乙型肝炎病毒前C区/BCP区突变对血清HBV DNA含量的影响

目的利用基因芯片技术检测乙型肝炎病毒(HBV)前C区/BCP区基因突变,探讨前C区/BCP区基因突变后血清HBV DNA水平的变化。方法应用基因芯片技术检测95例慢性乙肝的HBV前C区1896、1814及HBV C基因启动子(BCP)1762、1764四位点突变,同时用荧光定量PCR检测血清HBV DNA含量。结果95份血清标本中,有91份分别检测到了HBV前C区/BCP区基因突变,阳性率95.8%,其中G1896A突变33例(36.3%),A1762T G1764A联合突变64例(70.3%),G1896A A1762T G1764A联合突变22例(24.2%),未检测到A1814C突变毒株。G1896A A1762T G1764A联合突变后,血清HBV DNA水平较未变异组降低(P<0.05),其它各组则无显著变化。结论应用基因芯片法可同时检测HBV多个突变位点。乙型肝炎病毒前C区/BCP区突变对血清HBV DNA含量有一定的影响。     

四川省两县(区)乙型肝炎病毒S基因和P基因变异分析

目的 分析四川省两县(区)乙型肝炎(乙肝)病毒(Hepatitis B Virus,HBV)S基因和P基因变异.方法 从乙肝病毒表面抗原、乙肝病毒e抗原均阳性的血清中提取病毒脱氧核糖核酸,经聚合酶链反应和核苷酸测序后,比较和分析S基因和P基因核苷酸和氨基酸变异.结果 47例慢性HBV感染者血清标本中,12份标本在"a"决定簇内出现氨基酸变异(25.53%),分别为T126A、I126T/S、P127T、T131N、M133L、M133T、T140I;变异主要集中在"a"决定簇的第1环上(92.86%,13/14).1份标本逆转录酶C域中出现rtV2071变异.结论 自然发生的氨基酸变异主要集中在"a"决定簇的第1环上,其抗原性已改变,对乙肝疫苗接种人群构成了潜在的威胁.拉米夫定耐药株也可在未接受核苷(酸)类药物治疗的病人中发现.     

耐拉米夫定乙型肝炎病毒聚合酶P区变异与基因分型

目的：分析拉米夫定耐药的慢性乙型肝炎患者HBV P区变异和基因分型的关系,为制定针对不同HBV基因型抗病毒的个体化方案提供依据。方法应用聚合酶链反应（ PCR）和酶联免疫吸附试验（ ELISA）对187例拉米夫定耐药患者的HBV DNA载量及HBV-M进行检测,用特异的引物对待检标本HBV P区进行全序列测定,对不同性别、年龄的测序结果的基因分型作比较;并对C、B基因型发生实变模式作分析。结果187例慢性乙型肝炎患者感染的HBV均为B和C基因型,B基因型51例（27.27%）,C基因型134例（71.66%）,并存在混合感染2例（1.07%）;HBeAg含量在B基因型和C基因型间差异有统计学意义（ P〈0.05）;P区基因突变结果显示,B基因型以单纯YMDD突变为主,C基因型以YMDD突变＋ rtL180M 突变为主。结论本研究对象的HBV基因型主要是C型,不同基因型可能决定了与耐药相关的P区变异模式。     

乙型肝炎病毒隐匿性感染的分子机制研究

目的从基因变异角度研究HBV隐匿性感染的分子机制。方法不明原因的肝炎患者104例,收集临床资料及血清,套式PCR扩增HBVS、X、C基因,检测隐匿性HBV感染者。HBVS、X区扩增产物测序和变异位点分析,发现可能的和HBV隐匿性感染有关的变异位点,分析HBV基因变异在HBV隐匿性感染中的作用。结果检测出隐匿性HBV感染13例。S基因区未发现有义突变,X基因区及重叠核心启动子区、增强子Ⅱ区发现多位点变异,推测与HBV隐匿性感染有关。结论X基因区及重叠核心启动子区、增强子Ⅱ区多位点变异可能是HBV隐匿性感染的分子基础。     

