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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 explore the expression and clinical significance of immunosuppressive receptor T cell immunoglobulin and immunoreceptor tyrosine-based inhibitory motif domain (TIGIT) on the peripheral blood mononuclear cells (PBMC) in silicosis patients with Mycobacterium tuberculosis infection. Methods:August 2018, a total of 78 patients with silicosis (all were quarry workers in Sanmen County, Zhejiang Province) were enrolled and divided into silicosis combined with active pulmonary tuberculosis group (APTB group), silicosis combined with latent tuberculosis infection group (LTBI group), and simple silicosis with non-tuberculosis infection group (non-TB group). Flow cytometry was used to analyze the expressions of TIGIT, programmed death-1 (PD-1) and transcription factor T-bet on PBMC from patients. Mann-Whitney U test and Pearson correlations analysis were used for statistical analysis. Results:Among the 78 patients, eight were in the APTB group, 24 in the LTBI group, and 46 in the non-TB group. The expressions of PD-1 and TIGIT on CD8 + T cells in the APTB group (29.45%(16.78%) and 65.40%(12.12%), respectively) were significantly higher than those in the LTBI group (17.40%(11.17%) and 48.30%(28.75%), respectively; U=23.500 and 43.500, respectively, P=0.000 8 and 0.020 5, respectively) and non-TB group (15.95%(12.46%) and 45.30%(19.75%), respectively; U=64.000 and 69.000, respectively, P=0.002 3 and 0.003 8, respectively), and the differences were all statistically significant. The expression of TIGIT was positively correlated with PD-1 on CD8 + T cells in silicosis patients ( r=0.434 3, P<0.01). The proportion of PD-1 + TIGIT + CD8 + T cells in the APTB group (19.90%(22.67%)) was significantly higher than those in the non-TB group (11.55%(11.29%), U=76.500, P=0.007 1) and LTBI group (11.55%(10.53%), U=41.000, P=0.015 4), while the proportion of PD-1 -TIGIT -CD8 + T cells in the APTB group (30.60%(12.90%)) was significantly lower than non-TB group (48.90%(18.98%), U=58.000, P=0.001 3) and LTBI group (47.20%(24.59%), U=41.000, P=0.015 4). The differences were all statistically significant. The expression of T-bet on the peripheral blood CD8 + T cells in the APTB group (29.45%(16.78%)) was higher than that in the non-TB group (15.95%(12.46%)) and the LTBI group (17.40%(11.17%)), and the differences were both statistically significant ( U=46.500 and 46.000, respectively, P=0.000 3 and 0.028 3, respectively). The expression of T-bet on CD8 + T cells was positively correlated with TIGIT on CD8 + T cells ( r=0.456 7, P<0.01). The expression of T-bet on PD-1 + TIGIT + CD8 + T cells in the APTB group (65.40%(12.12%)) was higher than those in the LTBI group (48.30%(28.75%), U=23.500, P=0.000 8) and non-TB group (45.30%(19.75%), U=65.000, P=0.002 6), and the differences were both statistically significant. Conclusion:The immunosuppressive receptor PD-1 and TIGIT are highly expressed on CD8 + T cells in silicosis patients with active pulmonary tuberculosis, which indicates CD8 + T cells exhaustion in these population, while the highly co-expression of T-bet suggests the exhausted subsets may have reversed potentiality.
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Objective: To observe the effect of modified Erchentang on GATA-binding protein-3(GATA3) and T-box expressed in T cells(T-bet) in lung tissue of rats with chronic obstructive pulmonary disease (COPD). Method: Seventy SD rats were randomly divided into seven groups, namely normal group, model group, low, medium and high-dose modified Erchentang group(5,10,20 g ·kg-1), Xiaokechuan group(5 g ·kg-1) and Erchentang group(5 g ·kg-1), with 10 in each group. The rat model of COPD was established by smoking combined with intratracheal dripping of lipopolysaccharide (LPS). After successful modeling, the treatment group was given intragastric administration, and the normal group and the model group were given intragastric administration of equal volume of saline. Enzyme-linked immunosorbent assay (ELISA) was used to determine the concentrations of interleukin-10 (IL-10) and interleukin-12 (IL-12) in rat serum. The expressions of GATA3 and T-bet were detected by real-time fluorescence quantitative polymerase chain reaction (Real-time PCR). The expressions of GATA3 and T-bet in lung tissue were detected by immunohistochemistry (IHC). Result: Compared with the control group, the serum levels of IL-10 in the model group was significantly decreased, while the IL-12 level was significantly increased (PPPPConclusion: Modified Erchentang may reduce the inflammation of lung tissue and improve lung function in COPD rats by reducing IL-12, increasing the content of IL-10, inhibiting the protein and gene expressions of T-bet, and stimulating the protein and gene expressions of GATA3.
