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Background The goal of this study is to evaluate the effectiveness of S-1/Oxaliplatin vs. Doxifluridine/Oxaliplatin regimen and to identify miRNAs as potential prognostic biomarkers in gastric cancer patients. The expression of candidate miRNAs was quantified from fifty-five late stage gastric cancer FFPE specimens. Experimental Design Gastric cancer patients with KPS>70 were recruited for the trial. The control group was treated with 400 mg/twice/day Doxifluridine plus i.v. with Oxaliplatin at 130 mg/m2/first day/4 week cycle. The testing group was treated with S-1 at 40 mg/twice/day/4 week cycle plus i.v. with Oxaliplatin at 130 mg/m2/first day/4 week cycle. Total RNAs were extracted from normal and gastric tumor specimens. The levels of miRNAs were quantified using real time qRT-PCR expression analysis. Results The overall objective response rate (CR+PR) of patients treated with S-1/Oxaliplatin was 33.3% (CR+PR) vs. 17.6% (CR+PR) with Doxifluridine/Oxaliplatin for advanced stage gastric cancer patients. The average overall survival for patients treated with S-1/Oxaliplatin was 7.80 month vs. 7.30 month with patients treated with Doxifluridine/Oxaliplatin. The expression of miR-181b (P = 0.022) and miR-21 (P = 0.0029) was significantly overexpressed in gastric tumors compared to normal gastric tissues. Kaplan-Meier survival analysis revealed that low levels of miR-21 expression (Log rank test, hazard ratio: 0.17, CI = 0.06–0.45; P = 0.0004) and miR-181b (Log rank test, hazard ratio: 0.37, CI = 0.16–0.87; P = 0.018) are closely associated with better patient's overall survival for both S-1 and Doxifluridine based regimens. Conclusion Patients treated with S-1/Oxaliplatin had a better response than those treated with Doxifluridine/Oxaliplatin. miR-21 and miR-181b hold great potential as prognostic biomarkers in late stage gastric cancer.

Cell-mediated adaptive immunity is very important in tumor immune surveillance and tumor vaccination. However, the genetic program underlying an effective adaptive antitumor immunity is elusive. T-bet and Eomesodermin (Eomes) have been suggested to be master regulators of Th1 cells and CD8+ T cells. However, whether they are important for T cell-mediated antitumor immunity is controversial. In this paper, we show that the combined germline deletion of T-bet and T cell-specific deletion of Eomes resulted in profound defects in adaptive antitumor immune responses. T-bet and Eomes drive Tc1 differentiation by preventing alternative CD8+ T cell differentiation to Tc17 or Tc2 cells. Surprisingly, T-bet and Eomes are not critical for the generation of systemic CTL activities against cancer cells. Instead, T-bet and Eomes are crucial for tumor infiltration by CD8+ T cells. This study defines T-bet and Eomes as critical regulators of T cell-mediated immune responses against tumor.

BackgroundPhenobarbital (PB) is the most well-known among numerous non-genotoxic carcinogens that cause the development of hepatocellular carcinoma (HCC). PB activates nuclear xenobiotic receptor Constitutive Active/Androstane Receptor (CAR; NR1I3) and this activation is shown to determine PB promotion of HCC in mice. The molecular mechanism of CAR-mediated tumor promotion, however, remains elusive at the present time. Here we have identified Growth Arrest and DNA Damage-inducible 45beta (GADD45B) as a novel CAR target, through which CAR represses cell death.Methodology/principal findingsPB activation of nuclear xenobiotic receptor CAR is found to induce the Gadd45b gene in mouse liver throughout the development of HCC as well as in liver tumors. Given the known function of GADD45B as a factor that represses Mitogen-activated protein Kinase Kinase 7 - c-Jun N-terminal Kinase (MKK7-JNK) pathway-mediated apoptosis, we have now demonstrated that CAR interacts with GADD45B to repress Tumor Necrosis Factor alpha ( TNFalpha)-induced JNK1 phosphorylation as well as cell death. Primary hepatocytes, prepared from Car(+/+), Car(-/-), Gadd45b(+/+) and Gadd45b(-/-) mice, were treated with TNFalpha and Actinomycin D to induce phosphorylation of JNK1 and cell death. Co-treatment with the CAR activating ligand TCPOBOP (1,4 bis[2-(3,5-dichloropyridyloxy)]benzene) has resulted in repression of both phosphorylation and cell death in the primary hepatocytes from Car(+/+) but not Car(-/-) mice. Repression by TCPOBOP was not observed in those prepared from Gadd45b(-/-) mice. In vitro protein-protein interaction and phosphorylation assays have revealed that CAR interacts with MKK7 and represses the MKK7-mediated phosphorylation of JNK1.Conclusions/significanceCAR can form a protein complex with GADD45B, through which CAR represses MKK7-mediated phosphorylation of JNK1. In addition to activating the Gadd45b gene, CAR may repress death of mouse primary hepatocytes by forming a GADD45B complex and repressing MKK7-mediated phosphorylation of JNK1. The present finding that CAR can repress cell death via its interaction with GADD45B provides an insight for further investigations into the CAR-regulated molecular mechanism by which PB promotes development of HCC.

An effective Th1 type cell-mediated immune response against cancer cells is critical in limiting cancer progression. Gadd45b, a signaling molecule highly up-regulated during Th1 type responses, is studied for its role in limiting tumor growth. Mouse B16 melanoma cells implanted into Gadd45b(-/-) mice grew faster than those in WT or Gadd45b(+/-) littermate controls. The defect of Gadd45b(-/-) mice in tumor immunosurveillance was attributed to the reduced expression of IFN-gamma, granzyme B, and CCR5 in Gadd45b(-/-) CD8(+) T cells at the tumor site. Activation of p38 MAP kinase, but not ERK or JNK, by either TCR-stimuli or IL-12 and IL-18 is diminished in Gadd45b(-/-) CD8(+) T cells, resulting in reduced production of IFN-gamma. In addition, mRNA of T-bet and Eomes were reduced in Gadd45b(-/-) CD8(+) T cells, supporting a critical role of Gadd45b in shaping the Th1 fate. More importantly, the tumor vaccination, which is effective in WT mice, failed in Gadd45b/Gadd45g doubly deficient mice. Collectively, these data demonstrate that members of the Gadd45 gene family are important for anti-tumor immune responses.

