抗原致敏及白细胞介素27基因修饰的树突状细胞诱导抗食管癌免疫的体外研究

目的:探讨食管癌细胞抗原致敏、白细胞介素27(interleukin27,IL-27)基因修饰的树突状细胞(dendritic cells,DCs)疫苗的特异性抗食管癌免疫反应。方法:采用重组逆转录病毒介导IL-27基因修饰食管癌患者外周血DCs;反复冻融法提取食管癌细胞裂解产物,致敏经IL-27基因转染的DCs;ELISA法检测各组DCs和细胞毒性T淋巴细胞(cytotoxic T lymphocytes,CTLs)上清液中IL-27、IL-12和干扰素-γ(interferon-γ,IFN-γ)的分泌水平;FCM法分析各组DCs表面分子CD1a、CD83、CD80和CD86的表达水平;MTT法检测DCs刺激T细胞增殖的活性和DCs诱导CTLs杀伤食管癌细胞的效能。结果:经IL-27基因修饰和抗原致敏后的DCs高表达CD83、CD80和CD86分子,其表达量分别为(82.67±7.92)%、(78.33±7.31)%和(85.33±4.32)%;DCs上清液中IL-12、IL-27及IFN-γ分泌量明显提高,分别为(107.85±7.20)ng/L、(430.39±10.12)ng/L及(411.97±17.80)ng/L;并且,DCs明显刺激同源T细胞增殖,增强了CTLs上清液中IFN-γ水平[(751.50±31.30)ng/L]。诱导产生的CTLs对食管癌细胞有强大的杀伤作用,显著高于其他组(P<0.01)。结论:IL-27基因修饰增强了DCs在体外诱导自体T淋巴细胞产生特异性抗食管癌免疫的能力。其机制可能与IL-27基因修饰和抗原致敏活化了DCs抗原提呈第二信号,促进了DCs高分泌IL-27和IL-12因子,活化了T淋巴细胞,并致使CTLs分泌IFN-γ的能力增强等密切相关。     

rhHSP70联合冻融抗原修饰树突状细胞诱导的抗乳腺癌作用

  目的   利用rhHSP70联合树突状细胞递呈肿瘤抗原的特性提高细胞毒T淋巴细胞（CTLs）对乳腺癌细胞的杀伤活性。   方法   外周血单个核细胞体外经GM-CSF和IL-4诱导产生树突状细胞,负载冻融抗原肽的同时加入新型热休克蛋白（rhHSP70）,不同分组分别诱导自体CTLs产生。ELISA测定CTLs杀伤活性和细胞因子的分泌。   结果   冻融抗原肽致敏的DCs促进CTLs增殖,上调CTLs中CD3+和CD8+T细胞群及Th1型细胞因子的分泌;体外实验中具有对人乳腺癌细胞MCF-7的杀伤活性,在加入rhHSP70后效果更加明显,并能显著增强CTLs对肿瘤细胞的杀伤率。   结论   hHSP70联合肝癌冻融抗原修饰DCs,能够促进DCs的成熟,增强DCs刺激淋巴细胞增殖的能力,诱导的CTLs在体外对乳腺癌细胞能产生高效杀伤力。rhHSP70增强DCs抗肿瘤能力的机制可能与其促进DCs成熟有关。       