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Objective This study is aimed to investigate the possible role of Th1/Th2 cell in the pathogenesis of primary gout arthritis. Methods The peripheral blood of 21 acute gout patients (AG), 20 intermittent gout patients (IG) and 20 healthy controls (HC) were collected. The clinical data and laboratory indicators of them were enrolled. The percentages of Th1 and Th2 cells were detected by flow cytometry (FCM). The expression of GATA-3, T-bet, IL-4 and interferon (IFN)-γ mRNA in Peripheral blood mononuclear cells (PBMCs) were measured using Real time quantitative polymerase chain reaction (PCR). The protein expression levels of IL-4 and IFN-γ in serum were detected by enzyme-linked immuno sorbent assay (ELISA). The measurement data were compared by one factor analysis of variance test. The correlation between variables was used by Spearman correlation analysis. Results The percentage of Th1 cells in peripheral blood of the AG group was [(23.2 ±8.3)％], and the IG group was [(20.5 ±9.3)％], which were significantly higher (F=6.520, P<0.05) than the percentage of HC group [(14.8±3.8)％]. There was no significant difference between AG group and IG group. The percentage of Th2 cells in peripheral blood of AG group were [(1.9 ±0.7)％], which was significantly lower (F=8.267, P<0.05) than and the percentage of HC group [(3.4±1.8)％] and IG group [(3.3± 1.2)％]. There was no significant difference between the IG group and the HC group. And the proportion of TH1/Th2 cells in the AG group was higher than that of the IG group and the HC group (F=10.406, P<0.01). The expression of T-bet mRNA and IFN-γ mRNA in the AG group and IG group were higher than that in the HC group (F=4.942, P=0.010)、(F=4.458, P=0.016). However, there was no significant difference between IG group and AG group. The expression of GATA-3 mRNA and IL-4 mRNA was significantly lower (F=3.564, P=0.035) (F=5.385, P=0.007) in the AG group and IG group when compared to the HC group. And there was no significant difference between the AG and the IG group. The IFN-γ level increased in the AG and IG group compared to the HC group (F=7.659, P=0.001). The IL-4 levels in the AG group was lower (F=7.099, P=0.002) than those of the IG and HC group. Conclusion Th1 cells in the peripheral blood of patients with GA are increased and accompanied with the decrease of Th2 cells. The results of this study suggest that the imbalance of Th1/Th2 cells in the inflammatory and immune response plays a critical role in the pathogenesis of GA.
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Objective To investigate the intervention of Chinese herb Shengjiang San(SJS)for the imbal-ances of serum Th1/Th2 cells and related regulatory factors in sepsis patients. Methods Fifty-five sepsis patients were randomly divided into two groups:conventional treatment group of 27 cases and SJS group of 28 cases. Nine cases of healthy volunteers were enrolled as control group in the study. Cases of conventional group were treated with western medicine only and cases of SJS group were treated with both western medicine and Chinese herb SJS (100 mL twice one day). The therapy course of both groups was 3 days. Score of Chinese medical syndromes ,se-rum level of leukocyte count,C reactive protein(CRP)and procalcitonin(PCT),serum T-bet,GATA-3,Th1&Th2 cells in the proportion of whole CD4+Th cells and Th1/Th2 ratio were compared respectively in each group be-fore and after treatment. Results There were significant differences between SJS group and conventional group in score of Chinese medical syndromes,serum level of leukocyte count,T-bet,GATA-3,Th1&Th2 cells in the pro-portion of whole CD4+Th cells and Th1/Th2 ratio(P<0.05 or P<0.01). There were no significant differences be-tween the two groups in serum level of CRP,PCT and GATA3. Compared with control group,there were signifi-cant differences in all indicators except GATA3 of the two groups(P<0.01). Conclusion Chinese herb Shengji-ang San has an effective benefits to Chinese medical syndromes,inflammatory reaction,the imbalance of serum Th1/T2 and related regulatory factors(T-bet&GATA-3)in sepsis patients.