乙型肝炎病毒前C区及基本核心启动子区变异与基因型的临床相关性研究

目的探讨乙型肝炎病毒前C区G1896A突变、基本核心启动子区A1762T、G1764A双突变与基因型、拉米夫定疗效的相关性。方法采用基因测序法对接受拉米夫定治疗的81例慢性重型乙型肝炎患者的血清标本进行前C区、基本核心启动子区(BCP区)、基因型及逆转录酶区(P区)检测,并对前C区、BCP区的变异与基因型、逆转录酶区的变异进行相关性分析。结果 81例慢性重型乙型肝炎患者送检标本中,68份标本可检出前C区G1896A变异或BCP区A1762T/G1764A变异;单纯G1896A突变31例,单纯A1762T/G1764A突变24例,前C区联合BCP区变异13例;81例慢性重型乙型肝炎患者经拉米夫定治疗后,有45例发生变异,其中180M和204V变异株35例,204I变异株10例;前C区变异在野生型组、变异型组中的检出率分别为38.9%、66.7%,两者差异有统计学意义(P<0.05);BCP区变异在野生型组、变异型组中的检出率分别为33.3%、55.6%,两者差异有统计学意义(P<0.05);在81例慢性重型乙型肝炎患者血清标本中,B基因型18例,C基因型63例;前C区变异在B、C基因型中的检出率分别为72.2%、33.3%,两者差异有统计学意义(P<0.05);BCP区变异在B、C基因型中的检出率分别为16.7%、54.0%,两者差异有统计学意义(P<0.05);G1896A突变在B基因型感染者中检出率明显高于C基因型感染者;A1762T/G1764A双突变在B基因型感染者中检出率明显低于C基因型感染者。结论发生G1896A突变和A1762T/G1764A突变的患者,在接受拉米夫定治疗后更易发生逆转录酶区的变异;G1896A突变易出现在B基因型感染者中,A1762T/G1764A突变易出现在C基因型感染者中。     

HBeAg阴性的慢性HBV感染者乙肝病毒前C区G1896A变异分析

目的:了解HBeAg阴性的慢性HBV感染者乙肝病毒前C区G1896A变异情况,为临床乙肝诊治提供科学依据。方法:北京市海淀医院HBeAg阴性的慢性HBV感染者496例,其中HBeAg阴性慢性乙型肝炎患者380例,非活动期HBV感染者196例,取血液及肝组织样本进行HBV血清学标志检测(HBsAg、抗-HBs、HBeAg、抗-HBe和抗-HBc)、丙氨酸谷丙转移酶(ALT)、HBV DNA定量检测;HBV前C区G1896A变异检测;肝组织学检查;采用SPSS13.0软件进行数据分析。结果:两组患者血清中均检测到HBV DNA,其中HBeAg阴性慢性乙型肝炎患者组检出率100%(296/296),平均值为4.90±1.49log10拷贝数/毫升,非活动期HBV感染者组检出率42.0%(84/200)平均值为3.22±0.59 log10拷贝数/毫升,两组比较HBV DNA载量差异有显著性;在HBV DNA载量>1000拷贝/ml的病例中HBeAg阴性慢性乙型肝炎组的乙肝病毒存在前C区G1896A变异,发生率为83.8%(248/296),非活动期HBV感染组乙肝病毒前C区G1896A也存在变异,发生率为33.3%...     

质谱法检测乙型肝炎病毒YMDD变异

目的 研究基质辅助激光解吸附飞行时间质谱法(MALDI-TOF MS)检测乙型肝炎病毒(HBV)YMDD变异的价值.方法 应用聚合酶链反应(PCR)和MALDI-TOF MS基因分型法测定HBV YMDD基序变异位点寡核苷酸的含量.结果 MALDI-TOF MS基因分型法检测HBV变异的灵敏度为100 copies/mL血清,对HBV野生株和变异株共生变异的检测具有高度特异性.应用该法对40例患者血清进行检测,能准确鉴定已用先前方法 测知的病毒变种及未能检测到的病毒准种.结论 MALDI-TOF MS检测HBV YMDD变异具有高度的敏感性和特异性,且能适应接受拉米夫定治疗乙肝患者高通量筛查的需要.     

内江市东兴区乙型肝炎病毒基因型、基本核心启动子和前C区变异分析

目的 了解内江市东兴区乙型肝炎病毒（hepatitis B virus,HBV）慢性感染者中的HBV基因型和基本核心启动子（basal core promoter,BCP）、前C(Precore,PC)区变异。方法 从慢性HBV感染者血清标本中提取病毒DNA,进行核苷酸测序,构建进化树确定基因型,分析BCP和前C区位点变异。结果 S基因序列进化树分析显示,64份标本为B基因型,7份为C基因型,1份为I基因型;获取的53份BCP和PC序列标本中,25份标本发生A1762T/G1764A或G1896A变异;7份C基因型标本中6份发生A1762T/G1764A变异;B基因型标本以G1896A变异为主,并与A1762T/G1764A联合变异,HBeAg阳性组和阴性组之间G1896A和A1762T/G1764A变异率有差异（P均<0.05）。其他包括HBeAg前体启动子变异、1846位点核苷酸缺失等。结论 内江市东兴区慢性HBV感染者以B基因型感染为主,1例为I基因型。B基因型HBV以G1896A变异最常见,并与A1762T/G1764A联合变异,C基因型HBV更容易发生A1762T/G1...     

