Inhibition of CD83 Alleviates Systemic Inflammation in Herpes Simplex Virus Type 1-Induced Behçet's Disease Model Mouse.

Behçet's disease (BD) is an autoinflammatory disease that can lead to life- and sight-threating complications. Dendritic cells (DCs) are the most potent antigen-presenting cells that can regulate multiple inflammatory pathways. The objective of this study was to investigate the association of the DC stimulatory molecule CD83 with BD. Frequencies of costimulatory molecules expressing DCs in peripheral blood leukocytes (PBL) were measured by flow cytometry (FACS). The severity of symptoms in HSV-1-induced BD symptomatic mice was also assessed. Frequencies of CD83-positive cells were significantly increased in mice exhibiting BD symptoms, compared to those in asymptomatic mice. Abatacept, a CD80/86 blocker, significantly decreased the frequencies of CD83-positive cells in a time- and dose-dependent manner. BD symptomatic mice treated with Abatacept showed gradual reduction in the severity score of symptoms. Intraperitoneal injection of CD83 siRNA significantly reduced the frequencies of CD83-positive cells in PBL and peritoneal macrophages. After CD83 siRNA injection, BD symptoms of mice were improved and disease severity was decreased. Discontinuation of CD83 siRNA deteriorated symptoms while readministration of CD83 siRNA again improved BD symptoms of mice. These results clearly indicate the involvement of CD83-expressing cells in the inflammatory symptoms of BD. Therefore, CD83 might be useful as a therapeutic target for BD.

Differences in Expression of Human Leukocyte Antigen Class II Subtypes and T Cell Subsets in Behçet's Disease with Arthritis.

It has been reported Human Leukocyte Antigen (HLA) gene polymorphism is a risk factor for the development of Behçet's disease (BD). In this study, the association of HLA class II subtypes HLA-DP, DQ, DR, and T cell subsets in BD patients with arthritis was evaluated. Frequencies of HLA-DP, DQ, DR positive cells, and T cell subsets in peripheral blood leukocytes (PBL) were measured by flow cytometric analysis in BD, and compared to rheumatoid arthritis as disease controls and healthy controls. Frequencies of HLA-DQ were significantly decreased in whole PBL and granulocytes of BD active patients as compared to healthy controls. In monocytes populations, proportions of HLA-DR positive cells were significantly increased in BD active patients as compared to healthy controls. Proportions of CD4+CCR7+ and CD8+CCR7+ cells were significantly higher in BD active patients than in BD inactive in whole PBL. Frequencies of CD4+CD62L- and CD8+CD62L- cells in lymphocytes were significantly decreased in active BD than those in inactive BD. There were also correlations between disease activity markers and T cell subsets. Our results revealed HLA-DP, DQ, and DR expressing cell frequencies and several T cell subsets were significantly correlated with BD arthritis symptoms.

Characteristic patterns of HLA presentation and T cell differentiation in adult-onset Still's disease.

We examined the expression of human leukocyte antigen (HLA) and composition of differentiated T cells in the peripheral blood to understand the characteristics of the immune changes in patients with adult-onset Still's disease (AOSD). This study enrolled patients with AOSD (n = 14), patients with rheumatoid arthritis (RA, n = 20), and healthy controls (HC, n = 20). The percentage of surface-stained cells with HLA-DP, DQ, and DR alleles and the composition of differentiated T cells in peripheral blood leukocytes (PBLs) were evaluated by flow cytometry. AOSD patients exhibited significantly higher percentages of lymphocytes presenting HLA-DP and HLA-DR, and lower percentages of cells presenting HLA-DQ, than RA patients or HC. The proportions of CD4+, CD4+CCR7+, CD4+CD62L-, and CD8+CD62L- cells from PBLs were decreased in AOSD patients relative to RA patients or HCs. By contrast, AOSD patients had higher proportions of CD8+naïve T cells in whole blood relative to RA patients or HC. The proportions of CD4+ effector memory T cells, CD8+ naïve T cells, and CD8+ effector memory T cells in whole blood cells and CD4+ effector memory T cell in lymphocytes were significantly associated with the systemic score. While the proportions of CD4+, CD8+, CCR7+, CD4+CCR7+, CD4+CD62L-, and CD8+CD62L- cells were significantly decreased in AOSD patients, and the proportion of CD8+naïve T cells was elevated in AOSD and correlated with the systemic score. Further studies of a large cohort of AOSD patients will be necessary to evaluate these markers in the pathogenesis of AOSD.