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Aim To explore the protective effects and underlying mechanisms of Liu weiwuling Tablets (LW-WL)in concanavalin A (ConA)induced acute immu-nological liver injury in mice.Methods Mice were randomly divided into control,model,Bicyclol,LW-WL low dose (8 g·kg-1 )and LWWL high dose (16 g ·kg-1 )group.The medicattion was performed once daily for seven consecutive days,then the model of im-munological liver injury was prepared by intravenous injection of ConA (15mg·kg-1)in the tail of mice in each group except for the control group one hour after the last treatment.The pathological changes of liver tissues of mice were evaluated by HE staining with, and the levels of alanine amino transferase (ALT),as-partate aminotransferase (AST),and total bilirubin (TBIL)in serum were analyzed by colorimetric meth-od;the level of interleukin 12 (IL-12 ),interferon-γ(IFN-γ),tumor necrosis factor-α(TNF-α),interleu-kin 4 (IL-4)and interleukin 10 (IL-10)in liver was measured by real-time quantitative polymerase chain reaction (RT-qPCR);the changes of Th1 (IFN-γ) and Th2 (IL-4)cells were observed by flow cytometric (FCM)analysis;the expression of Th1/Th2 transcrip-tion factor T-bet/GATA-3 in liver tissue was detected by Western blot.Results Compared with normal con-trol group,the serum ALT,AST and TBIL were signif-icantly increased in model group, the pathological damage of the liver tissue was severe,and the necrosis and apoptosis of hepatic cells were large, which showed that the model was successful .Compared with model group,both low and high dose of LWWL could significantly reduce ALT,AST,TBIL levels in serum induced by ConA;Th1 cells in the spleen decreased, while Th2 cells increased;the expressions of IL-12, IFN-γand TNF-αmRNA were significantly inhibited with IL-4 and IL-10 mRNA expression elevated in mouse liver tissue;the expression of GATA-3 protein was up-regulated,T-bet protein expression showing no significant changes.Conclusion LWWL could regu-late Th1/Th2 balance,thus inhibiting the acute immu-nity hepatic injury induced by ConA.
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Objective:To discuss the influence of dexamethasone(DXM) to T-bet/GATA-3 of athma rats.Methods: 30 SD rats were randomly divided into normal group,model group and DXM group,10 rats in each group.Ovalbumin was intraperitoneal injected on the 1th and 8th day and aerosol inhaled from the 15th day,once a day,14 days altogether.Dexamethasone was intraperitoneal injected in 0.5 mg/kg from the 15th day,once per day,a total of 14 times.Flow cytometry tested the content of Th1 and Th2 in spleen and peripheral blood,immunohistochemistry and Western blot measured T-bet and GATA-3 protein expression and RT-PCR tested T-bet and GATA-3 mRNA expression.Results: Compared model group with normal group,the contents of Th1,Th2 in spleen and peripheral blood had a very significant increase(P<0.01);expression of T-bet,GATA-3 protein and mRNA in the lung tissue also had very significant risen(P<0.01).While compared DXM group with model group,T-bet/GATA-3 rose significantly(P<0.05)or very significantly(P<0.01).Conclusion: T-bet/GATA-3 is the key transcription factors that influence the balance of Th1/Th2,while DXM can raise the ratio of T-bet/GATA-3,which reduce the severity of asthma.
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Objective To investigate the effects of cardiopulmonary bypass (CPB) on the differentiation of T lymphocyte subsets and the expression of specific transcription regulator T-bet/GATA binding protein 3 (GATA3). Methods A prospective double-blind study was conducted. Patients with CPB pulmonary repair of ventricular septal defect (observation group) or off-pump ligation of ductus arteriosus (control group) with 20 cases each in the 150th Military Hospital from February 2015 to February 2016 were enrolled. The blood sampled was collected on the time of before operation, at the end of CPB or operation, 4 hours after operation, and 24 hours after operation. T lymphocytes were isolated, the helper T cell 1 (Th1) specific transcription factor T-bet mRNA, helper T cell 2 (Th2) specific transcription factor GATA3 mRNA expression and cytokine γ-interferon (IFN-γ) mRNA, interleukin-4 (IL-4) mRNA expression were measured by Northern Blot. Results Compared with before operation, expression levels of T-bet mRNA [integral gray values: (1.39±0.52)×105vs. (2.92±0.88)×105], IFN-γ mRNA [integral gray values: (3.68±0.65)×105vs. (6.10±0.93)×105] were decreased transiently at the end of CPB in the observation group (both P < 0.05), returned to preoperative levels at 24 hours after operation [integral gray values: (2.77±0.74)×105, (6.22±1.25)×105, respectively, both P > 0.05]; expression levels of GATA3 mRNA [integral gray values:(4.96±0.88)×105vs. (3.21±0.68)×105], IL-4 mRNA [integral gray values: (3.52±1.13)×105vs. (1.85±0.63)×105] were increased (both P < 0.05), recovered to the preoperative levels at 24 hours after operation [integral gray values: (3.11±0.51)×105, (1.93±0.84)×105, respectively, both P > 0.05]. There were no significant differences in the expressions of T-bet, GATA3, IFN-γ and IL-4 mRNA in the control group at each time points (all P >0.05). Conclusions CPB causes the imbalance of Th1, Tc1/Th2, Tc2 and pro-inflammatory and anti-inflammatory reactions specially, which participate the complication occurrence after CPB. The changing of T-bet/GATA3 may be the internal mechanism for these changes.