268例慢性乙肝患者拉米夫定治疗不同时间耐药突变株变化分析

目的:检测慢性乙肝患者在拉米夫定治疗过程中耐药突变株变化。方法:采用实时荧光PCR方法对接受拉米夫定治疗不同时间的患者进行YMDD耐药突变检测,及血清HBV DNA水平检测,分析耐药突变发生的时间节点。结果:通过检测拉米夫定治疗不同阶段血清HBV DNA水平和YMDD变异情况,发现在170例HBV DNA阳性的患者中有117例发生YMDD变异,变异率为69%。根据治疗时间进行分组,治疗时间小于6个月的患者YMDD变异率为13%,治疗时间超过24个月的患者全部出现YMDD变异(P<0.05)。结论:对接受拉米夫定治疗的患者从第6~7个月开始检测YM-DD变异,可以尽早发现耐药变异,为临床治疗方案的选择提供依据。     

乙型肝炎肝硬化患者与乙型肝炎病毒携带者乙型肝炎病毒X基因序列比较研究

目的研究乙型肝炎肝硬化患者与乙型肝炎病毒（HBV）携带者HBV X基因序列的差异。方法对20例标本的PCR产物进行直接测序;并对3例乙型肝炎肝硬化患者和3例无症状HBV携带者HBV DNA的扩增片段进行克隆测序,并分析比较。结果肝硬化患者中X基因核心启动子双突变T1762/A1764、G1719T、T1727G/A、G1730C、T1753C等变异高于HBV携带者;前者X启动子区变异率明显高于后者;乙型肝炎肝硬化同一患者X区不同克隆之间的碱基序列同源性为91.3%-99.7%,而HBV携带者为96.0%-100.0%。结论乙型肝炎肝硬化患者,HBV DNA X区的CP启动子区以及X启动子区变异程度明显高于HBV携带者,同时乙型肝炎肝硬化患者体内存在变异程度更大的HBV准种群。     

Egr-1基因启动子介导肿瘤基因放疗的研究进展

Egr-1基因是一种即刻早期基因，其启动子可感受自由基、电离辐射等理化刺激，继而诱导Egr-1基因或其下游基因表达。对转染了Egr-1基因启动子启动的肿瘤杀伤基因的肿瘤细胞．实施局部照射可诱导基因表达，通过射线与基因的双重作用杀伤肿瘤。此方法既解决了外源基因靶向表达的难题，又降低了照射剂量．减少了正常组织的损伤。     

Association of a polymorphism in the apo AI gene promoter with hyperalphalipoproteinemia

The distribution of a polymorphism due to an Adenine to Guanine transition in the ApoAI gene has been studied in 136 women and 108 men, through amplification of the promoter region of the gene and allele-specific oligonucleotide hybridization. The allelic frequencies for the A allele were 0.10, 0.14 and 0.27 in women and 0.08, 017 and 0.14 in men for the lowest decile, intermediate group and the highest decile of HDL-cholesterol levels, respectively. Statistical analysis showed that the A allele was associated with high HDL-cholesterol levels in women, but not in men.     

Association of a polymorphism in the apo AI gene promoter with hyperalphalipoproteinemia

The distribution of a polymorphism due to an Adenine to Guanine transition in the ApoAI gene has been studied in 136 women and 108 men, through amplification of the promoter region of the gene and allele-specific oligonucleotide hybridization. The allelic frequencies for the A allele were 0.10, 0.14 and 0.27 in women and 0.08, 017 and 0.14 in men for the lowest decile, intermediate group and the highest decile of HDL-cholesterol levels, respectively. Statistical analysis showed that the A allele was associated with high HDL-cholesterol levels in women, but not in men.Corresponding author.     

肥胖儿童食欲调节因子基因启动子区DNA甲基化探讨

目的 探讨肥胖儿童瘦素、瘦素受体及促阿片-黑素细胞皮质素原(proopiomelanocortin,POMC)基因启动子区CpG位点的甲基化改变。方法 选取北京酒仙桥地区45例4~6岁单纯性肥胖儿童及按性别、年龄1∶1 配对的45例正常儿童为研究对象。利用外周血提取基因组DNA,然后应用亚硫酸盐修饰直接测序法(BSP)和半巢式PCR检测儿童瘦素基因启动子区(324 bp,-228到+96)25个CpG位点、瘦素受体基因启动子区(271 bp,-411到-141)20个CpG位点及POMC基因启动子区(275 bp,-341到-67)18个CpG位点的甲基化状态;同时采用酶联免疫分析测定血浆瘦素含量。结果 单纯性肥胖儿童血浆瘦素含量(21.24±4.02) μg/L显著高于正常儿童(2.95±0.53) μg/L。肥胖儿童瘦素基因启动子区CpG位点的平均甲基化程度(0.43±0.13)显著低于正常儿童(0.52±0.15);对该启动子区单个CpG位点比较发现,9个位点的甲基化程度在肥胖组低于对照组。瘦素受体基因启动子区各CpG位点在两组儿童中均呈现完全去甲基化状态。POMC基因启动子区各CpG位点甲基化程度在肥胖与正常儿童之间均未见显著差异。结论 肥胖儿童瘦素基因启动子区CpG位点存在甲基化改变,可能与瘦素表达增加、瘦素抵抗发生有关。     