Frequencies of IL-15Rα+ cells in patients with Behçet's disease and the effects of overexpressing IL-15Rα+ on disease symptoms in mice.

It has been suggested higher serum levels of IL-15 and lower expression levels of IL-15 receptor alpha (IL-15Rα) are correlated with pathogenesis of Behçet's disease (BD). However, whether overexpressing IL-15Rα could be used as a therapeutic candidate for BD is currently unclear. Therefore, the purpose of this study was to determine whether overexpressing IL-15Rα could affect BD symptoms in a mouse model. IL-15/IL-15Rα complex expressing vector or protein complex of IL-15/IL-15Rα-Fc was used to treat BD mice. Frequencies of IL-15Rα+ cells in peripheral blood leukocytes (PBL) and lymph node cells were determined using a flow cytometer. BD symptoms in mice improved after treatment with IL-15/15Rα expression vector or IL-15/IL-15Rα-Fc protein complex. In addition, treatment with pIL-15/15Rα significantly (p = .016) decreased disease severity score of BD mice compared to treatment with control vector. Frequencies of IL-15Rα+ cells were also significantly (p = .01) higher in peritoneal macrophages of pIL-15/15Rα treated BD mice than those of mice treated with control vector. Frequencies of IL-15Rα+ PBL were also significantly higher in BD mice treated with IL-15/IL-15Rα-Fc protein complex than those in the control group. These results suggest up-regulating IL-15Rα+ cells could be used as novel therapeutic strategies to control BD in the future.

Role of N-acetyl galactosamine-4-SO4, a ligand of CD206 in HSV-induced mouse model of Behçet's disease.

CD206 is a macrophage mannose receptor involved in variety of autoimmune and inflammatory diseases. This study aimed to identify the pathogenic role of CD206 in a herpes simplex virus (HSV) induced Behçet's disease (BD) mouse model. CD206 positive cells were detected in peripheral blood mononuclear cells and quantified by flow cytometry. Levels of cytokines were measured by ELISA. CD206 was found to be down-regulated both in vitro (10-6M) and in vivo (200µg/mouse) after treatment with N-acetylgalactosamine (GalNAc), a ligand for CD206. The down-regulation of CD206 was correlated with improvement in BD symptoms. Colchicine (2µg/mouse) or pentoxifylline (400µg/mouse) treated mice displayed improvement in BD symptoms with fewer CD206 positive cells. The prevalence of CD206-positive cells differed between ligand-responsive and non-responsive BD mice. Inhibition of CD206 was associated with down-regulated serum level of interleukin-17 in GalNAc-treated BD mice. These results suggest that the expression of CD206 is correlated with HSV-induced BD symptoms in mice, implicating that CD206 might have a pathogenic role in BD.

The Correlation of CD206, CD209, and Disease Severity in Behçet's Disease with Arthritis.

The purpose of this study was to clarify the role of pattern recognition receptors in Behçet's disease (BD). The frequencies of several pattern recognition receptors (CD11b, CD11c, CD32, CD206, CD209, and dectin-1) were analyzed in patients with BD by flow cytometry, and cytokine levels, interleukin- (IL-) 18, IL-23, and IL-17A, were compared in plasma. The analysis was performed in active (n = 13) and inactive (n = 13) stages of BD patients. Rheumatoid arthritis patients (n = 19), as a disease control, and healthy control (HC) (n = 19) were enrolled. The frequencies of CD11b+ and CD32+ cells were significantly increased in active BD patients compared to HC. Disease severity score was correlated to CD11c+, CD206+, and CD209+ in whole leukocytes and CD11b+, CD11c+, CD206+, CD209+, and Dectin-1+ in granulocytes. The plasma levels of IL-17A were significantly different between HC and active BD. IL-18 showed significant difference between active and inactive BD patients. From this study, we concluded the expressions of several pattern recognition receptors were correlated to the joint symptoms of BD.

