B7-H1及其受体PD-1分子在原发性肝-癌组织中的表达及临床意义

目的:分析B7-H1和PD-1分子在原发性肝癌组织中的表达,并探讨其在肝癌发生过程中的临床意义.方法:采用免疫组织化学染色和半定量逆转录聚合酶链反应(RT-PCR)检测42例原发性肝癌组织及对应的癌旁正常肝组织中B7-H1蛋白和mRNA表达,分析B7-H1分子与肝癌分化程度、有无侵袭转移的关系.分离肝组织内淋巴细胞.流式细胞仪分析PD-1分子在T细胞表面表达,以及B7-H1信号对肝癌组织内T细胞功能的影响.结果:原发性肝癌组织中B7-H1蛋白及mRNA表达水平均比癌旁组织和正常组织显著升高(85.71%vs 28.57%,7.14%:0.73±0.21vs0.35±0.12,0.23±0.07,均P<0.05),B7-H1阳性细胞与癌组织分化程度、有无侵袭转移相关(x2=7.876,8.492,均P<0.05),但与患者性别、年龄、肿瘤大小、肿块类型及AFP水平无相关关系.肝癌组织中T细胞表达PD-1水平比癌旁对照显著上高(20.15%±3.47%vs2.67%±0.53%,P<0.001).结论:肝癌中B7-H1分子表达可能抑制肝癌组织内T细胞功能,促进肝癌细胞逃避免疫监控,阻断肝癌细胞表面B7-H1/PD-1信号传导,可能成为肝癌免疫治疗的新途径.     

共刺激分子B7-H1和受体PD-1在溃疡性结肠炎患者结肠黏膜中的表达及意义

目的研究共刺激分子B7-H1和受体PD-1在溃疡性结肠炎患者结肠黏膜中的表达,探讨二者在结肠黏膜炎症反应中的作用及意义。方法收集溃疡性结肠炎患者45例及正常对照者10例,结肠镜下活检结肠黏膜行免疫组织化学染色,检测结肠黏膜中B7-H1和PD-1的表达,以阳性细胞的累积光密度值（IOD值）为评价指标,比较各组差异。结果 B7-H1和PD-1主要在结肠黏膜固有层单个核细胞中表达。UC活动期患者炎症最重部位及炎症不明显部位、UC缓解期患者原炎症部位B7-H1和PD-1的表达强度（IOD值）均明显高于正常对照组。其中B7-H1表达强度最高的为UC活动期患者炎症最重部位,其后依次为UC活动期患者炎症不明显部位和UC缓解期患者原炎症部位,各组间比较差异均有统计学意义（P〈0.05）;PD-1表达强度最高的为UC活动期患者炎症不明显部位,其后依次为UC活动期患者炎症最重部位和UC缓解期患者原炎症部位,各组间比较差异均有统计学意义（P〈0.05）。结论 UC患者结肠黏膜中B7-H1和受体PD-1表达上调,且表达水平与炎症程度有关,提示二者参与了肠道的炎症反应。     

聚乙二醇化干扰素α-2a治疗慢性乙型肝炎患者免疫细胞B7-H1表达的特点及其临床意义

目的 探讨慢性乙型肝炎患者在接受聚乙二醇化干扰素(PEG-IFN)α-2a治疗后免疫细胞B7-H1表达的变化特点及其临床意义.方法 采用流式细胞术检测14例慢性乙型肝炎患者在PEG-IFNα-2a治疗过程中外周血髓样树突细胞(mDCs)和CD4+T淋巴细胞上B7-H1表达的动态变化情况.体外培养慢性乙型肝炎患者外周血单核细胞,采用酶联免疫斑点法检测分泌干扰素(IFN)γ的T淋巴细胞频数.结果 PEG-IFNα-2a治疗24周,有效组和无效组外周血mDCs的B7-H1阳性表达率分别为10.37%±2.83%和19.27%±4.04%;有效组和无效组CD4+T淋巴细胞的B7-H1阳性表达率分别为19.71%±5.18%和40.43%±7.37%,有效组mDCs和CD4+T淋巴细胞上B7-H1的表达率明显低于无效组.有效组患者治疗前和治疗后每1×105个淋巴细胞可检测到分泌IFNγ的斑点数量分别为(113.33±31.41)个和(266.67±42.26)个,患者治疗后分泌IFNγ的T淋巴细胞频数明显高于治疗前;无效组患者治疗前和治疗后分泌IFNγ的斑点数量分别为(109.87±40.81)个和(55.63±10.27)个,治疗后分泌IFNγ的T淋巴细胞频数与治疗前相比,明显降低.阻断B7-H1途径可明显提高分泌IFNγ的抗原特异性T淋巴细胞频数.结论 接受PEG-IFNα-2a治疗的慢性乙型肝炎患者的疗效与其免疫细胞87-H1的表达有关,阻断B7-H1途径可提高机体清除病毒的特异性T淋巴细胞的免疫功能.     

