Changes of Regulatory T Cells in Graves' Disease

The immune mechanism of Graves' diseases (GD) and the roles of regulator T cells were investigated. In 32 patients with GD (GD group) and 20 healthy volunteers (control group), flow cy-tometry was used to detect the proportion of CD4 CD25 cells, MACS to isolate CD4 CD25 cells, RT-PCR to assay the expression of FOXP3, and ELISA to test the level of IL-10, respectively. It was found that there was no significant change in the proportion of CD4 CD25 T cells between GD group and control group (P>0.05), while secretion of IL-10 and expression of FOXP3 in GD group were lower than control group (P<0.01 and P<0.05, respectively). In conclusion, though the proportion of regulatory T cells of peripheral blood lymphocytes in the patients with GD, the functions of them were significantly weakened, which might be a pathogenic factor in GD.     

Involvement of CD4^＋CD25^＋ regulatory T cells in the pathogenesis of polycythaemia vera

Background Regulatory T cells （mreg） have been shown to play an important role in the regulation of hematopoietic activity. However, there is no information about the effect of Treg cells in the pathogenesis of polycythaemia vera （PV）. Methods In this study, we investigated the percentage and function of Treg cells in the peripheral blood of 21 PV patients and 25 healthy donors, mreg cells were identified and characterized as CD4^＋CD25^＋FOXP3^＋ by flow cytometry. The suppressive activity of CD4^＋CD25^＋ Treg cells was assessed by the proliferation and cytokine secretion of the co-cultured CD4^＋CD25^- fractions. Results The results showed that the percentage of Treg cells in the peripheral blood of PV patients significantly increased compared to healthy controls （（10.93±4.02）% vs （5.86±1.99）%, P 〈0.05）. Moreover, the mRNA and protein expression of FOXP3 was higher in CD4^＋CD25^＋ Treg cells. Coordinately, when co-cultured with the activated CD4^＋CD25^- cells, the CD4^＋CD25^＋ Treg cells showed enhanced suppressive function in PV. Yet, the underlying mechanism for the increased frequency and function of CD4^＋CD25^＋ Treg cells is still to be clarified. Conclusion Treg cells expansion might account for the abnormal T cell immunity in PV patients and thus contribute to the pathogenesis of PV.     

Role of interleukin-10 and transforming growth factor beta 1 in otitis media with effusion

Background Otitis media with effusion （OME） is a disease with complicated pathogeneses which are not clearly known Increasing interest has been focused on immunological cells, cytokines and their roles in chronic inflammatory states. This study was designed to disclose the existence and roles of interleukin-10 （IL-10） and transforming growth factor beta1 （TGF-β1） in the cause of OME in adults, and to investigate the probable role of Foxp3^＋CD4^＋CD25^＋ T cells in OME. Methods The concentrations of IL-10 and TGF-β1 in the middle ear effusions （MEEs） and plasmas of 36 adults （45 ears） with OME were measured by means of enzyme linked immunosorbent assay （ELISA）. As contrast, the concentrations of IL-10 and TGF-131 in the plasma of 30 normal volunteers were measured using the same method. Furthermore, the proportion of Foxp3^＋CD4^＋CD25^＋ T cells in CD4^＋ T cells of blood was tested by flow cytometry. Results （1） The concentrations of IL-10 in all MEEs and plasmas of the chronic OME patients were higher than those in patients with acute OME （both P 〈0.05）, so was TGF-131 （both P 〈0.01）. The concentration of IL-10 in MEEs was significantly higher than that in plasmas, not only in acute OME （P〈0.01）, but also in chronic OME （P〈0.01）. In chronic OME, the concentration of TGF-β1 in MEEs had no statistical difference with those in plasmas of the same patients. However, the concentration of TGF-β1 in plasmas of patients with chronic OME was significantly higher than that in plasmas of normal volunteers （P 〈0.01）. （2） The concentrations of IL-10 and TGF-β1 in MEEs of the patients who had been treated more than once were higher than those MEEs of the patients who were treated for the first time, respectively （P〈0.05, P〈0.01）. The level of TGF-β1 in plasmas of the patients who had been treated more than once was higher than in those of the patients who were treated firstly （P 〈0.05）, while the level of IL-10 in plasmas had no difference. The concentration of IL-10 in mucoid MEEs was higher than those in serous ones （P〈0.05）, while TGF-β1 had no statistical difference between mucoid and serous MEEs （P〉0.05）. The concentration of IL-10 in MEEs had a strong correlation with the duration of the illness （r=0.547, P〈0.01）. The same correlation was also found between the concentration of TGF-β1 in MEEs and the times patients being treated （r=0.579, P 〈0.01）. （3） The proportion of Foxp3^＋CD4^＋CD25^＋T/CD4^＋ T cells in the blood of chronic OME was not only significantly higher than that in the acute OME （P〈0.01）, but also higher than that in normal volunteers （P 〈0.01）. In chronic OME, there was a correlation between the proportion of Foxp3^＋CD4^＋CD25^＋ T/CD4^＋ T cells in the blood and the concentration of IL-10 in the plasmas （r=0.602, P 〈0.05）. Conclusions IL-10 and TGF-β1, as two important immunoregulatory mediators, participate in middle ear inflammatory response, especially in chronic course of OME in adults. Foxp3^＋CD4^＋CD25^＋ T cells may play some immunoregulatory roles in the course of this disease.     

