Phenotypic Knockout of CXCR4 on Molt-4 with SDF-1α/54 Attached with KDEL

Objective ： To investigate the mechanism of phenotypic knockout of CXCR4 on T-cell leukemia cell line Molt-4 via SDF-1α/54/KDEL intrakine technology, which the mutant SDF-1α/54, human stromal cell-derived Faceor-1 （SDF-1α） was deleted its C- terminal α-helix and attached with a endoplasimc reticulum retention signal 4-peptide- KDEL encoding gene, so that retain the newly synthesized receptor CXCR4 within the Molt-4 cells endoplasmic reticulum. Methods： The recombinant vector pEGFP-C3/SDF- 1α/54/KDEL were transfected into Cos-7 cells by liposome, SDF-1α/54/KDEL fusion protein was confirmed with western blot. The recombinant plasmids were transfected transiently into Molt-4 by electroporation. Results：Western blot confirmed SDF-1α/54/KDEL expression in Cos-7. A dramatic downregulation of CXCR4 expression on Molt-4 was demonstrated by flow cytometric （FCM） analysis. Conelusion：SDF-1α/54/KDEL and SDF- 1αKDEL have no significant deviation for phenotypic knockout of CXCR4. These suggest that the phenotypic knockout effects of SDF-1α/54 against CXCR4 are not influenced by deleting of SDF-1α helix in the C-terminal.     

人脐动脉平滑肌细胞培养及转染TFPI基因的实验研究

Objective The aim of the present study is to investigate the effect of tissue factor pathway inhibitor (TFPI) gene on cell cycle of human vascular smooth muscle cells.Methods Human vascular smooth muscle cells were separated from human umbilical artery and identified by immunohistochemical staining.The cells were transfected with various amount of pIRES-TFPI plasmid (1,2,and 3 μg/ml,respectively)and the TFPI expression in the cells were analyzed by RT-PCR.MTT assay was employed to detect the effect of TFPI gene on the proliferation of human vascular smooth muscle cells.Results The proliferation of vascular smooth muscle cells was inhibited in pIRES-TFPI group 5 and 7 days after gene transfection when compared with that of pIRES 1-neo transfection group.Conclusion The overexpression of TFPI gene in human umbilical artery vascular smooth muscle cells may contribute to the suppression of the proliferation of cells by gene transfection.     

Effect of PKC signalling pathway and aldose reductase on expression of fibronectin induced by transforming growth factor-β1 in human mesangial cells

Objective To study the effect of PKC signalling pathway and aldose reductase (AR) on the expression of fibronectin (FN) induced by transforming growth factor-β1 (TGF-β1). Methods Human mesangial cells (HMCs) were cultured and transfected with pcDNA3-AR, and subject to AR gene silencing with small interfering RNA (siRNA) and then the cell was treated with recombinant human TGF-β1. The AR mRNA expression in the HMCs was examined using real time RT-PCR and protein expression of AR and FN was detected by Western blotting. Results The cultured HMC treated with TGF-$l showed increased expression of AR and FN,the normal HMC showed not reduced expression of FN after incubation with single inhibitors of AR. Pre-incubation of cells with inhibitors of AR and PKC, then the different groups of cells were treated with TGF-$l ,and the induction effect on FN expression was suppressed (34%) in HMC. HMCs transfected with AR showed a strong protein expression of FN, which was increased by 3. 6-fold after treatment with TGF-pl (P <0. 05) , and the induction effect on FN expression was suppressed by G(O)6983 (42%) in HMCs (P < 0. 05) . The HMC with AR gene knock-down by siRNA showed a decreased expression of AR and 90% decrease of FN protein in HMCs(P <0. 01) , and TGF-β1-induced up-regulation of FN was significantly suppressed by siRNA (12%) in HMCs (P <0. 01). Conclusions AR is capable of regulating FN expression only in the presence of TGF-β1, and this reaction is possibly accomplished through the activation of PKC signalling pathway.     

