OBJECTIVETo study the correlation between airway inflammation and osteopontin (OPN) level in the lung tissue, and to study the effect of dexamethasone (DXM) on OPN expression.

METHODSFifty mice were randomly divided into 5 groups: normal control, ovalbumin (OVA)-challenged asthma groups (OVA inhalation for 1 week or 2 weeks) and DXM-treated asthma groups (DXM treatment for 1 week or 2 weeks). The mice were sensitized and challenged with OVA to prepare mouse model of acute asthma. Alterations of airway inflammation were observed by haematoxylin-eosin staining. Serum level of OVA-sIgE was evaluated using ELISA. OPN expression in the lung tissue was located and measured by immunohistochemistry and Western blot respectively. OPN mRNA level in the lung tissue was detected by real-time PCR.

RESULTSThe asthma groups showed more pathological changes in the airway than the normal control and the DXM-treated groups. Compared with the OVA-challenged 1 week group, the pathological alterations increased in the OVA-challenged 2 weeks group. The level of OVA-sIgE in serum increased in the asthma groups compared with the control and the DXM groups (P<0.01). Serum OVA-sIgE sevel increased more significantly in the OVA-challenged 2 weeks group compared with the OVA-challenged 1 week group (P<0.01). OPN protein and mRNA levels were significantly raised in the asthma groups compared with the normal control and the DXM groups (P<0.01), and both levels increased more significantly in the OVA-challenged 2 weeks group compared with the OVA-challenged 1 week group (P<0.01).

CONCLUSIONSThe increased OPN expression in the lung tissue is associated with more severe airway inflammation in asthmatic mice, suggesting that OPN may play an important role in the pathogenesis of asthma. DXM can alleviate airway inflammation possibly by inhibiting OPN production.

Animals ; Asthma ; drug therapy ; metabolism ; pathology ; Dexamethasone ; therapeutic use ; Enzyme-Linked Immunosorbent Assay ; Female ; Immunoglobulin E ; blood ; Lung ; metabolism ; pathology ; Mice ; Mice, Inbred BALB C ; Osteopontin ; analysis ; genetics ; physiology ; Ovalbumin ; immunology

The mechanism underlying T cell-mediated fulminant hepatitis is not fully understood. In this study, we investigated whether myeloid derived suppressor cells (MDSCs) could prevent the concanavalin A (ConA)-induced hepatitis through suppressing T cell proliferation. We observed an increase in the frequencies of MDSCs in mouse spleen and liver at early stage of ConA treatment, implicating that the MDSCs might be involved in the initial resistance of mice against ConA-mediated inflammation. Subpopulation analysis showed that the MDSCs in liver of ConA-induced mice were mainly granulocytic MDSCs. Adoptive transfer of the bone marrow-derived MDSCs into ConA-treated mice showed that the MDSCs migrated into the liver and spleen where they suppressed T cell proliferation through ROS pathway. In addition, the frequencies of MDSCs in mice were also significantly increased by the treatment with immune suppressor glucocorticoids. Transfer of MDSCs into the regulatory T cell (Treg)-depleted mice showed that the protective effect of MDSCs on ConA-induced hepatitis is Treg-independent. In conclusion, our results demonstrate that MDSCs possess a direct protective role in T cell-mediated hepatitis, and increasing the frequency of MDSCs by either adoptive transfer or glucocorticoid treatment represents a potential cell-based therapeutic strategy for the acute inflammatory disease.
Adoptive Transfer ; Animals ; Blotting, Western ; Bone Marrow Cells ; immunology ; CD11b Antigen ; immunology ; metabolism ; Cell Movement ; immunology ; Cell Proliferation ; Chemical and Drug Induced Liver Injury ; etiology ; immunology ; prevention & control ; Concanavalin A ; toxicity ; Dexamethasone ; pharmacology ; Flow Cytometry ; Glucocorticoids ; pharmacology ; Liver ; immunology ; pathology ; Male ; Mice, Inbred C57BL ; Mitogens ; administration & dosage ; toxicity ; Myeloid Cells ; immunology ; metabolism ; transplantation ; Receptors, Chemokine ; immunology ; metabolism ; Spleen ; immunology ; pathology ; T-Lymphocytes ; immunology ; T-Lymphocytes, Regulatory ; immunology

OBJECTIVETo investigate the change of CD4⁺CD25(high)CD127(low) regulatory T cells (Tregs) percentage in patients with primary immune thrombocytopenia (ITP) treated by different methods.