Highly Expression of CD11b and CD32 on Peripheral Blood Mononuclear Cells from Patients with Adult-Onset Still's Disease.

BACKGROUND: We investigated the potential role of several pattern-recognition receptors (PRRs; CD11b, CD11c, CD32, CD206, CD209, and dectin-1) in adult-onset Still's disease (AOSD). METHODS: The study included 13 untreated AOSD patients, 19 rheumatoid arthritis (RA) patients (as a disease control), and 19 healthy controls (HCs). The PRRs were quantified in peripheral blood using flow cytometry. The serum levels of interleukin-17 (IL-17), IL-18, and IL-23 were measured by enzyme-linked immunosorbent assay. RESULTS: Significantly higher mean frequencies of cells presenting CD11b and CD32 from whole blood were observed in patients with AOSD than in patients with RA or HC. The levels of IL-17, IL-18, and IL-23 were elevated in AOSD patients compared to HCs. CD11b frequencies from whole cells correlated with systemic scores, lactate dehydrogenase (LDH) levels, aspartate transaminase levels, interleukin-23 (IL-23) levels, and IL-18. Frequencies of CD209 from granulocytes were significantly correlated with systemic scores, and the erythrocyte sedimentation rate and levels of C-reactive protein, ferritin, LDH, IL-23, and interleukin-18 (IL-18). CONCLUSIONS: Elevated frequencies of circulating CD11b-positive cells and positive correlations with disease activity markers suggest that circulating CD11b-positive cells contribute to the pathogenesis of AOSD.

Chitosan as an Immunomodulating Adjuvant on T-Cells and Antigen-Presenting Cells in Herpes Simplex Virus Type 1 Infection.

Herpes disease caused by herpes simplex virus type 1 (HSV-1) is an intractable condition. It is a major concern in public health. Our purpose of this study was to verify the function of chitosan as an adjuvant for immune regulation specifically under herpes simplex virus type 1 (HSV-1) infection. Ahead of HSV infection, chitosan, heat inactivated green fluorescent protein expressing HSV (G-HSV), and a combination of chitosan and G-HSV were used to pretreat ICR mice followed by HSV-1 infection. Using flow cytometric analysis, the frequencies of T-cells, monocytes, dendritic cells (DCs), and natural killer (NK) cells were analyzed by surface expression of CD4+, CD8+, CD14+, CD11c+, NK1.1+, and DX5+ cells. In HSV infected mice, chitosan treatment significantly increased the frequencies of CD4+ T-cells (33.6 ± 5.78%) compared to those in the control group (24.02 ± 12.47%, p = 0.05). The frequencies of DC and NK cells were also significantly different between chitosan treated mice and control mice. In addition, anti-HSV IgG antibody was downregulated in chitosan treated mice. These results suggest that chitosan is a potential modulator or immune stimulator as an adjuvant in HSV-1 infected mice.

IL-2/IL-2 antibody immune complex regulates HSV-induced inflammation through induction of IL-2 receptor alpha, beta, and gamma in a mouse model.

The differences of serum IL-2 levels were not consistent between Behçet's Disease (BD) patients and healthy controls, however, the correlation of interleukin-2 receptor (IL-2R) and BD has not been investigated. IL-2R is composed of three subunits; alpha, beta, and gamma. The expression frequencies of IL-2R subunits were analyzed in the peripheral blood mononuclear cells, spleens, and lymph node (LN) cells. The expression of IL-2R subunits was different between BD mice and controls. IL-2R beta expressing cell frequencies were also different between BD patients and healthy controls. The IL-2/anti-mIL-2 antibody complex administration regulated the IL-2R subunits in mice. The change of expression in IL-2R was accompanied by the increase of CD8+CD44+ memory T cells, CD3-NK1.1+CD11b+CD27+ natural killer cells, and improvement of symptoms. In this study, we elucidated the role of IL-2R subunits on BD, a finding that can be connected to therapeutic strategy for patients based on the results from the treatment of BD mice.