B7-H1及其受体PD-1在慢性乙型肝炎患者淋巴细胞上的表达及意义

目的研究B7-H1及其受体PD-1在慢性乙型肝炎患者T淋巴细胞及髓样树突细胞（mDCs）上的表达及它们的表达水平与患者疾病状态的关系。方法流式细胞技术检测正常人和慢性乙型肝炎患者CD4^＋、CD8^＋T淋巴细胞及mDCs上B7-H1和PD-1的表达水平。实时定量PCR检测患者的HBV DNA。结果慢性乙型肝炎患者B7-H1及其受体PD-1的表达水平明显升高,健康对照mDCs、CD4^＋及CD8^＋T淋巴细胞B7-H1的阳性表达率分别为0.35%±0.10%、3.63%±0.70%和1.20%±0.19%,慢性乙型肝炎患者分别为7.88%±1.40%、24.28%±2.86%和10.78%±1.62%,慢性乙型肝炎患者B7-H1在mDCs和T淋巴细胞上的表达水平明显高于健康对照（P值均〈0.05）健康对照CD4^＋及CD8^＋T淋巴细胞PD-1的阳性表达率分别为5.92%±1.75%和5.98%±0.88%,慢性乙型肝炎患者分别为17.76%±2.47%和11.92%±2.21%,慢性乙型肝炎患者PD-1在T淋巴细胞上的表达水平也明显高于健康对照（P值均〈0.05）。且它们的表达与患者的ALT水平及HBV DNA载量呈明显的正相关（P值均〈0.05）。结论慢性乙型肝炎患者淋巴细胞上B7-H1和PD-1的表达水平与患者疾病状态密切相关。     

B7-H1在结肠癌组织中的表达及意义

目的:探讨结肠癌组织中B7-H1的表达及其与肿瘤转移的关系.方法:应用免疫组化技术检测64例新鲜结肠癌组织以及18例癌旁正常结肠黏膜组织中B7-H1的表达,分析B7-H1的表达与结肠癌患者临床病理指标的关系.结果:结肠癌组织中有B7-H1蛋白的原位表达,阳性表达率为48.44%,而癌旁正常结肠黏膜组织中B7-H1蛋白阳性表达率为22.22%,两者差异具有统计学意义( P<0.05).B7-H1蛋白的阳性表达率与结肠癌的淋巴结转移有关( P<0.05),而与浸润深度、分化程度、临床分期等无关.结论:B7-H1可作为一个预测结肠癌转移潜能的新的生物学指标.     

膀胱移行细胞癌组织中B7-H1和PTEN的表达及意义

目的探讨共刺激分子B7-H1和PTEN在膀胱移行细胞癌组织中的表达变化及意义。方法采用免疫组化SP法检测50例膀胱移行细胞癌及10例正常膀胱组织中的B7-H1和PTEN,分析其与膀胱移行细胞癌临床病理参数的关系及两者的相关性。结果正常膀胱组织中B7-H1不表达,膀胱移行细胞癌组织中B7-H1的阳性表达率为72%,且B7-H1的表达与肿瘤的病理分级、临床分期及复发密切相关（P〈0.05）;正常膀胱组织中PTEN呈阳性表达,膀胱移行细胞癌中PTEN蛋白阳性表达率为58%,随着肿瘤病理分级、临床分期的增加PTEN表达显著降低（P〈0.05）。B7-H1与PTEN的表达呈明显负相关（r=-0.44,P〈0.01）。结论膀胱移行细胞癌中B7-H1的高表达和PTEN的突变或缺失与肿瘤的发生发展和免疫逃逸密切相关。     

