幽门螺杆菌尿素酶B亚单位和热休克蛋白A亚单位的融合表达

目的将幽门螺杆菌尿素酶B亚单位基因和热休克蛋白A亚单位基因融合,并实现其在大肠杆菌中的表达,为构建该菌的基因工程二价苗打下基础.方法通过交叠延伸拼接PCR法将幽门螺杆菌ureB基因和hspA基因通过(Gly4Ser)3linker连接起来,经NcoⅠ和NotⅠ双酶切后定向克隆入表达载体pET22b+的pelB前导序列下游,经测序鉴定后转化大肠杆菌BL31(DE3),用1PTG诱导表达.结果幽门螺杆菌的尿素酶B亚单位基因和热休克蛋白A亚单位基因通过linker获得了融合,序列分析表明,此融合基因序列拼接正确,且是通读的.含有该融合基因表达载体的表达菌株BL31(pET-BA)在30℃经IPTG诱导表达4h后,用10%SDS-PAGE电泳分析表明,在细菌周质表达有约与预期融合基因表达蛋白大小一致的外源蛋白质表达带.结论幽门螺杆菌ureB与hspA成功地实现了在大肠杆菌中的融合表达,为进一步评价其对Hp的免疫保护效果和构建Hp二价基因工程苗打下了基础.

庄河16村镇胃癌高危人群幽门螺杆菌感染状况普查

目的幽门螺杆菌（Helicobacterpylori，Hp）感染与胃癌的发生关系密切．我们剖析胃癌高发区Hp感染状况及其与胃疾病及胃癌的相关性，旨在为Hp感染及胃癌的综合防治提供依据．方法胃癌高发区16村镇胃癌高危人群3033人，经血清胃蛋白酶原检测及X线初筛出1779例，接受了胃粘膜活检精查．其切片作HE及亚甲兰（MB）染色，观察病理组织学改变及Hp感染情况，同时利用PCR检测Hp．结果Hp总检出率为60．8％．其中男高于女；30岁～59岁组最高（62．4％）；山区（64．4％）高于沿海区（56．7％）；胃窦（36．2％）高于胃角（32．6％）及胃体（31．2％）；以上P＜0．01．胃癌Hp检出率为43．8％；各型胃疾病Hp检出率由高至低，前四位依次为胃粘膜糜烂（90．6％）、胃溃疡（87．0％）、中重度浅表性胃炎（79．8％）及萎缩性胃炎（75．3％），上述各组Hp检出率与轻度浅表性胃炎Hp检出率（6．8％）相比，P＜0．01．与未检出Hp的组织学基本正常胃粘膜0％相比，异型增生（80．0％）及肠上皮化生（77．7％）的Hp检出率皆明显升高，P＜0．01．结论庄河居民Hp感染与胃粘膜糜烂、胃溃疡、中重度浅表性胃炎、萎缩性胃炎等呈明显正相关．庄河不仅胃癌高发且Hp感染亦是高流行．文内尚对有关结果只行了初步探讨。

低剂量^（13）C－尿素呼吸试验检测幽门螺杆菌感染

对６０例有上消化道症状而就医的患者作１３Ｃ－尿素呼吸试验检测幽门螺杆菌感染。分别进行１３Ｃ－尿素１００，５０，２５ｍｇ３组不同剂量的呼吸试验，并以细菌培养、病理组织学、尿素酶试验作为参考标准以对比。结果该呼吸试验敏感性为１００％，特异性为９５．８％，３组之间无明显差异，与参考标准的符合率为９７．５％。研究结果显示低剂量１３Ｃ－尿素呼吸试验是检测幽门螺杆菌感染的快速、安全、准确的非侵入无创性方法。

幽门螺杆菌染色体DNA片段的分子克隆

目的幽门螺杆菌（Helichacterpylori,Hp）与慢性胃炎、消化性溃疡有关．构建含HP染色体DNA片段克隆做为阳性对照，在基础研究和临床检测中都有一定意义．方法用PCR扩增Hp染色体片段，克隆至BluescriptSK＋载体中．用IMR及DNA测序进行鉴定．结果用PCR可扩出203hp的Hp特异带，DNA测序证实序列完全正确，表明克隆成功．