幽门螺杆菌感染对晚期胃癌自体DC-CIK维持治疗的影响

目的：对比幽门螺杆菌（Helicobacter pylori,Hp）感染阴性和阳性的晚期胃癌患者接受自体树突状细胞联合细胞因子诱导的杀伤（dendritic cells-cytokine induced killer,DC-CIK）细胞维持治疗的疗效差异。方法：收集2010年6月至2012年6月中国人民解放军第174医院肿瘤中心收治的72例晚期胃癌患者,年龄29~90岁,中位年龄56岁,通过胃镜检查进行胃癌确诊并检测Hp,分为Hp阳性组（45例）及Hp阴性组（27例）,在接受手术或/和放化疗后,接受2疗程自体DC-CIK维持治疗,比较两组外周血培养的DC分化成熟情况及临床疗效差异。结果： 两组DC成熟过程形态变化无差异;Hp阳性组DC表面分子CD83、CD86表达显著高于Hp阴性组（P<0.01）,两组间HIA-DR表达无明显差异（P>0.05）。Hp阳性组治疗后生活质量评分（KPS）、外周血T细胞亚群（CD3+、CD4+、CD8+）比例较治疗前显著提高（P<0.05）,肿瘤标志物（CEA、CA199、CA724）表达较治疗前下降（P<0.05）。Hp阴性组治疗后肿瘤标志物表达、外周血T淋巴细胞比例较治疗前显著降低（P<005）,KPS评分较治疗前无明显变化（P>0.05）。Hp阳性组治疗后KPS评分、CEA和CA199表达以及CD3+和CD4+T细胞数量等的改善均优于Hp阴性组（P<0.05）,但瘤体稳定率、CA724、CD8+T细胞比例无显著差异（P>0.05）。共随访2年,阳性组患者中位生存期为12.64个月,长于阴性组的11.42个月（P<0.05）。结论： 携带Hp抗原信息的DC疫苗协同CIK治疗能够提高Hp感染的晚期胃癌患者的自身免疫功能,达到稳定瘤体、改善生活质量、延长生存期的目的。     

NY-ESO-1致敏树突状细胞诱导的CTL对肝癌细胞株的特异性杀伤作用

目的:探讨肿瘤睾丸抗原NY-ESO-1(New York-esophageal-1)致敏树突状细胞体外诱导特异性CTL对肝癌细胞株的杀伤作用.方法:重组质粒pGEX-ESO1经原核诱导表达并纯化GST-ESO1融合蛋白肽.重组人粒细胞-巨噬细胞集落刺激因子(rhGM-CSF)和白细胞介素4(rhIL-4)诱导培养人外周血来源的树突状细胞(dendritic cells, DCs),经GST-ESO1融合蛋白肽致敏后诱导特异性CTL增殖.以此CTL为效应细胞,分别以NY-ESO-1阳性表达的肝癌细胞株HepG2和不表达NY-ESO-1的肝癌细胞株H2P为靶细胞,MTT法检测CTL对肝癌细胞株的杀伤作用.结果:重组质粒pGEX-ESO1经IPTG诱导,在大肠杆菌中表达相对分子质量约36 000的GST-ESO1融合蛋白肽,纯化后的质量浓度为50 μg/ml;经rhGM-CSF和rhIL-4联合诱导成功培养人外周血DCs,其表型分子HLA-DR为91.4%、CD86为70.5%、CD83为71.2%、CD80为55.3%.NY-ESO-1致敏的DCs能明显诱导CTL增殖,此CTL对肝癌细胞株HepG2的杀伤率显著高于GST刺激组、未致敏DC组和无DC刺激组(均P<0.05),效靶比为50 ∶1时杀伤效应达到最高峰[(53.23±3.78)%,P<0.01];相同条件下CTL对H2P细胞无特异性杀伤作用.结论: NY-ESO-1抗原致敏的DCs在体外可诱导同种CTL产生和增殖,后者对NY-ESO-1阳性肝癌细胞株具有特异性杀伤效应,该方法为肝癌免疫治疗提供了一条新思路.     

抗原致敏树突状细胞诱导CIK对胃癌细胞杀伤作用的研究

[目的]探讨抗原致敏的树突状细胞（DC）诱导CIK（cytokine induced killer）的杀伤作用。[方法]联合应用粒/巨细胞集落刺激因子（GM-CSF）及白介素-4（IL-4）从外周血单个核细胞中培养DC，用人胃癌细胞SGC提取肿瘤抗原致敏DC，流式细胞仪检测致敏前后DC表型的变化，用人胃癌细胞SGC提取肿瘤抗原致敏DC诱导CIK，用MTT法检测淋巴细胞的增殖及原致敏DC诱导CIK对SGC的杀伤效应。[结果]①利用GM-CSF及IL-4可从外周血单个核细胞中获取DC，肿瘤抗原可促进DC的成熟，肿瘤抗原致敏可促进DC成熟，细胞表面分子HLA-DR、CD86、CD86升高，CD14下降。②混合淋巴细胞反应提示成熟的DC可促进CIK细胞增殖。③SGC肿瘤抗原致敏DC诱导CIK对胃癌细胞SGC有特异性的杀伤作用，随着效靶比的升高，杀伤效应随之增强。[结论]抗原致敏的DC可通过诱导特异性CIK细胞及促进CIK细胞增殖两方面显著提高CIK细胞的杀瘤效应。     