Effect of PKC signalling pathway and aldose reductase on expression of fibronectin induced by transforming growth factor-β1 in human mesangial cells

Objective To study the effect of PKC signalling pathway and aldose reductase (AR) on the expression of fibronectin (FN) induced by transforming growth factor-β1 (TGF-β1). Methods Human mesangial cells (HMCs) were cultured and transfected with pcDNA3-AR, and subject to AR gene silencing with small interfering RNA (siRNA) and then the cell was treated with recombinant human TGF-β1. The AR mRNA expression in the HMCs was examined using real time RT-PCR and protein expression of AR and FN was detected by Western blotting. Results The cultured HMC treated with TGF-$l showed increased expression of AR and FN,the normal HMC showed not reduced expression of FN after incubation with single inhibitors of AR. Pre-incubation of cells with inhibitors of AR and PKC, then the different groups of cells were treated with TGF-$l ,and the induction effect on FN expression was suppressed (34%) in HMC. HMCs transfected with AR showed a strong protein expression of FN, which was increased by 3. 6-fold after treatment with TGF-pl (P <0. 05) , and the induction effect on FN expression was suppressed by G(O)6983 (42%) in HMCs (P < 0. 05) . The HMC with AR gene knock-down by siRNA showed a decreased expression of AR and 90% decrease of FN protein in HMCs(P <0. 01) , and TGF-β1-induced up-regulation of FN was significantly suppressed by siRNA (12%) in HMCs (P <0. 01). Conclusions AR is capable of regulating FN expression only in the presence of TGF-β1, and this reaction is possibly accomplished through the activation of PKC signalling pathway.     

靶向性反向半胱氨酸天冬氨酸蛋白酶3重组腺病毒诱导肝癌细胞凋亡的实验研究

Objective To investigate the effect and specificity of adenovirus containing r-caspase-3 gene on apoptosis of hepatocellular carcinoma cells. Methods The vector α-fetoprotein enhancer-albumin promotor (pAdTrack-EAFP-PALB) was constructed and the r-caspase-3 gene was subcloned into the vector. The linearized shuttle plasmid was homogenously recombined with AdEasy-1 in BJ5183 cells. The candidate clone was analyzed by restriction endonuclease digestion and sequencing, and then pAdeasy-EAFP-PALB/r-caspase-3 vector was digested with Pac Ⅰ and transfected into AD293 cells for packaging and amplifying.Infection titer and rate of recombinant virus was monitored by green fluorescent protein (GFP) expression.The expression of r-caspase-3 was detected by RT-PCR and Western-blot. The apoptosis of HepG2, 7721, L-02 and MDA- MB- 231 cells was detected by FCM method. Results The sequence of shuttle vector pAdTrack-EAFP-PALB/r-caspase-3 was correct after digestion by restriction endonuclease. Vector pAdeasy-EAFP-PALB/r-caspase-3 was identificated by Pac Ⅰ restriction digestion and PCR. The expression of GFP was observed in the transfected AD293 cells. The expression of r-caspase-3 gene was detected in the infected HepG2 cells by RT-PCR and Western-blot. The cells were infected with recombinant r-caspase-3 after 24 hours, and their apoptotic index were as follows: HepG cells, 48.2%; 7721 cells, 17.7%; L-02 cells, 7.3%;M DA- MB- 231 cells, 0%. Conclusion The recombinant of hepatocellular carcinoma - targeting adenovirus containing r- caspase- 3 gene is constructed successfully and can induce the targeted apoptosis of hepatocellular carcinoma cells, which provides the evidences for future research in hepatocellular carcinoma.     

Functional study of the different haplotypes cDNA in the coding region of CⅡTA gene

Objective To study the function of 4 different haplotypes cDNA which are constructed by two non-homonymy single nueleotide polymorphism (SNP) sites C19170G (Leu45Val) and C30799G (Ala500Gly) in the coding region of human CⅡTA gene. Methods HeLa cells were transfeeted with eu-karyotic expression vectors containing four different haplotypes cDNA. C Ⅱ TA mRNA and HLA classⅡanti-gen (HLA-DR, DP, DQ) were respectively detected by RT-PCR and indirect cell immunofluoreseence tech-nique in the untransfected and transfeeted with four eukaryotic expression vectors and empty vectors HeLa cells. The quantity of HLA classⅡ antigen were analyzed by flow eytometry. Results No expression of CⅡTA mRNA and HLA class Ⅱ antigen were observed on original HeLa cells and empty vector transfected cells. CⅡTA mRNA expression was emerged, and the expression of HLA class Ⅱ antigen were observed in the HeLa cells transfected with eukaryotic expression vectors containing four different haplotypes cDNA. And there were not significantly different with the levels of HLA class Ⅱ antigen expression among HeLa cells transfected with eukaryotic expression vectors containing four different haplotypes cDNA ( P > 0.05 ). Con-dusion The SNP of Chinese at the sites C19170G(Leu45Val) and C30799G(Ala500Gly) in the coding site of C Ⅱ TA gene did not influence capability of CⅡTA trans-aetivating HLA class Ⅱgene expression.     

