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Objective To explore the changes in Th1 cell,B cell and related gene expression during the healing of diabetic foot ulcers(DFU).Methods Lesional skin tissues from DFU patients surgically treated in our hospital from January 2022 to January 2023 were selected.DFU associated gene expression data were collected from GSE80178 and GSE143735 datasets.The expressions of genes(DEGs)between ulcer and no-ulcer patients were identified.Bioinformatics methods were used to identify abnormal immune cells and key genes.Finally,the key results were detected by RT-qPCR and flow cytometry.Results A total of 548 common DEGs were identified in two datasets.In 14 co-expression modules,darkgrey and darkgreen were most related to ulcer,which were mainly associated with the regulation of enzyme activity,immune function and endocrine regulation.Flow cytometry showed that Th1 and B cells were highly infiltrated in ulcer tissue and decreased in healing tissue.MMP13,S100A9 and STAT4 were involved in immune signaling pathways.MMP13 and STAT4 were lowly expressed in healed ulcers,whereas S100A9 was highly expressed.Conclusion Reduced levels of Th1 and B cell infiltration may promote DFU healing.
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ObjectiveTo investigate the efficacy of Bushen Shengxue prescription and Yiqi Yangxue prescription in the treatment of chronic aplastic anemia and the effect on T cell subsets and the expression of T-box expressed in T cells （T-bet） and GATA binding protein 3 （GATA3）. MethodA total of 585 patients with chronic aplastic anemia who were treated in 19 hospitals in China from May 2018 to June 2021 were enrolled. With the prospective， double-blind and randomized control methods， the patients were randomized into three groups： kidney deficiency group， Qi and blood deficiency group， and control group. The three groups were respectively treated with Bushen Shengxue prescription granule， Yiqi Yangxue prescription granule， and Placebo （half the dose of Bushen Shengxue formula granules）. In addition， all of them were given oral cyclosporin and androgen. The treatment lasted 6 months， with 3 months as a course. The blood routine indexes， T cell subsets， and fusion genes T-bet and GATA3 before and after treatment were analyzed， and the safety indexes were monitored. ResultDuring the observation， a total of 75 cases dropped out and 18 were rejected. Finally， 161 cases in the kidney deficiency group， 164 in the Qi and blood deficiency group， and 167 in the control group were included. After 6 months of treatment， the total effective rate was 98.8% （159/161） in the kidney deficiency group， which was higher than the 79.9% （131/164） in the Qi and blood deficiency group （χ2=30.135， P<0.01） and the 61.7% （103/167） in the control group （χ2=70.126， P<0.01）. The total effective rate was higher in the Qi and blood deficiency group than in the control group （χ2=13.232， P<0.01）. After treatment， the hemoglobin （HGB） content increased significantly in three groups （P<0.05） as compared with that before treatment， particularly the kidney deficiency group （P<0.01）. After treatment， the white blood cell （WBC） count and platelet （PLT） count in the kidney deficiency group and the control group increased compared with those in the Qi and blood deficiency group （P<0.01）. There was no specific difference in neutrophils （ANC） after treatment among the three groups. At the same time point， the level of T helper type 1 （Th1） cells， Th1/Th2 ratio （P<0.05）， level of CD4+， and CD4+/CD8+ ratio （P<0.05） were significantly low in the kidney deficiency group among three groups. There was no significant difference in CD19-， HLA/DR+， and CD25+ between the kidney deficiency group and the other two groups， but the T-bet of the kidney deficiency group and the control group was lower than that of the Qi and blood deficiency group （P<0.05）. ConclusionBushen Shengxue prescription exerts therapeutic effect on the aplastic anemia by improving the immunoregulatory mechanism， inhibiting the activity of immune system， modulating T cell subsets， suppressing Th1 and CD4+， and promoting bone marrow hematopoiesis. Moreover， it is safe with little side effects， which is worthy of further promotion.