卵巢癌患者外周血来源树突细胞的生物学特性

背景与目的：树突细胞（dendritic cells,DCs）是体内功能最强的抗原递呈细胞．在抗肿瘤免疫治疗中起着重要的作用。既往对DCs生物学特性的研究数据大多来自健康个体,而来自肿瘤患者的DCs的特性尚不清楚。本研究旨在探讨卵巢癌患者外周血单核细胞来源的树突细胞（monocyte—derived dendritic cells,MoDCs）的生物学特性。方法：收集8名上皮性卵巢癌患者及13名健康女性志愿者的外周静脉血标本,从中分离出单核细胞,将其置于含人白细胞介素4（interleukin4,IL4）及粒细胞．巨噬细胞集落刺激因子（granulocyte-macrophag ecolony stimulating factor,GM-CSF）的RPMI-1640完全培养基中培养,用肿瘤坏死因子-α（tumornecrosisfactor-α,TNF-α）刺激其成熟。对诱导7d后的MoDCs进行细胞形态、表型及刺激异体淋巴细胞增殖能力分析。结果：卵巢癌患者及健康妇女外周血单核细胞培养7d后,均诱导出形态学上典型的成熟MoDCs。两组MoDCs均表达丰富的HLA-ABC（MHC-Ⅰ）、HLA-DR（MHC-Ⅱ）和大量共刺激分子CD86和CD80。卵巢癌组HLA-ABC、HLA-DR、CD86及CD80的平均荧光强度（mean fluorescence intensity,MFI）与健康妇女组差异无统计学意义（P〉0．05）。混合淋巴细胞反应（mixed leukocytes reaction,MLR）结果显示,卵巢癌各组（按MoDCs与淋巴细胞的比例分组）的吸光度值均明显低于健康妇女相应各比例组（P〈0．05）。结论：卵巢癌患者的MoDCs刺激异体淋巴细胞增殖能力较健康妇女降低,提示其存在一定程度的免疫学功能异常。     

DC活化淋巴细胞对自体乳腺癌细胞的杀伤作用

 目的探讨负载自体抗原DC诱导的CTLs对自体乳腺癌细胞的杀伤作用。方法以负载自体癌细胞抗原的DCs体外诱导CTLs,用ELISA法检测IFN-γ和IL-12的表达水平,用LDH法检测CTL对自体癌细胞的杀伤作用。结果负载自体抗原DC组IFN-γ和IL-12的浓度高于未致敏DC组、抗原组及单核细胞对照组(P<0.05),且负载抗原DC组所刺激的CTLs的杀伤作用也强于未致敏DC组、抗原组及单核细胞组(P<0.01)及对照的MCF-7组和HT-29组(P<0.01)。结论肿瘤裂解物致敏DC可持续刺激特异性CTLs从而可在体外杀伤乳腺癌患者的自体肿瘤细胞,说明肿瘤抗原致敏的DC疫苗对于治疗残留的和(或)对化疗耐药乳腺癌可能是适合的,因而有可能成为标准的补救措施。     