Functional study of the different haplotypes cDNA in the coding region of CⅡTA gene

Objective To study the function of 4 different haplotypes cDNA which are constructed by two non-homonymy single nueleotide polymorphism (SNP) sites C19170G (Leu45Val) and C30799G (Ala500Gly) in the coding region of human CⅡTA gene. Methods HeLa cells were transfeeted with eu-karyotic expression vectors containing four different haplotypes cDNA. C Ⅱ TA mRNA and HLA classⅡanti-gen (HLA-DR, DP, DQ) were respectively detected by RT-PCR and indirect cell immunofluoreseence tech-nique in the untransfected and transfeeted with four eukaryotic expression vectors and empty vectors HeLa cells. The quantity of HLA classⅡ antigen were analyzed by flow eytometry. Results No expression of CⅡTA mRNA and HLA class Ⅱ antigen were observed on original HeLa cells and empty vector transfected cells. CⅡTA mRNA expression was emerged, and the expression of HLA class Ⅱ antigen were observed in the HeLa cells transfected with eukaryotic expression vectors containing four different haplotypes cDNA. And there were not significantly different with the levels of HLA class Ⅱ antigen expression among HeLa cells transfected with eukaryotic expression vectors containing four different haplotypes cDNA ( P > 0.05 ). Con-dusion The SNP of Chinese at the sites C19170G(Leu45Val) and C30799G(Ala500Gly) in the coding site of C Ⅱ TA gene did not influence capability of CⅡTA trans-aetivating HLA class Ⅱgene expression.     

原发性肝癌患者PD-1基因拷贝数及其转录和蛋白表达水平的研究

目的 检测原发性肝癌患者PD-1基因拷贝数及其转录和蛋白表达水平,研究宿主免疫调节因子PD-1基因拷贝数差异与肝癌的相关性.方法 采用实时荧光定量PCR法检测24例原发性肝癌患者PD-1基因拷贝数和外周血单个核细胞PD-1 mRNA表达水平,采用流式细胞术检测外周血CD8+T细胞PD-1蛋白表达水平.对照组为26例血清抗-HBs阳性健康者.结果 肝癌组和对照组PD-1基因单拷贝率分别为34.62％和4.17％,多拷贝率分别为61.54％和95.83％,两组间分布差异有统计学意义(x2 =7.639,P=0.006),肝癌组人群PD-1基因多倍体率显著高于对照组.PD-1mRNA表达平均值对照组为2.35E-03,肝癌组为1.23E-03,Mann-whitey检验比较两组间表达差异有统计学意义,肝癌组明显低于健康对照组(U=153,P=0.009);CD8+T细胞PD-1蛋白表达频率对照组(3.72±0.32),肝癌组PD-1表达频率(16.13±1.68),肝癌组PD-1表达比健康对照组高,差异有统计学意义(t=-7.073,P=0.000).结论 PD-1基因多拷贝可能是HBV感染者发生肝癌的高危因素.PD-1基因拷贝数差异与原发性肝癌的关系值得进一步研究.     

慢性乙型肝炎患者血清HBV DNA水平与细胞毒性T淋巴细胞表面PD-1表达的关系和意义

目的 探讨慢性乙型肝炎（CHB）患者血清HBV DNA水平与HBV特异性CTL表面程序性死亡受体-1（PD-1）表达的关系和意义.方法 将133例CHB患者根据血清HBV DNA水平分为甲、乙两组,甲组70例（52.63％）,HBV DNA水平为104～105拷贝/ml,乙组63例（47.37％）,HBVDNA水平分107 ～ 108拷贝/ml,对两组患者作HBV特异性CTL、非特异性CTL,HBV特异性CTL表面PD-1表达和肝功能的比较.结果 甲组HBV DNA水平为（5.03±1.01）log10拷贝/ml,低于乙组（7.59±0.99） log10拷贝/ml,t=11.23,P＜0.01,HBV特异性CTL表面PD-1表达（26.32±2.56）％,低于乙组（39.35 ±2.86）％,t=18.69,P＜0.01,HBV特异性CTL （0.37±0.02）％,高于乙组（0.22±0.02）％,t=25.80,P＜0.01,非特异性CTL（16.26 ±2.17）％,低于乙组（19.94±2.47）％,t=3.19,P＜0.01,ALT （259.16 ±85.11） U/L,低于乙组（501.16±86.15） U/L,t=15.10,P＜0.01,TBil（27.99±6.13）μmol/L,低于乙组（44.86±9.51） μmol/L,t=3.92,P＜0.01.结论 CHB患者HBV DNA水平低者,HBV特异性CTL表面PD-1表达也低,HBV特异性CTL水平高,HBV DNA水平高者,HBV特异性CTL表面PD-1表达也高,导致HBV特异性CTL水平降低,并启动非特异性CTL,引起肝功能损害加重.     

