幽门螺杆菌感染对Fas/FasL表达的影响在胃癌发生中的作用

目的：观察幽门螺杆菌（Hp）及其不同毒力株（cagA阳性与cagA阴性株）感染对胃黏膜上皮细胞Fas/FasL表达的影响，进而探讨胃癌的发生机制。方法：胃镜下取胃窦黏膜标本，将研究对象按病理结果分为黏膜萎缩组，黏膜萎缩伴轻度不典型增生组，黏膜萎缩伴中度不典型增生组，胃腺癌组，黏膜大致正常组为对照组，再根据Hp感染情况为分Hp阳性组与阴性组，并将Hp阳性组进一步成cagA阳性组及阴性组，共9组80例。以快速尿素酶试验，PCR及组织学第三种方法检测Hp，用PCR方法对Hp进行分型。用免疫组化法检测Fas、FasL等表达情况。结果Hp感染率为60．0％，cagA阳性率为90．47％，非腺癌病人Hp阳性组Fas/FasL表达明显高于Hp阴性组（P＜0．05），腺癌组Fas/FasL表达明显高于大致正常组及黏膜萎缩，黏膜萎缩伴轻度不典型增生，黏萎缩伴中度不典型增生组（P＜0．01）。cagA阳性组Fas/FasL表达与cagA阴性组差异无显著性（P＞0．05）。结论：幽门螺杆菌感染后早期即黏 萎缩阶段已出现Fas、FasL等的表达增加，随细胞凋亡的增加，黏膜上皮细胞萎缩加重，细胞DNA不稳定性增加，并出现不典型增生加重， Fas、FasL的表达随之增强，一旦肿瘤细胞形成，Fas/FasL表达进一步增加，形成局部免疫豁免区，导致肿瘤的浸润生长。cagA阳性的菌株在促成肿瘤的发生过程中无明显作用。     

幽门螺杆菌促进程序性死亡配体1在胃癌中的表达

目的探讨幽门螺杆菌(Hp)引起程序性死亡配体(PD-L)1在胃癌细胞中表达变化情况,并试图阐明Hp诱导胃癌发生的机制。方法收集感染或未感染Hp患者胃黏膜标本,使用Hp同SGC-7901细胞系及人原代胃上皮细胞共培养,采用RTPCR法检测Hp感染和未感染患者胃黏膜PD-L1表达,干扰素(IFN)-γ和肿瘤坏死因子(TNF)-α对AGS细胞中PD-L1表达的影响。采用酶联免疫吸附实验测定Hp上调CD4+T细胞分泌IFN-γ和TNF-α表达及Hp刺激后AGS细胞促进CD4+T细胞的凋亡率。结果 Hp阳性组PD-L1表达显著高于Hp阴性组(P0.05)。与对照组相比,IFN-γ和TNF-α促进AGS细胞中PD-L1表达(P0.05)。与对照组相比,Hp上调CD4+T细胞分泌IFN-γ和TNF-α(P0.05)。与对照组相比,Hp刺激后的胃癌细胞促进T淋巴细胞凋亡(P0.05)。结论 Hp感染导致了PD-L1分子表达上升,可能是导致胃癌发生的免疫机制之一。     

幽门螺杆菌相关性胃癌患者局部Th1/Th2细胞免疫应答变化

背景:幽门螺杆菌(Hp)感染与胃癌发生相关,免疫应答可能是其主要致病机制之一。目的:探讨Hp相关性胃癌患者黏膜局部Th1/Th2细胞免疫应答变化。方法:选取2011年1～6月27例胃癌患者和28例对照者,采用14C-尿素呼气试验、Hp抗体检测Hp感染,免疫组化法检测胃黏膜局部干扰素(IFN)-γ和白细胞介素(IL)-4表达情况。结果:Hp总体感染率为61.8%。Hp阳性患者中,胃癌组IFN-γ表达明显低于对照组(P<0.05),IL-4表达明显高于对照组(P<0.05);Hp阴性患者中,胃癌组IFN-γ、IL-4表达与对照组均无明显差异(P>0.05)。Hp阳性胃癌患者IL-4表达明显高于Hp阴性胃癌患者(P<0.05),Hp阳性对照组IFN-γ表达明显高于Hp阴性对照组(P<0.05)。结论:Hp阳性对照者以Th1细胞介导的免疫应答为主,Hp阳性胃癌患者以Th2细胞为主。黏膜局部Th1/Th2细胞免疫应答变化可能与Hp感染相关疾病的进展,特别是胃癌的发生密切相关。     