不同方式负载AFP抗原的DC诱导抗肝癌HepG2细胞免疫效应的比较

目的：比较AFP抗原以不同方式负载DCs对后者生物学功能的影响及体外刺激CTL对肝癌HepG2细胞的杀伤作用。方法：分离健康供者外周血单个核细胞培养DC,分别用（A）人工合成AFP抗原肽、（B）HepG2细胞裂解物、（C）HepG2细胞分泌的外泌体（tumor exosome, T-exo）、（D）AFP抗原的重组腺相关病毒（recombinant adeno-associated virus expressing α-fetoprotein antigen, rAAV/AFP）致敏DC前体细胞及（E）未负载抗原的DC作为对照,经GM-CSF、IL-4及LPS联合诱导DCs分化成熟。流式细胞术检测rAAV/AFP病毒感染效率、各组DCs表型及其剌激初始T细胞的增殖效应,7-ADD/CFSE双染法流式细胞术检测各组DCs诱导CTL对AFP阳性HepG2细胞的杀伤作用。结果：几种抗原负载方式均可诱导DCs成熟、促进CTL增殖及特异性识别并杀伤HepG2细胞,但rAAV/AFP感染DCs后,其CD83、CD86、ICAM-1、CD58、CD40分子表达水平明显高于对照组（P<0.05）,rAAV/AFP+DC组和HepG2-Texo+DC组对HepG2细胞的杀伤作用均分别显著优于其他抗原负载（AFP/peptide+DC、HepG2 lysate+DC）组\[（44.92±4.12）% vs（28.42±3.29）%、（24.28±1.79)%;（41.40±2.87）% vs （28.42±3.29）%、（24.28±1.79）%;均P<0.05）\]。结论：rAAV/AFP高效感染DCs后能有效刺激初始T细胞增殖,并增强CTL对AFP阳性靶细胞的杀伤活性,而负载Texo的DCs也能诱导显著的抗肝癌效应,上述结果为基于DCs的肝癌疫苗的研发提供新的思路。     

肺癌胸腔积液树突状细胞培养与鉴别及其抗肿瘤作用的观察

目的:证实肺癌胸腔积液中存在树突状细胞(DCs)的前体细胞,体外培养可以获得成熟的DCs,并探讨DCs对自体癌细胞的杀伤作用.方法:通过Ficoll淋巴细胞分离液对15例肺癌患者的恶性胸腔积液进行密度梯度离心,以两步贴壁法分别获得DCs的前体细胞和自体癌细胞.前者加入粒细胞/巨噬细胞集落刺激因子(rhGM-CSF)、白细胞介素-4(rhIL-4)及肿瘤坏死因子-α(TNF-α)培养.显微镜观察细胞形态,流式细胞仪鉴定DCs表型.MTT法检测DCs对自体癌细胞的杀伤作用.结果:体外培养9d后获得大量成熟的DCs,具有典型树突状形态,且高表达HLA-DR[(84.2±6.3)％]、CD80[(75.8±8.8)％]、CD83[(58.0±9.4)％]和CD86[(83.3±7.5)％];MTT法结果显示,DCs对自体癌细胞有较强的杀伤作用,当效靶比为20∶1时杀伤率为(50.62±7.51)％.结论:肺癌恶性胸腔积液中的DCs前体细胞经诱导培养后可转化为功能健全的成熟DCs,成熟的DCs对自体癌细胞有直接杀伤作用,为恶性胸腔积液的免疫治疗提供理论依据.     