METHODSOne hundred and thirty-eight newly diagnosed adult ITP patients (57 male, median age 40 years, range 18-70 years) were enrolled in this study, who were randomly separated into three regiment groups, namely prednisolone (PSL, 1.5 mg/kg for 2-4 weeks and subsequently stepwise reduction) group enrolled 49 patients, dexamethasone [(one course of high-dose dexamethasone (HDD) 40 mg/day, d1-4] 45 patients, and rituximab plus HDD (rituximab 100 mg on days 7, 14, 21, 28 and HDD) group 44 patients. Peripheral blood was taken in ITP patients of each group before treatment, 14 d and 28 d after treatment. The percentages of CD4⁺CD25(high)CD127(low) Tregs in 30 healthy controls and 138 patients were analyzed by flow cytometry.

RESULTSOverall response (OR) rates of PSL, HDD and R+HDD groups at day 28 were 69.4%, 66.7% and 79.5% respectively with no difference. After the following 12 months, sustained response (SR) was more pronounced in R+HDD group compared to the other two groups (R+HDD vs PSL: 66.7% vs 37.8%, P<0.05; R+HDD vs HDD: 66.7% vs 22.7%, P<0.05). The percentage of CD4⁺CD25(high)CD127(low) Tregs in peripheral blood of ITP patients [(1.67±0.70)%] was significantly lower than in healthy control group; After treatment, the percentages of Tregs in peripheral blood of patients both at day 14 and 28 in R+HDD group remarkably decreased compared with before treatment [(4.28±1.09)% vs (1.68±0.68)%, P<0.05; (4.44±0.63)% vs (1.68±0.68)%]. The percentages of Tregs at day 14 in both other two groups decreased notably compared with before treatment. But the Tregs levels measured at day 28 in PSL and HDD groups were similar with before treatment.

CONCLUSIONThe percentage of CD4⁺CD25(high)CD127(low) Tregs in peripheral blood of ITP patients was lower than healthy individual. The higher SR of patients treated by R+HDD was related to its ability to up-regulate the percentage of CD4⁺CD25(high)CD127(low) Tregs.

Adolescent ; Adult ; Aged ; Antibodies, Monoclonal, Murine-Derived ; therapeutic use ; Dexamethasone ; therapeutic use ; Female ; Humans ; Male ; Middle Aged ; Prednisolone ; therapeutic use ; Rituximab ; T-Lymphocytes, Regulatory ; Thrombocytopenia ; blood ; drug therapy ; immunology ; Young Adult

This study was aimed to investigate the effect of dexamethasone (Dex) on immunosuppressive ability of mesenchymal stem cells (MSC) during expansion and differentiation of MSC. MSC were cultured in 96-well flat-bottom plates. Proliferation assays were performed by using the BrdU colorimetric ELISA Kit. To explore the effect of Dex on MSC immunosuppressive ability, MSC were firstly cultured in complete culture medium for 14 d with Dex (10 nmol/L), and then, peripheral blood mononuclear cells (PBMNC) were co-cultured with MSC in 96-well flat-bottom plates for 3 d. Phytohemagglutinin A (PHA, 10 µg/ml) was used to stimulate activation of PBMNC. The concentrations of IFN-γ in culture supernatants was detected by ELISA. The results indicated that there was no obvious difference in representative phenotypes of MSC between experimental and control groups after MSC were treated with low concentration of Dex (10 nmol/L) for 14 d, but the suppression of Dex-treated MSC on lymphocyte activation in same concentration of cells was significantly reduced as compared with control group. After the Dex-treated MSC were co-cultured with IFN-γ for 12 h, the immunoregulatory ability of MSC was recovered in a certain degree. It is concluded that the Dex impairs the immunosuppressive ability of MSC, the IFN-γ can protect and reverse the immunosuppressive ability of MSC impaired by Dex, so that, when the immunoregulatory activity of MSC is investigated, it is necessary to avoid adding Dex in the culture medium.
Cells, Cultured ; Dexamethasone ; adverse effects ; Humans ; Immune Tolerance ; drug effects ; Interferon-gamma ; immunology ; Leukocytes, Mononuclear ; Lymphocyte Activation ; immunology ; Mesenchymal Stromal Cells ; cytology ; drug effects ; immunology