抗Fas单克隆抗体诱导人胃癌细胞系SGC-7901细胞凋亡

目的探讨抗Fas单克隆抗体诱导胃癌细胞凋亡的规律及在胃癌治疗中的意义.方法应用细胞形态观察、琼脂糖凝胶电泳、流式细胞光度术检测抗Fas 单克隆抗体对胃癌细胞SGC-7901增殖周期的影响以及对细胞杀伤作用的方式,并检测了SGC -7901细胞表面Bcl-2的表达情况..结果抗Fas单克隆抗体有阻滞细胞周期、通过诱发凋亡而抑制肿瘤细胞生长的作用. 经抗Fas单克隆抗体处理后,SGC-7901细胞表面Bcl-2蛋白表达无明显变化. .结论抗Fas单克隆抗体可以诱导胃癌细胞系SGC-7901细胞凋亡. 抗Fas单克隆抗体诱导胃癌细胞凋亡与Bcl-2表达无关.     

IL-12B mRNA在儿童及青少年Hp相关性胃十二指肠疾病中的表达

目的 研究白细胞介素12B(IL-12B)mRNA在儿童及青少年幽门螺杆菌(Hp)相关性胃十二指肠疾病胃黏膜中的表达.方法 采用RT-PCR方法检测胃黏膜IL-12B mRNA表达.血清Hp-IgG抗体检测、PCR、尿素酶实验及组织学检查作为确定Hp感染状态的依据.结果 儿童及青少年Hp感染者胃窦黏膜IL-12B mRNA阳性表达率、相对表达水平均较无Hp感染者高.IL-12B mRNA相对表达水平与胃窦黏膜慢性炎症程度呈正相关.结论 儿童及青少年Hp相关性胃十二指肠疾病胃窦黏膜IL-12B mRNA表达上调可能促进Th1细胞免疫反应从而增加黏膜破坏.     

幽门螺杆菌相关胃炎中细胞因子分泌与Fas抗原的原位关系研究

目的　原位检测幽门螺杆菌 (Hp)感染时胃粘膜活检标本中肿瘤坏死因子 α(TNF α)和白细胞介素 1β(IL 1β)的产生及Fas抗原的表达 ,并分析它们之间的关系。 方法　采集 13例Hp阴性受检者和 3 2例Hp阳性受检者的胃粘膜活检标本 ,通过放免法、免疫组化法观察并比较两组胃粘膜活检标本中TNF α和IL 1β的产生及Fas抗原的表达。 结果　胃粘膜活检标本中TNF α和IL 1β的分泌在Hp阳性组显著高于Hp阴性组 (P <0 .0 1) ,但与胃粘膜的病理分型无显著相关性 (P >0 .0 5 ) ;Hp阳性组的胃粘膜组织切片中TNF α、IL 1β和Fas抗原表达显著高于Hp阴性组 ;Hp阳性组中TNF α和IL 1β产生与胃粘膜Fas抗原的表达呈显著正相关 (P <0 .0 5 )。 结论　通过原位检测发现Hp感染可以使局部细胞因子TNF α和IL 1β产生增加 ,造成局部的炎症微环境 ,从而上调Fas抗原表达 ,导致胃粘膜损伤     

幽门螺杆菌感染者胃粘膜癌前病变与Fas抗原表达的关系

目的研究幽门螺杆菌(Hp)感染后胃粘膜癌前病变中 Fas 抗原表达的情况,了解 Hp 在胃癌发生过程中的作用。方法采用免疫组织化学等方法检测83例经病理证实为慢性胃炎病人胃粘膜上皮细胞中 Fas 抗原的表达情况。结果在浅表性胃炎、萎缩性胃炎、肠化生及异型增生中,Fas 抗原表达率分别为20.00%、36.36%、73.33%、43.75%,Fas 抗原在肠化生中的表达率显著高于浅表性胃炎、萎缩性胃炎及异型增生(P<0.01及P<0.05)。Hp 感染者 Fas 抗原表达率为60.71%,显著高于 Hp 阴性者的22.22%(P<0.01)。在萎缩、肠化生及异型增生等癌前病变中,Hp 感染者与未感染者表达率分别为65.96%及28.57%(P<0.01)。结论 Hp 感染对 Fas 抗原表达有一定的影响,Hp 感染可促进 Fas 抗原表达增加,这可能是 Hp 感染诱导胃粘膜上皮细胞凋亡的机制之一。     