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Objective :To study the effect of glucose on mouse CD4+ T cell differentiation. Methods :Mouse naïve CD4+ T cells cultured in the regulatory T cell (Treg), Th1, Th17 or Th2 differention condition were treated with different concentrations of glucose for 5 days. Treg, Th1, Th17 or Th2 percentages were measured by flow cytometry. Quantitative real-time PCR was used to detect the gene expressions of related cytokines and transcriptional factors. Results :The proportions of Treg and Th2 as well as the gene expressions of transforming growth factor-β, interleukin-4 (IL-4) and IL-13, and transcriptional factors, Foxp3 (forkhead box P3) and Gata3 (GATA binding protein 3), were increased significantly with the treatment of increasing concentration of glucose. On the contrary, with the glucose treatment, the percentages of Th1 and Th17 were reduced, and the gene expressions of the related cytokines and cytokine receptors, such as interferon-γ, IL-17A, IL-17F, IL-22 and IL-23R, and the related transcriptional factors, Tbx21 (T-box transcription factor 21) and RORC (RAR related orphan receptor C), were decreased consistently. Conclusion :Glucose promotes Treg and Th2 differentiation while inhibits Th1 and Th17 differentiation in vitro.
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Objective The concentrations of Th1/Th2/Th17 cytokines were compared in acute and chronic patients with brucellosis in order to understand the immune characteristics of acute and chronic brucellosis. Methods Using forward-looking design, 35 acute and 35 chronic patients with brucellosis were selected before treatment from the Endemic Disease Prevention and Control Center of Wulanchabu City as the research subjects with 25 local healthy persons as the healthy control group from March to June 2018. Enzyme linked immunosorbent assay (ELISA) method was used to determine the serum levels of Th1 cell cytokines interferon-γ(IFN-γ), tumor necrosis factor-α(TNF-α), and interleukin-12 (IL-12); Th2 cell cytokines interleukin-4 (IL-4), interleukin-6 (IL-6), and interleukin-10 (IL-10);and Th17 cell cytokines interleukin-17 (IL-17). Results There were totally 25 patients in the control group, aged (46.48 ± 9.74) years, 15 males and 10 females;35 patients with acute brucellosis, aged(54.00 ± 7.65) years, 23 males and 12 females; and 35 patients with chronic brucellosis, aged (49.04 ± 8.24) years, 22 males and 13 females. The means of IFN-γ[(280.50 ± 40.48), (462.79 ± 47.94), (431.91 ± 40.39) ng/L], TNF-α[(292.17 ± 31.45), (481.21 ± 43.14), (433.70 ± 41.23) ng/L], IL-12 [(45.13 ± 5.55), (70.74 ± 7.58), (62.56 ± 5.73) ng/L], IL-4 [(383.24 ± 53.98), (606.11 ± 51.86), (550.66 ± 51.56) ng/L], IL-6 [(14.48 ± 2.17), (23.54 ± 2.39), (21.89 ± 2.26) ng/L], IL-10 [(140.48 ± 110.97), (464.49 ± 52.10), (404.73 ± 52.10) ng/L], and IL-17 [(13.71 ± 1.58), (23.79 ± 2.15), (21.37 ± 2.26) ng/L] were significantly different between control group, acute brucellosis group and chronic brucellosis group (F = 126.13, 152.58, 106.81, 139.28, 113.80, 155.86, 152.91, P < 0.05). Compared to control group, the levels of the seven cytokines in acute brucellosis and chronic brucellosis groups were significantly increased before treatment (P < 0.05). The levels of all of the seven cytokines in acute brucellosis group were significantly increased compared to chronic brucellosis group before treatment (P < 0.05). Conclusions There is a difference in the expression of Th1/Th2/Th17 cytokines in patients with acute and chronic brucellosis group. In the acute brucellosis group, the cell immunity induced by Th1 cells and the humoral immunity guided by Th2 cells are stronger than those in chronic brucellosis group.
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Objective:To observe clinical efficacy of Taohua Tang and Buzhong Yiqi Tang on Crohn's disease (CD) at active phase (deficiency-cold in spleen and stomach), in order to observed its effect on Th1 and Th17 cytokines. Method:According to random number table, 86 patients with CD were divided into control group (42 cases) and observation group (44 cases). The control group (mild) was given SASP, 3-4 g·d-1, Po, tid. The control group (moderate or poor efficacy of SASP) was given prednisone acetate, 0.75 mg·kg-1·d-1, Po, tid. Observation group was given Taohua Tang and Buzhong Yiqi Tang in addition to therapy of the control group, 1 dose·d-1. The course of treatment was 12 weeks. Before and after treatment, Best CDAI, SES-CD, IBDQ and deficiency syndrome were scored, and levels of CRP, ESR, ALB, HB, PLT, IFN-γ, TNF-α, IL-2 and IL-17 were measured before and after treatment. Result:After treatment, the effect of traditional Chinese medicine(TCM) syndromes in the observation group was better than that in the control group (Z=2.058, PPPPZ=2.112, PZ=2.288, PPPγ, TNF-α, IL-2 and IL-17 levels in the observation group were lower than those in the control group (PConclusion:In addition to the therapy of conventional western medicine, Taohua Tang and Buzhong Yiqi Tang in treatment of deficiency syndrome of Crohn's disease (CD) can control the activity degree of the disease, reduce the degree of illness and inflammation, and improve the remission rate and the quality of life, with a better clinical efficacy than the pure western medicine therapy.
