 production. Our results show that infiltrating eDCs together with LCs strongly contribute to the pro-inflammatory environment within the epidermal compartment of the skin. In contrast, LCs alone show the capacity to activate pathogenic tissue resident T cells in resolved psoriasis.

412 Characterization of the immunomodulatory effects of sebocytes on macrophages

412 Characterization of the immunomodulatory effects of sebocytes on macrophages M Lovaszi, M Mattii, K Eyerich, D Kovacs, C Zouboulis, M Stahle, S Eyerich and D Torocsik 1 Department of Dermatology, Faculty of Medicine, University of Debrecen, Debrecen, Hungary, 2 ZAUM e Center for Allergy and Environment, Technische Universitat and Helmholtz Center Munich, Munich, Germany, 3 Department of Dermatology and Allergy, Technische Universitat Munich, Munich, Germany, 4 Departments of Dermatology, Venereology, Allergology and Immunology, Dessau Medical Center, Dessau, Germany and 5 Unit of Dermatology and Venereology, Department of Medicine, Karolinska lnstitutet, Karolinska University Hospital, Stockholm, Sweden Macrophages surround the sebaceous glands both in the healthy and the inflamed pilosebaceous unit; however, the reason for this phenomenon and the possible role of sebocytes in it has not yet been investigated. By performing immunohistochemistry on healthy biopsies, we could demonstrate that macrophages in the close proximity of sebaceous glands are exclusively alternatively activated (CD163/FXIII-A/CD206/CD209). To investigate if sebocytes contribute to the differentiation, polarization and function of macrophages, human peripheral blood monocytes were differentiated and activated in the presence of either supernatant from the human SZ95 sebocyte cell line, or major sebum lipid components such as oleic-, linoleic-, palmitic-, stearicacids and squalene. Our results showed that with the secretion of CXCL8, sebocytes could exert a chemoattractant effect towards monocytes, while sebocytes derived lipids promoted monocyte differentiation into alternatively activated macrophages as marked by the up-regulation of CD206, CD209 and FXIII-A. Moreover, detection of the produced IL-1b, IL-6 and TNFa protein levels by Propionibacterium acnes activated macrophages revealed a selective inflammatory effect for the various sebum lipids. Our results altogether suggest a role for sebaceous glands in initiating and modulating innate immune responses via their proteins and lipids that are of possible pathologic and therapeutic relevance. 413 Epithelial resident and infiltrating dendritic cells amplify active and resolved psoriasis inflammation E Martini, M Wiken, S Cheuk, A Smed Sorensen, M Stahle and L Eidsmo Dept. of Medicine, Solna, Karolinska Institutet, Stockholm, Sweden In psoriasis, scaly skin lesions are caused by interactions between activated keratinocytes, T cells and dendritic cells present in the skin. Here we investigate the inflammatory profiles of skin resident Langerhans cells (LCs) and infiltrating epitdermal dendritic cells (eDCs) to determine if they have the potential to steer epithelial pathology in psoriasis. A population of LangerinCD11cCD1c eDCs with variable levels of CD1a and CD14 was strictly confined to epidermis in active psoriasis lesions and could not be detected in non lesional or resolved psoriasis or healthy skin. LCs and eDCs sorted from active psoriasis displayed shared expression of genes associated with T cell activation and neutrophil attraction. Although LCs from psoriatic lesions stimulated with Toll-like receptor (TLR) ligands produced the pro-inflammatory cytokines IL-1 beta and IL-23, eDCs were the major producer of these cytokines in the epidermis, and displayed comparable cytokine production to dDCs. In resolved psoriasis eDCs are absent but LCs promptly responded to TLR stimulation with increased IL-23 production. Our results show that infiltrating eDCs together with LCs strongly contribute to the pro-inflammatory environment within the epidermal compartment of the skin. In contrast, LCs alone show the capacity to activate pathogenic tissue resident T cells in resolved psoriasis.

Abstract A03: Effects of survivin and lymphoma cell-derived exosomes on natural killer cell function

