糖基化、非糖基化、重组PAI－1功能和性质的比较

对从ＨｅｐＧ２细胞培养液中分离得到植基化和非糖基化ＰＡＩ－１（１型纤溶酶原激活物抑制剂）,以及从ｐＹＺＨＢＩ－６６表达菌中纯化的非糖基化重组ＰＡＩ－１的某些性质和功能进行比较,结果显示,糖基化ＰＡＩ－１对ｔＰＡ（组织型纤溶酶原激活物）有较强的抑制,能较显著地被蛋白质变性剂所激活,对热有较强的稳定性,糖基化与非糖基化ＰＡＩ－１在ｐＨ２．５－９．０的范围内都相当稳定。纤维蛋白原和肝素能明显提高两者对ｔＰＡ的抑制作用。     

Growth reference charts and the nutritional status of Indian children

We evaluate the growth performance of Indian children of age 0-3 using data from the 1998-1999 National Family and Health Survey, making use of the new child growth standards developed by the World Health Organization’ Multicentre Growth Reference Study. We find that the new charts lead to an increase of 4.2 million in the estimated number of stunted children, and an increase of 2.3 million in the estimated number of wasted children. The estimated number of underweight children decreases instead by 2.1 million. We also use data on ethnic Indians living in the United Kingdom to provide evidence on the height genetic potential of Indians. We find that children of Indian ethnicity who live in the UK have anthropometric outcomes comparable to those in commonly used growth standards and that the height of ethnic South Asian in the sample is negatively related with the amount of time spent outside the United Kingdom.     

CYP1A1基因单核苷酸多态性与肺癌的相关性研究

目的:探讨代谢酶CYP1A1基因MspI位点多态性与新疆汉族人群肺癌遗传易感性之间的相关性.方法:应用聚合酶链式反应(PCR)-限制性片段长度多态性(RFLP)技术检测59例新疆汉族肺癌和84例新疆汉族健康人的CYP1A1基因MspI位点多态性分布频率,并分析了CYP1A1基因MspI位点多态性与新疆汉族人群肺癌遗传易感性和患者性别之间的相关性.结果:(1)CYP1A1基因MspI位点3种多态基因型分布频率在两组间比较差异有统计学意义(χ2=6.682,P=0.035),CC基因型在病例组的分布频率显著高于正常对照组.(2)携带突变CC基因型的个体较携带TT基因型的个体患肺癌的危险性增加(OR=3.759.95%CI=1.228-11.494,P=0.035).(3)男女肺癌患者的CYP1A1基因MspI位点基因型及等位基因频率的差异均无显著性(P>0.05).结论:(1)CC突变基因型可能是新疆汉族人群的肺癌易感因素.(2)CYP1A1基因MspI位点多态性可能与新疆汉族肺癌患者的性别无关.     

当归对大鼠缺血性脑损伤后Flt-1、Flk-1 mRNA表达的影响

目的：研究大鼠缺血性脑损伤后不同时间点Flt-1、Flk-1 mRNA的表达及当归对其表达的影响。方法：雄性Wistax大鼠,随机分为缺血损伤组和当归治疗组。采用线栓法制作大鼠短暂性大脑中动脉阻断（MCAO）与再灌模型。治疗组腹腔注射当归注射液（剂量5g／kg）。36只大鼠（每组各18只）在脑缺血／再灌后1d、3d、7d神经行为学评分完成后被处死,取大脑行氯化三苯四唑（TTC）染色以测脑梗死比;另取72只大鼠（每组各36只）在脑缺血／再灌后3h、6h、12h、1d、3d、7d分别被处死,应用半定量逆转录聚合酶链反应（RT-PCR）技术检测缺血侧Flt-1、Flk-1 mRNA的表达。结果：在同时间点神经功能缺损评分比较,缺血损伤组明显高于当归治疗组（P〈0．05）;在同时间点当归治疗组梗塞比明显小于缺血损伤组（P〈0．01）。RT-PCR检测表明,缺血损伤组Flt-1、Flk-1 mRNA在缺血／再灌后3h即开始表达增强,于3d达高峰,后逐渐降低;当归治疗组Flt-1、Flk-1 mRNA表达比缺血损伤组明显增加,于3d达到高峰后缓慢降低至第7d仍保持较高水平。缺血损伤组和当归治疗组中Flt-1 mRNA与Flk-1 mRNA的表达呈正相关性,相关系数为r=0．957（P〈0．01）。结论：当归可增强缺血性脑损伤后Flt-1、Flk-1 mRNA表达。Flt-1、Flk-1 mRNA的表达紧密相关。     

