| Objectives:T helper lymphocytes differentiate into two subsets, Th1 and Th2 cells, each with distinct functions and cytokine profiles. IFN-γ is the signature cytokine of Th1 cell. IL-4 is the corresponding signature cytokine of Th2 cell, which also secretes IL-5, IL-6, IL-9 and IL-13, but doesn't secrete IFN-γ. These two subgroups of helper T cells arise in response to different immunogenic stimuli and cytokines, and they constitute an immunoregulatory loop, cytokines from Th1 cells inhibit Th2 cells, and vice versa. An imbalance in this reciprocal arrangement may be the key to asthma: there is credible evidence that, a hypothesis of the imbalance between Th1 and Th2, higher levels of Th2 cytokines IL-4 , IL-5 and IL-13 and lower levels of Thl cytokine IFN-y, is prominent in the pathogenesis of asthma. More recent studies have found T-bet and GATA-3 may be "master regulators" of Th lineage determination when IL-12 and IL-4 can strongly drive differentiation of Th1 and Th2 cells, respectively. CpG ODN containing unmethylated cytosine guanine dinucleotides CG (CpG) motifs could induce Th1-like immune reaction and suppress Th2-like immune response. Our study aimes to investigate the relation between the ratio of T-bet/GATA-3 in PBMCs and the imbalance of Th1/Th2 cells from patients with asthma and to observe the effect ofCpG ODN on the ratio of T-bet/GATA-3 and the balance of Thl/Th2 cells.Methods:Peripheral venous blood 6ml were obtained from 30 cases with exacerbating asthma (asthma group) and 20 cases with chronic obstructive pulmonary disease ( COPD group) and 20 normal cases ( control group) ,each to add EDTA 0.8ml for anticoagulation. The plasma were isolated by centrifugation and stored in -80°C. PBMCs were isolated by density gradient centrifugation. The expression of T-bet mRNA and GATA-3 mRNA in PBMCs were measured by reverse transcription-polymerase chain reaction (RT-PCR) and the plasma levels of IFN-y, IL-4 , IL-5 and IL-13 were measured by Enzyme-linked immunoadsordent assay (ELISA) .Peripheral venous blood 10ml were obtained from 20 cases with exacerbating asthma, each to add EDTA 1.2ml for anticoagulation. PBMCs were isolated by density gradient centrifugation and incubated with phytohemagglutinin (PHA) in vitro for 48h in the absence (blank group) or presence (CpG group) of CpG ODN.Supernatant and sediment were collected, respectively. The expression of T-bet mRNA and GATA-3 mRNA in sediment were measured by RT-PCR.The levels of IFN-y, IL-4, IL-5 and IL-13 in supernatant were determined by ELISA.Data were analysed using the Statistical Package of the SPSS 10.0. Discriptive data were given as means±SD. Continuous variables were tested by analysis of t-test and the relations between the variables were tested by linear correlation. The p-values are two-tailed and a p-value of less than 0.05 being considered to be statistically significant.Results:1. The T-bet mRNA expression in PBMCs in asthma group was significantly attenuated than those in COPD group and control group ( P<0.05 ) , meanwhile the GATA-3 mRNA expression was significantly enhanced ( P<0.05 ) , respectively.2. The plasma levels of IL-4, IL-5 and IL-13 in asthma group were significantly higher than those in COPD group and control group ( P<0.05 ) , respectively. The plasma level of IFN-y in asthma group was significantly lowerthan that in COPD group ( P<0.05 ) .3. The plasma level of IFN-y in COPD group was significantly higher than those in asthma group and control group ( P<0.05 ) , meanwhile the plasma level of IL-13 was significantly lower than those in asthma group and control group( P<0.05 ) , respectively.4. The ratio of T-bet/GATA-3 in asthma group was significantly lower than those in COPD group and control group ( P<0.05 ) .5. The T-bet mRNA expression has positive correlation with the plasma level oflFN-y (r=0.675) ( P<0.05 ) , has negative correlations with the plasma levels of IL-4(r=-0.780), IL-5C r=-0.664), and IL-13(r=-0.546)( P<0.05 ), respectively.6. The GATA-3 mRNA expression has negative correlation with the plasma level of IFN-y( r=-0.446)( P<0.05 ), has positive correlations with the plasma levels ofIL-4 (r=0.518 ), IL-5(r=0.393), and IL-13 (r=O.35O) ( P<0.05 ), respectively.7. The ratio of T-bet/GATA-3 has positive correlation with the plasma level of IFN-y (r=0.833 ) ( P<0.05 ) . has negative correlations with the plasma levels of IL-4 (r=-0.779),IL-5(r=-0.631), and IL-13(r=-0.584)( P<0.05 ),respectively.8. The supernatant levels of IL-4, IL-5 and IL-13 in CpG group were significantly lower than those in blank group ( P<0.05 ) , respectively. The supernatant level of IFN-y in CpG group was significantly higher than that in blank group (/><(). 05 ) .9. The T-bet mRNA expression in sediment in CpG group was significantly enhanced than that in blank group( P<0.05 ),and the GATA-3 mRNA expression was significantly attenuated ( P<0.05 ) , respectively. The ratio of T-bet/GATA-3 in sediment in CpG group was significantly higher than that in blank group ( P<0.05 ).Conclusions:1. A hypothesis of the imbalance between Thl and Th2 cells, higher levels of Th2 cytokines IL-4, IL-5 and IL-13 and lower level of Thl cytokine IFN-y, is prominent in the pathogenesis of asthma.The overproduction of Th2 cytokines caninduce an asthma phenotype.2. The T-bet mRNA expression in asthma was significantly attenuated ,and has positive correlation with the plasma level of IFN-y and has negative correlations with the plasma levels of 1L-4 ,IL-5, and IL-13,respectively. The GATA-3 mRNA expression in asthma was significantly enhanced ,and has negative correlation with the plasma level of IFN-y and has positive correlations with the plasma levels of IL-4 , IL-5, and IL-13, respectively. T-bet and GATA-3 are principally required for the production of cytokines of Thl and Th2 cells, respectively, and may be the regulatory element of upstream in the pathogenesis of asthma.3. The ratio of T-bet/GATA-3 has positive correlation with the plasma level of IFN-y and negative correlations with the plasma levels of IL-4> IL-5 and IL-13,respectively. The ratio of T-bet/GATA-3 has more close correlation with cytokines than T-bet and GATA-3 , respectively .The result indicates that the ratio of T-bet/GATA-3 could represent the imbalance of Thl/Th2 cells and may be the important regulatory element of in the pathogenesis of asthma.The general steps of the production of inflammation factors could be controlled at a much higher level by regulating the ratio of T-bet/GATA-3, so that could be to reverse the imbalance of Thl/ Th2 cells, and the asthma could be treated more effectively.4. CpG ODN could influence the ratio of T-bet/GATA-3 by upregulating the T-bet mRNA expression and downregulating the GATA-3 mRNA expression , so that could be to reverse the imbalance of Thl/Th2 cells by upregulating the expression of IFN-y and downregulating the expression of IL-4, IL-5 and IL-13. CpG ODN could be a new immunoregulatory attempt and promising therapeutic target for asthma which aims at recovering the imbalance of Thl/Th2 cells.
|
PDF Full Text Request