Abstract Tumors alter their microenvironment to promote survival using methods such as angiogenesis promotion, growth signals, and immune suppression. The immune system, whose regular function is to recognize and eradicate infected or abnormal cells, often seems unresponsive to transformed neoplastic cells. Many of the methods employed by tumors to reduce immune function are unclear, but there is evidence for T cell suppression, increased myeloid-derived suppressor cells (MDSCs), and reduced natural killer (NK) cell activity. NK cells are among the first cells to recognize and destroy abnormal cells due to their inherent killing capabilities. However, many types of cancers inhibit the surveillance and cytotoxic abilities of NK cells by releasing exosomes, vesicles that can modulate tumor microenvironment and intercellular communication for the purpose of enhancing tumor malignancy. Exosomes as a means for immunomodulation of tumor microenvironment have been the focus of recent research. These 50-150 nm sized lipid bound vesicles are secreted by many cell types, including immune cells and tumor cells, and the unique protein, lipid, mRNA and miRNA contents contribute to the complex intercellular communication occurring between malignant and normal cells. Cancer patients often have increased numbers of exosomes circulating through their body, including patients with hematological malignancies, such as lymphoma. Recently, cancer cell exosomes have been found to contain Survivin, an Inhibitor of Apoptosis (IAP) protein that prevents cell death and induces Th2 polarity shift and decreases proliferation and cytotoxicity of CD8+ T lymphocytes. The purpose of this study is to explore the effect of Survivin and cancer cell-derived exosomes, which contain Survivin, on the immune functions of NK cells. The hypothesis is that Survivin and exosomes will decrease NK cell functions by suppressing NK cell production or release of cytokines and cytotoxic granules. NK cells were obtained from peripheral blood using magnetic NK cell isolation kit from Miltenyi Biotec and exosomes were obtained from conditioned media from two lymphoma cell lines (WSU-DLCL2 and WSU-FSCCL) using ExoQuickTM (System Biosciences). Flow cytometry methods were used to evaluate degranulation capacity by measuring levels of CD107a, as well as expression of activating receptor NKG2D, cytotoxic granules (perforin, Granzyme B) and cytokines (TNF-α, IFN-γ). RNA message expression was investigated using both block RT-PCR and qPCR. Results showed little significant difference in ability to degranulate when exposed to stimuli, and NKG2D levels did not change after exposure to Survivin or lymphoma exosomes. Although ImageJ analysis of block PCR showed some instances of decreased band densities in some donors treated with Survivin protein, RNA levels in general did not show a trend between NK donors in response to treatment. However, protein expression of Granzyme B, perforin, TNF-α, and IFN-γ did appear to decrease after Survivin treatment. This work is still in the early stages and results are inconsistent due to donor variability. More samples are required, as well as further studies on NK cell functional cytotoxicity. Citation Format: Heather R. Ferguson Bennit, Amber Gonda, Jenniffer Licero Campbell, Laura Oppegard, David Chi, Nathan R. Wall. Effects of survivin and lymphoma cell-derived exosomes on natural killer cell function. [abstract]. In: Proceedings of the AACR Special Conference: Function of Tumor Microenvironment in Cancer Progression; 2016 Jan 7–10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2016;76(15 Suppl):Abstract nr A03.

Uptake of poly-dispersed single-walled carbon nanotubes and decline of functions in mouse NK cells undergoing activation

The interaction of poly-dispersed acid-functionalized single-walled carbon nanotubes (AF-SWCNT) with NK cells undergoing activation was examined. Exposure to AF-SWCNT during NK activation in vitro by interleukin (IL)-2, and in vivo by Poly(I:C) significantly lowered cytotoxic activity generated against YAC-1 tumor cells. Recoveries of spleen NK1.1+ cells as well as the activated subset of NK cells (NK1.1+CD69+ cells) were significantly reduced by the AF-SWCNT exposure. The proportion of apoptotic NK cells (NK1.1+ phosphatidylserine+) in the spleen cell preparations activated in vitro was also significantly elevated. Expression levels of CD107a [for assessing NK cell degranulation] as well as of FasL marker [mediating non-secretory pathway of NK cell killing] were significantly lower in cells exposed to AF-SWCNT during the activation phase. Intracellular levels of interferon (IFN)-γ and tumor necrosis factor (TNF)-α in the cells were also significantly reduced. Fluorescent AF-SWCNT (FAF-SWCNT) were internalized by the NK cells and uptake was significantly greater in activated cells. Confocal microscopy indicated the internalized FAF-SWCNT were localized to the cytoplasm of the NK cells. These results indicated that AF-SWCNT were internalized by NK cells and caused a general down-regulation of a variety of parameters associated with NK cell cytotoxicity and other cellular functions.

CD1 Antigen Presentation and Autoreactivity in the Pregnant Human Uterus.

CD11c(HI) human decidual macrophages express several isoforms of CD1 molecules. Their expression pattern and function required investigation. CD11c(HI) macrophages were isolated from decidua. Expression of CD1 isoforms and their ability to present lipid antigens to T cells was studied. CD1a, CD1c, and CD1d were all expressed on CD11c(HI) dMϕ, a pattern differing from those previously observed. Exposure of peripheral monocytes and dendritic cells to lipid isolates from decidua led to increased surface CD1a levels only. The CD1a and CD1c on dMϕ were able to present the appropriate lipid antigens to lipid antigen-specific T cells. Finally, autoreactivity of decidual T cells to CD1a was observed. The unique pattern of expression of CD1 isoforms on CD11c(HI) dMϕ is consistent with organ-specific roles of CD1 in human T-cell responses. dMϕ are able to present lipid antigens to both peripheral and decidual T cells and are major antigen-presenting cells in human decidua.