Association of killer cell immunoglobulin-like receptor (KIR) genes and their HLA ligands with susceptibility to Behçet's disease.

OBJECTIVES: Behçet's disease (BD) is a systemic inflammatory disorder with remissions and exacerbations. It is thought that defects in the natural killer (NK) cell repertoire may be involved in BD through killer cell immunoglobulin-like receptors (KIRs). This study aimed to evaluate KIR and HLA genes, their interactions in BD patients, and their associations with clinical manifestations. METHOD: The presence or absence of KIR and HLA alleles and genotypes was analysed by polymerase chain reaction sequence-specific primer on genomic DNA of 397 BD patients and 300 healthy controls. RESULTS: None of the KIR genes showed significant effects on BD susceptibility. HLA-C1Asn80 showed a protective effect against BD, whereas HLA-C2Lys80, HLA-B-Bw4Ile80, HLA-B5, and HLA-B51 were associated with a susceptibility risk for BD. In the combination of KIR and HLA genes, the frequencies of HLA genotypes no. 2, 3, 5, and 8, and inhibitory KIR no. 4 were significantly higher in patients than in controls. The frequencies of KIR genotype no. 3 and HLA genotypes no. 1, 4, 6, 7, and 9 were significantly lower in patients than in controls. There were many associations between KIR and HLA genes with clinical features of BD. CONCLUSION: Differences in the frequency of HLA genes, KIR-HLA interactions, and genotypes between BD and healthy controls and their associations with clinical manifestations indicate that NK cells are involved in BD pathogenesis. The observed differences indicated an NK cell activity imbalance in BD patients, and suggest a role of the KIR-HLA repertoire in the development of BD.

The Immunomodulatory Effect of Bone-Marrow Mesenchymal Stem Cells on Expression of TLR3 and TLR9 in Mice Dendritic Cells.

BACKGROUND: Mesenchymal stem cells (MSCs) are multipotent cells with immunomodulatory effect on immune cells including dendritic cells (DCs). DCs are the most potent antigen-presenting cells (APC). MSCs have been found to modulate both differentiation and function of DCs. DCs express a broad range of Toll-like receptors (TLR), which play a critical role in DCs maturation and function. OBJECTIVE: To evaluate expression level of TLR3 and TLR9 transcripts in DCs following treatment with MSCs supernatant. METHODS: MSCs and DCs were derived from adult BALB/c mice bone marrow and spleen, respectively. MSCs supernatant was harvested following 24, 48, and 72 hours. Isolated DCs were treated with MSCs supernatant and incubated for 24 and 48 hours. TLR3 and TLR9 transcript levels were evaluated using real-time PCR. RESULTS: The results showed that 48 and 72 hours MSCs supernatant significantly decreased the expression of TLR3 in DCs following 24 and 48 hours incubation in comparison with untreated cells (p<0.01). Moreover, 48 hours of treatment with 24, 48 and 72 hours MSCs supernatant significantly decreased TLR9 transcript level (p<0.05). CONCLUSION: TLR3 and TLR9 mRNA expression decreases in DCs after incubation with MSCs culture supernatant. This confirms the immunomodulatory role of MSCs in cell-base therapy.

In Vitro Effects of Sodium Benzoate on Th1/Th2 Deviation in Patients with Multiple Sclerosis.

Interleukin 4 (IL-4) can improve the clinical manifestations in experimental autoimmune encephalomyelitis (EAE), the animal model of multiple sclerosis (MS). Sodium benzoate (NaB) deviates the cytokine profile to Th2 (or IL-4 producing) cells in EAE and thus might be effective in the treatment of MS. Therefore, in this study the effect of different concentrations of NaB on the percentage and mRNA levels of IL-4 and interferon gamma (IFN-γ)-producing peripheral blood mononuclear cells (PBMCs) of 20 Relapsing-remitting multiple sclerosis (RR-MS) patients and eight healthy controls was evaluated in the presence of mitogen (phytohemagglutinin, PHA) or specific antigen (myelin basic protein, MBP). Our results showed that in the patient's group the percentage of CD4(+)IL-4(+) cells was significantly increased in the presence of all concentrations of NaB when PBMCs were stimulated by MBP (p = 0.001) or PHA (p < 0.03). The same results were obtained for normal donors in the highest concentration of NaB, 1000 µg/ml (p = 0.02). Moreover, in the patient's group the percentage of CD4(+)IFN-γ(+) cells was decreased significantly when the PBMCs were stimulated by PHA and NaB (p < 0.004) or by MBP and 1000 µg/ml of NaB (p < 0.03). The effect of NaB on IL-4 and IFN-γ production was also documented at the mRNA levels. In conclusion, our data suggest that NaB is able to induce IL-4 production by human PBMCs and therefore might be a useful candidate for conjunctive therapy in RR-MS.

