幽门螺杆菌疫苗免疫小鼠的Th免疫应答及作用

目的 研究幽门螺杆菌(Helicobacter pylori,Hp)疫苗诱导小鼠保护性免疫应答的机制。方法 以霍乱毒素(CT)加Hp全菌超声粉碎抗原免疫小鼠，并设CT组和生理盐水(NS)组为对照。免疫4周后以Hp攻击；在攻击前后不同时间点分批处死小鼠，比较免疫组和对照组胃黏膜唧定植及各项免疫指标的变化。结果 (1)攻击后5、26周后免疫组和CT组、NS组相比胃黏膜Hp定植量明显降低。(2)各时间点免疫组IgG1、IgG2a、IgA均高于NS组、CT组。(3)攻击前和攻击后5周免疫组和CT组胃黏膜Th1型细胞因子表达较NS组明显升高。攻击后26周免疫组和CT组除白细胞介素(IL)-12外，干扰素(IFN)-γ和IL-2表达均明显降低，而NS组Thl型细胞因子表达明显升高。(4)Th2型细胞因子在攻击前免疫组和CT组胃黏膜均有轻度表达；攻击后5周免疫组和CT组IL-4和IL-10均无表达，IL-6表达低于NS组；攻击后26周免疫组IL-4和IL-10再次表达，CT组仅表达IL-10，IL-6表达各组无明显差异。(5)各细胞因子在所有组脾脏攻击前后各时间点表达无差异。(6)与NS组相比，攻击后5周免疫组鼠胃黏膜出现明显的炎症反应，CT组也有轻度炎症反应；攻击后26周NS组炎症加重而免疫组则有所减轻。结论 (1)以CT为佐剂的Hp疫苗可诱导Thl和Th2混合型初始免疫应答；(2)Hp攻击后早期增强的Thl反应起主要免疫保护作用同时导致免疫后胃炎；晚期随Th2反应的增强Thl反应和免疫后胃炎强度减弱。     

幽门螺杆菌空泡毒作用

目的研究幽门螺杆菌（Ｈｅｌｉｃｏｂａｃｔｅｒｐｙｌｏｒｉ,Ｈｐ）空泡毒作用。方法用体外细胞毒试验,结合细胞化学及荧光染色、透射电镜技术研究Ｈｐ空泡毒作用及其病理机制。结果７８．２６％的Ｈｐ相关消化性溃疡患者感染的是空泡毒作用阳性Ｈｐ（Ｔｏｘｉｎ＋）,而只有４２．８６％的胃炎患者感染的是Ｈｐ（Ｔｏｘｉｎ＋）。Ｈｐ（Ｔｏｘｉｎ＋）引起的细胞空泡样变,吖啶橙及酸性磷酸酶染色阳性;透射电镜显示空泡为双层膜结构。结论Ｈｐ（Ｔｏｘｉｎ＋）感染与Ｈｐ相关消化性溃疡密切相关。Ｈｐ（Ｔｏｘｉｎ＋）引起的空泡是自噬体,是细胞在毒性物质作用下表现的病理现象。     

东西方幽门螺杆菌鞭毛素A差异分析

目的：对东西方幽门螺杆菌鞭毛素A（flagellin A,FlaA)的差异进行对照研究，并探讨这些差异对FlaA免疫学特性的影响。方法：借助软件分析FlaA的结构信息和理化性质；从西京医院门诊病人分离了的6株幽门螺杆菌基因组DNA中扩增FlaA可变区的序列并与Genbank中的1－株幽门螺杆菌的相应氨基酸序列进行差异比较。结果软件分析显示幽门螺杆菌FlaA具有典型的鞭毛丝蛋白的结构特征，在中央可变区发现三个亲水峰和两处N乙酰糖基化位点，分别位于第一和第二个亲水峰。对16株幽门螺杆菌的差异分析发现三个特征性的差异，种系分析结果显示按序列同源性大致分为两类，并与差异分析的结果吻合。结论：幽门螺杆菌FlaA是一个具有特征性二级结构和高级结构的分子，东西方FlaA中央可变区氨基酸序列无明显特征性的差异，16株幽门螺杆菌按种类分析结果和FlaA中央可主为区氨基酸序列的特征性差异可分为两类，结构和理化性质分析提示这些特征性差异可能对FlaA分子的免疫学特征有潜在的影响。     