β萘黄酮对大鼠谷氨酰半胱氨酸合成酶催化亚单位基因表达的影响

目的 通过了解外界刺激因子β萘黄酮(β-NF)对大鼠谷氨酰半胱氨酸合成酶催化亚单位(GCLC)基因转录的影响,研究大鼠γ-谷氨酰半胱氨酸合成酶(γ-GCS)基因转录的调节机制.方法 利用大鼠GCLC基因调控序列驱动的GCLC-PGL3-enhancer-Luciferase报道载体(GCLC-Luc)转染大鼠支气管上皮细胞(RTE)和肝癌细胞(H4ⅡE),分DMSO空白对照组和不同浓度的各刺激因子组,比较各组荧光素酶值的差异以筛选影响GCLC基因转录调控的刺激因子,并比较筛选到的刺激因子β-NF对GCLC-Luc在RTE和H4ⅡE细胞内表达影响的异同.2种转染后的细胞均分为β-NF实验组(10μmol/L)和DMSO空白对照组,以荧光定量PCR检测各组的细胞内源性γ-GCS转录水平变化.分别将构建的r系列缺失报道载体、GCLC-Luc和激活蛋白1(AP-1)、NF-κB定点突变报道载体转染2种细胞,分β-NF实验组(10 μmol/L)与DMSO空白对照组,比较各组荧光素酶值的变化,分析β-NF作用的调节位点和转录调控作用相关元件.结果 1、10、100μmol/L的β-NF均强烈抑制RTE中GCLC基因表达,荧光素酶值显著低于DMSO空白对照组(16 135±1456、2752±218、1579±294比25 971±1662,均P＜0.01).在H4ⅡE中,1、10、100 μmol/L的β-NF则促进其表达,荧光素酶值显著高于DMSO空白对照组(5686±441、13 601±746、13978±164比3645±367,均P＜0.01).荧光定量PCR显示10 μmol/L的β-NF作用下,RTE内源性γ-GCS的mRNA表达水平是DMSO空白对照组的0.73倍,在H4ⅡE细胞中则是1.98倍.GCLC-Luc、r系列缺失报道载体转染2种细胞后,RTE中各载体β-NF实验组荧光素酶值均低于DMSO空白对照组(P＜0.01),而H4ⅡE中则高于DMSO空白对照组(P＜0.05).β-NF作用的调节位点定位于转录起始位点上游-390～+2 bp范围内.用针对该区域的AP-1、NF-κB定点突变报道载体转染细胞后,转染GCLC-Luc载体的β-NF实验组RTE荧光素酶值较DMSO空白对照组下降(91.50±0.32)%,与之相比,转染AP-1、NF-κB定点突变报道载体的细胞下降幅度并没有减少(P＞0.05).在H4ⅡE,β-NF作用下转染GCLC-Luc载体的细胞荧光素酶值上升幅度则高于转染AP-1定点突变报道载体的细胞[(3.81±0.19)倍比(2.08±0.19)倍,P＜0.05],而与转染NF-κB定点突变报道载体的细胞荧光素酶值上升幅度[(4.1±1.01)倍]相比差异无统计学意义(P＞0.05).结论 在大鼠RTE和H4ⅡE,β-NF影响GCLC基因的表达具有细胞特异性.在大鼠H4ⅡE,β-NF通过激活抗氧化应答元件(ARE)类似的AP-1调控GCLC基因表达.     