Changes of CD4＋ CD25＋ Regulatory T Cells in Peripheral Blood in Patients with Hepatocellular Carcinoma Before and after TACE

This study investigated the changes of CD4＋ CD25＋ regulatory T cells （Tregs） in periph-eral blood of patients with hepatocellular carcinoma before and after transcatheter arterial chemoem-bolization （TACE）. The proportion of CD4＋ CD25＋ Tregs among CD4＋ T lymphocytes in peripheral blood of 33 patients with hepatocellular carcinoma was determined by flow cytometry before, 1 week and 1 month after TACE. And 25 healthy volunteers served as control. One month after TACE, the patients were divided into two groups： 22 in group A, who were in stable condition or getting better; and 10 in group B, who were deteriorating. One patient died and was excluded. The results showed that the percentage of CD4＋CD25＋ Tregs among CD4＋ T lymphocytes did not significantly change in the 33 patients 1 week after TACE as compared with that before TACE, however, the difference was significant （P〈0.01） between the patients with hepatocellular carcinoma and the healthy subjects. The percentage of CD4＋ CD25＋ Tregs among CD4＋ T lymphocytes in group A 1 month after TACE was decreased significantly in comparison with that before and 1 week after TACE （P〈0.01）, whereas, that in group B was increased significantly 1 month after TACE （P〈0.01）. It was concluded that patients with hepatocellular carcinoma had a higher proportion of CD4＋CD25＋ Tregs in peripheral blood. TACE did not significantly affect the level of CD4＋ CD25＋ Tregs within short time （such as 1 week）. The proportion of CD4＋CD25＋ Tregs in peripheral blood 1 month after TACE was related to the prognosis of hepatocellular carcinoma.     

Correlation between elevated FOXP3 expression and increased lymph node metastasis of gastric cancer

Background FOXP3 was thought to express in the T-cell lineage exclusively until recently when FOXP3 was shown to be expressed by cancer cells. It was indicated that FOXP3 may play a wider role in biology by endowing tumor cells with immune suppressive activity. However, researches between FOXP3 and lymph node metastasis of gastric cancer were relatively infrequent, so the present work was aimed to investigate the relationship between FOXP3 expression and lymph node metastasis in human gastric cancer.Methods A total of 122 gastric cancer patients were enrolled in this study, and gastric tumor specimens and lymph nodes were acquired. Thirty patients who had chronic superficial gastritis diagnosed by gastroscopy contemporaneously in the Peking University People＇s Hospital were chosen randomly as the control group. Immunohistochemistry was performed to evaluate FOXP3 expression. A survival analysis on the 122 patients was then performed. Then, NCI-N87cell lines were used to confirm FOXP3 expression in gastric carcinoma cells. Finally, evaluation of FOXP3 expression in gastric tumor and peritumor tissues in 12 patients were conducted using immunohistochemistry and Western blotting. A X2 test or Fisher＇s exact test （bilateral） was conducted to compare the percentage of positive percentage staining between groups. Kaplan-Meier analysis was performed for survival analysis.Results FOXP3 was expressed by gastric cancer cells and peritumor epithelial cells. FOXP3 expression was increased in primary tumors （58.2%） than that in control group （26.7%）. In the lymph-node metastasis group, the incidence of lymph node metastasis which was less than 60% had a significant upregulation of FOXP3 in primary tumors and lymph nodes.However, the frequency of FOXP3 expression had no relationship with survival.Conclusion FOXP3 probably has a relationship with lymph node metastasis of gastric cancer.     