Dendritic cells (DCs) are responsible for the initiation of immune responses. Our study demonstrates a new pathway for generating a large quantity of stimulatory monocyte-derived DCs (Mo-DCs) from human monocytes using anti-4-1BB ligand (4-1BBL) mAb to trigger reverse signaling. The anti-4-1BBL-driven Mo-DCs (DCs(alpha-4-1BBL)) not only express higher levels of CD86, CD83 and HLA-DR, when compared with the Mo-DCs matured by tumor necrosis factor alpha, but also exhibit a unique phenotype that expresses lower levels of PD-L1. High levels of GM-CSF, M-CSF and Flt3 ligand (FL) were found in the anti-4-1BBL-differentiation culture. Neutralizing M-CSF, GM-CSF and FL inhibited Mo-DC proliferation stimulated by anti-4-1BBL mAb, suggesting that M-CSF, GM-CSF and FL are involved in cell proliferation stimulated by anti-4-1BBL. Further analysis of the DCs(alpha-4-1BBL) showed increased secretion of T(h)1-type cytokines IL-12 and IFN-gamma and decreased secretion of IL-10. DCs(alpha-4-1BBL) induced much stronger proliferative responses in the mixed lymphocyte reaction assay when compared with DCs derived by GM-CSF. Moreover, DCs(alpha-4-1BBL) preferentially induced T(h)1 responses. We have further demonstrated that anti-4-1BBL antibody stimulated nuclear translocation of NF-kappaB from the cytoplasm in monocytes, suggesting that reverse signaling by 4-1BBL is likely responsible for mediating DC differentiation. Collectively, we have found that reverse signaling of 4-1BBL promotes the differentiation of potent T(h)1-inducing DCs from human monocytes.

CXCR4/SDF-1alpha signal is essential for cell development, hematopoiesis, organogenesis, and vascularization as well as leukocyte trafficking. Many published reports have highlighted CXCR4 as a target in HIV infection and in cancer metastasis. In this study, we generated two specific monoclonal antibodies (MAbs 6H7 and 7D4) against human CXCR4 and found that they could recognize different antigen epitopes identified by 12G5, a commercially available anti-CXCR4 MAb. With the two MAbs, we detected the expression pattern of CXCR4 on T lymphocytes and human tumor cell lines by FCM (flow cytometry), as well as glioma tissues by immunohistochemical staining. The results showed widespread CXCR4 expression patterns in tumor cell lines and tissues and an inducible expression in CD4(+) T lymphocytes. Furthermore, we demonstrated that both of the MAbs have different functions on glioma cell line proliferation and migration in vitro. MAb 6H7 could synergistically enhance glioma cell line U251 cell proliferation induced by SDF-1alpha, while MAb 7D4 showed a blocking effect. Despite the difference in proliferation, both antibodies could attenuate chemotaxis of U251 cell induced by SDF-1alpha. Taken together, the two novel antibodies may be of great value to explore the mechanisms of SDF-1alpha CXCR4 signal in tumor cells metastasis.

Expression of membrane CD276 (mB7-H3) has been reported on dendritic cells (DCs), monocytes, activated T cells, and various carcinoma cells. However, reports concerning its in vivo function have been inconsistent. Moreover, whether there is a soluble form of this protein is not known. In this study, using a sensitive dual monoclonal antibody sandwich enzyme-linked immunosorbent assay (ELISA) to detect the soluble form of B7-H3 (sB7-H3), we demonstrated the release of sB7-H3 by monocytes, DCs, activated T cells, and various mB7-H3+ but not mB7-H3- carcinoma cells. Release from cells was blocked by addition of a matrix metalloproteinase inhibitor (MMPI), which concomitantly caused the accumulation of B7-H3 on the cell surface. To determine the level of circulating sB7-H3, more than 200 serum samples were included in the study. The results indicated that sB7-H3 was present at high levels in all serum samples. Western blotting of sB7-H3 from cell culture supernatants or sera of healthy donors indicated that the molecular size was approximately 16 kDa. Soluble B7-H3 was able to bind to the B7-H3 receptor (B7-H3R) on activated T cells, which showed that sB7-H3 is a functionally active form. These results indicate that release of sB7-H3 from the cell surface is mediated by a matrix metalloproteinase and probably regulates B7-H3R/B7-H3 interactions in vivo. Cleavage of sB7-H3 to an active soluble form would alter both proximal and distal cellular responses.

Cancer patients mount adaptive immune responses against their tumors. However, tumor develops many mechanisms to evade effective immunosurveillance. T-cell death caused by tumor plays a critical role in establishing tumor immunotolerance. Chronic stimulation of T cells by tumors leads to activation-induced cell death. Abortive stimulation of T cells by tolerogenic antigen-presenting cells loaded with tumor antigens leads to autonomous death of tumor-specific T cells. Therapeutic approaches that prevent T-cell death in the tumor microenvironment and tumor draining lymph nodes, therefore, should boost adaptive immune responses against cancer.

In this work, we report elastocaloric effect (eCE) in Ni-Fe-Ga polycrystalline alloys. By application of a uniaxial compressive stress of 170 MPa at 298 K, a large and recoverable temperature change of 4 K has been obtained in precipitates-containing Ni54Fe19Ga27 alloys. The degradation of eCE is insensitive to 100 loading-unloading cycles. In comparison, the loss of mechanical integrity occurs in the single-phase alloy suffering from only 10 superelastic cycles, implying a short fatigue life. Good mechanical properties, large and equivalent adiabatic temperature change (ΔT) under loading-unloading cycle render dual-phase Ni-Fe-Ga promising candidate materials for solid-state mechanical cooling application at ambient conditions.

Rui Xia1, Xi Yang1, 3, Binfeng Lu1, 2 1 Department of Immunology, School of Medicine, University of Pittsburgh, EBST E1047, 200 Lothrop Street, Pittsburgh PA 15261, USA. 2 University of Pittsburgh Cancer Institute 3 School of Medicine, Tsinghua University, HaiDian, Beijing P.R. China 100084 Abstract: T cell metabolism is strongly influenced by the extracellular tissue microenvironment such as oxidizing status and amino acid availability. The underlying molecular mechanisms are not well understood. Here, we showed that increased extracellular oxidation and amino acid starvation induced a unique ATF4-regulated genetic program in T cells. ATF4 deficient T cells had a severe defect in metabolism, particularly in the oxidizing as well as amino acid-deprived environments. Lack of ATF4 resulted in T cell autonomous defects in the function of Th1 cells. In vivo, ATF4 deficiency resulted in decreased antitumor immune responses and associated with drastic changes of the tumor microenvironment. Our study establishes that ATF4 plays an important role in T cell metabolism and is important for the Th1 type immune response.

