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Background: Ulcerative colitis (UC) is a type of inflammatory bowel disease (IBD) that is associated with immune dysfunction. Recent studies have indicated that the neurosecretory hormone oxytocin (OXT) has been proven to alleviate experimental colitis. Methods: We investigated the role of OXT/OXT receptor (OXTR) signalling in dendritic cells (DCs) using mice with specific OXTR deletion in CD11c+ cells (OXTRflox/flox×CD11c-cremice) and a dextran sulfate sodium (DSS)-induced colitis model. Results: The level of OXT was abnormal in the serum or colon tissue of DSS-induced colitis mice or the plasma of UC patients. Both bone marrow-derived DCs (BMDCs) and lamina propria DCs (LPDCs) express OXTR. Knocking out OXTR in DCs exacerbated DSS-induced acute and chronic colitis in mice. In contrast, the injection of OXT-pretreated DCs significantly ameliorated colitis. Mechanistically, OXT prevented DC maturation through the phosphatidylinositol 4,5-bisphosphate 3-kinase (Pi3K)/AKT pathway and promoted phagocytosis, adhesion and cytokine modulation in DCs. Furthermore, OXT pre-treated DCs prevent CD4+ T cells differentiation to T helper 1 (Th1) and Th17. Conclusions: Our results suggest that OXT-induced tolerogenic DCs efficiently protect against experimental colitis via Pi3K/AKT pathway. Our work provides evidence that the nervous system participates in the immune regulation of colitis by modulating DCs. Our findings suggest that generating ex vivo DCs pretreated with OXT opens new therapeutic perspectives for the treatment of UC in humans. [ABSTRACT FROM AUTHOR]

Background: MicroRNAs have been shown to be important regulators of the immune response and the development of the immune system. It was reported that microRNA-125b (miR-125b) was down-regulated in macrophages challenged with endotoxin. However, little is known about the function and mechanism of action of miR-125b in macrophage activation. Macrophages use L-arginine to synthesize nitric oxide (NO) through inducible NO synthase (iNOS), and the released NO contributes to the tumoricidal activity of macrophages.Methods: Luciferase reporter assays were employed to validate regulation of a putative target of miR-125b. The effect of miR-125b on endogenous levels of this target were subsequently confirmed via Western blot. Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) was performed to determine the expression level of miR-125b in macrophage. MTS assays were conducted to explore the impact of miR-125b overexpression on the cell viability of 4T1 cells.Results: Here, we demonstrate that mmu-miR-125b overexpression suppresses NO production in activated macrophages and that LPS-activated macrophages with overexpressed mmu-miR-125b promote 4T1 tumor cell proliferation in vitro and 4T1 tumor growth in vivo. CCNA2 and eEF2K are the direct and functional targets of mmu-miR-125b in macrophages; CCNA2 and eEF2K expression was knocked down, which mimicked the mmu-miR-125b overexpression phenotype.Conclusions: These data suggest that mmu-miR-125b decreases NO production in activated macrophages at least partially by suppressing eEF2K and CCNA2 expression. [ABSTRACT FROM AUTHOR]

Background: Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) could induce apoptosis of HIV-1-infected monocyte-derived macrophage (MDM), but the molecular mechanisms are not well understood. Methodology/Principal Findings: By using an HIV-1 Env-pseudotyped virus (HIV-1 PV)-infected MDM cell model we demonstrate that HIV-1 PV infection down-regulates the expression of TRAIL decoy receptor 1 (DcR1) and 2 (DcR2), and cellular FLICE-inhibitory protein (c-FLIP), but dose not affect the expression of death receptor 4 and 5 (DR4, DR5), and Bcl-2 family members in MDM cells. Furthermore, recombinant soluble TRAIL and an agonistic anti-DR5 antibody, AD5-10, treatment stimulates reactive oxygen species (ROS) generation and JNK phosphorylation. Conclusions/Significance: HIV infection facilitates TRIAL-induced cell death in MDM by down-regulating the expression of TRAIL decoy receptors and intracellular c-FLIP. Meanwhile, the agonistic anti-DR5 antibody, AD5-10, induces apoptosis synergistically with TRAIL in HIV-1-infected cells. ROS generation and JNK phosphorylation are involved in this process. These findings potentiate clinical usage of the combination of TRAIL and AD5-10 in eradication of HIV-infected macrophage and AIDS. [ABSTRACT FROM AUTHOR]