B7-H1与肝脏免疫

B7家族除经典B7-1、B7-2分子外，迄今又发现了B7-homologl（B7-H1）、B7-homolog2t、B7-DC、B7-homolog3、B7-superfamily member-1、B7-homolog4等新成员。作为共刺激分子，它们的分工又有所不同，在免疫反应过程中，表达于抗原递呈细胞表面的B7-1／B7-2、B7-homolog2、B7-homolog3可与T淋巴细胞上的相应受体CD28／细胞毒性T淋巴细胞抗原（cytotoxic T lymphocyte antigen-4，CTLA-4）、诱导性共刺激分子（inducible costimulator，ICOS）、结合并传递正性共刺激信号，从而促进T细胞活化、增殖和细胞因子分泌。     

乙型肝炎病毒转基因小鼠B7-H1表达水平与免疫耐受的相关性

目的 探讨HBV转基因(Tg)小鼠脾脏树突状细胞(DC)及肝脏B7-H1表达水平与HBV免疫耐受的相关性. 方法 制备小鼠脾脏DC,混合淋巴细胞反应检测其同种抗原刺激能力,流式细胞仪检测DC表面主要组织相容性复合物-Ⅱ(MHC-Ⅱ)及CD80、CD86、B7-H1等共刺激分子表达水平,酶联免疫吸附法检测白细胞介素-2(IL-2)、干扰素γ、IL-10水平,逆转录聚合酶链反应法和Western blot法检测肝组织B7-H1表达水平.计量数据组间比较采用f检验.结果 DC和T淋巴细胞比例分别为1:1、1:10、1:100时,HBV Tg小鼠脾脏DC刺激同种小鼠T淋巴细胞增殖数量(每分钟放射细胞数)分别为(865.4±39.3)个、(680.2±34.8)个和(320.0±12.7)个,正常小鼠刺激同种小鼠T淋巴细胞增殖数量分别为(22 385.6±99.7)个,(17 850.6±79.4)个和(760.0±32.1)个,HBV Tg小鼠脾脏DC刺激同种小鼠T淋巴细胞增殖能力明显均弱于正常小鼠DC,t值分别为16.674、19.674和21.712,P值均<0.01,差异有统计学意义.同时,HBV Tg小鼠MHC-Ⅱ,CD80表达下调,而CD86、B7-H1表达差异无统计学意义.HBV Tg小鼠分泌IL-2、干扰素γ、IL-10水平均降低,而HBV Tg小鼠和正常小鼠肝组织表达B7-H1的差异无统计学意义. 结论 HBV免疫耐受与MHC-Ⅱ、CD80下调表达导致的HBV Tg小鼠脾脏DC功能缺陷有关,而与负性共刺激分子B7-H1表达无关.     

Mechanism of T cell hyporesponsiveness to HBcAg is associated with regulatory T cells in chronic hepatitis B

AIM: To study the mechanisms of hyporesponsiveness of HBV-specific CD4 T cells by testing TH1 and TH2 commitment and regulatory T cells. METHODS: Nine patients with chronic hepatitis B were enrolled. Peripheral blood mononuclear cells were stimulated with HBcAg or HBsAg to evaluate their potential to commit to TH1 and TH2 differentiation. HBcAg-specific activity of regulatory T cells was evaluated by staining with antibodies to CD4, CD25, CTLA-4 and interleukin-10. The role of regulatory T cells was further assessed by treatment with anti-interleukin-10 antibody and depletion of CD4 CD25 cells. RESULTS: Level of mRNAs for T-bet, IL-12R p2 and IL-4 was significantly lower in the patients than in healthy subjects with HBcAg stimulation. Although populations of CD4 CD25highCTLA-4 T cells were not different between the patients and healthy subjects, IL-10 secreting cells were found in CD4 cells and CD4 CD25 cells in the patients in response to HBcAg, and they were not found in cells which were stimulated with HBsAg. Addition of anti-IL-10 antibody recovered the amount of HBcAg-specific TH1 antibody compared with control antibody (P < 0.01, 0.34%±0.12% vs 0.15%±0.04%). Deletion of CD4 CD25 T cells increased the amount of HBcAg-specific TH1 antibody when compared with lymphocytes reconstituted using regulatory T cells (P < 0.01, 0.03%±0.02% vs 0.18%±0.05%). CONCLUSION: The results indicate that the mechanism of T cell hype-responsiveness to HBcAg includes activation of HBcAg-induced regulatory T cells in contrast to an increase in TH2-committed cells in response to HBsAg.     