The lack of co-stimulation, as well as the dominance of co-inhibition, of type 1 lymphocytes, such as CD8+ T cells and NK cells, is a principal immunosuppressive mechanism in the tumor microenvironment. Conversion of the tumor immunosuppressive microenvironment to one that favors cancer-eradicating immune responses is therefore a goal for the development of more effective cancer immunotherapy. Targeting co-stimulatory pathways represents one attractive therapeutic strategy. T cell Ig mucin (TIM) proteins are structurally related to type I membrane glycoproteins. Previous studies have shown that TIM-1 is highly expressed on activated CD4+ T cells, facilitates the activation of Th1 cells, Th17 cells, and Th2 cells, and is implicated in the pathogenesis of human allergy and autoimmune diseases. These data support an important role for TIM-1 in regulating CD4+ T cell function. However, the role for TIM-1 in CD8+ T cell- or NK cell-mediated immune responses, including antitumor immune responses, is unknown. Here we showed that tumoral expression of antitumor cytokines such as IL-12 and IL-33 upregulated TIM-1 on tumor-infiltrating CD8+ T cells and NK cells. In addition, we demonstrated that TIM1 mAbs greatly inhibited tumor growth, resulting in greatly elevated cellular immune responses against tumor. The in-depth mechanistic insights and implication in tumor immunity are gained by our studies. Our study establishes TIM-1 as a potential target for enhancing antitumor immunity.

Introduction of an appropriate inflammation to tumor should greatly increase tumor immunogenicity and thereby help breaking immune tolerance to tumor antigens and increase response rates of immunotherapy. Cytokines play a pivotal role in regulating tumor immunogenicity and antitumor immunity. IL-36γ is important for the IL-23/IL-17-dominated autoimmune diseases as well as anti-BCG Th1 immune responses. However, the impact of IL-36γ on tumor immunity is unknown. Here, we found IL-36γ stimulated IFN-γ production by CD8 T cells synergistically with TCR signaling and/or IL-12. Importantly, IL-36γ exerted profound antitumor effects in vivo and transformed the tumor microenvironment in favor of tumor eradication. Furthermore, IL-36γ strongly increased the efficacy of tumor vaccination. Our work also showed that IL-36γ expression in human cancers inversely correlated with cancer progression. Our study establishes a novel role of IL-36γ in promoting antitumor immune responses and suggests its potential clinical translation in cancer immunotherapy.

The growth arrest and DNA damage-inducible 45beta (GADD45beta) gene product has been implicated in the stress response, cell cycle arrest, and apoptosis. Here we demonstrated the unexpected expression of GADD45beta in the embryonic growth plate and uncovered its novel role as an essential mediator of matrix metalloproteinase-13 (MMP-13) expression during terminal chondrocyte differentiation. We identified GADD45beta as a prominent early response gene induced by bone morphogenetic protein-2 (BMP-2) through a Smad1/Runx2-dependent pathway. Because this pathway is involved in skeletal development, we examined mouse embryonic growth plates, and we observed expression of Gadd45beta mRNA coincident with Runx2 protein in pre-hypertrophic chondrocytes, whereas GADD45beta protein was localized prominently in the nucleus in late stage hypertrophic chondrocytes where Mmp-13 mRNA was expressed. In Gadd45beta(-/-) mouse embryos, defective mineralization and decreased bone growth accompanied deficient Mmp-13 and Col10a1 gene expression in the hypertrophic zone. Transduction of small interfering RNA-GADD45beta in epiphyseal chondrocytes in vitro blocked terminal differentiation and the associated expression of Mmp-13 and Col10a1 mRNA in vitro. Finally, GADD45beta stimulated MMP-13 promoter activity in chondrocytes through the JNK-mediated phosphorylation of JunD, partnered with Fra2, in synergy with Runx2. These observations indicated that GADD45beta plays an essential role during chondrocyte terminal differentiation.

Fibrosis and apoptosis are juxtaposed in pulmonary disorders such as asthma and the interstitial diseases, and transforming growth factor (TGF)-beta(1) has been implicated in the pathogenesis of these responses. However, the in vivo effector functions of TGF-beta(1) in the lung and its roles in the pathogenesis of these responses are not completely understood. In addition, the relationships between apoptosis and other TGF-beta(1)-induced responses have not been defined. To address these issues, we targeted bioactive TGF-beta(1) to the murine lung using a novel externally regulatable, triple transgenic system. TGF-beta(1) produced a transient wave of epithelial apoptosis that was followed by mononuclear-rich inflammation, tissue fibrosis, myofibroblast and myocyte hyperplasia, and septal rupture with honeycombing. Studies of these mice highlighted the reversibility of this fibrotic response. They also demonstrated that a null mutation of early growth response gene (Egr)-1 or caspase inhibition blocked TGF-beta(1)-induced apoptosis. Interestingly, both interventions markedly ameliorated TGF-beta(1)-induced fibrosis and alveolar remodeling. These studies illustrate the complex effects of TGF-beta(1) in vivo and define the critical role of Egr-1 in the TGF-beta(1) phenotype. They also demonstrate that Egr-1-mediated apoptosis is a prerequisite for TGF-beta(1)-induced fibrosis and remodeling.

The stress-inducible molecules GADD45beta and GADD45gamma have been implicated in regulating IFNgamma production in CD4 T cells. However, how GADD45 proteins function has been controversial. MEKK4 is a MAP kinase kinase kinase that interacts with GADD45 in vitro. Here we generated MEKK4-deficient mice to define the function and regulation of this pathway. CD4 T cells from MEKK4-/- mice have reduced p38 activity and defective IFNgamma synthesis. Expression of GADD45beta or GADD45gamma promotes IFNgamma production in MEKK4+/+ T cells, but not in MEKK4-/- cells or in cells treated with a p38 inhibitor. Thus, MEKK4 mediates the action of GADD45beta and GADD45gamma on p38 activation and IFNgamma production. During Th1 differentiation, the GADD45beta/GADD45gamma/MEKK4 pathway appears to integrate upstream signals transduced by both T cell receptor and IL12/STAT4, leading to augmented IFNgamma production in a process independent of STAT4.