幽门螺杆菌感染对循环Ghrelin及胃黏膜Proghrelin的影响

目的 研究循环Ghrelin及胃黏膜Proghrelin表达与幽门螺杆菌(Hp)感染及胃溃疡、胃腺癌等Hp感染相关疾病之间的关系.方法 使用放射免疫分析及RT-PCR测定10例Hp感染阴性健康志愿者及35例Hp感染阳性患者(其中胃溃疡11例,胃癌13例,无明显病灶者11例)的血浆Ghrelin水平及胃黏膜Proghrelin mRNA的表达.结果 在本研究中,各种Hp表达阳性的病例中循环Ghrelin水平及胃黏膜Proghrelin mRNA的表达要低于Hp阴性的健康志愿者(P＜0.05),与其他Hp感染患者相比,胃癌患者循环Ghrelin水平更低(P＜0.05),而胃黏膜Proghrelin mRNA的表达却无明显差异.结论 Hp能够降低胃黏膜Proghrelin的表达以及循环Ghrelin水平.     

幽门螺杆菌阳性胃癌P53和Fas的表达意义

胃癌是多种癌基因多阶段途径协同作用的结果,幽门螺杆菌(Hp)是胃癌发生的重要协同致病因子,我们观察了Hp阳性的胃癌组织的P53和Fas抗原的表达,探讨Hp在胃癌发生中与某些因素的作用。 1 材料和方法 1.1 材料 1996/1997本院手术切除胃癌25例,分2组,甲组:Hp阳性胃癌9例,男6例,女3例,年龄29岁～76岁,平均49.7岁,胃癌部位在窦部8例,贲门1例。乙组:Hp阴性胃癌16例,男14例,女2例,年龄29岁～81岁,平均53.5岁,胃窦部12例,胃体、贲门各2例。 1.2 方法胃癌组织标本常规制作,Hp检测用胃粘膜快速尿素酶试验和胃粘膜切片HE染色查菌。P53和Fas检测用免疫组化染色法,链霉菌抗生物蛋白—过氧化物酶连接法(S-P法)进行,Fas抗体、P53抗体和S-T试剂盒由美国Zymed和Santa     

幽门螺杆菌相关胃炎中TNF-α分泌和凋亡的原位关系研究

幽门螺杆菌（H．pylori）被认为是慢性胃炎、消化性溃疡、胃淋巴瘤和胃癌的主要致病因子。H．pylori侵袭、定居于胃黏膜，刺激机体产生细胞和体液免疫反应，可以造成局部炎症微环境，影响胃黏膜组织结构。本研究中，我们旨在观察H．pylori感染的胃黏膜标本局部细胞因子的产生，以及在相同活检标本组织切片中细胞因子TNF—α．和Fas抗原的表达以及胃黏膜细胞凋亡的关系，进一步证实麒pylori感染在胃黏膜局部通过免疫介导的凋亡反应导致胃上皮的损伤作用。     

Inhibition of apoptosis in human neutrophils by Helicobacter pylori water-soluble surface proteins.

BACKGROUND: Helicobacter pylori infection in humans causes persistent neutrophil infiltration into the gastric mucosa. It is believed that a prolongation of neutrophil life-span could contribute to the pathogenesis of H. pylori infection. We therefore examined whether the water-soluble surface proteins of H. pylori can influence the apoptosis of neutrophils. METHODS: After neutrophils were incubated with H. pylori water extract (HPWE), neutrophil apoptosis was evaluated by TUNEL assay, Hoechst 33342 staining, electron microscopy and ELISA for cytosolic oligonucleosome-bound DNA for up to 48 h. To investigate the regulatory mechanisms of neutrophil apoptosis associated with HPWE, mRNA expression and protein production of Fas, Fas ligand (FasL) and tumor necrosis factor receptor 1 (TNF-R1) were analyzed by RT-PCR, ribonuclease protection assay, Northern blot and Western blotting. Cell surface expression of these death factors was also measured by flow cytometry. RESULTS: HPWE inhibited neutrophil apoptosis and cytotoxicity for up to 48 h. The mRNA and protein expression of FasL and the cell surface expression of Fas, FasL and TNF-R1 in HPWE-treated neutrophils were suppressed compared with the controls. CONCLUSION: The water-soluble surface proteins of H. pylori could suppress neutrophil apoptosis. This may be caused by the suppression of FasL expression in neutrophils and Fas, FasL and TNF-R1 expression on the surface of neutrophils.     