Virulence and potential pathogenicity of coccoid Helicobacter pylori induced by antibiotics

AIM: To explore the virulence and the potential pathogenicity of coccoid Helicobacter pylori (H. pylori) transformed from spiral form by exposure to antibiotic. METHODS: Three strains of H. pylori, isolated from gastric biopsy specimens of confirmed peptic ulcer, were converted from spiral into coccoid from by exposure to metronidazole. Both spiral and coccoid form of H. pylori were tested for the urease activity, the adherence to Hep-2 cells and the vacuolating cytotoxicity to Hela cells, and the differences of the protein were analysed by SDS-PAGE and Western blot. The mutation of the genes including ureA, ureB,hpaA, vacA and cagA, related with virulence, was detected by means of PCR and PCR-SSCP. RESULTS: In the coccoid H. pylori,the urease activity, the adherence to Hep-2 cells and the vacuolating cytotoxicity to Hela cells all decreased. In strain F44, the rate and index of adherence reduced from 70.0% +/- 5.3% to 33% +/- 5.1% and from 2.6 +/- 0.4 to 0.96 +/- 0.3 (P < 0.01), respectively. The invasion of coccoid H. pylori into Hep-2 cell could be seen under electronmicroscope. SDS-PAGE showed that the content of the protein with the molecular weight over Mr 74000 decreased, and the hybriditional signal in band M(r) 125000 weakened, while the band M(r)110000 and M(r)63000 strengthened in coccoid H.pylori as shown in Western blot. The results of PCR were all positive, and PCR-SSCP indicated that there may exist the point mutation in gene hpaA or vacA. CONCLUSION: The virulence and the proteins with molecular weight over M(r)74000 in coccoid H.pylori decrease, but no deletion exists in amplification fragments from ureA, ureB, hpaA, vacA and cagA genes, suggesting that coccoid H.pylori may have potential pathogenicity.     

敲除PPK基因对幽门螺杆菌逃避巨噬细胞免疫清除的影响

目的:研究敲除多聚磷酸激酶(polyphosphate kinase,PPK)基因后对幽门螺旋杆菌(Helicobacterpylori,H.pylori)逃避巨噬细胞免疫清除的影响.方法:通过同源重组的原理构建H.pylori PPK敲除菌株.提取PPK敲除菌株体内多聚磷酸盐,转化为ATP进行定量,并与野生型菌株内多聚磷酸盐水平进行比较.将PPK敲除菌株及野生型菌株分别与小鼠巨噬细胞系RAW264.7共同培养,比较存活的H.pylori数量.结果:成功构建敲除PPK的H.pylori菌种.Western blot显示该菌无PPK蛋白表达.对敲除PPK的H.pylori菌株中的多聚磷酸盐定量结果显示,其内PP含量为0.46±0.25nmol Pi/mg Protein,显著低于G27野生菌种(175.33±21.22nmol Pi/mg Protein,P<0.01).将该菌株与小鼠巨噬细胞系RAW264.7共同培养,2h时间点,G27及G27ΔPPK菌种在巨噬细胞内的存活无显著差异;而在24h时间点,G27ΔPPK菌种在巨噬细胞内的存活率显著低于野生型G27菌种,巨噬细胞内的BacLightkit染色结果显示,G27ΔPPK菌种在巨噬细胞内获得染色的活菌显著低于野生型G27菌种.结论:PPK是H.pylori合成多聚磷酸盐的关键酶.敲除该基因后,H.pylori合成多聚磷酸盐的能力显著下降,其逃避巨噬细胞清除的能力明显减弱.     

Polymorphism of flagellin A gene in Helicobacter priori

AIM: To study the polymorphism of flagellin A genotype and its significance in Helicobacter pyiori ( H. Pylori).METHODS: As the template, genome DNA was purified from six clinical isolates of H. Pylori from outpatients, and the corresponding flagellin A fragments were amplified by polymerase chain reaction. All these products were sequenced. These sequences were compared with each other, and analyzed by software of FASTA program.RESULTS: Specific PCR products were amplified from all of these H. Pylori isolates and no length divergence was found among them. Compared with each other, the highest ungapped identity is 99. 10%, while the lowest is 94.65%.Using FASTA program, the alignments between query and library sequences derived from different H. Pylori strains were higher than 90%.CONCLUSION: The nucleotide sequence of flagellin A in H.pylori is highly conservative with incident divergence. This information may be useful for gene diagnosis and further study on flagellar antigen phenotype.     