CD 40激发肿瘤抗原特异性DCs对CIK细胞生物学活性的影响

目的:研究CD40激发凋亡肿瘤细胞致敏的树突状细胞(dendritic cells,DCs)对细胞因子诱导杀伤(cytokine-inducedkiller,CIK)细胞的细胞表型、增殖活性及细胞毒活性的影响。方法:常规方法从健康人外周血单个核细胞中诱导DCs和CIK细胞,采用凋亡肿瘤细胞负载DCs,并用或不用激发型CD40单克隆抗体(CD40mAb)激活DCs成熟;成熟DCs与同源的CIK细胞共育5d,分别获得DC40Ag-CIK细胞及DCAg-CIK细胞;观察细胞增殖活性;流式细胞仪检测细胞表型;酶联免疫吸附实验(enzyme-linked immunosorbent assays,ELISA)法检测细胞培养上清液中IFN-γ的含量;[3H]-TdR掺入法测定细胞杀伤活性。结果:凋亡肿瘤细胞负载激发可使DCs上调表达CD1a、CD80、CD86、CD83、HLR-DR,联合CD40mAb激发可进一步促进DCs成熟;培养至第14天时,DC40Ag-CIK细胞、DCAg-CIK细胞、CIK细胞分别扩增(18.2±1.7)倍、(15.0±1.2)倍、(9.3±1.8)倍;DC40Ag-CIK细胞中的CD3 CD56 比例较DCAg-CIK细胞和CIK细胞明显上调(P<0.05);DC40Ag-CIK细胞、DCAg-CIK细胞对A549杀伤活性强于CIK细胞(P<0.05),并且DC40Ag-CIK细胞强于DCAg-CIK细胞(P<0.05);CIK细胞、DCAg-CIK细胞、DC40Ag-CIK细胞培养上清液中IFN-γ的含量递增,分别为(1494.7±246.3)pg/mL、(2706.3±197.0)pg/mL、(3676.3±335.0)pg/mL。结论:与单独凋亡肿瘤细胞负载的DCs相比,CD40激发的凋亡肿瘤细胞负载的DCs可进一步提高CIK细胞的增殖活性及细胞毒活性。     

Is there a link between environmental factors and a genetic predisposition to cancer? A lesson from a familial cluster of gastric cancers

To determine the prevalence of gastric precancerous lesions and mucosal genetic alterations in relatives of a cluster of familial gastric cancer (FGC), we studied a kindred spanning two generations. The founder, daughter and niece underwent surgery for gastric cancer (GC); a son and other two daughters of the founder, presented with chronic dyspepsia. In all subjects, gastric mucosa samples were analysed for pathological features, Helicobacter pylori infection, microsatellite (MIN) and chromosomal (CIN) instability. The overexpression of mp53 and c-myc, and cytoplasmic beta-catenin delocalisation were found in the 2 younger cancer patients. All GC and gastritis patients had normal E-cadherin expression and were MIN-negative. Aneuploidy characterised all GC cases, and mixed euploid and aneuploid cell populations were present in the gastric biopsies from two of three 'at-risk' relatives. These two subjects, one of whom had severe active gastritis, and gastric mp53 and c-myc expression, were CagA-positive H. pylori-infected. DNA aneuploidy, p53 and c-myc expression disappeared after H. pylori eradication. In this FGC cluster, genetic abnormalities were found in first-degree relatives (3 patients) only in presence of H. pylori infection (2 cases H. pylori-positive versus 1 case H. pylori-negative) supporting the hypothesis that, besides the influence of a genetic profile, FGC may be, at least partly, mediated by intrafamilial clustering of H. pylori infection.     

Enhanced expression of inducible nitric oxide synthase and nitrotyrosine in gastric mucosa of gastric cancer patients.

Recent studies (K. Komoto et al., Am. J. Gastroenterol., 93: 1271-1276, 1998) have shown that Helicobacter pylori infection is associated with gastric cancer. However, the mechanism of H. pylori in carcinogenesis has not been clarified. H. pylori infection leads to a sustained production of reactive nitrogen species that may contribute to cause DNA damage. In this study, we examined the expression of inducible nitric oxide synthase (iNOS) and nitrotyrosine in gastric mucosa. The expression of iNOS and nitrotyrosine was examined by immunohistochemistry in 93 patients who initially underwent gastric biopsies between 1975 and 1992. Thirty-four individuals were later found to have gastric cancer at least 2 years after the initial biopsies (group A). The other 59 subjects have shown no evidence of gastric cancer during long-term follow-up. Fifty-one of these patients were positive for H. pylori (group B), and eight were negative for H. pylori (group C). The expression of iNOS and nitrotyrosine in the gastric mucosa was significantly higher in H. pylori-positive groups A and B than in H. pylori-negative group C. Among the H. pylori-positive patients, the expression of iNOS and nitrotyrosine was significantly higher in group A than in group B. These results suggest that high production of iNOS and nitrotyrosine in the gastric mucosa infected with H. pylori may contribute to the carcinogenesis of gastric cancer.     