6种中药单体对人孕烷X受体介导细胞色素酶P450 3A4转录的调节作用

目的研究6种中药单体野黄芩苷、丹皮酚、芍药苷、甘草苷、黄芪甲苷以及黄芩素是否可以通过诱导孕烷X受体（PXR）介导细胞色素酶P4503A4（CYP3A4）转录表达。方法采用脂质体瞬时转染的方法,将人PXR表达质粒、CYP3A4报告质粒以及内参质粒转入LS174T细胞中,建立报告基因体系,与不同浓度及不同给药时间的中药单体共同孵育,检测细胞中荧光素酶的活性。结果黄芩素（20μmol/L、40μmol/L和80μmol/L）分别诱导PXR介导CYP3A4表达（1.49±0.23）倍（P〈0.05）、（2.45±0.56）倍（P〈0.01）和（5.27±0.40）倍（P〈0.01）,野黄芩苷、丹皮酚、芍药苷、甘草苷和黄芪甲苷在实验的浓度范围内不能明显诱导LS174T细胞中PXR介导CYP3A4表达。在不同给药时间实验中,20μmol/L、40μmol/L和80μmol/L的黄芩素在12～48h能诱导PXR介导CYP3A4表达,诱导能力随时间的延长而呈增强的趋势,各浓度的最大诱导倍数均在48h出现。结论黄芩素在LS174T细胞系中可以通过诱导PXR介导CYP3A4转录表达,而野黄芩苷、丹皮酚、芍药苷、甘草苷或黄芪甲苷无此作用。     

Accumulation of CD5<Superscript>+</Superscript>CD19<Superscript>+</Superscript> B lymphocytes expressing PD-1 and PD-1L in hypertrophied pharyngeal tonsils

Programmed death-1 (PD-1) is one of the most important inhibitory co-receptors expressed predominantly on activated T and B lymphocytes whose expression could be sustained by permanent antigenic stimulation accompanying chronic or recurrent tonsillitis. The expression of PD-1 and PD-1L was analyzed using flow cytometry on hypertrophied tonsils collected from 57 children. We observed high expression of PD-1 and PD-1L on certain lymphocytes subpopulations of hypertrophied tonsils; among T cells, the expression of PD-1 on protein level was higher on CD4⁺ cells (70.3 %) than on CD8⁺ cells (35 %). Interestingly, a limited expression of PD-1 was observed on CD19⁺ B lymphocytes (6.5 %), while CD5⁺CD19⁺ B cells overexpressed PD-1 (52.5 %). Moreover, the expression of PD-1L was also higher on CD5⁺CD19⁺ B cells (16.5 %) than on CD19⁺ B cells (3.5 %) and on CD4⁺ T cells (20 %) than on CD8⁺ T cells (10 %). PD-1 and PD-1L expressions correlated only on CD5⁺CD19⁺ cells. In conclusion, high expression of PD-1 and PD-1L on T and B cells could represent hallmark of immune system adaptation to chronic antigenic exposition in patients with tonsillitis.     

正常人PBMC中T细胞的PD-1表达及其表型分析

目的观察正常人外周血单个核细胞（PBMC）T细胞的PD-1表达,并对其表型进行分析。方法分离正常成年人PBMCs,利用流式细胞术检测CD4、CD8、CD45RA、CD27和PD-1等表面分子的表达;通过对比分析,了解PD-1＋T细胞表型与记忆T细胞的关系。结果正常成年人周围血中CD4＋PD-1＋T细胞和CD8＋PD-1＋T细胞的平均表达水平为（8.71±3.97）%,CD8＋PD-1＋T细胞的频率为（2.44±0.76）%。正常人外周血CD4＋PD-1＋T细胞可分成3个亚群,即初始T细胞（CD45RA＋CD27＋T细胞）、中央记忆型（CD45RA-CD27＋）T细胞和效应记忆型（CD45RA-CD27-）T细胞,其比例分别为（18.5±8.79）%、（65.0±13.84）%和（15.5±5.12）%,而在CD4＋PD-1-T细胞中3种T细胞的比例分别为（68.25±10.33）%、（27.75±9.04）%和（3.0±1.22）%。结论正常人周围血中存在CD4＋PD-1＋T细胞,并且多为中央记忆T细胞,而CD4＋PD-1-T细胞则多为初始T细胞。     