幽门螺杆菌感染对胃粘膜增殖活性的影响

目的观察幽门螺杆菌（Hp）感染与胃粘膜增殖活性的关系．方法慢性胃炎活检标本68例，用W－S银染法、0．25％复红法和HpDNAPCR技术检测Hp，用免疫组化法观察增殖细胞核抗原（PCNA），C-erb-B2和p53基因的表达，按标准化方案对核仁组成区嗜银蛋白（AgNOR）颗粒计数．结果Hp阳性组（30例）和阴性组（38例）中InyNAⅠ，Ⅱ，Ⅲ，Ⅳ级分别为5，7，4，14例和27，11，0，0例，AgNOR计数分别为3．44±1.20个和1．08±0．08个，差异有显著性（P＜0．05）．Hp阳性组中有4例C-erb-B2在新生腺体和不全肠化柱状细胞中局部阳性，Hp阴性组中未见阳性表达结论Hp感染时胃粘膜增殖活性增强，这可能是胃癌发生的高危因素之一．

幽门螺杆菌与胃癌的关系

目的为了探索幽门螺杆菌（Hp）与胃癌及其病理类型的关系．方法对住院的经内镜及病理证实的23例胃癌，21例慢性浅表性胃炎及26例十二指肠溃疡患者进行冒窦粘膜Hp检测（采用快速尿素酶试验）及血清Hp-IgG抗体测定（采用ELISA法）．结果Hp在慢性浅表性胃炎、十二指肠溃疡、中晚期胃癌的检出率分别为52．4％，88．5％及78．3％．Hp在十二指肠溃疡及慢性浅表性胃炎的感染率具有明显差异（P＜0．01），而Hp在中晚期胃癌及慢性浅表性胃炎的感染率无差异（P＞0．05）．胃腺癌的Hp检出率高于印戒细胞癌的Hp检出率（P＜0．05）．结论①Hp与中晚期胃癌无明显相关性．②胃腺癌患者的Hp检出率较胃印戒细胞癌者的高．

胃癌高发区cagA^(+)幽门螺杆菌的感染

目的研究胃癌高发区不同人群幽门螺杆菌（Ｈｐ）ｃａｇＡ＋亚型感染状况，探讨ＨｐｃａｇＡ＋亚型感染与胃癌高发流行的关系．方法用ＥＬＩＳＡ方法检测了全部受检者的抗Ｈｐ抗体及抗ｃａｇＡ抗体，并用１３Ｃ尿素酶呼吸实验（１３Ｃ－ＵＢＴ）方法检测了７０名儿童Ｈｐ感染状况．结果胃癌高发区成人Ｈｐ感染率为７４７％，儿童Ｈｐ感染率为６８６％；成人ｃａｇＡ＋亚型感染率为６５％，儿童为７１４％．７７名癌症患者的Ｈｐ和ｃａｇＡ＋亚型感染率均为５９７％，其中胃癌患者（ｎ＝３８）Ｈｐ和ｃａｇＡ＋亚型感染率分别为５０％和５５３％，均低于其他癌症患者（ｎ＝３９）的感染率（６９２％和６４１％）．结论胃癌高发区成人、儿童及癌症患者ＨｐｃａｇＡ＋亚型感染率相当高，表明Ｈｐ感染，尤其是ｃａｇＡ＋的Ｈｐ感染与当地胃癌高发有明显的关系．