Different T helper type immune responses induced by Helicobacter pylori vaccine in mouse model

OBJECTIVE: To study the protective immune response induced by Helicobacter pylori vaccine in mice. METHODS: Mice were orally immunized with cholera toxin (CT) plus H. pylori whole‐cell sonicate antigen (group A), CT only (group B) or normal saline (group C), and challenged with H. pylori 4 weeks later. Mice from each group were killed at different time points before and after challenge. Gastric H. pylori colonization, serum anti‐H. pylori IgG1, IgG2a and IgA antibodies, the mRNA expression of both Th1‐type and Th2‐type cytokines in the gastric mucosa and splenic tissue, and mucosal inflammation at different time points were compared among the groups. RESULTS: Colonization of H. pylori in group A decreased significantly compared with groups B and C at both 5 and 26 weeks after challenge. The serum concentrations of anti H. pylori IgG1, IgG2a and IgA in group A were significantly higher than those of groups B and C at all time points. The mRNA expressions of Th1‐type cytokines in the gastric mucosa of groups A and B were higher than those of group C both before and 5 weeks after H. pylori challenge. At 26 weeks after challenge, Th1‐type cytokines were increased in the gastric mucosa of group C, but decreased in the gastric mucosa of groups A and B, except for IL‐12. Before challenge, Th2‐type cytokines were expressed at a similar level in the gastric mucosa of both group A and group B; 5 weeks after challenge, IL‐4 and IL‐10 expressions were not detectable in either group and the expression of IL‐6 in both groups was lower than that in group C; 26 weeks after challenge, the expression of both IL‐4 and IL‐10 was detected in group A, but only IL‐10 in group B. The expression of IL‐6 was similar in all groups. Both Th1‐ and Th2‐type cytokines were expressed in the splenic tissue of all groups at different time points. Significant gastritis characterized by marked infiltration of mononuclear cells and neutrophils was found in group A at 5 weeks after challenge, but only mild inflammation was present in group B; 26 weeks after challenge, gastritis had increased in group C but decreased in the other two groups. CONCLUSIONS: The vaccine consisting of H. pylori whole‐cell sonicate antigen and CT induces both Th1 and Th2 immune responses at an early stage. The Th1 response increases early after H. pylori challenge, which contributes to both protective immunity and postimmunization gastritis, whereas the Th2 response increases in the gastric mucosa late after challenge when both the Th1 response and postimmunization gastritis have abated.     

幽门螺杆菌疫苗研究的现状和展望

幽门螺杆菌感染与慢性胃炎、消化性溃疡、胃癌及胃粘膜相关性淋巴瘤有关。疫苗是防治幽门螺杆菌感染切实可行的方法。近年来新的防治和根除幽门螺杆菌感染的疫苗正在开发并取得了很大的进展。本文对新近研制的蛋白组分疫苗、活菌疫苗、DNA疫苗、缓释微球疫苗及表位疫苗等幽门螺杆菌疫苗的进展作一综述。     

幽门螺杆菌双亚单位表位疫苗的构建及免疫原性研究

目的构建幽门螺杆菌(Helicobacter pylori,Hp)黏附素(HpaA)和尿素酶B亚单位(UreB)双亚基多表位疫苗,并对其免疫原性进行研究。方法设计引物采用PCR法将黏附素(HpaA)的一个B细胞表位与尿素酶B亚单位的三个TH表位及一个B细胞表位串联起来(HUepi),表位之间用两个赖氨酸(KK)间隔,T-A克隆后构建融合基因表达质粒pET-22b( )-HUepi,在大肠杆菌BL21(DE3)中表达,重组蛋白采用阳离子和疏水层析进行纯化,经鉴定后皮下免疫BALB/c小鼠,检测细胞免疫应答及体液免疫应答。结果PCR法扩增出了约233bp的目的片段;原核表达质粒pET-22b( )-HUepi经酶切及测序鉴定,目的基因序列与设计序列一致;重组蛋白HUepi表达率约20.0%,PAGE初步测定目的蛋白的相对分子质量(Mr)约8.38×103Dr,破菌后电泳证实目的蛋白以可溶形式表达,纯化后蛋白纯度大于97.6%,经N端测序证实为设计的表位疫苗蛋白,TH表位多肽、表位疫苗蛋白及rUreB均能够刺激表位疫苗致敏的小鼠淋巴细胞增殖(SI>2),并且此疫苗能够刺激机体产生特异性抗体。结论Hp HpaA、UreB双亚基表位疫苗HUepi经基因克隆获得了较高的表达量,并初步显示了较好的免疫原性。为新一代Hp疫苗的研制奠定实验基础。     