Berry intake increases the activity of the gamma-glutamylcysteine synthetase promoter in transgenic reporter mice

A diet rich in fruit and vegetables is associated with decreased risk of disease. One possible mechanism for this is that dietary antioxidants positively regulate protective genes. Toward our goal to identify bioactive compounds with such functions in plants, we developed transgenic mice that express luciferase controlled by the gamma-glutamylcysteine synthetase heavy subunit (GCS(h)) promoter. Mice that consumed a nonpurified diet ad libitum were supplemented with juices or extracts of antioxidant-rich berries for 42 h or 3-4 wk. The treatments generally increased luciferase activity in brain and skeletal muscle and decreased it in liver compared with controls fed water. The same overall pattern was also found in mice fed ellagic acid (EA), a phenolic acid found in many berries. This change in GCS(h) promoter activity after berry treatment occurred in only approximately 50% of the mice, indicating that they were either responders or nonresponders. Our results demonstrate for the first time that berry extracts rich in polyphenols and EA can induce GCS(h) in vivo. The induction of protective enzymes may be important for the chemopreventive effects of fruits and vegetables.     

miR-34启动子甲基化在胃癌发病过程中的作用研究

目的探讨胃癌组织中miR-34基因启动子甲基化状态对miR-34的表达影响及其与患者预后的相关性。方法采用实时荧光定量PCR法检测50例胃癌组织及癌旁组织中成熟miR-34的表达。采用甲基化特异性PCR法检测miR-34基因启动子的甲基化状态,运用统计学分析miR-34基因启动子甲基化状态与miR-34表达量之间的关系,并随访患者复发和转移情况。结果胃癌组织中成熟miR-34的表达水平比癌旁组织明显降低(P<0.05)。miR-34甲基化水平与淋巴结转移有相关性(P<0.05)。miR-34基因启动子非甲基化组的成熟miR-34表达水平明显高于甲基化阳性单一模式组和甲基化阳性混合模式组(P<0.05)。与非甲基化组和甲基化阳性混合模式组相比,miR-34基因甲基化阳性组的胃癌患者复发和转移率明显升高(P<0.05)。结论在胃癌患者中,miR-34基因启动子甲基化可导致miR-34表达下调。miR-34基因启动子的甲基化状态与胃癌的复发与转移明显相关。     

Evaluation of an intracellular-activated promoter for the generation of live Salmonella recombinant vaccines

Salmonella enterica serovar Typhimurium is a versatile host organism for the generation of recombinant live vaccines for mucosal immunization. We investigated the performance of an intracellular-activated promoter for expression of heterologous antigens by Salmonella carrier strains. We observed that intracellular expressed antigens were more efficient in elicitation of humoral and cellular immune responses than constitutively expressed antigens. Furthermore, the intracellular-activated promoter was more efficient in combination with a carrier strain deficient in SPI2 gene sseC than with the conventional aroA strain. We propose that intracellular-activated promoters will be useful tools for construction of efficient recombinant live vaccines.     

Breast-feeding: main promoter of infant health.

Unique immunological factors in breast milk are responsible for the protection it affords infants against pathogenic agents. This protection is particularly effective against the infectious agents which invade or colonize the small and large intestine. Factors interfering with enterotoxigenic and enteroinvasive bacteria, and with entero- and rotaviruses, have already been detected in colostrum and mature human milk. Moreover, the biochemical composition of human milk makes it the best infant food during the first 6 months of life, and with supplementation, for months thereafter. At the same time, optimal maternal-infant interactions are effected during breast-feeding which strengthen infant care in health, in illness and during convalescence. The low cost of breast-feeding when compared to other types of alimentation, coupled with its unparalleled nutritive, anti-infectious, and behavior stimulating properties makes it the main promoter of infant health. There remains a need for investigation of the factors that have brought about a reduction in breast-feeding throughout the world. New methodologies to increase the incidence of breast-feeding, particularly in the developing nations, are critical. The availability of such methodologies could facilitate delivery of other health interventions ideally delivered by the mother, the most important agent for improving the health of infants and children.