胃癌高发区儿童幽门螺杆菌感染及传播途径

目的研究胃癌高发区儿童幽门螺杆菌（Ｈｐ）和ｃａｇＡ＋亚型感染状况及细菌传播方式，探讨儿童早期感染Ｈｐ与胃癌的关系．方法用１３Ｃ尿毒酶呼吸实验（１３ＣＵＢＴ）方法检测（山东省临朐县）９８名儿童Ｈｐ感染状况，用ＥＬＩＳＡ方法检测７０名儿童抗ＣａｇＡ抗体及成人抗Ｈｐ抗体．结果胃癌高发区儿童Ｈｐ感染率为６９４％，其中３岁～４岁儿童Ｈｐ感染率为５２６％．９岁～１０岁上升为８５％，１１岁～１２岁下降为６６７％，接近于成人水平．儿童ｃａｇＡ＋亚型感染率为７１４％．Ｈｐ感染有明显的家族聚集性，其中父母双方或一方Ｈｐ感染阳性的子女Ｈｐ感染率（８５％）明显高于父母Ｈｐ感染均阴性者（２２％，Ｐ＜００５）．结论胃癌高发区儿童Ｈｐ、特别是ｃａｇＡ＋亚型感染率很高，且有家族聚集性，表明儿童阶段即可感染Ｈｐ，并且与当地胃癌高发有明显关系．

The Effect of Helicobacter Pylori Infection on Expression of hMSH2, hMLH1 and p53 in Gastric Carcinogenesis

Objective： To detect the expression of hMSH2, hMLH1 and p53 in gastric epithelial cells with and without Helicobactcr pylori （H. pylori） infection and investigate the relationship between H. pylori infection and these genes in gastric carcinogenesis. Methods： H. pylori infection was detected by rapid urease tests. Expression of hMSH2, hMLHland p53 in gastric cancer （GC） tissue, its adjacent mucosa, gastritic mucosa and normal mucosa was assessed by immunohistochemistry SP method. Results： Positive expression rate of hMSH2 in GC tissue （62.7%） was higher than those in adjacent mucosa （29.4%）, gastritic mucosa （32.4%） and normal mucosa （30.0%） （P〈0.001）. Positive rate of hMSH2 in poorly differentiated adenocarcinoma （76.4%） was higher than those in other carcinomas （54.3%, 53.1%） （P〈0.05）. Positive expression rate of hMLH1 in GC tissue （64.3%） mucosa （82.4%） and normal mucosa （80.0%） was lower than those in adjacent mucosa （84.4%）, gastritic （P〈0.01）. Positive rate of hMLH1 in mucoid carcinoma （43.7%） was lower than those in other carcinomas （78.2%, 64.7%） （P〈0.01）. Positive expression rate of p53 in GC tissue （51.9%） was higher than those in adjacent mucosa （3.1%）, gastritic mucosa （0.0%） and normal mucosa （0.0%） （P〈0.001）. Positive rate of p53 in well differentiated adenocarcinoma （32.6%） was lower than those in other carcinomas （58.8%, 68.7%） （P〈0.01）. Positive rates of hMSH2 and hMLH1 in GC with H. pylori infection were lower than those without the infection, respectively （P〈0.05）. Positive rate of p53 in GC with H. pylori infection （61.4%） was higher than that without the infection （40.6%） （P〈0.05）. Conclusion： Gastric carcinogenesis may be associated with abnormal expression of hMSH2, hMLH1 and p53; H. pylori infection affecting expression of these genes may be one of its carcinogenic mechanisms.

中国胃癌高发区庄河16村镇5万自然人口胃癌普查报告

目的为完成国家“九五”科技攻关计划──胃癌高发现场高危人群综合防治研究，对庄河地区16个村镇约5万自然人口进行了胃癌普查．方法普查对象为35岁以上，有胃病史及胃癌家族史者．普查方法包括临床流行病学调查，X－ray气钡双重造影，血清胃蛋白酶原检测，内镜及粘膜活检病理检查．结果实际接受各种检查者总计为3303人．结果发现，各型胃病患者检出率占受检入群的81．84％．查出胃癌患者32例，检出率为1．06％，其中早期胃癌18例，占胃癌检出总数的56．25％．经确诊的胃癌患者尤其是早期胃癌患者已陆续接受旱期治疗．除胃癌外，查出一批不同类型的胃病患者，包括胃粘膜上皮异型增生，肠上皮化生，萎缩性胃炎，浅表性胃炎，糜烂性胃炎及溃疡等，为下一步分组投药，干预治疗打下了良好基础．本次普查结果发现，庄河居民食用成猪肉相当普遍，食用者达93．97％．特别是在偏远山区，几乎达到100％，且持续数年受检群众冒粘膜幽门螺杆菌感染率60％以上结论庄河地区胃病及胃癌高发，庄河居民食用成猪肉及胃粘膜幽门螺杆菌感染相当普遍，针对上述胃癌危险因素进行行为干预及综合防治，切实可行．