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AIM:To investigate the effect of arsenic trioxide (ATO) on T-bet/GATA3 signal pathway in MRL/lpr mice.METHODS:MRL/lpr mice and C57BL/6J mice at the age of 20 weeks were chosen and then divided in 2 different sub-groups,respectively.The mice in 2 sub-groups received ATO (0.4 mg · kg-1.d-1) and sodium chloride (NS,volume weight-determined) by intraperitoneal injection respectively for 2 months.Afterward,the spleens were isolated fron the MRL/lpr and C57BL/6J mice under pathogen-free condition and the suspensions were prepared.The mRNA level of T-bet,GATA3,IFN-γ,IL-4 and the mRNA ratio of T-bet/GATA3 were detected by RT-qPCR.The protein expression of T-bet and GATA3 was determined by Western blot.The serum levels of IFN-γ and IL-4 were measured by ELISA.RESULTS:The mRNA and protein levels of T-bet,IFN-γand the mRNA ratio of T-bet/GATA3 in NS group of MRL/lpr mice were higher than those in NS group of C57BL/6J mice (P ＜0.05).However,the GATA3 and IL-4 were lower in NS group of MRL/lpr mice in both mRNA and protein level (P ＜ 0.05).In MRL/lpr mice,the mRNA and protein levels of T-bet,IFN-γ and the mRNA ratio of T-bet/GATA3 were lower in ATO group compared with NS group (P ＜ 0.05),no difference was found in GATA3 and IL-4.No difference of the indexes mentioned above between ATO group and NS group in C57BL/6J mice was observed.CONCLUSION:ATO may affect the signaling pathway of T-bet/GATA3 to down-regulate the mRNA expression and the protein secretion of IFN-γ by decreasing the expression of T-bet in MRL/lpr mice.
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AIM To study the improvement of quercetin on Pseudomonas aeruginosa-induced lung infection in rats.METHODS Forty SPF SD rats were randomly divided into five groups,eight rats in each group:normal group,P.aeruginosa infection group,quercetin group,levofloxacin group,levofloxacin combined with quercetin group (combined group),the rats were anesthetized and then injected with P.aeruginosa in bronchus.The pathological changes of lung tissue in rats were observed,the contents of IL-4 and IFN-γcytokines,changes of transcription factors T-bet and Gata-3 in lung tissue were detected,and then semi-quantitative RT-PCR was used for the detection of IL-4,IFN-γ,T-bet and Gata-3 mRNA expressions.RESULTS The content of IL-4,levels of IL-4 and Gata-3 mRNA in lung tissue in the quercetin group,the levofloxacin group and the combined group were lower than those in the P.aeruginosa infection group,but the opposite was true in the levels of IFN-γ and T-bet mRNA.CONCLUSION Quercetin and levofloxacin can induce the differentiation from type Th2 to type Th1 for rat organism,but there is no synergistic effect between them.
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AIM:To investigate the effect of arsenic trioxide (ATO) on T-bet/GATA3 signal pathway in MRL/lpr mice.METHODS:MRL/lpr mice and C57BL/6J mice at the age of 20 weeks were chosen and then divided in 2 different sub-groups,respectively.The mice in 2 sub-groups received ATO (0.4 mg · kg-1.d-1) and sodium chloride (NS,volume weight-determined) by intraperitoneal injection respectively for 2 months.Afterward,the spleens were isolated fron the MRL/lpr and C57BL/6J mice under pathogen-free condition and the suspensions were prepared.The mRNA level of T-bet,GATA3,IFN-γ,IL-4 and the mRNA ratio of T-bet/GATA3 were detected by RT-qPCR.The protein expression of T-bet and GATA3 was determined by Western blot.The serum levels of IFN-γ and IL-4 were measured by ELISA.RESULTS:The mRNA and protein levels of T-bet,IFN-γand the mRNA ratio of T-bet/GATA3 in NS group of MRL/lpr mice were higher than those in NS group of C57BL/6J mice (P ＜0.05).However,the GATA3 and IL-4 were lower in NS group of MRL/lpr mice in both mRNA and protein level (P ＜ 0.05).In MRL/lpr mice,the mRNA and protein levels of T-bet,IFN-γ and the mRNA ratio of T-bet/GATA3 were lower in ATO group compared with NS group (P ＜ 0.05),no difference was found in GATA3 and IL-4.No difference of the indexes mentioned above between ATO group and NS group in C57BL/6J mice was observed.CONCLUSION:ATO may affect the signaling pathway of T-bet/GATA3 to down-regulate the mRNA expression and the protein secretion of IFN-γ by decreasing the expression of T-bet in MRL/lpr mice.