The relevance of miRNA-21 in HSV-induced inflammation in a mouse model.

The purpose of this study was to clarify the correlation between microRNA-21 (miR-21) expression and inflammation in a herpes simplex virus (HSV)-induced Behçet's Disease (BD) mouse model. miR-21 was compared between BD patients and healthy controls in peripheral blood mononuclear cells (PBMC). For miR-21 inhibition, miR-21 antagomir was applied to BD mice. The change of symptoms was monitored. The levels of cytokines and related molecules were determined by ELISA and real time qPCR. Treatment with colchicine or pentoxifylline down-regulated the level of miR-21 with improved symptoms in mice. miR-21 inhibition was accompanied by down-regulated serum levels of IL-17 and IL-6. The expression levels of PDCD4, RhoB, PD-1, IL-12p35, and toll-like receptor-4 were also regulated by miR-21 inhibition. miR-21 was correlated with HSV-induced BD-like inflammation in mice and BD patients. The expression of miR-21 was regulated by antagomir in mice.

The role of classical and alternative macrophages in the immunopathogenesis of herpes simplex virus-induced inflammation in a mouse model.

BACKGROUND: The exact mechanism of the inflammatory changes occurring during the development of Behçet's disease (BD) remains unclear. OBJECTIVE: We investigated the role of classical (M1) and alternative (M2) activation of macrophages in a herpes simplex virus (HSV)-induced BD mouse model. METHODS: The classical vs. alternative activated macrophage ratio (M1/M2 ratio) was calculated by analyzing the surface markers CD16/32 and CD23 as M1 and M2 markers, respectively, by flow cytometry. mRNA expression of interferon (IFN)-γ and interleukin (IL)-6 as M1 and arginase-1, FIZZ-1, and MHC-II as M2 markers were analyzed by reverse transcription-polymerase chain reaction. Cytokine levels were assessed by enzyme-linked immunosorbent assay. RESULTS: The M1 phenotype was upregulated in BD mice, and an increased M1/M2 ratio was observed compared to that in asymptomatic BD normal and normal healthy mice. Recombinant (r)IFN-γ significantly increased the M1/M2 ratio (1.74±0.42) compared with that of rIL-4 (0.83±0.20). BD mice treated with rIL-4 showed a decreased M1/M2 ratio (1.2±0.3) compared to that of the rIFN-γ- (2.1±2.3) treated group and also showed ameliorated BD symptoms accompanied by downregulation of IL-17 and IL-6 and up-regulation of IL-4. CONCLUSION: Therefore, modulation of macrophage phenotypes could be an effective therapeutic approach for treating BD in the future.

Tocotrienols enhance melanosome degradation through endosome docking/fusion proteins in B16F10 melanoma cells.

Vitamin E inhibits tyrosinase activity and acts as a melanogenesis inhibitor in epidermal melanocytes in vitro. However, there is no direct evidence indicating that melanosomes are degraded in lysosomes in the presence of vitamin E. To determine whether vitamin E-induced melanosome disintegration is related to the expression of endosome docking/fusion proteins in B16F10 melanoma cells, electron microscopy, reverse transcription-polymerase chain reaction (RT-PCR), and real-time PCR were used to observe the effects of tocomin (α-tocopherols and α,γ,δ-tocotrienols in palm oil) on B16F10 melanoma cells. Melanosomal integrity was lost in lysosomes of B16F10 melanoma cells when treated with tocomin, indicating that tocomin caused the degradation of melanosomes in the lysosomal compartment. RT-PCR and real-time PCR analysis demonstrated mRNA expression of tyrosinase and the endosome docking/fusion proteins (syntaxin7, Rab7, Vps11, Vps16, Vps33, Vps39, and Vps41). Expression of syntaxin7, Vps16, Vps33, and Vps41 mRNA increased significantly in cells treated with tocomin compared with that in controls. These results indicate that the tocomin-induced degradation of melanosomes in the lysosomal compartment occurs with an increase in endosome docking/fusion proteins (syntaxin7, Vps16, Vps33, and Vps41) in cultured B16F10 melanoma cells.