幽门螺杆菌感染对胃黏膜环氧合酶表达的影响

目的评估幽门螺杆菌（Hp）急、慢性感染对小鼠胃黏膜环氧合酶（COX）表达的影响.方法清洁级BALB/C小鼠共40只,分为阴性对照组（10只）予0.4ml生理盐水灌喂,急性感染组（10只）予0.4 ml Hp菌液灌喂,共5 d,该两组小鼠在最后一次灌喂后2周处死;另20只小鼠予0.4ml Hp菌液灌喂,共5d,在最后一次灌喂2个月后将小鼠均分为两组,治疗组予克拉霉素按每天13.5 mg/kg分2次灌喂,连续7d（本组有1只小鼠死亡）;慢性感染组予生理盐水,1个月后分别处死两组动物.取胃黏膜标本,用免疫组化半定量法测定上述各组胃窦黏膜COX-1及COX-2的蛋白表达.结果四组小鼠胃黏膜小凹上皮均可见COX-1蛋白表达,各组阳性率差异无统计学意义（P＞0.05）.与阴性对照组相比,急、慢性感染组COX-2蛋白表达均明显增加,其中慢性感染组治疗前COX-2阳性率明显高于急性感染组.慢性感染组治疗后阳性率与治疗前相比有所下降,但差异无统计学意义（P＞0.05）.结论Hp感染导致胃黏膜COX-2表达增加,对COX-1表达无明显作用,清除Hp后COX-2表达仍高于正常.     

Omeprazole attenuates neutrophil-endothelial cell adhesive interaction induced by extracts of Helicobacter pylori

In this study, the influence of omeprazole on the adhesive activity of neutrophils, provided by an extract of Helicobacter pylori, was determined. Human neutrophils were collected from peripheral blood and labelled with a fluorochrome. Helicobacter pylori (NCTC 11637) was cultured and its water extract was obtained by centrifugation of the bacterial suspension. Neutrophils were incubated with the extract in a plastic plate. Percentage adherence was calculated by measuring the fluorescence of floating and adherent cells. Rat mesenteric venule was prepared on an intravital microscope and the number of neutrophils which adhered to venular endothelium was counted. Neutrophil adherence to the plastic plate was increased by the presence of H. pylori extract. Pretreatment with omeprazole significantly decreased this adherence in a dose-dependent manner (10(-6)-10(-4)mol/L). Neutrophil adherence to the mesenteric venule was also increased by H. pylori extract and significantly inhibited by omeprazole. These results indicate that the neutrophil-endothelial adhesive interaction is inhibited by omeprazole, suggesting that omeprazole prevents neutrophil recruitment to the gastric mucosa associated with H. pylori infection.     

幽门螺杆菌OmpP1蛋白的生物信息学分析

目的应用生物学软件预测幽门螺杆菌(Helicobacter pylori,Hp)ompP1基因编码蛋白(OmpP1)的结构与功能。方法从NCBI数据库中获取ompP1基因的相关信息,应用软件MEGAX 10.0.5构建Hp进化树,应用生物信息学软件分析OmpP1蛋白的氨基酸序列、理化性质、跨膜区、信号肽、糖基化和磷酸化位点、空间结构以及抗原表位。结果 ompP1基因核苷酸序列编码区长1 764 bp,编码587个氨基酸;生物信息学分析显示ompP1基因在不同菌株间的同源性为96.2%~97.17%,其编码的OmpP1蛋白是一种性质稳定、偏碱性的亲水蛋白;该蛋白无跨膜区和信号肽,含有糖基化和磷酸化位点;OmpP1二级结构中含α-螺旋178个,延长链162个,β转角37个,无规则卷曲210个,分别占30.32%、27.60%、6.30%和35.78%;三级结构分析显示OmpP1含有OM_channels蛋白家族特征性结构域;BepiPred1.0和SYFPEITHI9预测该蛋白含有22个B淋巴细胞和28个CTL淋巴细胞相关抗原表位。结论 Hp OmpP1为碱性亲水性蛋白,含有T、B细胞抗原表位,可为其抗原性研究和高效表位疫苗的研发提供理论参考。     