This study was purposed to investigate the effect of high-dose dexamethasone (DXM) on function and Toll like receptor 9 (TLR-9) expression of plasmacytoid dendritic cells (pDC) in peripheral blood of patients with immune thrombocytopenic purpura (ITP). 15 newly diagnosed patients with ITP received high dose DXM at single daily doses of 40 mg for 4 consecutive days. The peripheral blood plasmacytoid dendritic cells from 13 remission patients and 15 normal controls were separated by immunomagnetic beads and then induced by CpG-OND2216. 24 h later, the levels of IFN-α, IL-6 and TNF-α in the supernatant were detected by enzyme linked immunosorbent assay (ELISA). The expression of TLR9 mRNA of pDC was detected by real-time quantitative PCR. The results indicated that the levels of IFN-α, IL-6 and TNF-α produced by pDC in ITP patients were significantly higher than those in normal controls (P < 0.05). After high dose DXM treatment, the levels of IFN-α, IL-6 and TNF-α decreased without significant difference compared with normal controls (P > 0.05). The expression of TLR9 mRNA in pDC of untreated patients was significantly higher than that in control group (P < 0.05), and significantly reduced after treatment without difference from that in control group (P > 0.05). It is concluded that pDC may play an important role in ITP by their TLR9 and secreted cytokines; dexamethasone may down regulate the expression of TLR9, inhibit pDC function, and thus play a therapeutic role.
Adolescent ; Adult ; Case-Control Studies ; Dendritic Cells ; immunology ; metabolism ; Dexamethasone ; administration & dosage ; therapeutic use ; Female ; Humans ; Male ; Middle Aged ; Purpura, Thrombocytopenic, Idiopathic ; blood ; drug therapy ; immunology ; RNA, Messenger ; genetics ; Toll-Like Receptor 9 ; genetics ; metabolism ; Young Adult

OBJECTIVETo study the anti-inflammatory effect of the combination of Epimedii Folium, Ligustri Lucidi Fructus and dexamethasone on asthma and its effect in inhibiting adverse reaction against hormone.

METHODDexamethasone had been injected intraperitoneally into asthmatic model rats for 2 weeks, together with the oral administration of Epimedii Folium and Ligustri Lucidi Fructus. The contents of IL-4, INF-gamma, IL-5 and GCR in BALF, serum COR, plasma ACTH and hypothalamic CRH were observed.

RESULTThe combination of Epimedii Folium, Ligustri Lucidi Fructus and dexamethasone can significantly decrease the contents of IL-5 and IL-4 in BALF and ACTH and CRH content in plasma, increase the content of IFN-gamma and GCR in BALF and balance Th1/Th2.

CONCLUSIONThe combination of Epimedii Folium, Ligustri Lucidi Fructus and dexamethasone has a better anti-inflammatory effect on asthmatic model rats, and play a protective role in the pathway of endogenous dexamethasone.