Chronic inflammatory infiltrates that take on the form of organized lymphoid organs are called tertiary lymphoid organs (TLOs). The inflammatory cytokine interleukin-17 is now found to be the critical participant driving the development of a type of TLO called inducible bronchus-associated lymphoid tissue (iBALT) in an animal model of neonatal pulmonary inflammation.1 The highly organized architecture of secondary lymphoid organs (SLOs), characterized by distinct B-cell and T-cell areas, is evolved to optimize the efficiency of cellular and humoral immune responses.2 The organization of SLOs is formed during development as a result of the highly orchestrated interplay between hematopoietic cells, stromal cells and naive lymphoid cells. TLOs arise from chronic inflammation in non-lymphoid tissues and bear morphological resemblance to SLOs. TLOs are composed of discrete T-cell and B-cell areas, follicular dendritic cells, resident dendritic cells (DCs), high endothelial venules and lymphatics. However, unlike lymph nodes, TLOs are not encapsulated, suggesting an intimate interaction with the inflamed tissues. The presence of TLOs is thought to allow antigen presentation at the local mucosal tissue with high efficiency and reduce the possibility of disseminating infection. Many molecules that drive the formation of TLOs, a process known as lymphoid neogenesis, are the same as those involved in the development of conventional lymph nodes.3 The development of SLOs is initiated by lymphoid tissue inducer (LTi) cells, which express lymphotoxin-α1β2 (LT-α1β2).4 LT-α1β2 stimulates the expression of CXCL13 and CCL21 in stromal cells, which are required for the homing of B and T cells to the SLOs.5 Transgenic expressed LT-α, under the rat insulin promoter, induced chronic inflammatory infiltrates characterized by the presence of T cells, B cells and high endothelial venules, and an increased expression of adhesion molecules mediating the homing of naive lymphocytes to SLOs.3 Subsequent studies also demonstrated that overexpression of lymphoid homeostatic chemokines such as CCL21 or CXCL13 promoted SLO generation.6, 7 Despite these studies, the initiation of lymphoid neogenesis is poorly understood. Randall and colleagues1 recently identified IL-17 produced by T cells as being required for the formation of iBALT, a type of TLO that forms in the lungs during chronic inflammation and infection. This study provides exciting novel mechanistic insights into the initiating events and the cell components that participate in iBALT formation. Randall and colleagues induced chronic pulmonary inflammation and iBALT by repeatedly administering lipopolysaccharide (LPS) to the lungs of neonatal C57BL/6 mice. Because LTi cells are involved in the development of conventional lymph nodes, the authors studied whether LTi cells are also involved in the generation of iBALT, by using RORγ-deficient mice and Id2-deficient mice, both of which fail to develop LTi cells. Interestingly, both RORγ-deficient mice and Id2-deficient mice developed normal iBALT areas, suggesting that LTi cells are not involved in the generation of iBALT. The authors also confirmed previous findings that DCs are critical for the maintenance of iBALT.8, 9 Using various knockout mice, they further showed that LT-α and homeostatic chemokines, such as CXCL13, CCL19 and CCL21a, were required for the organization of iBALT. In searching for candidate genes for the induction of iBALT, the authors discovered that, after repeated challenge of the lungs with LPS, more IL-23p19 and IL-17A were expressed in neonatal lungs than in adult lungs. They further found that IL-17 influenced the number and size but not the architecture of the iBALT, and they uncovered the source of IL-17 as being CD4+ αβ- and γδ-T cells. Using an adoptive transfer system, they established that either αβ- or γδ-T cells were sufficient to establish B-cell aggregates in their model of lung inflammation. Of all of the αβ-T cell subsets, which include Th1, Th2, follicular helper T cells and Th17 cells, Th17 cells were found to be the most potent inducers of iBALT. In addition, they found that IL-17 induced the expression of CXCL13 and CCL19 but not CCL21 in isolated pulmonary fibroblasts and that IL-17 was required for the production of these homeostatic cytokines in the LPS-challenged lung tissues. This study reveals a novel role of IL-17 in iBALT formation. The proposed function of IL-17 is to directly stimulate the expression of CXCL13 and CCL19 in stromal cells. The exact sequence of events that initiates iBALT formation, however, remains to be carefully examined. Both this study and previously published studies have shown that DCs are required for iBALT formation. Therefore, the initiating events might involve DCs being activated by LPS within the lungs and then migrating to the lung-draining lymph nodes, where they initiate the generation of Th17 cells. This local activation of myeloid cells would be followed by the activation of γδ-T cells, which are expected to be the first cells that produce IL17 in LPS-treated lung tissue. The second cells to produce IL-17 would be pre-activated Th17 cells, which have been generated in the lung-draining lymph nodes and subsequently recruited to the lungs. The findings of this study are consistent with the role of IL-17 in the initiation of chronic inflammation.10 In addition to homeostatic chemokines, such as CXCL13 and CCL19, IL-17 induces a wide range of inflammatory chemokines, such as CXCL9, CXCL10 and CXCL11. Blockade of IL-17 reduces the number and size of iBALTs but does not affect the architecture of iBALTs. Thus, IL-17 is more important for inducing tissue inflammation than for organizing the structure of iBALTs. Randall and colleagues' data also show that IL-17 is not required for the maintenance of iBALT, despite its crucial function in iBALT formation. From a therapeutic point of view, this implies that IL-17 blockade might not affect established TLOs but might inhibit the generation of new TLOs, which is responsible for the relapse of autoimmune diseases such as multiple sclerosis. In the future, it will be interesting to determine whether other T-cell subsets are involved in TLO maintenance and organization.

By direct measurements, the elastocaloric effect at various temperatures in a [001]-oriented Ni45.7Mn36.6In13.3Co5.1 metamagnetic shape memory polycrystal has been investigated. A reversible temperature change of ±3.5 K, due to the relatively low stress (100 MPa) induced martensitic transformation, was observed at room temperature. A theoretical analysis based on the Debye description has revealed that the adiabatic temperature change arising from the lattice vibration plays a dominant role in the large elastocaloric effect for Ni-Mn-In-Co alloys.

In this paper, ( Cr , Fe )7 C 3( M 7 C 3)/γ- Fe composite layer has been in situ fabricated on a low carbon steel surface by vacuum electron beam irradiation (VEB). Three kinds of powder mixtures were placed on a low carbon steel substrate, which was then irradiated with electron beam in vacuum condition. The microstructure and wear resistance of the composite layers has been studied by means of optical microscope (OM), X-ray diffraction (XRD), scanning electron microscope (SEM), microhardness tester and tribological tester. The chemical composition of all specimens were carefully analyzed using energy-dispersive X-ray spectroscopy (EDAX) technique. Depending on three different powder mixtures, hypereutectic and hypoeutectic microstructures were obtained on surface composite layers. No pores and cracks were found on the coatings. The amount of carbides formed in the surface composite layer was mainly determined by carbon concentration. The microstructure close to the fusion line was largely primary austenite dendrite. The hardness and wear resistance of the surface composite layer has been greatly improved due to the extensive distribution of carbides.