幽门螺杆菌根除前后血清抗体滴度变化的临床意义

目的探讨幽门螺杆菌（Hp）血清抗体细胞毒素相关蛋白A（CagA）、尿素酶（Ure）滴度的变化在Hp根除疗效中的临床意义。方法采用经胃镜及快速尿素酶试验确诊为Hp相关性消化性溃疡患者118例,以蛋白芯片方法检测血清CagA、Ure抗体,观察其根除前、根除后8周、24周抗体滴度变化。结果118例患者有102例Hp根除,根除率为86.4%。根除治疗成功组在治疗后8周CagA、Ure抗体滴度下降无显著性,24周后则显著下降（P〈0.05）,下降幅度分别为63.5%、41.3%,CagA及Ure抗体的下降幅度比较差异有统计学意义（P〈0.05）。而根除治疗失败组的CagA、Ure抗体滴度无明显变化（P〉0.05）。结论蛋白芯片法动态观察血清CagA、Ure抗体水平8～24周以上,可作为评估根除Hp疗效的一种参考方法,且CagA抗体可能更有意义。     

粪便抗原检测诊断幽门螺杆菌现症感染

目的 评价应用免疫酶联吸附试验(ELISA)检测粪便中幽门螺杆菌(Helicobacter pylori)抗原诊断H.pylori现症感染的敏感性和特异性。方法 应用^14C呼气试验以及幽门螺杆菌粪便抗原(HpSA)试验，对100例因上消化道不适就诊，怀疑有H.pylori感染的患者进行检测，观察两种检查的符合率。结果 ^14C呼气试验和HpSA同时阳性者38例，^14C呼气试验阳性而HpSA阴性者4例；^14C呼气试验和HpSA同时阴性者57例，^14C呼气试验阴性而HpSA阳性1例。以^14C呼气试验作为金标准计算，HpSA检测方法的敏感性为90．48％，特异性为98．28％。结论 幽门螺杆菌抗粪便原检测与^14C呼气试验有较高的符合率，而且简便易行，不需特殊设备，解决了无法进行呼气试验的婴幼儿和有肺部疾患者的非侵人性幽门螺杆菌现症感染诊断问题，是一种非侵入性幽门螺杆菌现症感染诊断的新方法。     

定量检测幽门螺杆菌细胞毒素相关蛋白基因A方法的建立

目的建立定量检测幽门螺杆菌（Ｈｐ）细胞毒素相关蛋白基因Ａ（ｃａｇＡ）的方法。方法以含ＨｐｃａｇＡ片段的质粒（ｐＭＣ３）作为外参照品，在微孔板中对聚合酶链反应（ＰＣＲ）扩增ｃａｇＡ的产物进行辣根过氧化物酶标记探针的杂交及酶促显色以定量检测ｃａｇＡ阳性Ｈｐ。结果当ＰＣＲ反应体系中原始模板为１～１０５拷贝，循环次数小于或等于２５时，原始模板以相对固定的指数形式增加，在该范围内适宜定量分析。通过对２０份已知Ｈｐ菌株ｃａｇＡ的检测，符合率达１００％，观察结果的敏感度较电泳法提高了２００倍。定量检测不同胃黏膜组织中ｃａｇＡ阳性Ｈｐ，检出底线为６拷贝／毫克组织。结论本方法特异性强，敏感度高，对研究Ｈｐ与消化道疾病的关系及建立定量检测其他微生物的方法均有实用价值     