Helicobacter pylori enhances tumor necrosis factor-related apoptosis-inducing ligand-mediated apoptosis in human gastric epithelial cells

AIM: To investigate the relations between tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and Helicobacter pylori (H pylori) infection in apoptosis of gastric epithelial cells and to assess the expression of TRAIL on the surface of infiltrating T-cells in H pylori-infected gastric mucosa. METHODS: Human gastric epithelial cell lines and primary gastric epithelial cells were co-cultured with H pylori in vitro, then recombinant TRAIL proteins were added to the culture. Apoptosis of gastric epithelial cells was determined by a specific ELISA for cell death. Infiltrating lymphocytes were isolated from H pylori-infected gastric mucosa, and expression of TRAIL in T cells was analyzed by flow cytometry. RESULTS: The apoptosis of gastric epithelial cell lines and primary human gastric epithelial cells was mildly increased by interaction with either TRAIL or H pylori alone. Interestingly, the apoptotic indices were markedly elevated when gastric epithelial cells were incubated with both TRAIL and H pylori (Control vs TRAIL and H pylori: 0.51+/-0.06 vs 2.29+/-0.27, P = 0.018). A soluble TRAIL receptor (DR4-Fc) could specifically block the TRAIL-mediated apoptosis. Further studies demonstrated that infiltrating T-cells in gastric mucosa expressed TRAIL on their surfaces, and the induction of TRAIL sensitivity by H pylori was dependent upon direct cell contact of viable bacteria, but not CagA and VacA of H pylori. CONCLUSION: H pylori can sensitize human gastric epithelial cells and enhance susceptibility to TRAIL-mediated apoptosis. Modulation of host cell sensitivity to apoptosis by bacterial interaction adds a new dimension to the immunopathogenesis of H pylori infection.     

Homing commitment of lymphocytes activated in the human gastric and intestinal mucosa

BACKGROUND: Gastric infection with the human pathogen Helicobacter pylori results in a large accumulation of IgA and IgM secreting cells in the gastric mucosa. The molecular mechanisms resulting in B cell migration to the gastric mucosa in H pylori infection are however not known. AIMS: To examine expression of the mucosal homing receptor integrin alpha4beta7 and the homing receptor for secondary lymphoid tissues, L-selectin, on lymphocytes activated by gastric, intestinal, or systemic antigens. Furthermore, to examine gastric expression of the mucosal addressin cellular adhesion molecule 1 (MAdCAM-1), the endothelial counter-receptor to integrin alpha4beta7. SUBJECTS AND METHODS: H pylori infected individuals were immunised by either gastric (n=8) or intestinal (n=8) delivery of an inactivated cholera vaccine. The resulting circulating vaccine specific B cells were sorted according to alpha4beta7 and L-selectin expression and assayed for production of IgA and IgG using an enzyme linked immunospot assay. In addition, circulating CD4+ T cells from seven H pylori infected individuals were fractionated according to alpha4beta7 and L-selectin expression. The resulting T cell fractions were then assayed for specific proliferation against H pylori or the systemic antigen tetanus toxoid. Finally, gastric expression of MAdCAM-1 was determined by immunohistochemistry in H pylori infected (n=16) and uninfected (n=8) individuals. RESULTS: Virtually all B cells induced by both gastric and intestinal antigen delivery expressed alpha4beta7 whereas less then half coexpressed L-selectin. Furthermore, H pylori reactive T cells were mainly found in the alpha4beta7+L-selectin+ T cell fraction whereas tetanus specific T cells were largely alpha4beta7-L-selectin+. MAdCAM-1 was present in similar amounts in gastric mucosa from H pylori infected and uninfected individuals. CONCLUSIONS: B cells and T cells activated by antigens delivered to the gastric mucosa express the mucosal homing receptor integrin alpha4beta7, as do cells activated in the intestine. Together with the observation that gastric endothelial cells express MAdCAM-1, this may partly explain the homing of lymphocytes activated in the stomach or in the small intestine to the gastric mucosa.     