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AIM To study the improvement of quercetin on Pseudomonas aeruginosa-induced lung infection in rats.METHODS Forty SPF SD rats were randomly divided into five groups,eight rats in each group:normal group,P.aeruginosa infection group,quercetin group,levofloxacin group,levofloxacin combined with quercetin group (combined group),the rats were anesthetized and then injected with P.aeruginosa in bronchus.The pathological changes of lung tissue in rats were observed,the contents of IL-4 and IFN-γcytokines,changes of transcription factors T-bet and Gata-3 in lung tissue were detected,and then semi-quantitative RT-PCR was used for the detection of IL-4,IFN-γ,T-bet and Gata-3 mRNA expressions.RESULTS The content of IL-4,levels of IL-4 and Gata-3 mRNA in lung tissue in the quercetin group,the levofloxacin group and the combined group were lower than those in the P.aeruginosa infection group,but the opposite was true in the levels of IFN-γ and T-bet mRNA.CONCLUSION Quercetin and levofloxacin can induce the differentiation from type Th2 to type Th1 for rat organism,but there is no synergistic effect between them.
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<p><b>OBJECTIVE</b>To analyze the immunological characteristics of 2,4,6-trinitrobenzene sulfonic acid (TNBS)-induced colitis model and examine the therapeutic effects and mechanisms of Astragalus polysaccharides (APS) treatment.</p><p><b>METHODS</b>Thirty-two male specific pathogen free Spragne-Dawley rats were randomly equally assigned to four groups: control, TNBS, APS and prednisone groups. Experimental colitis was induced by enema administration of TNBS. Then rats were treated with APS (0.5 g•kg•day, once daily) or prednisone (1.0 mg•kg•day, once daily) by gavage for 14 days. Macroscopic lesion and histological damage were determined, and activity of myeloperoxidase (MPO) was measured in the colonic tissues. Expressions of T-box expressed in T-cells (T-bet) and GATA-binding protein-3 (GATA-3) were determined by immunohistochemistry analysis and western blot.</p><p><b>RESULTS</b>Both macroscopic lesion and histological colonic damage induced by TNBS were reduced by APS and prednisone treatment. These were accompanied by significant attenuation of MPO activity (P=0.03). TNBS intervention enhanced the expression of both GATA-3 and T-bet, but the expression of T-bet was significantly enhanced than that of GATA-3, resulting in significant reduction of GATA-3/T-bet ratio (P=0.025). APS administration enhanced the expression of T-bet (P=0.04) and GATA-3 (P=0.019) in comparison to TNBS group, and resulting in an up-regulated GATA-3/T-bet ratio. Prednisone treatment inhibited both expressions; however it also resulted in up-regulation of the GATA-3/T-bet ratio.</p><p><b>CONCLUSIONS</b>These results demonstrated that APS exerted a beneficial immune regulatory effect on experimental colitis. It promoted the expression of T helper cell 1 (Th1) and T helper cell 2 (Th2) specific transcription factors but ultimately favor a shift toward Th2 phenotype, suggesting that APS possessed therapeutic potential in experimental colitis.</p>
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Animals , Male , Astragalus Plant , Chemistry , Blotting, Western , Colitis , Drug Therapy , Pathology , Colon , Pathology , GATA3 Transcription Factor , Metabolism , Immunohistochemistry , Immunomodulation , Peroxidase , Metabolism , Polysaccharides , Pharmacology , Therapeutic Uses , Rats, Sprague-Dawley , T-Box Domain Proteins , Metabolism , Trinitrobenzenesulfonic AcidABSTRACT
Objective To investigate the effects of a novel synthetic immunostimulator CH2b containing thiazolidin-4-one on the function of invariant nature killer T (iNKT) cells isolated from patients with active rheumatoid arthritis (RA).Methods Peripheral blood mononuclear cells (PBMCs) isolated from patients with active RA were in vitro cultured with α-Galcer and IL-2.The iNKT cells were separated by using magnetic activated cell sorting (MACS) method.The effects of CH2b on the proliferation of iNKT cells were analyzed by using MTT assay.MILLIPLEX MAP Human Cytokine/Chemokine kit was used to measure the levels of IFN-γ and IL-4 in the supernatants of iNKT cell culture.The expressions of IFN-γand IL-4 at mRNA level in iNKT cells were analyzed by RT-PCR.Western blot assay was used to detect the levels of T-bet and GATA-3 in iNKT cells.Results CH2b significantly enhanced the proliferation of IL-2 activated iNKT cells isolated from the patients with active RA.CH2b promoted the secretion of IL-4,resulting in a decrease in the ratio of IFN-γ/IL-4.Moreover,CH2b promoted the expressions of GATA-3 and IL-4 at mRNA level in iNKT cells.Conclusion The novel immunostimulator,CH2b,might enhance the immunoregulatory effects of iNKT cells by promoting the GATA-3 pathway-mediated secretion of Th2-1ike cytokines and inducing the differentiation of Th0 to Th2 cells.