Assessment of the number and function of macrophages in the placenta of gestational diabetes mellitus patients

In order to assess the number and function of macrophages in the placenta of pregnancy complicated with gestational diabetes mellitus （GDM） as well as those of normal pregnancies, placenta samples were collected from 15 GDM patients （GDM group） and 10 normal pregnant women （control group）. The expression levels of macrophage markers （CD68/CD14） and inflammatory cytokines （IL-6/TNF-α） in placenta were detected using immunohistochemistry and PCR. The results showed that the number of CD68＋ or CD14＋ cells in the GMD group was remarkably higher than that in the control group （P〈0.05）, indicating that the number of macrophages in the GDM group was significantly greater than that in the control group. The mRNA expression levels of CD68＋, IL-6 and TNF-α were higher in the GMD group than in the control group. In conclusion, more macrophages accumulate in placenta of pregnancy complicated with GDM, and the expression levels of pro-inflammation factors are also in- creased in GDM pregnancies, suggesting that macrophages and inflammatory mediators （IL-6 and TNF-α） mav olav an imoortant role in GDM.     

Study on clinical effect and mechanism of Jianpi Qingre Huayu Recipe (健脾清热化瘀方) in treating patients with gastric ulcer

Objective： To study the effects of Jianpi Qingre Huayu Recipe （健脾清热化瘀方, JQH） in curing gastric ulcer and to preliminarily probe into its pathogenic mechanism, Methods： Fifty patients with gastric ulcer of Pi （脾）-insufficiency and stasis-heat syndrome type were assigned to the treated group （30 patients） and the control group （20 patients）. They were treated respectively with JQH and Ranitidine. At the same time, another group consisting of 20 healthy persons was set up for normal control. The clinical effect on gastroscopic figure and traditional Chinese medicine （TCM） syndrome were observed. Changes of T-cell subsets and interleukin-8 （IL-8） in serum as well as IL-8 in mucosa around the gastric ulcer were determined before and after treatment by flow cytometry and ELISA. Results： Comparison of the total effective rate on gastroscopic figure in the treated group and the control group （86.7% vs 80.0%） showed insignificant difference, but the cure rate and markedly effective rate in the former （50.0% and 20.0%） was higher than that in the latter （40.0% and 15.0%） respectively. Comparison of the total effective rate on TCM syndrome in the treated group and in the control group （96.7% vs 70.0%） showed insignificant difference, but the cure rate and markedly effective rate in the former （63.3% and 23.3%） was higher than that in the latter （50.0% and 20.0%） respectively. Serum levels of CD3^＋, CD4^＋, CD8^＋ got restored to normal range in the treated group after treatment but it was not so in the control group. IL-8 level in gastric mucosa was improved in both groups but the improvement in the treated group was better. Cenclusien： JQH could effectively treat gastric ulcer and partly reduce its recurrence through improving patients＇ immune function.     

Influence of Danshen Injection on airway inflammation and CD4^＋CD25^＋ regulatory T cells of asthmatic rats

Objective： To investigate the influence of Danshen Injection on airway inflammation and CD4^＋CD25^＋ regulatory T cells（CD4^＋CD25^＋ Tr） of asthmatic rats, and elucidate the possible mechanism of Danshen Injection in treatment of asthma. Methods： 30 Wister rats were randomly divided into control group, asthma group and Danshen Injection treated group. Bronchoalveolar lavage fluids （BALF） were collected, and cytology studies were conducted. Lung tissues were obtained and pathologic analyses were done with hematoxylin and eosin stain （HE）. Flow cytometry was used to detect the CD4^＋CD25^＋ Tr ratio in peripheral blood mononuclear cells （PBMCs）. Results： Total cell, the percentage of lymphocytes, neutrophils and eosinophils （Eos） in BALF of Danshen Injection-treated group were lower than that in asthma group （P〈0.05, P〈0.01）. Compared with asthma group, less infiltration of inflammatory cells in lung tissues was observed in Danshen Injection-treated group. CD4^＋CD25^＋ Tr of asthma group was lower than that of control and Danshen Injection treated group （P〈0.05）. Conclusion： Danshen Injection can suppress airway inflammation of asthmatic rats, probably by increasing the number of CD4^＋CD25^＋ Tr.     