Exploring the limits of sequence and structure in a variant betagamma-crystallin domain of the protein absent in melanoma-1 (AIM1)

βγ-Crystallins belong to a superfamily of proteins in prokaryotes and eukaryotes that are based on duplications of a characteristic, highly conserved Greek key motif. Most members of the superfamily in vertebrates are structural proteins of the eye lens that contain four motifs arranged as two structural domains. Absent in melanoma 1 (AIM1), an unusual member of the superfamily whose expression is associated with suppression of malignancy in melanoma, contains 12 βγ-crystallin motifs in six domains. Some of these motifs diverge considerably from the canonical motif sequence. AIM1g1, the first βγ-crystallin domain of AIM1, is the most variant of βγ-crystallin domains currently known. In order to understand the limits of sequence variation on the structure, we report the crystal structure of AIM1g1 at 1.9 Å resolution. Despite having changes in key residues, the domain retains the overall βγ-crystallin fold. The domain also contains an unusual extended surface loop that significantly alters the shape of the domain and its charge profile. This structure illustrates the resilience of the βγ fold to considerable sequence changes and its remarkable ability to adapt for novel functions.     

家蝇细胞凋亡起始酶Caspase-1基因的克隆及在不同虫态的表达

为了研究Caspases家族在昆虫发育变态中的作用,通过RT-PCR扩增并结合RACE技术,克隆得到家蝇Musca domestica Caspase-1基因1条,命名为Mdom-Caspase-1（GenBank中cDNA序列号为EU854472, 氨基酸序列号为ACF71490）。该基因全长1 295 bp,阅读框序列870 bp,共编码289个氨基酸,理论分子量32.83 kDa,等电点8.67。 Mdom-Caspase-1蛋白有5个保守的半胱氨酸位点QACQG, 具有Caspase的典型特征; 整个分子呈现亲水性, 有8个区域共89个氨基酸为亲酯性, 蛋白质二级结构主要由11个α螺旋区、7个β-折叠区、17个β-转角区组成。昆虫间Caspase-1分子具有明显的保守性, Mdom-Caspase-1与黑腹果蝇Drosophila melanogaster、埃及伊蚊Aedes aegypti和致倦库蚊Culex quinquefasciatus的Caspase-1氨基酸序列相似性为65%~77%。RT-PCR半定量分析结果表明, Mdom-Caspase-1基因在家蝇各个虫态中均有表达, 但在卵期、3龄幼虫、预蛹、蛹和羽化5 d的雌虫中的表达量明显高于其他虫态。这些结果提示Caspase-1可能与昆虫发育变态关系密切, 为进一步研究昆虫Caspase-1功能、设计Caspase-1抑制剂提供了分子基础。     

HIV-1辅受体配体的融合表达载体的构建及表达

应用PCR扩增RANTES-KDEL基因,鉴定后与真核表达载体pCMV-S/K连接,构建成HIV-1辅受体的配体、趋化因子RANTES和SDF-1的融合表达载体pCMV-R-K-S-K,酶切鉴定和测序证明成功构建了pCMV-R-K-S-K融合表达载体.脂质体介导pCMV-R-K-S-K转染HeLa细胞,间接免疫荧光证实RANTES和SDF-1可高效表达于HeLa细胞.结果表明构建的pCMV-R-K-S-K融合表达载体能在HeLa细胞中高效表达,可用于下一步的HIV-1感染实验.     

JunD的结构功能特征及其在肿瘤发展中的作用

JunD是一种属于多功能激活剂蛋白-1（activating protein-1,AP-1) 家族的转录因子,可以激活或抑制多种靶基因的表达.在生长发育过程中,在各种细胞类型中都呈现出组成性表达.近20年的临床数据及分子生物学研究表明,JunD蛋白的功能受多个复杂过程调控,包括转录控制、转录后调节、蛋白质翻译后修饰及蛋白-蛋白相互作用等.JunD基因表达的精细调控及JunD蛋白与其它蛋白之间的相互作用可调节细胞增殖、分化和凋亡等过程.JunD蛋白活性异常会导致肿瘤、代谢及病毒类疾病的发生.JunD蛋白的转录激活及抑制受1个复杂调控网络调控,在这个网络调节下,JunD蛋白在细胞的生长调控过程中发挥重要作用.本文就JunD基因表达的调控机制及其与肿瘤之间关系的最新研究进展做一综述.     