重组幽门螺杆菌疫苗免疫保护机制的研究

目的 研究以减毒鼠伤寒沙门菌为载体构建的重组幽门螺杆菌（Hp)疫苗诱导小鼠产生保护性免疫应答的机制。方法 将表达Hp尿素酶B亚单位（UreB)，黏附素（HpaA)及尿素酶B亚单位/黏附素融合蛋白（UreB/HpaA)的减毒鼠伤寒沙门菌（Salmonella typhimurium)给小鼠分别灌胃，另设单纯减毒鼠伤寒沙门菌和生理盐水免疫鼠为对照，免疫4周后以Hp活菌攻击，观察各组小鼠的免疫保护率，攻击前后血清中抗Hp抗体IgC1,IgG2a和IgA的变化。小鼠脾脏和胃黏膜中γ干扰素（IFN-γ）和白介素-4（IL-4）mRNA表达变化。结果 UreB,HpaA及UreB/HpaA组的免疫保护率分别为50%，41%和77%，和生理盐水组相比，攻击前各鼠伤寒沙门菌免疫组IgG1,IgG2a均轻度升高而IgA无变化，攻击后各鼠伤寒沙门菌免疫组IgG2a升高显著并以UreB/HpaA组为最，而IgG1和IgA的升高无统计学差异。胃黏膜攻击前生理盐水组无IFN-γ表达，其余各组均100%表达；攻击后生理盐水组IFN-γ轻度表达，但仍明显低于各鼠伤寒沙门菌免疫组，IL-4在攻击前后各组均无表达，脾IFN-γ和IL-4在所有组攻击前后均全部表达。结论 以减毒鼠伤寒沙门菌为载体构建的Hp疫苗在小鼠体内诱导出以TH1反应为主的保护性免疫应答。     

Helicobacter pylori infection in Icelandic children

Objective: The prevalence of Helicobacter pylori (HP) infection is decreasing in the western world. The seroprevalence among 25–50-year-old Icelandic adults was recently shown to be 30–40%. Information on the seroprevalence in Nordic children is limited. We aimed at ascertaining the infection prevalence among healthy Icelandic children.

Methods: The infection status in stored frozen blood samples from two cross-sectional studies on the health of 7–9-year-old children (n?=?125) and 16–18-year-old adolescents (n?=?80) was determined by enzyme-linked immunosorbent assay (ELISA). Information on family demographics and GI symptoms was obtained by standardized questionnaires.

Results: Overall, 3.4% (7/205) of the children were infected with H. pylori. The prevalence was 2.6% (5/190), missing data n?=?3, among children with both parents born in a low prevalence country compared to 17% (2/12) among those with at least one parent born in a high prevalence area (p?=?.026). When at least one parent was born in a high prevalence country, the odds ratio for being H. pylori seropositive was 2.2 (95% CI, 1.02–54.67), when adjusted for the educational status of the mother. There was no significant association between H. pylori infection and gastrointestinal symptoms.

Conclusion: Prevalence of H. pylori infection in Iceland has become very low, suggesting a great reduction in transmission from older generations. There was an association between H. pylori infection and origin from high prevalence areas but not with gastrointestinal symptoms. The results mirror recent studies of children of Scandinavian ancestry.     

幽门螺杆菌鞭毛基因PCR-RFLP分析及其与胃十二指肠疾病关系的研究

目的探讨幽门螺杆菌(Helicobacter pylori)鞭毛基因(flaA和flaB)PCR—RFLP分型及其与胃十二指肠疾病的关系一方法用PCR技术分别扩增46株从消化性溃疡(25株)和非消化性溃疡(21株)患者胃黏膜中分离得到的H.pylori flaA和flaB基因，并用HpaⅡ和HindⅢ消化PCR扩增产物，琼脂糖凝胶电泳分离酶切片段，将酶切片段编码后输入SPSS10．0软件进行聚类分析．并比较不同的基因型别在溃疡组和非溃疡组的分布。结果46株H．pylori菌株flaA基因HpaⅡ和HindⅢ酶切后分别产生4种和3种酶切图谱；flaB基因HpaⅡ和HindⅢ酶切后分别产生1种和2种酶切图谱：综合flaA和flaB基因酶切结果，46株菌株可分为四个基因型别：但这四个基因型别在溃疡组和非溃疡组的分布差异无统计学意义(P=0．513)。结论应用PCR—RFLP方法可以将H.pylori fla基因分为不同的基因型别．但胃十二指肠溃疡和非溃疡组的菌株fla基因型别无差别。     

Serum IgG response to differentiated antigens of Helicobacter pylori

ＳｅｒｕｍＩｇＧｒｅｓｐｏｎｓｅｔｏｄｉｆｅｒｅｎｔｉａｔｅｄａｎｔｉｇｅｎｓｏｆＨｅｌｉｃｏｂａｃｔｅｒｐｙｌｏｒｉＨＵＡＪｉｅＳｏｎｇ１,ＫＨＩＮＭａｒＭａｒ１,ＺＨＥＮＧＰｅｎｇＹｕａｎ１,ＹＥＯＨＫｈａｙＧｕａｎ２,ＮｇＨａｎＣｈｏｎ...