特发性血小板减少性紫瘢患者外周血淋巴细胞亚群和调节性T细胞的变化及临床意义

 Objective To study the changes and clinical significance of lymphocyte subsets and regulatory T cells in peripheral blood of patients with idiopathic thrombocytopenic purpura （PITP）. Methods Peripheral blood lymphocyte subsets and regulatory T cells of 40 ITP patients and 40 normal controls were measured with flow cytometry. Results Compared with the normal group,the percentage of CD3+ T lymphocyte,CD4+T lymphocyte and CD16+ /56+ NK cell and ratio of CD4+ /CD8+ decreased markedly in ITP group （P < 0.01）,while the percentage of CD8+T lymphocyte increased slightly（P > 0.05）and CD19+B lymphocyte increased significantly（P < 0.01）. In ITP patients,the proportion of CD4+ CD25+ T cells in CD4+T cells was higher than that of the control group （P < 0.05）and the proportion of CD4+CD25high T cells was slightly higher（P > 0.05）,while the proportion of CD4+ CD25+ Foxp3+ regulatory T cells was lower（P < 0.05）. Con-clusion Cellular immune function in patients with ITP is obviously abnormal and the decreased proportion of CD4+CD25+ Foxp3+ regulatory T cells may be related to the immune pathogenesis of ITP. Detection of lymphocyte subsets and regulatory T cells is of some clinical value in evaluation of therapeutic effect and prognosis of ITP.     

Analysis of Epstein-Barr viral DNA load,EBV-LMP2 specific cytotoxic T-lymphocytes and levels of CD4^＋CD25^＋ T ceils in patients with nasopharyngeal carcinomas positive for IgA antibody to EBV viral capsid antigen

Background Epstein-Barr virus （EBV） is a herpesvirus commonly associated with several malignant diseases including nasopharyngeal carcinoma （NPC）, which is a common cancer in Southeastern Asia. Previous studies showed that plasma levels of EBV-DNA might be a sensitive and reliable biomarker for the diagnosis, staging and evaluating of therapy for NPC. There are a few analyses of the levels of EBV-latent membrane protein 2 （LMP2）-specific cytotoxic T-lymphocytes （CTLs） in patients with NPC. This study was conducted to investigate the levels of EBV-LMP2-specific CTLs, EBV-DNA load and the level of CD4^＋CD25^＋T cells in such patients.
Methods From February 2006 to April 2006, 62 patients with NPC, 40 healthy virus carriers positive for EBV viral capsid antigen （EBV-IgA-VCA） and 40 controls were enrolled in the study. We used a highly sensitive ELISPOT assay, real-time polymerase chain reaction （PCR） and flow cytometry to measure the EBV-LMP2-specific CTL response, the EBV DNA load and the level of CD4^＋CD25^＋T cells, respectively.
Results The EBV-LMP2-specific CTL responses of the samples from the control, healthy virus carriers and patients with NPC were significantly different from the LMP2 epitopes, with the control and healthy virus carrier samples displaying a stronger response in three cases. There were significant differences in EBV DNA load in serum between NPC and the healthy groups; patients with NPC at stages Ⅲ or Ⅳ had significantly higher viral loads compared with those at stages Ⅰ or Ⅱ. A significantly higher percentage of CD4^＋CD25^＋ T lymphocytes were detected in the patients, compared with healthy virus carriers and healthy controls. Moreover, patients with advanced stages of NPC （Ⅲ and Ⅳ） had significantly higher percentages than the patients with early stages （Ⅰ and Ⅱ）. Conclusions Patients with NPC are frequently unable to establish or maintain sufficient immunosurveillance to control proliferating B cells harboring EBV and to destroy the tumor cells that express immunodominant LMP2 proteins.
Controlling the activity of CD4^＋CD25^＋T cells and elevating CD8^＋ cells specific for LMP2 epitopes could be an effective immunotherapy for patients with NPC.     