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Objective@#To investigate the inhibitory effects and mechanism of Triticum aestivum (TA) on anaphylaxis in ovalbumin (OVA)-sensitized mice.@*Methods@#Twenty-five female BALB/c mice were randomly divided into five groups (n=5), including control group, OVA group, 100 mg/kg TA group, 200 mg/kg TA group and DEX (dexamethasone) group. OVA (20 μg) and aluminum hydroxide (1 mg) were dissolved in 100 μl of PBS solution and injected into the abdominal cavity of each experimental mouse, while the mice in the control group were injected with 100 μl of 0.9% normal saline. All mice were given drinking water. The above processes were repeated two weeks later, and different concentrations of experimental drugs were diluted in 1% CMC (carboxymethyl cellulose) solution to feed mice. Allergic symptoms and behaviors of each mouse were scored in the process of feeding. On the 12th day after first sensitization, mice in the control and experimental groups were subcutaneously injected in the auricle with 20 μl of 0.9% saline solution and OVA, respectively. The thickness of each mouse′s auricle was measured 6 and 24 hours later. Histopathologic changes in auricle tissues were observed by hematoxylin-eosin(HE) staining 24 hours later. The mice were euthanized by cervical dislocation in the 6th week after first sensitization. Abdominal aorta serum samples were collected and tested for specific inflammatory cytokines (IgE, IgG1, TNF-α, IL-4) and anti-OVA antibodies (IgE, IgG1) by ELISA. ELISA and RT-PCR were used to detect the expression of IFN-γ, IL-4, IL-5, IL-12 and IL-13 after spleen lymphocytes were stimulated with different concentrations of drugs.@*Results@#Both the score of allergic symptoms and behavior and the thickness of the auricle of the OVA group were the highest among all groups (P<0.05). A TA dose-dependent decrease was found in both of the two parameters, which was confirmed by histopathological analysis. ELISA results showed that the plasma levels of IgE, IgG1, IL-4 and TNF-α in the experimental groups were significantly higher than those in the control group, but all of them were decreased in a TA dose-dependent manner. TA at the high concentration of 200 mg/kg was similar to DEX in inhibiting the expression of IgE and TNF-α. RT-PCR results showed that Th1 cytokines (IFN-y and IL-12) in the experimental groups increased significantly as compared with those in the control group (P<0.05). Expression of IFN-y and IL-12 was not significantly affected by TA at various concentrations, but markedly inhibited by DEX. The levels of Th2 cytokines (IL-4, IL-5 and IL-13) in the experiment groups were significantly higher than those in the control group (P<0.05). Compared with the OVA group, the levels of IL-4, IL-5 and IL-13 were significantly reduced in TA groups, especially in the TA 100 μg/ml group in which the expression of the three cytokines was decreased by about 60%, 18% and 20%, respectively.@*Conclusion@#TA helps to maintain immune balance by selectively inhibiting the expression of Th2 cytokines (IL-4, IL-5, IL-13), IgE and IgG and not influencing Th1 cytokines (IFN-γ and IL-12). It has an anti-allergic effect as it effectively suppresses allergic responses. This research provides both theoretical and experimental basis for further development of novel anti-anaphylactic treatment with TA.