幽门螺杆菌感染的胃黏膜上皮细胞环氧合酶-2表达及意义

目的　探讨幽门螺杆菌(Hp)感染的胃黏膜上皮细胞环氧合酶-2 (COX -2 )的表达及其在胃黏膜癌变过程中的意义。方法　采用快速尿素酶试验和组织学碱性品红染色法检测胃黏膜Hp感染状况;应用免疫组化法检测胃黏膜上皮细胞COX- 2表达状况。结果　32例胃癌中,COX -2表达阳性2 2例(6 8.7%)。1 2例Hp阴性的胃黏膜中,有1例(8.3%)COX -2低表达;1 0例Hp阳性正常胃黏膜中,仅1例(1 0 .0 %)COX -2低表达;9例Hp阳性的胃黏膜充血水肿糜烂者中,有5例(5 5 .6 %)COX 2表达阳性,与Hp阴性者和Hp阳性正常胃黏膜者比较,差异有统计学意义(P <0 .0 5 ) ;1 0例Hp阳性的轻度萎缩性胃炎伴轻度肠化生者中,COX -2表达阳性5例(5 0 .0 %) ;1 0例Hp阳性的中重度萎缩性胃炎伴中重度肠化生者中,COX -2表达阳性8例(80 .0 %) ;8例Hp阳性的中重度不典型增生者中,COX 2表达阳性6例(75 .0 %)。中重度萎缩性胃炎伴中重度肠化生和不典型增生者的COX 2表达高于轻度萎缩性胃炎伴轻度肠化生者(P <0 .0 5 )。结论　Hp感染诱导慢性浅表性胃炎黏膜上皮细胞的COX -2表达与黏膜损伤形成有关;根据胃癌发生模式,COX -2表达上调与Hp感染胃黏膜癌变发生相关,且可能在癌前病变形成早期阶段起作用。     

CIK分泌的细胞因子对逆转耐顺铂肺腺癌细胞系A549/DDP耐药性的研究

  目的   探讨细胞因子诱导的杀伤细胞(cytokine induced killer, CIK) 分泌的细胞因子在逆转耐顺铂(DDP) 人肺腺癌细胞系A549/DDP顺铂耐药性中的可能作用。   方法  采用transwell非接触共培养CIK与A549/DDP细胞, 收集不同时间点的共培养上清, 酶联免疫试剂方法(ELISA) 检测IFN-γ、TNF-α、IL-2的分泌情况, 四氮甲唑兰比色法(MTT法) 检测DDP耐药性的变化, 实时定量PCR检测GST-π基因表达水平的变化。   结果  分析发现A549/DDP细胞的耐药逆转与IFN-γ的分泌水平显著相关, 与TNF-α、IL-2的分泌量无显著相关。给予anti-IFN-γ中和抗体后, 共培养20h A549/DDP的DDP耐药性显著上升(P < 0.05), GST-π基因水平的表达量较未加中和抗体组也显著增加(P < 0.05)。而加入anti-TNF-α、anti-IL-2中和抗体组, A549/DDP细胞的DDP耐药性及GST-π基因水平的表达量较未加入中和抗体组无明显变化(P > 0.05)。   结论  CIK通过分泌IFN-γ下调A549/DDP细胞中GST-γ的表达来实现其逆转DDP耐药的作用。      

Higher methylation levels in gastric mucosae significantly correlate with higher risk of gastric cancers.

BACKGROUND: Helicobacter pylori infection potently induces methylation of CpG islands in gastric mucosae, which is considered to decrease to a certain level after active H. pylori infection discontinues. Noncancerous gastric mucosae of H. pylori-negative cases with a gastric cancer had higher methylation levels than those of H. pylori-negative healthy individuals. Here, using cases with multiple gastric cancers, we analyzed whether the higher methylation levels correlated with the higher risk of gastric cancers. METHODS: Twenty-six healthy volunteers (HV), 30 cases with a single well-differentiated gastric cancer (S cases), and 32 cases with multiple well-differentiated gastric cancers (M cases) were recruited. H. pylori infection status was analyzed by the culture method. Methylation levels were quantified by real-time methylation-specific PCR of seven CpG islands. RESULTS: In H. pylori-negative individuals, significant increasing trends were present in the order of HV, S cases, and M cases for FLNc and HAND1 methylation levels (P < 0.01, Spearman's rank-order test). Furthermore, the FLNc methylation level of M cases was significantly higher than that of S cases (P < 0.01, t test). Even adjusted by the extent of gastric atrophy, the FLNc methylation level retained a significant increasing trend (P = 0.03). In contrast, methylation levels in H. pylori-positive individuals were increased to various degrees in all the three groups. CONCLUSIONS: In H. pylori-negative individuals, methylation levels in gastric mucosae significantly increased in cases with a single gastric cancer and more in cases with multiple gastric cancers. Quantitative analysis of methylation levels is a promising risk marker for gastric cancers.     