Expression of surface markers on peripheral CD4＋CD25^high T cells in patients with atopic asthma： role of inhaled corticosteroid

Background CD4^＋CD25^＋ regulatory T cells （Tregs） mediate immune suppression through cell-cell contact with surface molecules, particularly cytotoxic T lymphocyte-associated antigen 4 （CTLA-4）, glucocorticoid-induced tumor necrosis factor receptor family-related protein （GITR）, and transforming growth factor β （TGF-β）, but little is known about the exact role of Tregs in the pathogenesis of asthma. This study sought to characterize the expression of surface markers on peripheral blood mononuclear cells-derived Tregs in patients with atopic asthma and healthy subjects, and to investigate the effect of inhaled corticosteroid on them.
Methods The expression of surface molecules on CD4^＋CD25^high Tregs was detected by flow cytometry. The effect of inhaled corticosteroid on expression of the surface molecules on Tregs was determined in vivo and in vitro. Total serum immunoglobulin E （IgE） and high-sensitivity C-reactive protein were measured by enzyme linked immunosorbent assay and latex enhanced immunoturbidimetric assay, respectively. Results Equivalent numbers of peripheral Tregs were found in patients with atopic asthma （stable and acute） and healthy subjects. Tregs preferentially expressed CTLA-4, GITR, toll-like receptor 4 （TLR4）, latency-associated peptide （LAP/FGF-β1）, and forkhead box P3 （FOXP3）. Patients with acute asthma had decreased numbers of CD4^＋CD25^highLAP^＋ T cells compared to healthy subjects and stable asthmatics. Inhaled corticosteroid enhanced the percentage of Tregs expressing LAP in vivo and in vitro dose-dependently. Furthermore, the percentages of Tregs expressing LAP were negatively correlated with total serum IgE levels and severity of asthma, but positively correlated with forced expiratory volume in one second percentage of the predicted value in patients with asthma.
Conclusions The results suggest that membrane-bound TGF-β1 is a potential candidate for predicting the severity of asthma, and may contribute to the sustained remission     

Expression of CD123 and CD114 on the bone marrow cells of patients with myelodysplastic syndrome

Background Recent studies have shown that interleukin-3 receptor α （CD123） is highly expressed on leukemia stem cells of patients with acute myeloid leukemia, and is correlated with tumor load and poor prognosis.The expression of CD123 may also be high in patients with myelodysplastic syndrome （MDS）.In this study, the expression and clinical significance of CD123 and granulocyte colony stimulating factor （G-CSF） receptor （CD114） on the bone marrow cells of patients with MDS were investigated to explore the molecular marker of the malignant clone of MDS.Methods Forty-two patients with MDS, who were diagnosed in the Hematological Department of General Hospital of Tianjin Medical University from 2008 to 2009, and twelve normal controls were enrolled in this study.Fluorescence activiated cell sorter （FACS） was used to measure the expression of CD123 on CD34＋CD38- cells and CD114 on CD34＋cells of the bone marrow of these patients and controls and the clinical significance was analyzed.The expression of CD114 on CD123＋CD34＋CD38- cells was further measured to explore the molecular marker of the malignant clone in MDS.Results MDS patients displayed significantly higher proportion of CD34＋CD38-/CD34＋ （（14.03±5.27）%） than normal controls （（7.70±4.36）%, P 〈0.05）.The expression rate of CD123＋CD34＋CD38-/CD34＋CD38- was significantly higher in MDS patients （（48.39±28.15）%） than that in normal controls （（8.75±11.71）%, P 〈0.01）.The expression level of CD123 was significantly correlated with the proportion of bone marrow blasts （r=0.457, P 〈0.05）.The expression rate of CD114＋CD34＋/CD34＋ was lower in MDS patients （（33.05±21.71）%） than that in normal controls （（38.99±19.07）%） but was not statistically significant （P 〉0.05）.The expression of CD114 on CD123＋CD34＋CD38- cells （（34.82±29.58）%） was significantly lower than that on CD123-CD34＋CD38- cells （（53.48±27.41）%） of M DS patients （P 〈0.05）.Conclusions MDS patients displayed higher proportion of CD34＋CD38-/CD34＋ than normal controls.CD123 was highly expressed in the bone marrow of the patients with MDS, significantly correlated with the proportion of bone marrow blasts, and thus might be the marker of MDS malignant clone.CD123＋CD34＋CD38- cells exhibited lower expression of G-CSF receptors, which might partly explain why MDS clone responds worse to G-CSF in vitro and in vivo.     