Reactive oxygen species (ROS) play a major role in the pathogenesis of chronic inflammatory and autoimmune diseases. Little is known about underlying molecular mechanisms regulating T cell immune responses to ROS. Here, we demonstrated that ROS induced ATF4 in activated T cells. We found, in the high ROS condition, ATF4-/- T cells failed to proliferate and produced much less IFN-γ when cultured in the Th1 polarization condition. Inhibition of ROS by addition of antioxidants significantly recovered proliferation and IFN-γ production by ATF4-/- CD4 T cells in the Th1 condition, suggesting a crucial role of ATF4 in preventing oxidative stress during T cell activation and Th1 differentiation. In addition, we demonstrated that ATF4-/- mice develop exacerbated experimental autoimmune encephalomyelitis (EAE), which is associated with elevated levels of IL17 but reduced IFN-gamma production. The in-depth molecular mechanisms are further studied and the result will be presented. Our study establishes that the ROS/ATF4 axis promotes Th1 and suppresses Th17 immune responses and likely plays an important role in autoimmunity and cancer immunology.

Introduction: The impact of anesthetic exposure on the immune system is relatively unknown at both mechanistic and clinical levels. In this in vitro study, we employ immunological assays and understanding of immune cell function to evaluate the effect of volatile anesthetic (VA) exposure on bone marrow dendritic cells (BMDCs). BMDCc play a critical role in antigen presentation and link the innate and adaptive arms of the immune response. Hypothesis: We believe that VA exposure may modulate immune responses, specifically BMDC function. Methods: Bone marrow cells were harvested from C57BL/6 mice and cultured for six days to generate BMDCs. They were stimulated with low and high dose lipopolysaccharide (LPS). Simultaneously, the cells were exposed to 1.5% isoflurane and incubated for 5 hours. Upon completion of VA exposure, the BMDCs were lysed with Trizol, RNA extracted, and reverse transcript PCR used to generate cDNA. RT-qPCR was performed to evaluate gene expression of common BMDC cytokines. Results: BMDC exposure to isoflurane resulted in decreased gene expression of IL-10, IL-6, and IL-1β. RT-qPCR data indicated 2-4 fold decreases in gene expression of these cytokines. Conclusions: BMDCs exposed to VA in vitro displayed decreased gene expression of inflammatory cytokines. Significance: Better understanding of immunomodulation by anesthetics is critical for optimizing patient outcomes and improved perioperative management.

One major approach of cancer immune therapy is to convert the tumor immunosuppressive microenvironment to one that favors antitumor immune responses. We have recently demonstrated that Interleukin 33 (IL-33) promotes effector functions of CD8 T cells, suggesting a potential function in antitumor immunity. Here we showed that overexpression of IL-33 in two tumor cell lines, 4T1 and B16, potently inhibited tumor growth in vivo. CD45+, CD8+ T cells, NK cells, IFN-γ+ CD8+ T cells and IFN-γ+ NK cells were greatly increased in the IL-33-expressing tumors when compared with those in the control tumors. We further demonstrated that the antitumor effect of IL-33 was dependent on NK cells and CD8 T cells. In contrast, MDSC were greatly decreased in the IL-33-expressing tumors when compared to those in the control tumors. Our results indicate that the intratumoral delivery of IL-33 can be a new strategy of tumor immunotherapy by promoting an immunogenic microenvironment.

CD4+CD25+Foxp3+ regulatory T cells (Tregs) can inhibit cytotoxic responses. Though several studies have analyzed Treg frequency in the peripheral blood mononuclear cells (PBMCs) of pancreatic ductal adenocarcinoma (PDA) patients using flow cytometry (FCM), few studies have examined how intratumoral Tregs might contribute to immunosuppression in the tumor microenvironment. Thus, the potential role of intratumoral Tregs in PDA patients remains to be elucidated. In this study, we found that the percentages of Tregs, CD4+ T cells and CD8+ T cells were all increased significantly in tumor tissue compared to control pancreatic tissue, as assessed via FCM, whereas the percentages of these cell types in PBMCs did not differ between PDA patients and healthy volunteers. The percentages of CD8+ T cells in tumors were significantly lower than in PDA patient PBMCs. In addition, the relative numbers of CD4+CD25+Foxp3+ Tregs and CD8+ T cells were negatively correlated in the tissue of PDA patients, and the abundance of Tregs was significantly correlated with tumor differentiation. Additionally, Foxp3+ T cells were observed more frequently in juxtatumoral stroma (immediately adjacent to the tumor epithelial cells). Patients showing an increased prevalence of Foxp3+ T cells had a poorer prognosis, which was an independent factor for patient survival. These results suggest that Tregs may promote PDA progression by inhibiting the antitumor immunity of CD8+ T cells at local intratumoral sites. Moreover, a high proportion of Tregs in tumor tissues may reflect suppressed antitumor immunity.

In this paper, (Cr, Fe)7C3(M7C3)/γ-Fe composite layer has been in situ fabricated on a low carbon steel surface by vacuum electron beam irradiation (VEB). Three kinds of powder mixtures were placed on a low carbon steel substrate, which was then irradiated with electron beam in vacuum condition. The microstructure and wear resistance of the composite layers has been studied by means of optical microscope (OM), X-ray diffraction (XRD), scanning electron microscope (SEM), microhardness tester and tribological tester. The chemical composition of all specimens were carefully analyzed using energy-dispersive X-ray spectroscopy (EDAX) technique. Depending on three different powder mixtures, hypereutectic and hypoeutectic microstructures were obtained on surface composite layers. No pores and cracks were found on the coatings. The amount of carbides formed in the surface composite layer was mainly determined by carbon concentration. The microstructure close to the fusion line was largely primary austenite dendrite. The hardness and wear resistance of the surface composite layer has been greatly improved due to the extensive distribution of carbides.

Stra13, a basic helix-loop-helix transcription factor, is up-regulated upon activation of CD4+ T cells. Here we show that Stra13-deficient mice exhibit defects in several phases of CD4+ T cell activation. In vivo, Stra13 deficiency results in ineffective elimination of activated T and B cells, which accumulate progressively, leading to lymphoid organ hyperplasia. Consequently, aging Stra13-/- mice develop autoimmune disease characterized by accumulation of spontaneously activated T and B cells, circulating autoantibodies, infiltration of T and B lymphocytes in several organs and immune complex deposition in glomeruli. Our studies identify Stra13 as a key regulator of lymphocyte activation that is vital for maintenance of self-tolerance and for constraint of autoimmunity.