Compartment theory in Helicobacter pylori-associated gastric carcinogenesis

BACKGROUND: The compartment theory has not been well investigated in gastric carcinogenesis. This study was aimed at examining the compartment alterations through the Helicobacter pylori (H. pylori)-related chronic gastritis-intestinal metaplasia-carcinoma sequence, and investigating the long-term effect of bacterial eradication on the compartment changes. PATIENTS AND METHODS: Gastric biopsy specimens were obtained from subjects with H. pylori-negative normal mucosa (N = 12), H. pylori-positive non-metaplastic gastritis (N = 42), H. pylori-positive intestinal metaplasia (N = 21) and intestinal-type adenocarcinoma (N = 20). The specimens were immnostained for monocloncal antibodies against the proliferating cell nuclear antigen (PCNA) for proliferating analysis. Additionally, 50 patients with H. pylori-positive gastritis were enrolled to investigate the long-term effect of bacterial eradication on the compartment changes of gastric epithelium. RESULTS: The mean PCNA labeling indices (L.I.) of non-metaplastic gastritis, intestinal metaplasia and adenocarcinoma were significantly higher than that of normal mucosa (31.1, 49.2 and 40.7 vs. 21.4; p < 0.01, 0.001 and 0.001, respectively). The proliferating zone was principally located in the lower compartment of normal mucosa. In patients with intestinal metaplasia, there was a full expansion (phase 1 change) of proliferating zone to the middle compartment of gastric pits (ratio of L.I. between middle and lower compartment = 1.00). The proliferating cells were evenly distributed in adenocarcinoma (complete loss of compartmentalization). Eradiation of H. pylori led to a reversion of compartment changes of gastric epithelium in patients with chronic gastritis. CONCLUSION: H. pylori-related gastric carcinogenesis is a multistep process involving progressive alterations of proliferating activity as well as loss of compartmentalization. Eradication of H. pylori reverses the changes in growth kinetics of gastric epithelium.     

Helicobacter pylori infection as an independent prognostic factor for locally advanced gastric cancer patients treated with adjuvant chemotherapy after curative resection

A few studies reported the association between negative Helicobacter pylori infection and poor clinical outcome in resected gastric cancer patients. We investigated the H. pylori infection status and its association with the clinical outcome in 274 locally advanced gastric cancer patients (American Joint Committee on Cancer stage IB: 25, II: 82, IIIA: 80, IIIB: 39 and IV: 48) who underwent adjuvant chemotherapy after curative resection (≥ D2 dissection). H. pylori infection status in hematoxylin and eosin stained corporal and antral mucosa of non-tumor tissue was graded according to the updated Sydney System and categorized as H. pylori negative (normal or mild infection) and H. pylori positive (moderate or marked infection). Eighty-one patients received 5-fluorouracil (5-FU) and doxorubicin-based chemotherapy, while 193 patients underwent 5-FU, mitomycin-C and polysaccharide-K chemotherapy. The median follow-up duration of survivors was 144 (120-184) months. In univariate analysis, patients with H. pylori negative status (108 patients) demonstrated significantly poor 10-year overall survival (OS) compared to those with H. pylori-positive status (166 patients; 21.3% vs. 71.1%, p < 0.0001). H. pylori negative status was associated with poor outcome in all stages except stage IIIB. In multivariate analysis, H. pylori-negative status was the most significant independent prognostic factor of poor OS (hazard ratio: 3.45, 95% confidence interval: 2.43-4.89, p < 0.0001) followed by old age (>54 years, p < 0.0001), advanced stage (stage III or IV, p = 0.001), and Borrmann type IV (p = 0.027). H. pylori infection status seems to have strong prognostic significance in locally advanced gastric cancer. H. pylori-negative patients may need careful follow-up after curative resection.     