B7-H4与肿瘤及T细胞免疫的研究进展

肿瘤是严重危害人类健康的恶性疾病,发病率和病死率均较高,预后差.B7家族的新成员B7-H4是T细胞免疫应答的负性调控因子,能通过抑制T细胞的增殖,活化,细胞因子的分泌和细胞周期的进程来负性调控T细胞的免疫应答,在多种肿瘤组织中高表达,其与肿瘤的形成及免疫有着重要的关系,可作为肿瘤诊治的新靶点.     

B7-H1在HBV感染导致肝细胞癌中的调控作用

目的探讨B7-H1（B7-homolog1）在乙型肝炎病毒（HBV）感染导致的肝细胞癌（HCC）中的作用机制。方法通过慢病毒lentivirus．B7．H1转染人正常肝细胞系L02、人肝癌细胞系HepG2、HepG2．2．15后,应用MTT法观察细胞活性,流式细胞仪检测细胞的凋亡情况,Westernblot技术检测蛋白B7-H1、Survivin蛋白的变化情况。结果抑制B7-H1表达水平后,通过MTT检测表明细胞系HepG2．2．15细胞活性比HepG2细胞、L02细胞活性降低,流式细胞仪检测凋亡增加,同时Westernblot显示Survivin蛋白的表达减少。结论通过慢病毒转染抑$1JB7．H1的表达可以促进细胞系HepG2．2．15凋亡,B7-H1可能是通过调控Survivin的变化影响HBV感染的肿瘤细胞的增殖与凋亡。     

Relationship between co-stimulatory molecule B7-H3 expression and gastric carcinoma histology and prognosis

AIM:To investigate the expression of co-stimulatorymolecule B7-H3 in gastric carcinoma and adenomatissue as well as normal gastric tissue and to explore therelationship between B7-H3 expression and pathologicalfeatures and prognosis of gastric carcinoma.METHODS:B7-H3 expression was detected in 102samples of human gastric carcinoma and 10 samples ofgastric adenoma and 10 samples of normal gastric tissueby immunohistochemical assay.Correlation betweenthe expression of B7-H3 and the patients'age,sex,gastric carcinoma locus,tumor size,tissue type,tumorinfiltration depth,differentiation degree,lymph nodemetastasis,and survival time was analyzed.RESULTS:B7-H3 was expressed in all gastric adenomasamples and in 58.8% samples of gastric carcinoma.B7-H3 expression in gastric carcinoma samples wasnot related with the patients'age,sex,lymph nodemetastasis,and tumor size(P>0.05),but with thesurvival time,infiltration depth of tumor and tissue type.CONCLUSION:Detection of B7-H3 expression in gastriccarcinoma tissue is beneficial to the judgment of theprognosis of gastric carcinoma patients and the choice oftreatment.     

B7-H1 expression is upregulated in peripheral blood CD14+ monocytes of patients with chronic hepatitis B virus infection, which correlates with higher serum IL-10 levels