Helicobacter Pylori-induced Gastritis Model in BALB/c Mice Infected With Fresh Isolates from a Human Complex Ulcer Patient

A useful helicobacter pylori-induced gastritis model using BALB/c mice was established for mimicking of human gastritis induced by Helicobacter pylori (H. pylori). The H. pylori isolates were obtained freshly from a human complex ulcer patient. BALB/c mice were fasted for 24 h and then 0.25 mL of 0.2 mol·L -1 NaHCO 3 was administered after by gavage to each mouse and 0.5 mL of 10 9 colonies formation unit per milliliter (CFU/mL) of H. pylori was administered 15 min. On the 3 rd day and 5 th day, the H. pylori inoculations were repeated. The inoculated mice were sacrificed in batch on the 5 th day, in the 2 nd week, 3 rd week and 4 th week. The gastric mucous membrane near pyloric portion was removed, treated and then cultured under microaerobic condition for detection of H.pylori. The remainders of the gastric membrane were fixed by 10% formaldehyde solution for pathological detection. The results showed that the H. pylori could be separated from the gastric membranes of inoculated mice. Obvious invasion of inflammatory cells in the gastric membranes of inoculated mice could be observed from pathological sections. It can be concluded that the inoculating fresh human H. pylori isolates can produce mouse gastritis. This model of BALB/c mice can be used for evaluating the therapeutic agents for the treatment of gastritis induced by H. pylori.

Reduced secretion of epidermal growth factor in duodenal ulcer patients with Helicobacter pylori infection

AIM To investigate the concentration changes of epidermal growth factor (EGF) in duodenal ulcer patients with H. pylori infection.

Ultrastructural observation on the relation of H.pylori to the gastric epithelia in chronic gastrictis and in peptic ulcer

AIMS The relationship between Helicobacter pylori (Hp) and gastric epithelia in chronic gastritis and in petic ulcer was studied by transmission electron mi- croscopy (TEM). METHODS Seventy-five gastric antral biopsy speci- mens from the patients examined by six other methods for Hp were fixed in glutaraldehyde and treated with tanin acid before OsO_4 staining than routinely prooessed for TEM studies (at least 4 semi- thin sections oriented for ultrathin sections in each sample). RESULTS The bacilli were detected by TEM within gastric mucosa in 53 of 55 patients infected with Hp. Ultrathin sections especially stained with tanin acid re- vealed clearly glycocalyx by which the bacillus was connected with the epithelium. As the bacilli grouped as colony and breed,the adjacent mucous cells degerated and characterized by erosion of the juxtalu- minal cytoplasm,vacuolation or blebs,even desqua- mation of cell. Evidence was accumulated to show that the baoilli were located in the lumen attracted neu- trophils which intended to migrate into intercellular space of epithelia or into the lumen to exert the effect of Hp phagocytosis. CONCLUSIONS The sensitivity and specificity of Hp diagnosis by TEM is respectively 96% and 95%. Tanin acid is suitable for the preservation of glycocalyx of cell. The colonized bacilli,usually with the wide periplasmic pools,contributed to the spectrum degen- eration of epithelia,including mucous neck cells. If Hp infection persists,the degeneration and regeneration of mucous neck cells alternatively carried on and ultimate- ly the generative stem cells were damaged,as the result,the chronic atrophy gastritis could occure.