Background: Adeno-associated virus-2 (AAV-2)-mediated gene therapy is quite suitable for local or regional application in head and neck cancer squamous cell carcinoma (HNSCC). However, its low transduction efficiency has limited its further development as a therapeutic agent. DNA damaging agents have been shown to enhance AAV-mediated transgene expression. Cisplatin, one of the most effective chemotherapeutic agents, has been recognized to cause cancer cell death by apoptosis with a severe toxicity. This study aims to evaluate the role of cisplatin in AAV-mediated tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) expression and the effect on HNSCC both in vitro and in vivo. Methods: Five human HNSCC cell lines were treated with recombinant soluble TRAIL (rsTRAIL) and infected with AAV/TRAIL to estimate the sensitivity of the cancer cells to TRAIL-induced cytotoxicity. KB cells were infected with AAV/EGFP with or without cisplatin pretreatment to evaluate the effect of cisplatin on AAV-mediated gene expression. TRAIL expression was detected by ELISA and Western blot. Cytotoxicity was measured by MTT assay and Western blot analysis for caspase-3 and -8 activations. Following the in vitro experiments, TRAIL expression and its tumoricidal activity were analyzed in nude mice with subcutaneous xenografts of HNSCC. Results: HNSCC cell lines showed different sensitivities to rsTRAIL, and KB cells possessed both highest transduction efficacy of AAV and sensitivity to TRAIL among five cell lines. Preincubation of KB cells with subtherapeutic dosage of cisplatin significantly augmented AAV-mediated transgene expression in a heparin sulfate proteoglycan (HSPG)-dependent manner. Furthermore, cisplatin enhanced the killing efficacy of AAV/TRAIL by 3- fold on KB cell line. The AAV mediated TRAIL expression was observed in the xenografted tumors and significantly enhanced by cisplatin. AAV/TRAIL suppressed the tumors growth and cisplatin augmented the tumoricidal activity by two-fold. Furthermore, Combination treatment reduced cisplatin-caused body weight loss in nude mice. Conclusion: The combination of AAV-mediated TRAIL gene expression and cisplatin had synergistic therapeutic effects on head and neck cancers and reduced the potential toxicity of cisplatin. These findings suggest that the combination of AAV/TRAIL and cisplatin may be a promising strategy for HNSCC therapy. [ABSTRACT FROM AUTHOR]

Abstract   Neuromyelitis optica (NMO) is an inflammatory demyelinating disease with generally poor prognosis that selectively targets optic nerves and spinal cord. Although diagnostic criteria for NMO are available, there is still a need for biomarkers, predicting disease development and progression to improve individually tailored treatment. CSF proteins were separated by two-dimensional electrophoresis and identified by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). The interaction between these proteins was further analyzed by Pathway Studio software. Seven protein spots in CSF were significantly altered in NMO patients compared with controls. Identification made by mass spectrometry revealed that the most significant protein was haptoglobin, which was increased in the NMO gels. The subsequent ELISA test were performed to validate it, which confirmed the results of proteomic analysis. Protein network was built, which showed some biological interactions among the seven proteins. These results support a correlation between the level of haptoglobin and NMO. Haptoglobin may be a potential useful biomarker for diagnosis or a medicine target for treatment of NMO. [ABSTRACT FROM AUTHOR]

The diagnosis of multiple sclerosis (MS) is challenging for the lack of a specific diagnostic test. Recent researches in quantitative proteomics, however, offer new opportunities for biomarker discovery and the study of disease pathogenesis. To find more potential protein biomarkers, we used two technologies, 2-dimensional fluorescence difference in-gel electrophoresis (2D-DIGE), followed by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) and ultra-performance liquid chromato-graph coupled with quadrupole time-of-flight mass spectrometry (UPLC/Q-TOF MS), to quantitatively analyse differential proteomic expression in the cerebrospinal fluid (CSF) between patients with MS (the experiment group) and patients with other neurological diseases (ONDs; the control group). Analysis by the former technology identified more than 43 different protein spots (39 proteins), of which 17 spots (13 proteins) showed more than 1.5-fold difference in abundance as analysed by DeCyder software (GE Healthcare, Piscataway. NJ, USA) between the MS and the ONDs groups. The expression of five protein spots was elevated and the expression of 12 protein spots was decreased in the MS group. Meanwhile, the latter method, UPLC/Q-TOF MS showed 68 different proteins. There were 45 proteins with a difference of more than 1.5 folds between the two groups, in which the expression of 20 proteins was elevated and the expression of 25 proteins was decreased in the MS group. Data provided by the two methods indicated that the proteins overlapped ratio was 27% in the 26 significant proteins that had the same regulation tendency. The differential CSF proteins were analysed further by biological network and it revealed interaction of them. The subsequent ELISA measuring the concentration of cystatin C (P < 0.01), which was one of the proteins discovered simultaneously with the two technologies, confirmed the results of the two quantitative proteomic analysis. The combination of the two quantitative proteomic technologies was helpful in discovering differentially expressed proteins that may have a connection with MS disease physiology and serve as useful biomarkers for diagnosis and treatment of MS diseases. [ABSTRACT FROM AUTHOR]