Therapeutic potency of Poly I:C in HSV-induced inflammation through up-regulation of IL-15 receptor alpha.

Interleukin-15 receptor alpha (IL-15Rα) forms stable complex with IL-15 on the cell surface of activated monocytes and mediates the proliferation of memory CD8+ T cells. Recent studies informed that polyinosinic:polycytidylic acid (Poly I:C) is an immunostimulant which boosts the generation of memory T cells through induction of IL-15Rα. The aim of this study is to evaluate the relevance of IL-15Rα in Herpes simplex virus (HSV)-induced Behçet's disease (BD) mouse model and BD patients. The frequencies of IL-15Rα expression in PBMCs of BD patients and BD-like symptomatic mice were analyzed by flow cytometry. In addition, Poly I:C supplementation could reduce inflammation through the up-regulation of memory T cells and IL-15Rα+ cells accompany with down-regulation of pro-inflammatory cytokine, IL-17A in BD mice. In BD patients, the frequencies of IL-15Rα expression in PBMCs were also significantly different between the inactive and active disease states. These results suggest that IL-15Rα is a relevant factor in BD.

CCL21 attenuates HSV-induced inflammation through up-regulation of CD8+ memory cells.

CCR7 and its ligand, CCL21, are known to establish microenvironments for the initiation of immune responses in secondary lymphoid tissue. It has also been reported that CCR7 ligand gene-deleted mice have defects in lymphocyte homing. In addition, the injection of the CCR7 ligand was shown to induce the expression of memory T cells. In this study, we analyzed the expression of CCR7 and its ligand in HSV-induced Behçet's disease (BD)-like inflammation of mice. Additionally, plasmids containing the CCR7 ligand CCL19 or CCL21, pcDNA3.1-CCL19 or pcDNA3.1-CCL21, respectively, were injected into symptomatic mice, and changes in the population of memory T cells were determined. After administration of pcDNA3.1-CCL21, the frequencies of CD8+CD44+, CD8+CD62L- memory T cells were significantly up-regulated and the symptoms were not deteriorated when compared to the control vector injected group. Specifically, the difference in frequencies of CCR7+ peripheral blood mononuclear cells between active BD patients and inactive BD patients was similar to that of HSV-induced BD-like mice. These results suggest that CCR7, its ligand, and CD8+ memory cells are correlated with the regulation of BD symptoms.

The role of T cell immunoglobulin mucin domains 1 and 4 in a herpes simplex virus-induced Behçet's disease mouse model.

The T cell immunoglobulin mucin (TIM) proteins regulate T cell activation and tolerance. TIM-1 plays an important role in the regulation of immune responses and the development of autoimmune diseases. TIM-4 is a natural ligand of TIM-1, and the interaction of TIM-1 and TIM-4 is involved in the regulation of T helper (Th) cell responses and modulation of the Th1/Th2 cytokine balance. Behçet's disease (BD) is a chronic, multisystemic inflammatory disorder with arthritic, intestinal, mucocutaneous, ocular, vascular, and central nervous system involvement. Tim-1 expression was lower in a herpes simplex virus-induced BD mouse model compared to that in asymptomatic BD normal (BDN) mice. Tim-4 expression was higher in BD mice than that in BDN mice. In this study, we investigated the Tim expression in a BD mouse model with BD-like symptoms. Tim-1 and Tim-4 expression was regulated by an expression vector or siRNA injected into the BD mouse model. The Tim-1 vector injected into BD mice resulted in changes in BD-like symptoms and decreased the severity score. Treatment with Tim-4 siRNA also improved BD-like symptoms and decreased the severity score accompanied by upregulation of regulatory T cells. We showed that regulating Tim-1 or Tim-4 affected BD-like symptoms in mice.