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Objective To explore the regulation role and expression level of thymosin for Th1/Th2 and T-bet , GATA-3 in asthma model rats respectively.Methods 60 SPF grade male rats (6-8w) were randomly divided into six groups:control group, asthma model group, dexamethasone (0.5 mg/kg) group, thymosin high dose group (5 mg/kg), thymosin middle dose group(2.5 mg/kg), thymosin low dose group (1.25 mg/kg).The asthma model was constructed with ovalbumin.The rat in the control group and asthma model group were injected with normal saline by intraperitoneal , while other rats were injected with corresponding drugs.Then the concentration of IL-4 and IFN-γin the serum of rats were tested by the method of enzyme linked immunosorbent assay(ELISA) and the protein expression level of GATA-3 and T-bet factor in the lung tissue were tested by immunohistochemistry in each group.Results The concentration of IL-4 in the serum was higher and IFN-γwas lower in the asthma model group than the other groups , while the protein expression level of GATA-3 was higher and T-bet was lower in the asthma model group than the other groups ( all P <0.05 ) . Conclusion Thymosin upregulation of IFN-γand inhibition of IL-4 may be achieved by upregulating T-bet and downregulating GATA-3, and thymosin has therapeutic effect on asthma rats.
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T-bet is a critical transcription factor that regulates differentiation of Th1 cells from CD4+ precursor cells. Since T-bet directly binds to the promoter of the IFN-gamma gene and activates its transcription, T-bet deficiency impairs IFN-gamma production in Th1 cells. Interestingly, T-bet-deficient Th cells also display substantially augmented the production of IL-2, a T cell growth factor. Exogenous expression of T-bet in T-bet deficient Th cells rescued the IFN-gamma production and suppressed IL-2 expression. IFN-gamma and IL-2 reciprocally regulate Th cell proliferation following TCR stimulation. Therefore, we examined the effect of T-bet on Th cell proliferation and found that T-bet deficiency significantly enhanced Th cell proliferation under non-skewing, Th1-skewing, and Th2-skewing conditions. By using IFN-gamma-null mice to eliminate the anti-proliferative effect of IFN-gamma, T-bet deficiency still enhanced Th cell proliferation under both Th1- and Th2-skewing conditions. Since the anti-proliferative activity of T-bet may be influenced by IL-2 suppression in Th cells, we examined whether T-bet modulates IL-2-independent cell proliferation in a non-T cell population. We demonstrated that T-bet expression induced by ecdysone treatment in human embryonic kidney (HEK) cells increased IFN-gamma promoter activity in a dose dependent manner, and sustained T-bet expression considerably decreased cell proliferation in HEK cells. Although the molecular mechanisms underlying anti-proliferative activity of T-bet remain to be elucidated, T-bet may directly suppress cell proliferation in an IFN-gamma- or an IL-2-independent manner.