INVESTIGATION OF REGULATORY T CELLS IN PATIENTS WITH NON-SMALL CELL LUNG CANCER

Objective To evaluate the prevalence of CD4 ^＋ CD25^high regulatory T cells （ Treg cells） in the peripheral blood mononuclear cells （PBMC） and tumor-infiltrating lymphocytes （TIL） of patients with non-small cell lung cancer （NSCLC） and to investigate immunosuppression to the progression of cancer. Methods Peripheral blood and tumor tissues were collected from 20 patients with NSCLC at the time of surgery. None of the patients received surgery, radiotherapy, chemotherapy, or other medical interventions before this study. Cancer stages of the patients were Ⅰ-Ⅲ A. Venous blood samples were obtained from 20 health donors. PBMC were isolated from blood samples by differential centrifugation over Ficoll-Hypaque. TILs were isolated from tumors by differential centrifugation over Ficoll-Hypaque and Percoll. Percentage of CD4^＋ CD25^highTr/CD4＋T in PBMC and TIL was assessed by the flow cytometry. Results The percentage of CD4^ ＋ CD25high Tr/ CD4 ^＋T in PBMC [ （4. 87 ± 1.22 ） % ] of NSCLC patients was significantly higher than that in healthy donors [ （ 2.36 ± 0. 72 ） % ] （ P 〈 0.01 ）. The percentage of CD4^＋ CD25^highTr/ CD4^＋ T in PBMC [ （5.40 ± 1.20） % ] of NSCLC patients in stage Ⅱ-Ⅲ A was significantly higher than that in stage Ⅰ [ （3. 87 ± 0. 22 ） % ] （ P 〈 0. 01 ）. The percentage of CD4 ＋ CD25hiShTr/ CD4 ＋ T in TIL[ （ 8. 66 ±0. 76） % ] of NSCLC patients in stage Ⅱ-Ⅲ A was significantly higher than that in stage Ⅰ [ （ 7. 04 ± 0. 80） % ] （ P 〈 0. 01 ）. Conclusion The prevalence of CD4 ^＋ CD25^highTreg cells in PBMC and TIL of NSCLC patients was significantly higher than that in healthy donors. These Treg cells may be preventing appropriate antitumor immune responses. The population of CD4^ ＋ CD25^highTreg cells in PBMC and TILs of NSCLC patients with Ⅱ-Ⅲ A stage was significantly higher than that of NSCLC patients with Ⅰ stage. These Treg cells may facilitate development of tumors.     

Expression of chemokine receptors CCR3, CCR5 and CXCR3 on CD4^＋ T cells in CBA/J×DBA/2 mouse model,selectively induced by IL-4 and IL-10, regulates the embryo resorption rate