Targeting fibroblast-like synovial cells at sites of inflammation with peptide targeted liposomes results in inhibition of experimental arthritis.

In this study we examined a synovium-specific targeted liposomal drug delivery system for its ability to localize and release its drug cargo to inflamed joints. Targeted liposomes were tested in vitro for binding to synovial fibroblast like (FLS) and endothelial cells using flow cytometry and in vivo for localization to joints using a rat model of adjuvant induced arthritis (AIA). Targeted liposomes were then loaded with anti-arthritic medications and examined for clinical efficacy in AIA. Targeted liposomes specifically bound to rabbit FLS and human FLS and showed a 7-10 fold increase in vivo localization in affected joints compared to unaffected joints. Histological sections from rats treated with prednisone and a new immunosuppressive peptide CP showed minimal inflammation. This report substantiates the ability of the novel FLS sequence to target liposomal drug delivery and offers an alternative therapeutic approach for the treatment of arthritis.

Tolerogenic dendritic cells produced by lentiviral-mediated CD40- and interleukin-23p19-specific shRNA can ameliorate experimental autoimmune encephalomyelitis by suppressing T helper type 17 cells.

Down-regulation of soluble or membrane-bound co-stimulatory molecules by RNAi in dendritic cells can prevent the activation of immune responses. Therefore, this study was designed to evaluate the therapeutic efficacy of bone marrow-derived DCs (BMDCs) transduced with lentiviral vectors to permanently expressed shRNA specific for CD40 (CD40LV-DCs) and/or p19 subunit of interleukin (IL)-23 (p19LV-DCs) mRNAs in experimental autoimmune encephalomyelitis (EAE). In-vitro studies showed that double-transduced BMDCs (CD40(+) p19LV-DCs) resemble tolerogenic DCs due to profound down-regulation of CD40, lower expression of proinflammatory cytokines (IL-6 and IL-12), increased IL-10 production and stronger stimulation of myelin oligodendrocyte glycoprotein (MOG)35-55 -specific T cells for production of IL-10 compared with CD40LV-DCs, p19LV-DCs and BMDCs transduced with control lentiviral vector (CoLV-DCs). Moreover, injection of transduced CD40(+) p19LV- BMDCs in EAE mice resulted in more reduction in clinical score, significant reduction in IL-17 or increased production of IL-10 by mononuclear cells derived from the lymph nodes or spinal cord compared with CoLV-DCs-treated EAE mice. In conclusion, simultaneous knock-down of CD40 and IL-23 production by BMDCs may represent a promising therapeutic tool for the treatment of IL-17-dependent autoimmune diseases, including multiple sclerosis.

Investigation of osteopontin levels and genomic variation of osteopontin and its receptors in Type 1 diabetes mellitus.

BACKGROUND: Failure in self-tolerance towards ß-cells in diabetes mellitus (DM) pathogenesis involves a series of complex events that are governed by environmental and genetic factors. Considering the importance of osteopontin (OPN) in T-helper-1 (Th1) cells development, the aim of this study was to evaluate the serum level and gene polymorphism of OPN in Iranian Type 1 diabetic (T1DM) children. METHODS: In this case-control study, 87 T1DM children and 86 healthy ones were enrolled. Blood samples of both groups were checked for OPN level. The single nucleotide polymorphisms (SNP) were genotyped by RFLP analysis for OPN rs1126772, its receptor integrin α4 (ITGA4) rs 1449263, and CD44 rs8193. RESULTS: Serum levels of OPN in diabetic children were significantly higher in cases compared to the control group (p=0.023), but there was no significant relationship between OPN rs1126772 (p=0.79), its receptor integrin α4 (p=0.31), and CD44 rs8193 (p=0.45), and T1DM. CONCLUSION: Higher amounts of OPN were seen in T1DM children. It is assumed that OPN might have inducing effects on T1DM development, particularly when genetically susceptible individuals are predisposed by an environmental insult. However, the 3 SNPs of OPN and its receptors did not show noticeable association with T1DM. The power of our study (~19%) was insufficient to observe any significant statistical difference between the groups; moreover, this study does not exclude the possibility of association of other SNPs of OPN and its receptors with this disease, and more studies are needed to clarify the issue.

Proteome differences of placenta between pre-eclampsia and normal pregnancy.