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Animals , Humans , Mice , Cell Proliferation , Ecdysone , Interleukin-2 , Kidney , Th1 Cells , Transcription FactorsABSTRACT
ObjectiveTo examine whether Shenfu injection (SFI) reduces post-resuscitation myocardial dysfunction in a pig model by modulating expression imbalance of transcription factors of regulatory T cell, namely GATA-3 and T-bet.Methods Thirty pigs were randomly divided into sham group (n = 6) and cardiopulmonary resuscitation (CPR) group (n = 24) according to the random number table method, and the pigs in the CPR group were randomly subdivided into normal saline (NS) group, epinephrine (EP) group, and SFI group (n = 8 per group). After 8minutes of untreated ventricular fibrillation (VF) followed by 2 minutes of CPR, animals in three groups respectively received central venous injection of either 20 mL SFI (1.0 mL/kg, SFI group), EP (0.02 mg/kg, EP group) or NS (NS group). Blood samples were obtained before VF and 0.5, 2, 6 hours after restoration of spontaneous circulation (ROSC), and the parameters of hemodynamics and oxygen metabolism were determined. Surviving pigs were sacrificed at 24 hours after ROSC, the pathological changes in myocardium were observed, the levels of interleukin-4 (IL-4), tumor necrosis factor-α (TNF-α) andγ-interferon (IFN-γ) were measured by enzyme linked immunosorbent assay (ELISA), and expressions of protein and mRNA of GATA-3 and T-bet were determined by Western Blot and quantitative real-time polymerase chain reaction (RT-qPCR), respectively.Results Six pigs of three resuscitation groups were successfully resuscitated. The CPR time, number of defibrillation, defibrillation energy, and ROSC time were significantly decreased in the EP and SFI groups compared with those in the NS group. Compared with the sham group, the parameters of left ventricular systolic function and oxygen metabolism were significantly decreased, myofibril organelles were extensively damaged, and progressive and severe deterioration of the myocardium was found, and mitochondrial structure was not recognizable in the NS group; the level of IL-4 in myocardium were markedly decreased, while that of TNF-α, IFN-γand IFN-γ/ IL-4 [reflecting helper T cell 1/2 (Th1/Th2)] were significantly increased. Protein and mRNA expressions of GATA-3 were markedly reduced in the myocardium of pigs in the NS group compared with that of the sham group at 24 hours after ROSC, while T-bet was significantly increased. Compared with the NS group, animals treated with SFI had minimal myocardial intracellular damage, with decreased heart rate (HR, bpm: 90.33±3.79 vs. 106.83±5.36) and increased mean arterial pressure (MAP), cardiac output (CO), oxygen delivery (DO2), and oxygen consumption (VO2) at 6 hours after ROSC [MAP (mmHg, 1 mmHg = 0.133 kPa): 107.67±1.96 vs. 86.83±1.85, CO (L/min): 2.47±0.08 vs. 2.09±0.04, DO2 (mL/min): 364.31±4.21 vs. 272.33±3.29, VO2 (mL/min): 95.00±2.22 vs. 82.50±2.28, allP<0.05]. Compared with the NS groups at 24 hours after ROSC, level of IL-4 was markedly increased in myocardial cells (ng/L: 33.80±3.06 vs. 16.15±1.34,P< 0.05), while the levels of TNF-α, IFN-γ and IFN-γ/IL-4 were lowered significantly [TNF-α (ng/L): 18.16±0.71 vs. 29.64±1.89, IFN-γ (ng/L): 373.75±18.36 vs. 512.86±27.86, IFN-γ/IL-4: 16.15±1.34 vs. 33.80±3.06, allP< 0.05], and myocardial T-bet protein and mRNA expressions were reduced [T-bet protein (gray value): 0.41±0.07 vs. 0.59±0.11, T-bet mRNA (2-ΔΔCt): 4.37±0.21 vs. 7.57±0.55, bothP< 0.05], furthermore, myocardial GATA-3 protein and mRNA expressions were significantly up-regulated in SFI group [GATA-3 protein (gray value): 0.25±0.07 vs. 0.16±0.07, GATA-3 mRNA (2-ΔΔCt): 0.63±0.07 vs. 0.34±0.05, bothP< 0.05]. The parameters in SFI group were significantly improved compared with those of the EP group.ConclusionsMyocardial immune dysfunction is induced by Th1/Th2 imbalance following myocardial injury subsequent to CPR in pigs. SFI can attenuate myocardial injury and regulate myocardial immune disorders, protect post-resuscitation myocardial injury by modulating expression imbalance of transcription factors GATA-3 and T-bet.