Background Chemokines and their receptors have been a research focus in transplantation immunology. Chemokines and their receptors play a role in lymphocyte recruitment and differentiation process. This study aimed to observe whether IL-4 and IL-10 may regulate the expression of chemokine receptors CCR3, CCR5 and CXCR3 on CD4^＋ T cells in CBA/J×DBA/2 mouse model and to explore the role of CCR3, CCR5, CXCR3 in immune tolerance in pregnancy. Methods The mouse model of spontaneous abortion （CBA/J×DBA/2） and the normal pregnant mouse model （CBA/J×BALB/c） were used. CBA/J×DBA/2 mice were injected with IL-4 （CBA/J×DBA/2-IL-4）, IL-4 and IL-10 （CBA/J×DBA/2-IL-4＋IL-10）, or normal saline （CBA/J×DBA/2-NS） as a control. The expression of CCR3, CCR5 and CXCR3 on CD4^＋ T cells from mouse peripheral blood was measured by the double-labelled FCM method, and the embryo resorption rate was also examined. Results The embryo resorption rate in the CBA/J×DBA/2 group without any treatment was significantly higher than that in the CBA/J×BALB/c group （17.9% vs 3.7%, P 〈0.01）. The embryo resorption rate in the CBA/J×DBA/2 group immunized with IL-4 or IL-4 together with IL-10 was significantly decreased, compared with that in the control and NS groups respectively. CCR3 expression on CD4^＋ T cells in the CBA/J×DBA/2 group without any treatment was significantly lower than that in the CBA/J×BALB/c group （0.3738±0.3575 vs 1.2190±0.2772, P 〈0.01）; both CCR5 （3.0900±1.5603 vs 1.2390±0.6361, P〈0.01） and CXCR3 （2.4715±0.9074 vs 0.9200±0.5585, P 〈0.01） expressions on CD4^＋ T cells of the CBA/J×DBA/2 group without any treatment were significantly higher than those of the CBA/J×BALB/c group. Significant up-regulation of CCR3 and down-regulation of CXCR3 were found in the CBA/J×DBA/2 group treated with IL-4 （CCR3： 2.0360±0.6944, CXCR3： 1.3510±0.5263, P〈0.01） or IL-4 and IL-10 （CCR3： 1.8160±1.0947, CXCR3：1.0940±0.7168, P〈0.01）. Because of the CCR5, IL-4 and IL-10 （1.9400±0.8504 vs 3.0900±1.5603, P 〈0.05）, but IL-4 alone （2.5310±1.3595 vs 3.0900±1.5603, P 〉0.05） treatment significantly decreased the expression of CCR5 in CBA/J×DBA/2. Conclusions The abnormal expression of CCR3, CCR5 and CXCR3 on CD4^＋ T cells may play an important role in the pathogenesis of spontaneous abortion. The pregnancy immune tolerance may be induced through selective induction of CCR3, CCR5 and CXCR3 expressions by IL-4 together with IL-10.     

Effect of pretreatment with different concertrations of morphine, fentanyl and tramadol on the differentiation of human helper T cells in vitro

Objective： To investigate and compare the .effects of different concentrations of morphine, fentanyl and tramadol on the differentiation of human adult helper T cells in vitro. Methods： Twenty out-patients without immune disease were selected and their peripheral blood was collected. Then the Whole blood of peripheral blood mononuclear cells （PBMCs） were pretreated with different concentration of morphine, fentanyl and tramadol for 24 h. The level of CD4^＋ IFN-γ^＋ IL-2^＋/CD4^＋ IL-4^＋ IL-10^＋ was analyzed by three-color flow cytometry, and the CD4^＋ CCR5^＋ and CD4^＋ CCR3 ^＋ cells were counted to observe the imbalance of Th2/Th2. Results： The number of Th2 increased significantly and the ratio of Th2/Th2 decreased dramatically compared with the control group, and there was a dose-dependent fashion in all drugs. Conclusion： Morphine, fentanyl and tramadol can direct Th0 cells toward Th2 differentiation, especially morphine and fentanyl.     

Expression of transcription factor Oct4 in bladder cancer cell line T24 and its effects on the biological characteristics of the cells

The expression of octamer binding factor 4 （Oct4） gene in bladder cancer cell line T24 and its effects on the biological characteristics of the cells were investigated. RT-PCR and Western blot were employed to detect the expression of Oct4 in T24 cells. The changes of biological characteristics in T24 cells were analyzed before and after gene-silencing by Boyden chamber and MTT. The results showed that the expression of Oct4 gene was detectable in T24 cells by RT-PCR and Western blot. The expression of Oct4 gene and protein was down-regulated by siRNA, and average number of transwell cells in interference group, negative control group and blank control group was 101.40±54.56, 104.20± 10.03 and 111.00±11.90, respectively. There was significant difference in the proliferation abilit,） of the cells from 48 h, 72 h to 96 h after the interference by siRNA between interference group and negative group or blank control group （P〈0.05）. It was suggested that Oct4 gene was related with proliferation ability of T24 cells, but not with invasive capability.