There have been two previous conflicting reports that the development of T-cell-mediated autoimmune diabetes (type 1 diabetes) was respectively unaffected or inhibited in NOD mice genetically deficient in the T-helper (Th) 1 cytokine interferon (IFN)-gamma or the alpha-chain subunit of its receptor. Our goal was to resolve this conundrum by congenically transferring, from a 129 donor strain to the NOD background, a functionally inactivated gene for the beta-chain signaling (located on chromosome 16) rather than the alpha-chain ligand binding domain (located on chromosome 10) of the IFN-gamma receptor. These NOD.IFNgammaRBnull mice were characterized by normal patterns of leukocyte development and T-cells that produced greatly enhanced levels of the putatively type 1 diabetes-protective Th2 cytokine interleukin (IL)-4. However, despite being unable to respond to the primary Thl cytokine IFN-gamma and having T-cells that produce greatly enhanced levels of IL-4, NOD.IFNgammaRBnull mice remained highly susceptible to type 1 diabetes. This result indicated that the previously reported inhibition of type 1 diabetes in NOD mice carrying a functionally inactivated IFN-gamma receptor alpha-chain gene may have been due to a closely linked and previously unidentified diabetes resistance allele. Furthermore, our results indicate that the pathogenicity of diabetogenic T-cells in NOD mice is not dampened by an inability to respond to IFN-gamma and a concurrent shift to greatly enhanced Th2 cytokine production. This finding calls into question whether clinical protocols designed to shift beta-cell autoreactive T-cells from a Thl to Th2 cytokine production profile will truly be safe and efficacious in blocking the development of type 1 diabetes in humans.

Using human cyclin D1 as the “bait” in a yeast two-hybrid system, together with a HL60 cDNA library, we identified a novel human nuclear protein designated DIP1. This protein is expressed in a variety of cell types, and in fibroblasts its level remains constant throughout the cell cycle. However, the level of this protein increases severalfold during the differentiation of HL60 cells. The DIP1 protein can be phosphorylated in vitro by a cellular kinase and this activity reaches its maximum in extracts obtained from cells in the G1 phase of the cell cycle. DIP1 contains a helix–loop–helix motif but lacks an adjacent basic DNA-binding domain, thus resembling the Id family of proteins. The dip1 gene is located on human chromosome 16p11.2-12, a locus that is amplified in several types of human cancer. These results suggest that DIP1 may be involved in the control of gene expression and differentiation, but its precise function remains to be determined.

This review features two areas of our research interests. First, our laboratory is interested in elucidating the molecular mechanisms that control generation, effector function, and fate of TH1 cells during inflammatory responses. We have been particularly interested in the role of the growth arrest and DNA damage-inducible protein 45 (GADD45) gene family in TH1-mediated immune responses. We and others have shown that, in TH1 cells, Gadd45b and Gadd45g are induced by TCR signaling or IL-12 and IL-18. Gadd45b and Gadd45g are very important for the activation of p38 MAP kinases in TH1 cells and for effector functions of TH1 cells. We have found that deletion of Gadd45b and Gadd45g genes in mice results in a drastically reduced number of TH1 cells against Listeria monocytogenes. The critical role of Gadd45b and Gadd45g in TH1 responses in this acute infectious model has led us to hypothesize that they promote TH1-mediated autoimmune diseases. We therefore decided to test this hypothesis in experimental allergic encephalomyelitis (EAE), a mouse model of multiple sclerosis (MS). Expecting low numbers of TH1 cells, we were surprised to see that Gadd45b deletion resulted in exacerbated chronic phase of EAE. In contrast to the reduction of TH1 cells we saw in acute L. monocytogenes infection, during the chronic phase of EAE we observed increased TH1 cells in the inflamed CNS when Gadd45b was deleted. Gadd45b deletion also resulted in enlarged spleens in older mice. Gadd45b/Gadd45g greatly enlarged spleens in older mice compared with Gadd45b single deletion. The enlargement of spleens was due to the accumulation of CD4+ T cells with an activated phenotype and B cells. In addition, we have found that Gadd45b and Gadd45g inhibit proliferation and promote apoptosis of activated CD4+ T cells. Therefore, Gadd45b and Gadd45g play a critical role as a molecular "doubleedged sword" in TH1-type immune responses to ensure a prompt, robust but self-limiting TH1 response. The second area of research in our laboratory focuses on the role of autophagy in T cells. We have recently discovered that autophagy is induced in TH1 and TH2 cells. There are more effector TH2 cells than TH1 cells that undergo autophagy. We have used RNAi strategy to "knockdown" autophagy in TH2 cell lines and found that autophagy is required for growth factor-withdrawal cell death.

The population size of the T cells is tightly regulated. The T cell number drastically increases in response to their specific antigens. Upon antigen clearance, the T cell number decreases over time. Apoptosis, also called type I programmed cell death, plays an important role in eliminating T cells. The role of autophagic cell death, also called type II programmed cell death, is unclear in T cells. Our recent work demonstrated that autophagy is induced in both Th1 and Th2 cells. Both TCR signaling and IL-2 increase autophagy in T cells, and JNK MAP kinases are required for the induction of autophagy in T cells, whereas caspases and mTOR inhibit autophagy in T cells. Autophagy is required for mediating growth factor withdrawal-dependent cell death in T cells. Here, we hypothesize that autophagic cell death plays an important role in T cell homeostasis.

To study the role of the interferon- (IFN) γR2 chain in IFN-γ signaling and immune function, IFN-γR2-deficient mice have been generated and characterized. Cells derived from IFN-γR2 −/− mice are unable to activate either JAK/STAT signaling proteins or gene transcription in response to IFN-γ. The lack of IFN-γ responsiveness alters IFN-γ-induced Ig class switching by B cells from these mice.In vitrocultures of T cells demonstrate that the T cells from the IFN-γR2 −/− mice have a defect in Th1 cell differentiation. The IFN-γR2 (−/−) mice also produce lower amounts of IFN-γ in response to antigenic challenge. In addition, IFN-γR2 −/− mice are defective in contact hypersensitivity and are highly susceptible to infection byListeria monocytogenes. These results demonstrate that the IFN-γR2 is essential for IFN-γ-mediated immune responsesin vivo.