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Objective To study the role of imbalance between transcription factors GATA-3 and T-bet expressions in causing acute lung injury after resuscitation in cardiac arrest model of swine.Methods Mter swine model of electrically induced cardiac arrest was established for 8 minutes,animals were resuscitated to get restoration of spontaneous circulation (ROSC).The swine with ROSC were randomly assigned to be sacrificed at 12 and 24 h after ROSC (n =8 in each group).CD3 +,CD4+ and CD8 + lymphocyte subsets were determined by flow cytometry,and the levels of serum IL-4,TNF-α,and IFN-γ were measured by using ELLSA.The protein levels and expressions of GATA-3/T-bet mRNA were detected in lung tissue by western blotting and quantitative real-time PCR device,respectively.Results Pulmonary function was significantly impaired after ROSC.CD4 + lymphocyte subsets (28.4 ± 2.3) ％,(24.1 ± 1.6) ％ and CD4 +/CD8 + (1.7 ±0.9),(1.5 ± 1.0) were significantly lower in the post-ROSC group compared with the sham-operated group (48.4±2.9)％,(51.1±5.4)％ (2.5±1.3),(2.7±1.1) (P＜0.05) at 12 h and 24 h after ROSC.The levels of serum IL-4 and TNF-α were markedly increased,while IFN-γ and IFN-γ/IL-4 were significantly decreased in the post-ROSC group compared with the sham-operated group (P ＜0.05) at 2-12 h after ROSC.Protein level and expression of GATA-3 mRNA in lung tissue were markedly increased,while those of T-bet were significantly reduced in the post-ROSC group compared with the sham-operated group (P ＜0.05) at 12 and 24 h after ROSC.Conclusions The lung immune dysfunction induced by imbalance between transcription factors GATA-3 mRNA and T-bet mRNA expressions may complicate in the process of post-resuscitation lung injury in a porcine model of cardiac arrest.
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Aplastic anemia (AA) is a Tlymphocyte-mediated autoimmune disease. The research on AA was focused on three areas, which were the pathogenesis of immune dysfunction, hematopoietic stem cell damage and abnormal hematopoietic microenvironment. In recent years, more attentions have been paid on abnormal immune mechanisms in the blood. There are complex intracellular cytokine and signal transduction pathway of pathogenesis in AA. And T-bet/IFN-γ signaling pathway plays an important role in development and progression of AA. This article aimed to review T-bet/IFN-γ signaling pathways in AA.
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Objective To explore the effects of mesenchymal stem cells ( MSC ) treatment on platelet counts in mice with immune-mediated thrombocytopenia ( ITP) and the possible mechanism .Meth-ods ITP was induced by daily intraperitoneal injection of anti-platelet membrane CD 41 antibody (MWReg30) into BALB/c mice.The mice were then divided into experiment and control groups with 20 mice in each.Each mouse in experimental group was injected with 2×107 mesenchymal stem cells (MSC) through the tail vein .The numbers of blood platelets in mice from two groups were counted on days 5, 7 and 14 after MSC injection .Reverse transcriptase polymerase chain reaction ( RT-PCR) was performed to meas-ure T-bet and GATA-3 gene expression in peripheral blood mononuclear cells ( PBMCs ) at mRNA level on day 14.The levels of IFN-γ, IL-2, IL-4 and IL-10 in serum were detected by ELISA .Results The platelet counts in experimental group were significantly higher than those in control group on days 7 and 14 after MSC injection [(588.0±81.6)×109/L and (623.0±78.9) ×109/L vs.(317.0±90.1) ×109/L and (288.0± 87.8)×109/L ] (P<0.05).On day 14 after MSC injection, the T-bet expression at mRNA level in PBMCs from mice in experimental group was significantly lower than that in control group [(0.04±0.03) vs.(0.27 ±0.05)] (P<0.05), while the GATA-3 expression at mRNA level was higher than those in control group [ (0.14±0.04) vs.(0.07±0.05)] (P<0.05).Compared with control group, the concentrations of Th1 type cytokines such as IFN-γand IL-2 were remarkably down-regulated in experimental group [(3.1±1.7) pg/ml and (3.2±2.1) pg/ml vs.(10.3±4.8) pg/ml and (16.3±5.7) pg/ml](P<0.05), while the con-centrations of Th2 type cytokines such as IL-4 and IL-10 were up-regulated in experimental group [(88.6± 15.2) pg/ml and (38.3±11.8) pg/ml vs.(32.7±5.7) pg/ml and (22.1±3.4) pg/ml ] (P<0.05). Conclusion MSC treatment can effectively increase platelet counts in mice with immune-mediated thrombo-cytopenia, which may be associated with the suppression of Th 1-dominant response mediated by abnormal ex-pression of T-bet and GATA-3.