Animals ; Asthma ; drug therapy ; immunology ; Bronchoalveolar Lavage Fluid ; immunology ; Cytokines ; analysis ; Dexamethasone ; administration & dosage ; Drug Therapy, Combination ; Drugs, Chinese Herbal ; administration & dosage ; Ligustrum ; Male ; Rats ; Rats, Sprague-Dawley ; Th1 Cells ; immunology ; Th2 Cells ; immunology

Carboxylesterases (CESs) play important roles in the metabolism of endogenous and foreign compounds in physiological and pharmacological responses. The aim of this study was to investigate the effect of dexamethasone at different doses on the expression of CES1 and CES2. Imidapril and irinotecan hydrochloride (CPT-11) were used as special substrates for CES1 and CES2, respectively. Rat hepatocytes were cultured and treated with different concentrations of dexamethasone. The hydrolytic activity of CES1 and CES2 was tested by incubation experiment and their expression was quantitated by real-time PCR. A pharmacokinetic study was conducted in SD rats to further evaluate the effect of dexamethasone on CESs activity in vivo. Western blotting was performed to investigate the regulatory mechanism related to pregnane X receptor (PXR) and glucocorticoid receptor (GR). The results showed that exposure of cultured rat hepatocytes to nanomolar dexamethasone inhibited the imidapril hydrolase activity, which was slightly elevated by micromolar dexamethasone. For CES2, CPT-11 hydrolase activity was induced only when dexamethasone reached micromolar levels. The real-time PCR demonstrated that CES1 mRNA was markedly decreased by nanomolar dexamethasone and increased by micromolar dexamethasone, whereas CES2 mRNA was significantly increased by micromolar dexamethasone. The results of a complementary animal study showed that the concurrent administration of dexamethasone significantly increased the plasma concentration of the metabolite of imidapril while the ratio of CPT-11 to its metabolite SN-38 was significantly decreased. PXR protein was gradually increased by serial concentrations of dexamethasone. However, only nanomolar dexamethasone elevated the level of GR protein. The different concentrations of dexamethasone required suggested that suppression of CES1 may be mediated by GR whereas the induction of CES2 may result from the role of PXR. It was concluded that dexamethasone at different concentrations can differentially regulate CES1 and CES2.
Animals ; Carboxylic Ester Hydrolases ; genetics ; Dexamethasone ; pharmacology ; Gene Expression ; drug effects ; immunology ; Male ; Rats ; Rats, Sprague-Dawley ; Receptors, Cytoplasmic and Nuclear ; metabolism

OBJECTIVE: To determine the role of miR-155 in the pathogenesis of generalized myasthenia gravis (GMG) and the effect of dexamethasone (DXM) on miR-155. METHODS: The expression of miR-155 in B cells from the GMG patients and healthy controls was analyzed by qPCR. The B cells were cultured with DXM and PBS. The B cell proliferation was examined by MTT; CD80 and CD86 frequencies were detected by flow cytometry; and anti-AChRIgG and isotypes anti-AChR-IgG1, 2, 3 in the supernatant were detected by ELISA. RESULTS: qPCR revealed that the expression of miR-155 in the B cells was much higher than that in the controls, and the miR155 expression decreased after DXM treatment. flow cytometry showed that there was no significant difference in the proliferation and the expressions of CD80 and CD86 in the B cells between the DXM group and the PBS group. The concentration of anti-AChR-IgG1 was obviously lower in the DXM group than in the PBS group, but the concentration of anti-AChRIgG, anti-AChR-IgG2, and anti-AchR-IgG3 was similar. CONCLUSION high expression of miR-155 may be associated with myasthenia gravis progression. DXM may disturb the antibody class switch of B cells by suppressing the expression of miR-155 and improve the symptom of MG patients.
Adult ; B-Lymphocytes ; cytology ; immunology ; metabolism ; B7-1 Antigen ; metabolism ; Cell Proliferation ; Cells, Cultured ; Dexamethasone ; therapeutic use ; Female ; Humans ; Immunoglobulin G ; immunology ; Male ; MicroRNAs ; genetics ; metabolism ; Middle Aged ; Myasthenia Gravis ; drug therapy ; genetics ; immunology ; Receptors, Cholinergic ; immunology ; Tetraspanin 28 ; metabolism ; Young Adult