Reactive oxygen species (ROS) play a major role in the pathogenesis of chronic inflammatory and autoimmune diseases. It is well established that ROS modulate T cell-mediated immune responses. However, little is known about underlying molecular mechanisms of how ROS regulate T cell immune responses and how T cells adapt to high levels of ROS. We demonstrated that ROS induced ATF4 in activated T cells. ATF4 is a basic leucine-zipper (bZip) transcription factor, which regulates cellular redox state and amino acid metabolism. We found that, in the high ROS condition, ATF4-/- Th1 cells failed to produce IFN-γ whereas WT Th1 cells still made large amounts of IFN-γ. Inhibition of ROS by addition of antioxidants recovered the IFN-γ production by ATF4-/- Th1 cells, suggesting ATF4 is critically required for the function of Th1 cells in the presence of high levels of ROS. ATF4-/- antigen presenting cells (APC) promoted IL-17 production, suggesting ATF4 plays an important role in APCs in suppressing Th17-driving cytokines. In addition, we demonstrated that experimental allergic encephalomyelitis (EAE) was exacerbated in ATF4 deficient mice compared with wild type (WT) mice. The number of autoreactive Th1 cells was decreased whereas that of autoreactive Th17 cells was elevated in ATF4-/- mice compared with WT mice. Our study establishes that ATF4 promotes Th1 and suppresses Th17 immune responses and likely plays an important role in autoimmunity and cancer immunology.

Growth arrest and DNA-damage-inducible, beta (GADD45β) has been reported to inhibit apoptosis via attenuating c-Jun N-terminal kinase (JNK) activation. We demonstrated here that GADD45β mediated its anti-apoptotic effect via promoting p53 protein degradation following arsenite treatment. We found that p53 protein expression was upregulated in GADD45β−/− cells upon arsenite exposure as compared with those in GADD45β+/+ cells. Further studies showed that GADD45β attenuated p53 protein expression through Src/protein phosphatase 2A/murine double minute 2-dependent p53 protein-degradation pathway. Moreover, we identified that GADD45β-mediated p53 protein degradation was crucial for its anti-apoptotic effect due to arsenite exposure, whereas increased JNK activation was not involved in the increased cell apoptotic response in GADD45β−/− cells under same experimental conditions. Collectively, our results demonstrate a novel molecular mechanism responsible for GADD45β protection of arsenite-exposed cells from cell death, which provides insight into our understanding of GADD45β function and a unique compound arsenite as both a cancer therapeutic reagent and an environmental carcinogen. Those novel findings may also enable us to design more effective strategies for utilization of arsenite for the treatment of cancers.

Aim: CD11c is a marker for dendritic cells which help initiating adaptive antitumor immunity. The study was designed to investigate correlation between the frequency of CD11c+ cells in the gastric cancer tissue and the clinical and pathological parameters of gastric cancer patients. Methods: CD11c expression in gastric cancer tissues was examined by immunohistochemistry. Surface CD11c expression on the leukocytes within cancer tissues was examined by flow cytometry. Results: The number of CD11c+ cells was higher in gastric cancer tissues than that in gastritis or gastric polyps tissues. The number of CD11c+ cells in tumor specimens from the early-stage gastric cancer was significantly higher than that in specimens from the late-stage gastric cancer (t=2.77,P=0.0064). Flow cytometry found that CD11c surface expression was stronger in specimens from the early stages than those from the late stages of gastric cancer. The Univariate analysis revealed that the frequency of intratumoral CD11c+ cells showed association with gender and tumor location, and was reversely correlated with tumor size, depth of infiltration, nodal metastasis, tumor histological grade and cancer stages. The median survival time (MST) for the subgroups of patients with stronger and weaker CD11c expression were 67.0 months (95%CI, 39.0-112.0 months) and 29.0 months (95%CI, 21.0-39.0 months), respectively. And MSTs were significantly different between the CD11c higher and lower subgroups (χ2=11.6425,P=0.0006). Among patients who suffered from late-stage diseases, those with higher numbers of intratumoral CD11c cells had a significantly lower risk of death than those with lower numbers of CD11c cells (χ2=8.549,P=0.0035), and the lowered risk of death was statistically significant after adjusting gender, age, tumor size and infiltrating depth (RR=0.52,95%CI=0.29-0.94). Conclusions: The number of CD11c+ cells within gastric cancer tissue is a favorable prognostic marker for gastric cancer. Our study suggests gradual elimination of dendritic cells is characteristic of gastric cancer progression. Citation Format: Jingting Jiang, Changping Wu, Binfeng Lu. The frequency of CD11c+ cells within the gastric cancer tissue is associated with favorable prognosis. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1258. doi:10.1158/1538-7445.AM2013-1258 Note: This abstract was not presented at the AACR Annual Meeting 2013 because the presenter was unable to attend.

Interleukin-33 is a member of the IL-1 family of cytokines and is released by stressed cells, necrotic cells or activated innate immune cells such as macrophages during trauma or infection. Thus, IL-33 is thought to serve as an endogenous “danger signal” to trigger inflammation and promote cell mediated immune responses. Prior studies have supported the role of IL-33 in promoting type 2 immune responses but little was known for its role in type 1 immune responses such as the anti-tumor immune response. We have found that that ST2, a receptor for IL-33, is highly expressed in activated CD8 T cells. We have further revealed that, T-bet, a critical transcriptional factor for Th1 differentiation and effector functions, is required for ST2 expression by CD8 T cells, suggesting that ST2 is an integral part of the Th1 gene network in CD8 T cells. Furthermore, we have demonstrated that IL-33 synergized with TCR or IL-12 in increasing both human and mouse CD8 T cell functions such as IFN-γ production. Thus IL-33 can greatly enhance type 1 cell-mediated immune responses in both human and mice. The in vivo relevance of this pathway is examined in a preclinical model of tumor immunity. Our results support a potent function of IL-33 in inhibiting tumor growth and provide a novel therapeutic approach for tumor immunity. Citation Format: Xin Gao, Gang Li, Bo Teng, Qianting Yang, Yibei Zhu, Xueguang Zhang, Olivera J. Finn, Binfeng Lu. IL-33 promotes CD8 T cell effector function and has a potent antitumor activity in vivo. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology: Multidisciplinary Science Driving Basic and Clinical Advances; Dec 2-5, 2012; Miami, FL. Philadelphia (PA): AACR; Cancer Res 2013;73(1 Suppl):Abstract nr A66.

Interleukin-33 is a member of the IL-1 family of cytokines and is released by stressed cells, necrotic cells or activated innate immune cells such as macrophages during trauma or infection. Thus, IL-33 is thought to serve as an endogenous “danger signal” to trigger inflammation and promote cell-mediated immune responses. Prior studies have supported the role of IL-33 in promoting type 2 immune responses but little was known for its role in type 1 immune responses such as the anti-tumor immune response. We have found that that ST2, a receptor for IL-33, is highly expressed in CD8 T cells cultured in Th1 conditions. We have further revealed that, T-bet, a critical transcriptional factor for Th1 differentiation and effector functions, is required for ST2 expression by CD8 T cells, suggesting that ST2 is an integral part of the Th1 gene network in CD8 T cells. Furthermore, we have demonstrated that IL-33 synergized with TCR or IL-12 in increasing both human and mouse CD8 T cell functions such as IFN-γ production. Thus IL-33 can greatly enhance type 1 cell-mediated immune responses in both human and mice. The in vivo relevance of this pathway is examined in a preclinical model of tumor immunity. Our results support a potent function of IL-33 in inhibiting tumor growth and provide a novel therapeutic approach for tumor immunity.