Transactivation of the EGFR by AP-1 is induced by Helicobacter pylori in gastric cancer

BACKGROUND: Helicobacter pylori infection of the gastric mucosa is strongly associated with gastritis, peptic ulcer disease, and gastric cancer. However, the mechanisms by which H. pylori causes cancer are currently unknown. Binding of epidermal growth factor (EGF) to its receptor (EGFR) may be important in the development of gastric cancer. This interaction accelerates cell proliferation and migration, and triggers epithelial cell signaling. In this study, we investigated the effects of H. pylori on EGFR- and AP-1-mediated signal transduction pathways in the AGS gastric epithelial cell line and gastric tissue from humans. METHODS: Cells were treated with H. pylori and cell death was examined at a variety of time points using cell viability and trypan blue exclusion dye assay. To investigate the effects on EGFR regulation, AGS cells were transfected with a full-length and truncated EGFR luciferase (luc) reporter. Tissue microarray containing 44 samples of gastric biopsies from H. pylori-positive patients was analyzed for protein expression level of EGFR by immunohistochemistry. RESULTS: EGFR promoter activity was increased (twofold) 3 h after treatment with H. pylori commenced. Using a series of EGFR promoter deletion mutants, we identified a region that was crucial for transactivation of the EGFR by H. pylori. To determine whether AP-1 binding was altered, we transfected AGS cells with an AP-1 luciferase construct and then treated them with H. pylori for up to 6 h. We found that AP-1 activity was induced by H. pylori in gastric cells, while electrophoretic mobility shift assays confirmed that binding of AP-1 to the EGFR promoter site was increased following H. pylori treatment. Binding of c-Jun and c-Fos to the EGFR promoter region -1,062/-900 was induced eight- and six fold, respectively, using ChIP assay. Active EGFR staining was markedly increased in gastric mucosa from infected persons, compared to uninfected controls. CONCLUSIONS: We conclude that exposure of gastric cells to H. pylori induces increased production of EGFR through various signal transduction pathways, including those mediated by the EGFR and AP-1. Distinct effects on EGFR activation may specify the subset of AP-1 target genes that are selected, including those involved in proliferation and apoptosis. This is consistent with EGFR activation that was found in the gastric mucosa of humans infected with H. pylori. Hence, the balance between apoptosis and proliferation in these cells may be altered in response to injury caused by H. pylori infection, leading to an increased risk of cancer.     

Fas ligand expression in colon cancer： A possible mechanism of tumor immune privilege

AIM: To detect the expression of Fas ligand (FasL) in colon cancer tissues and cell lines and analyze the function of FasL-expressing colon cancer cells in inducing Fas-sensitive T lymphocyte apoptosis. METHODS: Ninety surgically resected colon cancer tissues and 15 hepatic metastasis specimens were investigated by immunohistochemical method with normal colon mucosa and colon adenoma as control. The relationship between FasL expression and pathologic features was also analyzed. FasL expression of 4 colon cancer cell lines, SW620, Lovo, LS-174T and SW1116, were detected by Western blotting assay. The function of FasL expressed on colon cancer cells was determined by coculture assay with Jurkat T lymphocytes, the apoptotic rate of which was detected by flow cytometry assay. RESULTS: Fifty-six (62.22%) cases of all the 90 colon cancer tissues and all (100%) the liver metastasis specimens expressed FasL, significantly higher than normal colon mucosa and colonic adenoma. Higher expression of FasL was found in more advanced stage of colon cancer and in cancer tissues with lymphatic or hepatic metastasis. All the colon cancer cell lines were found to express FasL. After coculture with the SW1116 cells for 24 h with an effector: target ratio 10:1, the rate of apoptosis of Jurkat cells rose from 1.9% to 21.0%. CONCLUSION: The expression of FasL is upregulated in colon cancer and the functionally expressed FasL can induce apoptosis of Fas-expressing T lymphocytes.