Pretreatment with interleukin-6 small interfering RNA can improve the survival rate of polymicrobial cecal ligation and puncture mice by down regulating interleukin-6 production.

Interleukin-6 (IL-6) is considered to be an early marker of severe sepsis that is associated with increased morbidity and mortality. Therefore, we pretreated male ICR mice with IL-6 small interfering RNA (siRNA) before cecal ligation and puncture (CLP) and observed the changes in their survival in response to down regulation of IL-6, as well as the role of Th subsets during sepsis. In addition, sham and CLP operated mice were sacrificed at different time points to determine the serum IL-6 levels during early and late sepsis. IL-6 siRNA pretreated septic mice showed markedly extended survival (23.3%) up to 10 days and significantly reduced serum IL-6 levels at day 5 (209.90 ± 0.50 pg/ml; P<0.0001) when compared to CLP mice at day 1. Furthermore, IL-6 mRNA and protein were highly expressed during early sepsis in CLP mice at day 1 and mRNA was not detected in IL-6 siRNA treated CLP mice at days 1 or 5 and serum level of IL-6 was also decreased significantly (P<0.01). In addition, the mRNA expression of C5aR, ROR-γt, PU.1 and protein expression of IL-17 were high at day 5 in IL-6 siRNA treated mice. Taken together, the results of the present study demonstrate that pretreatment with IL-6 siRNA improved CLP induced septic mice survival. Furthermore, the IL-6 level was down-regulated and the transcription factors ROR-γt and PU.1 were up-regulated by IL-6 siRNA at late sepsis. The results presented herein also suggest that IL-6 siRNA could be a potential molecular therapeutic strategy for the treatment of sepsis.

Vitamin D3 ameliorates herpes simplex virus-induced Behçet's disease-like inflammation in a mouse model through down-regulation of Toll-like receptors.

OBJECTIVES: This study was conducted to understand the role of vitamin D3 through the regulation of Toll like receptors (TLRs) and cytokines in herpes simplex virus-induced Behçet's disease (BD)-like mice. METHODS: Serum 25-hydroxyvitamin D (1,25(OH)2D3) levels were measured in BD-like mice, as well as virus injected and asymptomatic appearance BD normal mice (BDN). The frequencies of TLRs of peritoneal macrophages were compared by FACS. To determine the effect of vitamin D3 in vitro, peritoneal macrophages were isolated and then incubated with 1,25(OH)2D3. To identify the mechanism of improvement of BD-like symptoms induced by 1,25(OH)2D3, mice were orally administered 1,25(OH)2D3. RESULTS: The serum 25-hydroxyvitamin D levels in BD-like mice were 12.4 ± 5.4ng/ml, while they were 17.5 ± 7.2ng/ml in BDN mice. The frequency of TLR2 in BD-like mice was 32.91 ± 20.88%, while it was 12.73 ± 7.67% in BDN. The frequency of TLR4 was 26.09 ± 10.20% in BD-like mice and 9.72 ± 5.30% in BDN. In a 72 h culture of peritoneal macrophages in 10-8 M 1,25(OH)2D3, the frequency of TLR2 was 25.0 ± 2.7%, while it was 37.3 ± 5.8% in the control group. The frequency of TLR4 was 18.9 ± 5.3% with 1,25(OH)2D3, while it was 30.3 ± 0.1% in the control group. Treatment with 1,25(OH)2D3 improved the symptoms in six out of 11 BD-like mice and downregulated the frequency of TLR2 and TLR4. Moreover, 1,25(OH)2D3 influenced Interleukin-6 and TNF-alpha expression in the sera of BD-like mice. CONCLUSIONS: Vitamin D improved BD-like symptoms by down-regulating the expression of TLRs and pro-inflammatory cytokines in in vivo mouse models.

Synthesized pyridine compound derivatives decreased TNF alpha and adhesion molecules and ameliorated HSV-induced inflammation in a mouse model.