Risk for gastric neoplasias in patients with chronic atrophic gastritis:A critical reappraisal

Chronic atrophic gastritis （CAG） is an inflammatory condition characterized by the loss of gastric glandular structures which are replaced by connective tissue （non-metaplastic atrophy） or by glandular structures inappropriate for location （metaplastic atrophy）. Epidemiological data suggest that CAG is associated with two different types of tumors： Intestinal-type gastric cancer （GC） and type I gastric carcinoid （T I GC）. The pathophysiological mechanisms which lead to the development of these gastric tumors are different, It is accepted that a multistep process initiating from Helico- bacterpylori-related chronic inflammation of the gastric mucosa progresses to CAG, intestinal metaplasia, dysplasia and, finally, leads to the development of GC. The T I GC is a gastrin-dependent tumor and the chronic elevation of gastrin, which is associated with CAG, stimulates the growth of enterochromaffin-like cells with their hyperplasia leading to the development of T I GC. Thus, several events occur in the gastric mucosa before the development of intestinatype GC and/ or T I GC and these take several years. Knowledge ofCAG incidence from superficial gastritis, its prevalence in different clinical settings and possible risk factors as- sociated with the progression of this condition to gastric neoplasias are important issues. This editorial intends to provide a brief review of the main studies regarding incidence and prevalence of CAG and risk factors for the development of gastric neoplasias.

Third-line rescue therapy for Helicobacter pylori infection

H pylori gastric infection is one of the most prevalent infectious diseases worldwide. The discovery that most upper gastrointestinal diseases are related to Hpylori infection and therefore can be treated with antibiotics is an important medical advance. Currently, a first-line triple therapy based on proton pump inhibitor （PPI） or ranitidine bismuth citrate （RBC） plus two antibiotics （darithromycin and amoxicillin or nitroimidazole） is recommended by all consensus conferences and guidelines. Even with the correct use of this drug combination, infection can not be eradicated in up to 23% of patients. Therefore, several second line therapies have been recommended. A 7 d quadruple therapy based on PPI, bismuth, tetracycline and metronidazole is the more frequently accepted. However, with second-line therapy, bacterial eradication may fail in up to 40% of cases. When Hpylori eradication is striclly indicated the choice of further treatment is controversial. Currently, a standard third-line therapy is lacking and various protocols have been proposed. Even after two consecutive failures, the most recent literature data have demonsbated that Hpylori eradication can be achieved in almost all patients, even when antibiotic susceptibility is not tested. Different possibilities of empirical treatment exist and the available third-line strategies are herein reviewed.

Helicobacter pyloripromote gastric cancer cells invasion through a NF-κB and COX-2-mediated pathway

AIM: To examine the effects of Helicobacter pylori(Hpylori) infection on the invasiveness of gastric cancer cells,and to elucidate its mechanism. METHODS: Gastric carcinoma cells, MKN-45, were incubated with CagA-positive H pylori, and cell invasion was determined by Matrigel analysis.The expression of matrix metallopr-oteinase-9 (MMP-9),vascular endothelial growth factor (VEGF), and cyclooxygenase-2 (COX-2) were assessed by Western-blot analysis, and transcriptional activation of the COX-2 promoter was examined by measuring luciferase and β-galactosidase activities. Lastly,the proteinDNA interaction was confirmed by an electrophoretic mobility shift assay. RESULTS: The current studies showed that: (1) incubation of CagA-positive H pylori with MKN-45 cells significantly promotes gastric cancer cells invasion, and this effect is attenuated by pre-treatment with NS-398, a COX-2 inhibitor, or PDTC,a nuclear factor κB (NF-κB) inhibitor;(2) the induction of MKN-45 cells invasion by Hpylori is associated with increases in COX-2, MMP-9, and VEGF protein expression, and co-incubation of NS-398 or PDTC significantly reduces these effects;(3) H pylori infection transactivates COX-2 promoter activity and increases the binding of NF-κB to this promoter. CONCLUSION: Our data demonstrate that H pylori infection promotes gastric epithelial cells invasion by activating MMP-9 and VEGF expression. These effects appear to be mediated through a NF-κB and COX-2 mediated pathway, as COX-2 or NF-κB inhibitor significantly attenuate the invasiveness of gastric cancer cells and the expressions of MMP-9 and VEGF protein.