Abstract   Objective To better understand the pathophysiological mechanisms underlying Guillain-Barré Syndrome (GBS) and to ascertain the protein that presents with the most observable changes in the cerebrospinal fluid (CSF) of patients GBS. Methods we analyzed individually the proteomes of CSF of patients with GBS (the experiment group) and control subjects suffering from other neurological disorders (the control group) with two-dimensional gel electrophoresis (2-DE). The harvested gel images analyzed with software to ascertain the most significant differential protein between the two groups and identify it by matrix-assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF/TOF MS). We based the enzyme linked immuno-absorbent assay (ELISA) experiment, which followed, on the results of the first experiment. Results There are six of the protein spots had significant difference in expression between two group and identification made by mass spectrography revealed that the most significant disparity was cystatin C, which was decrease in the gels. The subsequent ELISA confirmed a considerable decrease in the level of cystatin C (P Conclusion The level of cystatin C decreases significantly in the CSF of GBS and calls for further studying the role in the pathogenesis of GBS. [ABSTRACT FROM AUTHOR]

In the present study we demonstrated Jurkat leukemia cell lines of TIB152 and TIB153 with different sensitivities to recombinant soluble TRAIL cytotoxicity. TRAIL receptor death receptor 5 (DR5) was constitutively localized in the rafts in both cell lines. FADD, caspase-8, and PI3K-p85 subunit were recruited into DR5 lipid rafts of TIB152 but not in TIB153 cells. The expression and enzyme activity of acid sphingomyelinase, which digests sphingomyeline to produce ceramide and plays an essential role in lipid raft assembling, were higher in the rafts of TIB152 than in TIB153. These data provide evidences that DR5-recruited raft components contribute to the different sensitivity of Jurkat leukemia cell lines to TRAIL-induced cell death and may throw some light on the development of better therapeutic strategies for the cancer cells resistant to TRAIL treatment. © 2009 IUBMB IUBMB Life, 61(3): 261–267, 2009 [ABSTRACT FROM AUTHOR]

Abstract   Objective The diagnosis of multiple sclerosis (MS) is still challenging recently due to the lack of a specific diagnostic test. Proteomics analysis was applied to biomarkers discovery and their pathways study. Methods First, the proteins of CSF from MS patients and control group were analyzed individually with 2D-DIGE technology (two-dimensional difference gel electrophoresis). Then, protein spots were found out with DeCyder6.0 software which showed different expression levels in the gel images between the two groups. The information regarding these proteins was collected based on MALDI-TOF/MS and related database searches. Lastly, interaction between these proteins was further analyzed by using Metacore software. Results There were 13 proteins that showed more than 1.5-fold difference in expression levels between the two groups. Furthermore, the identification made by MALDI-TOF/MS revealed that one of the most significant differential proteins was DBP (vitamin D-binding protein), which decreased in the experimental group. This result was confirmed by ELISA (P Conclusion These results support a correlation between the level of DBP and MS. DBP may be a potential useful biomarker for diagnosis or a medicine target for treatment of MS. [ABSTRACT FROM AUTHOR]

We have reported that Erbin expression was down-regulated in the Jurkat leukaemia T lymphocytes treated with the recombinant soluble tumour necrosis factor-related apoptosis-inducing ligand (rsTRAIL). Herein, we studied the expression and the regulation of Erbin and its binding partner, ErbB2, in the MCF-7 breast cancer cell line. We showed that the expressions of Erbin and ErbB2 were modulated by PKCδ inhibitor, rottlerin, in the TRAIL-resistant MCF-7 cell line. The affinity of Erbin–ErbB2 interaction was reduced by ErbB2 phosphorylation. Inhibiting the expression of Erbin facilitated the sensitivity of the MCF-7 cells to TRAIL via suppressing the ErbB2/AKT/NF-κB signalling pathway. [ABSTRACT FROM PUBLISHER]

Background: Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) functions as a cytokine to selectively kill various cancer cells without toxicity to most normal cells. Numerous studies have demonstrated the potential use of recombinant soluble TRAIL as a cancer therapeutic agent. We have showed previous administration of a recombinant adeno-associated virus (rAAV) vector expressing soluble TRAIL results in an efficient suppression of human tumor growth in nude mice. In the present study, we introduced Tet-On gene expression system into the rAAV vector to control the soluble TRAIL expression and evaluate the efficiency of the system in cancer gene therapy.Methods: Controllability of the Tet-On system was determined by luciferase activity assay, and Western blotting and enzyme-linked immunoabsorbent assay. Cell viability was determined by MTT assay. The breast cancer xenograft animal model was established and recombinant virus was administrated through tail vein injection to evaluate the tumoricidal activity.Results: The expression of soluble TRAIL could be strictly controlled by the Tet-On system in both normal and cancer cells. Transduction of human cancer cell lines with rAAV-TRE-TRAIL&rAAV-Tet-On under the presence of inducer doxycycline resulted in a considerable cell death by apoptosis. Intravenous injection of the recombinant virus efficiently suppressed the growth of human breast carcinoma in nude mice when activated by doxycycline.Conclusion: These data suggest that rAAV-mediated soluble TRAIL expression under the control of the Tet-On system is a promising strategy for breast cancer therapy. [ABSTRACT FROM AUTHOR]