鼻咽癌组织中PD-1＋淋巴细胞数升高的意义

目的探讨鼻咽癌组织局部CD8＋和PD-1＋淋巴细胞在鼻咽癌发生发展中的作用。方法采用免疫组化方法检测64例鼻咽癌肿瘤组织和20例慢性鼻咽炎组织的CD8＋和PD-1＋淋巴细胞的分布情况。结果鼻咽癌肿瘤组织中CD8＋淋巴细胞百分比为（10.68±6.81）%,与慢性鼻咽炎组织的（28.5±9.33）%比较,差异有统计学意义（P〈0.05）;鼻咽癌组织中PD-1＋淋巴细胞为（10.4±5.95）%,与鼻咽炎组织的（6.8±3.21）%比较,差异有统计学意义（P〈0.05）。鼻咽癌组织中PD-1＋/CD8＋淋巴细胞比率为（1.24±0.91）,与鼻咽炎组织（0.28±0.2）比较,差异有统计学意义（P〈0.001）,但PD-1＋/CD8＋淋巴细胞比率升高与鼻咽癌临床分期无关。结论鼻咽癌组织中CD8＋淋巴细胞减少,PD-1＋淋巴细胞增加,提示PD-1可能参与鼻咽癌免疫逃逸,与鼻咽癌的发生有关。     

SFRP5基因甲基化通过激活Wnt/β-catenin通路调节白血病细胞MDR1/P-gp的表达

 目的:研究分泌型卷曲相关蛋白5（SFRP5）基因甲基化对耐药白血病细胞多药耐药蛋白1/P-糖蛋白（MDR1/P-gp）表达的调节及其可能的信号通路。方法:采用甲基化特异性PCR（MSP）检测白血病患者及白血病细胞系中SFRP5基因启动子区甲基化状态。采用Western blotting检测非磷酸化β-catenin（NP-β-catenin）在SFRP5基因甲基化白血病患者及细胞系中表达。使用去甲基化试剂5-氮杂-2′-脱氧胞嘧啶（DAC）恢复SFRP5表达,采用RT-PCR和real-time PCR检测处理前后MDR1 mRNA表达,采用Western blotting检测P-gp表达。结果:5/12例白血病患者标本及4种白血病细胞系存在SFRP5基因完全甲基化。在5例患者标本中,有3例（L2、L7和L8）NP-β-catenin水平高于其它标本,在4种白血病细胞系中,KG1a中NP-β-catenin水平高于另外3个细胞系。存在NP-β-catenin高水平表达的3例患者（L2、L7和L8）中检出MDR1 mRNA及P-gp表达。在4种细胞系中,KG1a存在MDR1 mRNA及P-gp表达。使用去甲基化试剂DAC处理L2、L7、L8患者标本和KG1a细胞,恢复SFRP5表达,L2、L7、L8和KG1a中MDR1 mRNA水平均显著下降（P<0.01）,P-gp表达水平亦被下调。结论: SFRP5基因甲基化可导致Wnt/β-catenin信号通路被激活,活化的β-catenin激活下游靶基因MDR1转录,编码的药物外排泵P-gp表达增加,促进白血病多药耐药的形成。     

应用基因芯片筛选慢性乙型肝炎患者PD-1/PD-L通路基因差异表达

目的 应用基因芯片技术筛选慢乙肝病人PD-1/PD-L通路差异性表达的基因.方法 应用基因芯片技术筛选4例慢乙肝病人与2例健康人外周血,获得差异性表达的基因,结合初步构建的PD-1/PD-L信号通路,挑选出一致的转录因子或调控蛋白,并应用Western blot方法检测CD 40L、Bcl-10、GATA-3、NFAT蛋白的水平的表达.结果 与健康人相比,慢乙肝患者有838个异常表达的基因,其中高表达的基因有150个,低表达的基因有688个.在这些异常表达基因中,有13个基因属于所构建的PD-1/PD-L信号通路中的转录因子或调控蛋白,其中表达上调的6个分别是BCL-10、AP、SRY、MYOD1、ELK4、NFAT,表达下调的7个分别是CD 40L、GADS、PDK1、IL-4、GATA3、CREB、RARG.Westem blotting法证实慢乙肝组高表达BCL-10,低表达CD 40L、GATA3,NFAT但与病毒载量不相关.结论 CD40L、BCL-10、GATA3、NFAT在慢性乙型肝炎患者PD-1/PD-L信号传导通路中可能有重要的调控作用,可以做深入的功能分析研究.