Cellular and molecular aspects of gastric cancer

Gastric cancer remains a global killer with a shifting burden from the developed to the developing world. The cancer develops along a multistage process that is defined by distinct histological and pathophysiological phases. Several genetic and epigenetic alterations mediate the transition from one stage to another and these include mutations in oncogenes, tumour suppressor genes and cell cycle and mismatch repair genes. The most significant advance in the fight against gastric caner came with the recognition of the role of Helicobacter pylori （H pylori） as the most important acquired aetiological agent for this cancer. Recent work has focussed on elucidating the complex host/microbial interactions that underlie the neoplastic process. There is now considerable insight into the pathogenesis of this cancer and the prospect of preventing and eradicating the disease has become a reality. Perhaps more importantly, the study of H pylori-induced gastric carcinogenesis offers a paradigm for understanding more complex human cancers. In this review, we examine the molecular and cellular events that underlie Hpyloriinduced gastric cancer.

Serum pepsinogen levels and their influencing factors:A population-based study in 6990 Chinese from North China

AIM: To explore the essential characteristics of serum pepsinogen (PG) levels in Chinese people, by analyzing the population-based data on the serum levels of PG Ⅰ and Ⅱ and the PGⅠ/Ⅱ ratio, and their influencing factors in Chinese from North China. METHODS: A total of 6990 subjects, who underwent a gastric cancer screening in North China from 1997 to 2002, were collected in this study. Serum pepsinogen levels were measured by enzyme-linked immunosorbent assay (ELISA). H pylori status was determined by histological examination and H pylori-IgG ELISA. The cut-off point was calculated by using receiving operator characteristics (ROC) curves. Factors linked to serum PG Ⅰ/Ⅱ ratio were identified using a multivariate logistic regression. RESULTS: The serum PGⅠ and PGⅡ levels were significantly higher in males than in females (95.2 μg/L vs 79.7 μg/L, P < 0.01; 12.1 μg/L vs 9.4 μg/L, P < 0.01), PGⅠ/Ⅱ ratio was significantly lower in males than in females (7.9 vs 8.3, P < 0.01). The PG Ⅰ/Ⅱ ratio decreased significantly in the aged groups following the progression of gastric mucosa from normal to non-atrophic and atrophic lesions (10.4, 8.8, and 6.6, respectively). The serum PGⅠand Ⅱ levels were significantly higher in patients with H pylori infection than in those without H pylori infection (88.7 μg/L vs 81.4 μg/L, P < 0.01; 11.4 μg/L vs 8.4 μg/L, P < 0.01), while the PGⅠ/Ⅱ ratio was significantly lower in patients with H pylori infection than in those without H pylori infection (7.7 vs 9.6, P < 0.01). For patients with atrophic lesions, the area under the PGⅠ/Ⅱ ROC curve was 0.622. The best cut-off point for PGⅠ/Ⅱ was 6.9, with a sensitivity of 53.2%, and a specificity of 67.5%. Factors linked to PGⅠ/Ⅱ were sensitive to identified PG using a multinomial logistic regression relying on the following inputs: males (OR: 1.151, 95% CI: 1.042-1.272, P = 0.006), age ≥ 61 years (OR: 1.358, 95% CI: 1.188-1.553, P = 0.000), atrophic lesion (OR: 2.075, 95% CI: 1.870-2.302, P = 0.000), and H pylori infection (OR: 1.546, 95% CI: 1.368-1.748, P = 0.000). CONCLUSION: The essential characteristics of serum PG levels in Chinese are significantly skewed from the normal distribution, and influenced by age, sex, gastric mucosa lesions and H pylori infection. PGⅠ/Ⅱ ratio is more suitable for identifying subgroups with different influence factors compared with PGⅠor PGⅡ alone.