Synthesized pyridine compound derivatives (SK94, SK126) from a natural lead source were administered to mice to test for possible anti-TNF alpha and anti-inflammatory activities. Lipopolysaccharide (LPS)-induced TNF alpha production was analyzed in the endothelial cells, Raw 264.7 cells, and serum of normal mice after treatment with SK compounds. These compounds were also orally administered to a herpes simplex virus (HSV)-induced Behcet's disease mouse model to investigate their anti-inflammatory therapeutic effect. TNF alpha production was inhibited in a dose-dependent manner in the SK94 treated cells. E-selectin, VCAM-1, and ICAM-1 mRNA levels were also down-regulated. Treatment with 30mg/kg SK94 inhibited 55% of the TNF alpha production in LPS challenged Balb/c mice (n=8). SK94 and SK126 were administered to the Behcet's disease-like mice for five consecutive days and SK94 improved in five out of six mice (83%), while it only improved in one out of nine mice (11%) in the pH 1.2 saline (artificial gastric juice) group (P<0.005), four out of ten mice (40%) in the thalidomide group (P<0.05), and six out of seven (86%) in the SK126 group (P<0.005). Soluble ICAM-1 was inhibited by 23.8% in the sera of SK94 treated mice and by 34.6% in SK126 treated mice when compared to artificial gastric juice. Based on these findings, SK compounds could be candidates for clinical trials.

The role of Qa-2, the functional homolog of HLA-G, in a Behcet's disease-like mouse model induced by the herpes virus simplex.

BACKGROUND: It has been suggested that the HLA-G molecule is a genetic risk factor for Behcet's disease (BD). In this study, we evaluated the level of Qa-2, a murine nonclassical class I MHC molecule and possible functional homolog of HLA-G, to determine if it was associated with various symptoms of BD-like mice. In addition, we investigated siRNA (small interfering RNA) treatment to determine if it inhibited Qa-2 expression, thereby changing the symptoms of mice. METHODS: RNA interference (RNAi) and vector transfection were employed to manipulate gene expression in vivo in mice. siRNA (small interfering RNA) or Qa-2 expression vector was applied to inhibit or up-regulate Qa-2 expression, respectively. RESULTS: The Qa-2 levels in granulocytes were lower in BD-like mice than in normal controls. The silencing of Qa-2 by intravenous injection of siRNA (500 nmol/mouse, 4 times at 3-day intervals) specifically reduced the Qa-2 levels and worsened the BD-like symptoms. CONCLUSIONS: Silencing Qa-2 by injecting siRNA into mice resulted in deterioration of symptoms in BD-like mice.

Rebamipide affects the efficiency of colchicine for the herpes simplex virus-induced inflammation in a Behcet's disease mouse model.

Rebamipide inhibits free radicals derived from activated neutrophils and decreases the inhibiting inflammatory cytokine. Behcet's disease (BD) is a chronic, multi-systemic inflammatory disorder with arthritic, gastrointestinal, mucocutaneous, ocular, vascular, and central nervous system involvement. This disease has a chronic course with periodic exacerbations and progressive deterioration. To study the effect of rebamipide treatment to BD-like mice, combination treatment with rebamipide and colchicine was compared to colchicine treatment. Colchicine is one of the most frequently prescribed medicine to the patients with BD. For each BD mouse, 200 microl gastric fluid or 2 microg colchicine or 150 microg rebamipide or 2 microg colchicine plus 150 microg rebamipide was treated orally once per day. Treatment was done for 5 consecutive days. Two hour or 20 days after last administration, spleens were isolated for RT-PCR and real time PCR, and serum was collected for ELISA. In the combination treated group, TNF alpha, MIP-1 alpha, p22 phox, p47 phox, and gp91 phox mRNA expressions were lower than rebamipide treated or colchicine treated groups by reverse transcriptase PCR. NADPH oxidase subunits mRNA were markedly downregulated compared to the colchicine treated group by real time PCR. At 20 days after administration, combination treatment decreased 23.5% of the severity score compared to before administration. In contrast, colchicine treatment decreased 14.3% of the severity score compared to before administration. Rebamipide helped the function of colchicine to improve the HSV induced BD-like symptoms by inhibiting the expression of NADPH oxidase in vivo mouse model.