Chronicity in hepatitis B virus (HBV) infection is maintained by increased type 2 T-helper cell response, possibly because of increased interleukin-10 (IL-10) productions. B7-H1 can negatively regulate T-cell responses via its receptor, programmed death 1. Ligation of B7-H1 to T-cells can result in the preferential secretion of IL-10. In this study, we investigated whether there was an upregulated expression of B7-H1 in peripheral blood mononuclear cells in patients chronically infected by HBV and further explored the correlation between B7-H1 expression and serum interleukin 2, interferon-gamma, IL-10, HBeAg, alanine aminotransferase (ALT) levels and viral load. Fifty-five patients with chronic HBV infection and 20 healthy controls (HCs) were enrolled in the present study. The results showed that in patients with chronic hepatitis B CD14+ monocytes but not CD3+ and CD19+ cells had a significantly increased expression of B7-H1 compared with HCs, which positively correlates with serum IL-10 levels and the presence of HBeAg and negatively correlates with serum ALT levels. In conclusion, chronic HBV patients harbour an increased B7-H1 expression in CD14+ monocytes compared with controls, which may be responsible for the increased serum IL-10 levels. This might be an important way by which HBV evades an adequate immune response, leading to viral persistence and disease chronicity.     

调节性T细胞、调节性B细胞与支气管哮喘

支气管哮喘(简称哮喘)是一种由多种细胞及细胞组分参与的气道慢性炎症性疾病,目前经典的Th1/Th2理论并不能完全解释哮喘的免疫失衡.调节性T细胞(Tregs)和调节性B细胞(Bregs)是一类具有免疫抑制作用的淋巴细胞亚群,通过多种免疫调节通路调控免疫应答.本文就Tregs、Bregs的生物学特性与哮喘发病机制及其免疫治疗最新研究进展作一综述.     

Expression of B7-H4 and hepatitis B virus X in hepatitis B virus-related hepatocellular carcinoma

AIM: To investigate the expression and clinical significance of B7-H4 and hepatitis B virus X(HBx) protein in hepatitis B virus-related hepatocellular carcinoma(HBV-HCC).METHODS: The expression of B7-H4 in the human HCC cell lines Hep G2 and Hep G2.2.15 were detected by western blot, flow cytometry, and immunofluorescence. The expression of B7-H4 and HBx in 83 HBV-HCC was detected by immunohistochemistry, and the relationship with clinicopathological features was analyzed. Paraffin sections were generated from 83 HBV-HCC patients(22 females and 61 males) enrolled in this study. The age of these patients ranged from 35 to 77 years, with an average of 52.5 ± 11.3 years. All experiments were approved by the Ethics Committees of the Second Affiliated Hospital, Zhejiang University School of Medicine.RESULTS: B7-H4 was significantly upregulated in Hep G2.2.15 cells compared to Hep G2 cells. Specifically, the protein expression of B7-H4 in the lysates of Hep G2 cells was more than that in Hep G2.2.15 cells. In addition, HBx was expressed only in Hep G2.2.15 cells. Similar data were obtained by flow cytometry. The positive rates of B7-H4 and HBx in the tissues of 83 HBV-HCC patients were 68.67%(57/83) and 59.04%(49/83), respectively. The expression of HBx was correlated with tumor node metastases(TNM) stage, and the expression of B7-H4 was positively correlated with HBx(rs = 0.388; p 0.01). The expression level of B7-H4 in HBx-positive HBV-HCC tissues was substantially higher than that in HBx-negative HBV-HCC tissues. The expression level of B7H4 was negatively related to tumor TNM stage.CONCLUSION: Higher expression of HBx and B7-H4 was correlated with tumor progression of HBV-HCC, suggesting that B7-H4 may be involved in facilitating HBV-related hepatocarcinogenesis.     

强化剂量阿托伐他汀对不稳定型心绞痛患者PCI围手术期B7-H3、B7-H4的影响

目的探讨强化剂量阿托伐他汀对不稳定型心绞痛患者经皮冠状动脉介入治疗(PCI)围手术期B7-H3和B7-H4表达的影响。方法将80例不稳定型心绞痛患者随机分为常规剂量组(阿托伐他汀20 mg/d,n=40)和强化剂量组(阿托伐他汀80 mg/d,n=40),分别于PCI术前和术后18~24 h收集外周静脉血,采用ELISA检测外周血白细胞介素4(IL-4)、白细胞介素10(IL-10)、干扰素γ(IFN-γ)、血清可溶性B7-H3(sB7-H3)、可溶性B7-H4(sB7-H4)水平,用实时荧光定量PCR检测外周血单核细胞B7-H3、B7-H4 mRNA相对表达量。结果两组患者PCI术后IL-10、sB7-H3、sB7-H4水平和B7-H3、B7-H4的mRNA表达水平均较术前升高,其中强化剂量组升高更显著(P0.05)。相反,两组患者PCI术后IL-4、IFN-γ水平均降低,且强化剂量组较常规剂量组下降更明显(P0.05)。结论强化剂量阿托伐他汀可能通过促进B7-H3、B7-H4表达,从而降低不稳定型心绞痛患者PCI术后免疫炎症反应。     