Beclin 1, the mammalian homologue of the yeast Atg6, regulates macroautophagy (autophagy) in eukaryotic cells and plays a critical role in the regulation of cell death and survival in various types of cells. Beclin 1 is expressed in T cells. However, its role in T cell-mediated immune responses remains unknown. We have found that mice with Beclin-1 deletion in T cells have profound deficiencies in mounting T dependent immune responses. Autophagy is abolished in Beclin 1 deficient T cells. We observed that, upon activation, Beclin 1 deficient T cells undergo programmed cell death. Mice with Beclin 1 deletion in T cells fail to mount T cell-dependent humoral immune responses to KLH and are resistant to experimental autoimmune encephalomyelitis. Our work establishes a crucial role of Beclin 1 in autoimmunity.

Cell-mediated adaptive immunity is very important in tumor immune surveillance and tumor vaccination. However, the genetic program underlying an effective adaptive anti-tumor immunity is elusive. T-bet and Eomes have been suggested to be master regulators of Th1 cells and CD8+ T cells. Whether they are important for anti-tumor immunity is controversial. Here we show that lack of T-bet and Eomes has resulted in profound defects in adaptive anti-tumor immune responses in both prophylactic and therapeutic settings. T-bet and Eomes are also important for CD8+ T cell activation without affecting T cell clonal expansion, the gaining of effector phenotype and cytolytic function. Unexpectedly, T-bet and Eomes are dispensable for IFNg production by CD8+ T cells under optimal Th1 differentiation conditions. However, they are necessary for suppressing alternative fates of CD8+ T cells. They drive Tc1 differentiation by preventing alternative CD8+ T cell differentiation to Tc17 or Tc2 cells. Surprisingly, T-bet and Eomes are not critical for the generation of systemic CTL activities against tumor cells. Instead, T-bet and Eomes are crucial for T cell migration and retention in the tumor site and elimination of solid tumors by CD8+ T cells through regulating chemokine receptors such as CXCR3 and IFNg expression in tumor microenvironment. Therefore, our study establishes T-bet and Eomes as key regulators of adaptive cell-mediated immunity against cancer.

AIM: To analyze the correlation between cytokine-induced killer (CIK) cells adoptive immunotherapy and cancer-related death in gastric cancer patients. METHODS: One hundred and fifty-six gastric cancer patients after operation at the Third Affiliated Hospital of Soochow University were enrolled in this study. Their clinical data including demographic characteristics, operation time, tumor size, pathological type and staging, tumor metastasis, outcome of chemotherapy or CIK cells adoptive immunotherapy, survival time or time of death were collected with a standard structured questionnaire. Kaplan-Meier method was used to estimate the median survival time, and the 2- and 5- year survival rates. Hazard risk (HR) and 95% confidence interval (95% CI) of CIK cells adoptive immunotherapy for gastric cancer were calculated using the two-stage time-dependent covariates Cox model. RESULTS: The survival time of gastric cancer patients was longer after CIK cells adoptive immunotherapy than after chemotherapy (χ2 = 10.907, P = 0.001). The median survival time of gastric cancer patients was also longer after CIK cells adoptive immunotherapy than after chemotherapy (49 mo vs 27 mo, P < 0.05). The 2- and 5-year survival rates of gastric cancer patients were significantly higher after CIK cells adoptive immunotherapy than after chemotherapy (73.5% vs 52.6%, 40.4% vs 23.9%, P < 0.05). A significant difference was observed in the survival curve for patients who received CIK cells adoptive immunotherapy (0, 1-10, 11-25, and over 25 frequencies) (χ2 = 14.534, P = 0.002). The frequencies of CIK cells adoptive immunotherapy were significantly related with the decreasing risk of death in gastric cancer patients after adjustment for sex and age of the patients, tumor stage and relapse (HR = 0.54, 95% CI: 0.36-0.80) when the first stage Cox model was used to define the subjects who remained alive beyond 36 mo as survivors. However, no correlation was observed between the frequencies of death in CIK cells adoptive immunotherapy and the risk of gastric cancer patients (HR = 1.09, 95% CI: 0.63-0.89) when the second stage Cox model was used to define the subjects who survived for more than 36 mo as survivors. CONCLUSION: The survival time of the gastric cancer patients treated with chemotherapy combined with CIK cells adoptive immunotherapy is significantly longer than that of the patients treated with chemotherapy alone and increasing the frequency of CIK cells adoptive immunotherapy seems to benefit patients more.

Abstract 917 Post-transcriptional regulation such as RNA editing in hematopoiesis and lymphopoiesis is poorly understood. ADAR1 (adenosine deaminase acting on RNA-1) is a RNA editing enzyme essential for embryonic development. Disruption of the ADAR1 gene was shown to cause defective embryonic hematopoiesis (Wang Q et al, Science 2000). Moreover, we have recently obtained direct evidence for the preferential effect of ADAR1 deletion on adult hematopoietic progenitor cells as opposed to the more primitive cells via a RNA-editing dependent mechanism by different conditional gene deletion strategies (Xufeng R et al PNAS 2009, in press). To further determine the role of ADAR1 in T cell development, we generated a mouse model in which ADAR1 was deleted specifically in T lymphocytes by interbreeding ADAR1lox/lox mice with Lck-Cre transgenic mice. In our current study, we report that ADAR1 is essential for T cell differentiation at the late progenitor stage in the thymus, coincident with T cell receptor-α/β expression. In ADAR1lox/loxLck-Cre mice, mature T cells decreased dramatically in peripheral blood, spleen and lymph nodes in comparison to littermate controls. In the thymus, the production of CD4+/CD8+ double positive cells was severely impaired and massive cell death was observed in pre-T cell populations. Within the pro-T cells, ADAR1 deletion resulted in a significant decrease of late progenitor cells but not early progenitor subsets. In both pro-T and pre-T cell stages, defective T cell development preferentially occurred in the beta chain positive cells, but was not apparent in gamma/delta T cells. Our data demonstrated an indispensable role of ADAR1 in early T cell differentiation that correlated with T cell receptor beta chain expression, thereby indicating that RNA editing by ADAR1 is an essential event in T cell development. Disclosures: No relevant conflicts of interest to declare.