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Objective To explore the role of signal transducer and activator of transcription 3 (STAT3) phosphorylation in the pathogenesis of Behcet's disease (BD),and to investigate the association between STAT3 phosphorylation and disease activity in BD patients.Methods Peripheral blood mononuclear cell (PBMC) was isolated from 15 mL peripheral boood of 10 active BD patients (BD-A),10 BD patients in remission (BD-R) and 10 healthy controls (HC) respectively.The blockade of STAT3 phosphorylation was performed by Stattic.The PBMC was divided into Stattic subgroup (treated with 2.5 μmol/L stattic and 1 640 medium,5 mL) and blank control subgroup (treated with 5 mL 1 640 medium),respectively.The protein levels of phosphorylated STAT3 (pSTAT3) and STAT3 were examined by flow cytometry and Western blot.The protein and mRNA levels of TNF-α,IFN-γ and IL-17 were tested by RT-PCR and ELISA.Two-way ANOVA and Bonferroni post hoc test were used to analyze the results.Results Compared with HC,the BD patients showed higher protein levels of pSTAT3 and STAT3,and higher protein and mRNA levels of TNF-α,IFN-γ and IL-17;compared with blank control subgroup,the protein levels of pSTAT3 and STAT3,and protein and mRNA levels of TNF-α,IFN-γ and IL-17 decreased in Stattic subgroup.In the BD-A group,the protein level of pSTAT3,and protein and mRNA levels of TNF-α,IFN-γ and IL-17 were significantly higher than those in the BD-R group.Conclusions An increased activation of the STAT3 pathway may contribute to the pathogenesis of BD and relate to disease activity in BD patients by inducing TH1 and Th17 cells activation.
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Objective To investigate the effect of doxycycline on the Th1/Th2 cell balance in experimental allergic encepha-lomyelitis(EAE)rats.Methods Forty female Wistar rats were randomly divided into the EAE control group,low,medium and high does DOX treatment groups,10 cases in each group.The onset situation in rats was observed.The IL-4 and IFN-γlevels secreted by peripheral blood mononuclear cells (PBMC)at the peak stage were detected.The levels of IL-1β,IL-10,TNF-αin brain tissue,and the albumin content in cerebrospinal fluid and serum were detected.The QA value was calculated.Results In each DOX group,the clinical symptoms of rats were alleviated compared with the EAE control group.In each DOX group,the PBMC secreting IFN-γlev-el and IFN-γ/IL-4 ratio in the onset peak stage were lower than those in the EAE control group,while the IL-4 level was higher than that in the EAE control group(P 0.05),but the difference between other DOX groups had statistical significance(P <0.01).The IL-1βand TNF-αlevels of brain tissue and QA value during onset peak stage in various doses DOX groups were decreased compared with the EAE control group,while the IL-10 level was increased compared with the EAE control group(P <0.05).With the DOX dose increasing,the levels of IL-1β,TNF-αand QA value in various doses DOX groups became lower,the IL-10 level became high-er,there was statistically significant difference among various doses DOX groups (P <0.05 ).Conclusion DOX can obviously alle-viate the clinical symptoms of EAE rats,its mechanism may be related with that DOX could decrease the level of Th1 cytokine and increase the level of Th2 cytokine,correct the Th1/Th2 cell balance,thus protect the blood brain barrier(BBB).
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Background Mooren 's ulcer is an immune-related corneal inflammatory disease,and its pathogenesis remains below understood.Previous studies showed that the imbalance of T helper cell type 1 (Th1) and Th2 cell play important roles in the development of some autoimmune diseases.Thereby the influence of Th1/Th2 cells on the pathogenesis of Mooren's ulcer is being concerned.Objective This study was to investigate the change of Th1 and Th2 subsets in periphery blood of patients with active Mooren's ulcer.Methods Eleven consecutive patients with active Mooren's ulcer and 8 age-and gener-matched healthy controls were included in Zhongshan Ophthalmic Center,Sun Yat-sen University from January 2012 to July 2013 under the approval of Ethic Committee of this hospital and informed consent of each subject.The peripheral blood samples of all the subjects were obtained separately and periphery blood mononuclear cells (PBMCs)were isolated.The percentages of Th1 and Th2 in the PBMCs were assayed by flow cytometer.The relative expressions of T-bet mRNA,GATA-3 mRNA and signal transducer and activator of transcription 5 (Stat5) mRNA in the PBMCs were examined and compared by real-time fluorescence quantitative PCR (RT-qPCR) Results The percentage of Th1 cells in CD4+ T cells and Th1/Th2 value was 0.21％ (0.11％,0.31％) and 8.01 (4.49,12.01) respectively in the Mooren's ulcer group,which were significantly lower than 0.35％ (0.22％,0.71％)and 23.90 (22.49,33.49)in the normal control group,respectively (Z =-2.01,P =0.04 ; Z =-3.06,P =0.00).However,no significant difference was found in the percentage of Th2 between the two groups (Z=-1.98,P＞0.05).The relative expressions of T-bet mRNA and GATA-3 mRNA in PBMCs were significantly lower in the Mooren's ulcer group than those in the normal control group (Z =-3.47,-3.06,both at P=0.00) ;While the relative expression of Stat5 mRNA in PBMCs was insignificant changed between the two groups (Z =-1.05,P =0.33).Conclusions Th1 and Th2 cells are unbalanced in the active Mooren's ulcer patients.In addition,the down-expression of relevant transcription factors in peripheral blood also is seen in these patients.It is inferred that Th1 and Th2 cells may participate in the progress of Mooren's ulcer.