B7 molecule mRNA expression in colorectal carcinoma

AIM: To observe the status of tumor-associated B_7 molecule mRNA expression in human colorectal cancer tissue by in situ hybridization. METHODS: The mRNA expression patterns of cancerassociated B_(7-1),B_7H_1,B_7H_2,ICOS in 22 specimens of human colorectal cancer tissue were monitored by in situ hybridization (ISH) with digoxin-iabeled oligonucieotide probes. RESULTS: B_(7-1)B_7H_1,B_7H_2,ICOS mRNA were detected in both cancer cells and tumor infiltrating lymphocytes (TIL). The mRNA expression level of these molecules in tumor cells was higher than that in TIL (0.76±0.54-1.62±0.82 vs 0.38±0.19-0.65±0.33, P<0.001). There was no relationship between expression level of tested B_7 family molecules and patients'sex, age, differentiation status of cancer and regional lymph node metastasis. CONCLUSION: Th2 cytokine predominant in tumor microenvironment might be related to the expression of B_7H_1,B_7H_2 co-signal molecules in tumor cells and TIL.     

Immunology of B7-H1 and Its Roles in Human Diseases

B7-H1 was originally identified by homology analysis in comparison with B7-1 and B7-2, two molecules with important immunoregulatory functions. B7-H1, however, was broadly induced in the majority of peripheral tissues as well as hematopoietic cells. Upon binding to an as yet unidentified costimulatory receptor on primed T-cells, B7-H1 costimulates T-cell proliferation and preferentially induces interleukin 10 and interferon gamma. The costimulatory function of B7-H1 may be critical for enhancing maturation and differentiation of T-cells in lymphoid organs. Conversely, by binding to programmed death 1 receptors on activated T-cells and B-cells, B7-H1 may inhibit ongoing T-cell responses in peripheral tissues by inducing apoptosis and arresting cell-cycle progression. Although a positive regulatory role of B7-H1 has been demonstrated in vitro and in various animal models, a negative regulatory role of B7-H1 has also been documented in human diseases, including cancer, rheumatoid arthritis, and human immunodeficiency virus infection. Delineation of the complex interactions between B7-H1 and its receptors as well as its interplay with other ligands is critical for understanding this new immunoregulatory system. Precise manipulation of B7-H1 and its receptors may provide unique opportunities for designing new disease treatments.     

大肠癌中nm23-H1的表达及其与临床病理特征的关系

目的:观察nm23-H1的表达与临床病理特征的关系,探讨nm23-H1的表达与大肠癌发生、发展、转移的作用.方法:选取行结、直肠癌根治术的患者63例,并取腺瘤组织16例做对照.应用免疫组化S-P法检测大肠癌患者癌组织、同源癌旁组织及正常黏膜组织和大肠腺瘤组织nm23-H1的表达.结果:nm23-H1在癌组织中表达(58.7%)显著低于癌旁组织(90.5%)、正常黏膜组织(96.0%)及腺瘤组织(93.8%)(P＜0.05).在大肠癌组织中,无淋巴结转移者nm23-H1表达(73.3%)显著高于有淋巴结转移者(45.5%)(P＜0.05).大肠癌中nm23-H1表达与肿瘤分化程度呈正相关(r=0.192,P＜0.05),与肿瘤浸润深度呈负相关(r=-0.263,P＜0.05).nm23-H1表达程度与患者年龄、性别、肿瘤大体分型、组织学类型、同期肿瘤大小均无关.结论:nm23-H1表达下调是大肠癌发生、发展中的重要分子生物学事件,可作为提示肿瘤分化、侵袭和淋巴结转移的一项指标.