抗幽门螺杆菌治疗在肿瘤化疗止吐中的作用研究

  目的   评估抗幽门螺杆菌（Helicobacter pylori,HP）治疗及联合常规止吐药物对肿瘤患者化疗所致胃肠道不良反应的疗效。   方法   2010年1月至2011年1月,共纳入176例患者,其中伴有幽门螺杆菌感染患者86例。将HP感染阳性者随机分为A组和B组。A组43例给予抗幽门螺杆菌联合止吐药物（奥美拉唑20 mg+克拉霉素500 mg+替硝唑500 mg,2次/d,口服7 d;联合托烷司琼5 mg,静脉滴注,1次/d;疗程共4周）治疗,B组43例仅给予止吐药物（托烷司琼5mg,疗程共4周）治疗,两组患者在性别、年龄、临床表现无差别。应用WHO胃肠道反应分度标准进行观测和评价,应用14C-尿素呼气试验评估幽门螺杆菌根除率。   结果   接受化疗并伴有幽门螺杆菌感染的患者呕吐程度比未感染者严重,其中Ⅲ~Ⅳ度呕吐（χ2=21.92,P < 0.001）,Ⅰ~Ⅱ度呕吐（χ2=9.73,P < 0.01）差异均有统计学意义。联合抗菌治疗患者的恶心、呕吐等症状比常规止吐患者减轻,A组治疗恶心、呕吐的总有效率为81.39%（35/43）,明显高于B组的58.14%（25/43）,差异有统计学意义（χ2=4.46,P < 0.05）。   结论   针对幽门螺杆菌阳性患者给予抗幽门螺杆菌联合常规止吐治疗,可以有效减轻和缓解化疗所致的胃肠道不良反应。      

Higher gastric cycloxygenase-2 expression and precancerous change in Helicobacter pylori-infected relatives of gastric cancer patients.

PURPOSE: This study was conducted to determine whether relatives of gastric cancer patients (GCF) showed greater gastric cycloxygenase-2 (COX-2) expression or a greater incidence of precancerous lesions after Helicobacter pylori infection and whether H. pylori eradication could reduce COX-2 expression. EXPERIMENTAL DESIGN: Three hundred subjects were enrolled in this study: half were relatives of 50 H. pylori-infected gastric cancer patients, and half were relatives of 50 H. pylori-infected duodenal ulcer (DU) patients (controls). Each relative underwent endoscopy to detect H. pylori infection and related gastric histology. One hundred and twenty GCFs were found to have H. pylori infection. After H. pylori eradication, 90 of the 120 GCFs were followed up with annual endoscopy examinations over the next 2 years. Gastric COX-2 intensity in all of the specimens collected from these patients was immunochemically stained and graded from 0 to 4. RESULTS: H. pylori infection, gastric atrophy, and intestinal metaplasia (IM) were more prevalent in GCFs than in relatives of H. pylori-infected patients with DUs (P < 0.05). H. pylori-infected GCFs also showed a greater COX-2 intensity than H. pylori-infected relatives of patients with DUs (89.1% versus 62.7%, P < 0.001; relative risk: 4.9; 95% confidence interval: approximately 2.34-10.29). Among the H. pylori-infected GCFs, COX-2 intensity correlated with atrophy and IM (P < 0.001). After H. pylori eradication, gastric COX-2 expression disappeared only in those relatives without IM (P < 0.001). CONCLUSIONS: GCFs are more likely to show greater gastric COX-2 expression and a higher incidence of precancerous lesions after H. pylori infection than the relatives of H. pylori-infected patients with only DUs. H. pylori eradication can reverse gastric COX-2 expression in patients without IM but not in patients with IM.