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Objective To explore the polarization and proliferation of naive CD4+TT cells after preincubation with interleukin-2. Methods The expression of suppressor of cytokine signal (SOCS)-3 was detected by the real-time PCR in the naive T lymphocytes of DO11.10 TCR transgenic and C57BL/6N mice after they were preincubated by 50 U/ml of IL-2. The naive CD4+T T lymphocytes preincubated for 4 h by IL-2 were stimulated with ovalbumin (OVA) and inactivated BALB/c spleen cells, respectively. After 14 d, IL-12Rβ1, IL-12β2 and cytoplasm IL-4 (cyIL-4) were detected by flow cytometry. Results SOCS-3 in the naive CD4+T lymphocytes of DO11. 10 TCR transgenic and C57BL/6N mice reached the peak after preincu- bated by IL-2 for 6 h. The polarization to TH1 and the proliferation ability of naive CD4+ T lymphocytes stimulated with specific antigen and allogeneic antigen were inhibited conspicuously during the time of the peak of SOCS-3. Conclusion The expression of SOCS-3 in naive CD4+T lymphocytes can be upregulated by IL-2. The upregulation of SOCS-3 can suppress the polarization to TH1and the proliferation ability of naive CD4+T lymphocytes stimulated by specific antigen and allogeneic antigen.
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GITR (glucocorticoid-induced TNF receptor) is a recently identified member of the TNF receptor superfamily. The receptor is preferentially expressed on CD4+CD25+ regulatory T cells and GITR signals break the suppressive activity of the subset. In this study, we wanted to reveal the in vivo function of GITR in herpes simplex virus type 1 (HSV-1) infection. A single injection of anti-GITR mAb (DTA-1) immediately after viral infection significantly increased the number of CD4+ and CD8+ T cells expressing CD25, an activation surface marker, and secreting IFN-gamma. We confirmed these in vivo observations by showing ex vivo that re-stimulation of CD4+ or CD8+ T cells with a CD4+ or CD8+ T-cell-specific HSV-1 peptide, respectively, induced a significant elevation in cell proliferation and in IFN-gamma secretion. Our results indicate that GITR signals play a critical role in the T-cell immunity to HSV-1.
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Animals , Female , Mice , Antibodies, Monoclonal/pharmacology , CD4-Positive T-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/immunology , Cell Proliferation , Glucocorticoids/pharmacology , Herpes Simplex/immunology , Herpesvirus 1, Human/pathogenicity , Immunity, Cellular , Interferon-gamma/metabolism , Lymphocyte Activation , Mice, Inbred BALB C , Peptide Fragments/metabolism , Receptors, Interleukin-2/metabolism , Receptors, Nerve Growth Factor/genetics , Receptors, Tumor Necrosis Factor/genetics , T-Lymphocytes/immunologyABSTRACT
Acupuncture could modulate the immune responses and is clinically used for the treatment of some allergicand immunological disorders. But the benefits of the clinical use of acupuncture for the treatment of theimmunoligical diseases have not been fully established. Further, the mechanisms by which acupuncture affects the immune responses have not been clarified. Here, we review the reports about the clinical effects of acupuncture on immunological diseases and about the basic mechanisms of immune modulation by the acupuncture. From these reports and our experimental data, acupuncture could influence the amount of cytokines produced by immunocompetent cells, so we present the hypothetical mechanisms of immunoregulatory action of acupuncture.
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Objective To investigate the TH1/TH2 patterns by calculating the percentage of TH1, TH2 cells, and the TH1/TH2 ratio of peripheral blood from patients with pregnancy-induced hypertension (PIH). Methods The percentage of TH1 and TH2 cells in peripheral blood from 15 normal pregnant controls and 20 PIH patients (including 12 moderate and 8 severe cases) were calculated using flow cytometry for the analysis of both the surface marker CD3 CD8 and intracellular cytokines, interleukin-4 (IL-4), and interferon-? (IFN-?). Results The percentage of TH1 cells and the ratio of TH1/TH2 in PIH patients were significantly higher than those in the normal third-trimester pregnant controls [(20.50?4.02)% vs (12.57?2.18)%, P