Placenta is a tissue unique to pregnancy and despite its major role in pregnancy, little is known about the proteome changes within placenta during pregnancy-related diseases such as pre-eclampsia (PE). Therefore, the aim of this study is the analysis of proteome differences between pre-eclamptic and normal full-term placentas. To achieve this goal, five normal and five severe pre-eclamptic placentas were included in this study. Total placental proteins were extracted and subjected to two-dimensional polyacrylamide gel electrophoresis (2D-PAGE). After staining, the gels were scanned and the protein spots were analysed using Image Master 2D Platinum Software. Non-parametric Mann-Whitney test was used for analysis of the mean intensity differences of the spots between normal and pre-eclamptic placentas. Statistical analysis indicated that 17 spots were differently expressed in pre-eclamptic compared with normal placentas (p<0.05). Using Matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI TOF/TOF) mass analysis, 11 out of 17 spots were identified. Among them, four proteins (chloride intracellular channel 3, apolipoprotein A-I, transthyretin (TTR) and protein disulphide isomerase) were up-regulated while seven (peroxiredoxin 2, peroxiredoxin 3, Hsc 70, Cu/Zn-superoxide dismutase (SOD-1), actin gamma 1 propeptide, chain A of enoyl-coenzyme A hydratase and HSP gp96) showed decreased expression in PE in comparison with normal placentas. In conclusion, down-regulation of proteins with anti-oxidant activities (peroxiredoxin 2 and peroxiredoxin 3) and altered expression of stress-response proteins (Hsc 70, Hsp gp96 and protein disulphide isomerase) might play an important role in the pathogenesis of PE.

Association of TNF-alpha -308 G/A and IL-4 -589 C/T gene promoter polymorphisms with asthma susceptibility in the south of Iran.

BACKGROUND: Both tumor necrosis factor alpha (TNF-alpha) and interleukin (IL) 4 have been implicated in the pathogenesis of asthma. Furthermore, a G/A substitution at position -308 of the TNF-alpha gene promoter and a C/T substitution at position -589 of the IL-4 gene promoter have been associated with increased production of TNF-alpha and IL-4, respectively. OBJECTIVE: The aim of the present study was to analyze the association between TNF-alpha-308 G/A and IL-4-589 C/T polymorphisms and susceptibility to asthma in a group of patients from southern Iran. METHODS: We analyzed the frequency of TNF-alpha -308 G/A and IL-4-589 C/T polymorphisms in a total of 203 asthmatic patients compared to 113 nonasthmatic control subjects. RESULTS: An association was observed between the TNF-alpha -308 G/A polymorphism and susceptibility to asthma in patients with a ratio between forced expiratory volume in 1 second and forced vital capacity of less than 75% compared with normal subjects; however, the association did not achieve statistical significance (P = .054). The IL-4-589 C/T polymorphism was associated with asthma susceptibility (P = .02). In addition, the association between this polymorphism and asthma severity approached statistical significance (P = .07). CONCLUSION: These results provide further evidence for a role of TNF-alpha-308 G/A and IL-4-589 C/T polymorphisms in susceptibility to and severity of asthma. Further studies involving a larger number of patients may help to confirm our observations.

Association of interleukin-8 (IL-8 or CXCL8) -251T/A and CXCR2 +1208C/T gene polymorphisms with breast cancer.

A case-control study involving 257 breast cancer patients with invasive ductal carcinoma and 233 healthy women was carried out to explore if the IL-8 -251T/A polymorphism and the CXCR2 +1208 C/T polymorphism have a role in breast cancer susceptibility. Genotypic analysis showed an increased frequency of high producer IL-8 -251AA genotype (p=0.016) in the patient group as compared to controls while CXCR2 +1208 C/T polymorphism did not show any differences between studied groups. However, in contrast to IL-8 -251 polymorphism, the percentage of CXCR2 +1208 TT genotype was significantly lower in patients with NPI3.4 (2% and 12%, respectively; p=0.03). Also, ER- tumors showed an approximately significant higher CXCR2 +1208 TT genotype compared to ER+ tumors (18.6% and 7.1%, respectively; p=0.07). In conclusion, IL-8 -251T/A polymorphism is associated with development of invasive ductal carcinoma type of breast cancer while CXCR2 +1208C/T polymorphism may affect the disease progression.

TNF-alpha, TNF-beta and IL-4 gene polymorphisms in Iranian patients with multiple sclerosis.

OBJECTIVES: To investigate the association between tumor necrosis factor-alpha (TNF-alpha) G-308A, tumor necrosis factor-beta (TNF-beta) G+252A and interleukin-4 (IL-4) C-590T polymorphisms and susceptibility to multiple sclerosis (MS) development and clinical course of the disease. MATERIALS AND METHODS: Two hundred and seventy patients with MS and 542 sex and ethnic matched controls were enrolled in the present study. An allele-specific oligonucleotide polymerase chain reaction was used to detect the polymorphism at position -308 of the TNF-alpha gene. The genotypes of TNF-beta and IL-4 were determined by polymerase chain reaction-restriction fragment length polymorphism. RESULTS: Allelic and genotypic frequencies for these polymorphisms were similar in patients with MS and population controls or among different types of the disease. CONCLUSION: The results of the present study suggest that the three mentioned functional polymorphisms are not likely to cause susceptibility to MS in the Iranian population.