To develop a HBV infection mouse model by hydrodynamic-based transfection and further to optimize the method of development of HBV infection mouse model. We first developed a construct which contained inverted terminal repeat elements (ITR) of adeno-associated virus (AAV) and 1. 3 copies of HBV genome (ayw subtype). The pAAV-HBV1. 3 DNA was then injected hydrodynamically into the tail veins of C57BL/6 mice in 5 seconds. The virus load in serum and liver was assayed by ELISA and Real-time PCR. The expression of virus antigen and the pathologic changes of liver were analyzed by HE and immunohistochemical staining. Meanwhile, to develop HBV transfected immunosuppressied mouse, mice were injected intraperitoneally triple with 0.2 ml dexamethason (50 mg/kg) every two days before HBV transfection. The levels of HBsAg and HBeAg were assayed by ELISA. Our data showed: (1) HBsAg and HBeAg were positive (100%) in serum and liver of experimental normal mouse at day 10 after HBV transfection, and became negative at day 30 and day 60. Meanwhile the viral load in serum and liver in experimental group was significantly higher than that in control group at day 10, 30 and 60 after HBV transfection (P < 0.01, P < 0.05, respectively). (2) HBsAg and HBeAg in serum in immunosuppressed mouse model were positive until 60 days. In conclusion, a HBV infection mouse model was developed successfully by hydrodynamic-based transfection. By suppressing the immune status of mice injected with dexamethasone, the expression of HBV antigens was extended longer than that in normal adult mice. These models pave a way for HBV research and evaluation of HBV vaccine and drug development.
Animals ; Dependovirus ; genetics ; metabolism ; Dexamethasone ; administration & dosage ; immunology ; Disease Models, Animal ; Female ; Gene Expression Regulation, Viral ; drug effects ; Genetic Vectors ; genetics ; metabolism ; Hepatitis B ; immunology ; virology ; Hepatitis B Antigens ; genetics ; metabolism ; Hepatitis B virus ; drug effects ; genetics ; physiology ; Humans ; Immunosuppressive Agents ; administration & dosage ; immunology ; Liver ; immunology ; virology ; Mice ; Mice, Inbred C57BL ; Transfection ; methods

OBJECTIVETo investigate the change of B-cell activating factor of the TNF family (BAFF) and regulatory T-cells (Tregs) before and after high-dose dexamethasone(HD-DXM) therapy and assess the effect of BAFF on Treg cells in immune thrombocytopenic purpura (ITP).

METHODSThe plasma BAFF concentration was measured by ELISA, and Treg cell numbers by flow cytometry.

RESULTSThe plasma BAFF level \[(599.70 +/- 199.40) pg/ml\] was significantly increased (P < 0.05), and the percentage of Treg cells \[(1.56 +/- 0.73)%\] was significantly decreased (P < 0.01) in ITP patients before treatment as compared with that in controls \[(454.5 +/- 132.5) pg/ml and (4.08 +/- 1.08)%, respectively\]. After treatment with HD-DXM, the plasma BAFF level \[(296.9 +/- 119.7) pg/ml\] was significantly decreased (P < 0.01), and the percentage of Treg cells \[(5.94 +/- 2.22)%\] was significantly increased (P < 0.01). The BAFF level and Treg proportion had no significant correlation with platelets count (P > 0.05). In in vitro assays, no difference was found in the number of Treg cells between rhBAFF0 group and rhBAFF20 group \[(1.53 +/- 0.69)%, (1.49 +/- 0.67)%, P = 0.89)\].

CONCLUSIONBAFF level was increased and Treg cells decreased in ITP patients. HD-DXM might play a role in ITP treatment by down-regulating BAFF expression and up-regulating Treg cells number. BAFF had no influence on the number of Treg cells.

B-Cell Activating Factor ; Dexamethasone ; administration & dosage ; Humans ; Interleukin-4 ; Purpura, Thrombocytopenic, Idiopathic ; immunology ; T-Lymphocytes, Regulatory ; immunology