植物光敏色素PHYA、PHYB研究进展

光是植物生长发育过程中一个重要的环境信号,光敏色素能够把外界的红光和远红光转变成生物体内的信号。介绍近年来光敏色素结构和其相互作用蛋白的研究进展。     

Genetic variants of glutathione S-transferase and the risk of acute myeloid leukemia in a Saudi population

ObjectiveThis study aims to investigate the genetic association of acute myeloid leukemia and glutathione S-transferase (GST) gene polymorphisms in a Saudi population.Method100 AML cases and 100 healthy controls were recruited from the Riyadh regional hospital. In the GST gene, GSTM1 and GSTT1 variants were genotyped by multiplex PCR, and GSTP1 variants were genotyped by PCR-RFLP analysis. Statistical analysis between AML cases and controls included anthropometric measurements and evaluation of the genotypic and allelic frequencies.ResultThe null genotypes of GSTM1 and GSTT1 showed no association with AML [OR 0.56 (0.26–1.19); p = 0.31 and OR 0.65 (0.37–1.16); p = 0.14]. Similarly, the GSTP1 genotype and allele frequencies did not indicate any association with AML [GG + AG vs. AA: OR 0.75 (0.43–1.31) and p = 0.32; GG vs. AA: OR 1.73 (0.55–5.44) and p = 0.34; G vs. A: OR 0.95 (0.61–1.46) and p = 0.82]. Further, a haplotype analysis between AML cases and controls did not show any positive association (p < 0.05).ConclusionIn conclusion, there was no statistical association of the genotypes and alleles in GSTM1, GSTT1, and GSTP1 with AML. Our results confirm the negative association of the investigated genetic markers with susceptibility to AML. Further association studies would be required in different ethnic populations to facilitate a meta-analysis in the future. Our findings suggest that the GST gene has no role in the pathogenesis of AML in patients from Saudi Arabia.     

NOD1真核表达质粒的构建及表达

目的:构建天然免疫胞内识别受体核苷酸寡聚域1(NOD1)真核表达质粒。方法:NOD1基因片段经PCR扩增获得,经酶切后连接到真核表达载体pcDNA3/flag中,对挑选出的阳性克隆测序,将序列正确的重组质粒pflag-NOD1转染293T细胞,用Western印迹检测目的蛋白的表达,同时用NF-κB的萤光素酶报告基因检测NOD1蛋白的活性。结果:pflag-NOD1可以在真核细胞293T中表达,并可以增强NF-κB报告基因的转录活性。结论:构建了重组质粒pflag-NOD1,在细胞中表达NOD1后能够提高NF-κB转录的生物活性,为进一步研究NOD1的功能奠定了基础。     

FXR1真核表达载体的构建与表达及其对神经节苷脂浓度的影响

目的:构建脆性X相关基因1(FXR1)的真核表达载体并检测其对神经节苷脂(GM1)浓度的影响。方法:以pYESTrp3-FXR1为模板,利用PCR扩增FXR1基因,PCR产物经EcoR I和Xho I双酶切后插入真核表达载体pcDNA3.1(-)中,获得的阳性克隆进一步酶切及测序鉴定;将构建成功的pcDNA3.1(-)-FXR1转染SH-SY5Y细胞后,采用Western blot检测FXR1的表达情况,同时采用ELISA试剂盒检测细胞内GM1的浓度。结果:PCR扩增产物为1.9 Kb的片段,与FXR1基因大小相符,阳性克隆经双酶切后获得两条分别为5.4 Kb和1.9 Kb的片段,测序结果与GeneBank中序列相同。构建成功的重组质粒pcDNA3.1(-)-FXR1转染SH-SY5Y细胞后,细胞中FXR1的表达增加,同时有效提高了细胞内GM1的浓度(P0.05)。结论:成功构建了真核表达载体pcDNA3.1(-)-FXR1,FXR1的表达增加可以提高SH-SY5Y细胞中的GM1浓度,这些为后续深入研究FXR1基因在神经组织发育中的调控功能及其在脆性X综合征(FXS)中的作用机制奠定了基础。     

Molecular and cellular events at the site of myocardial infarction: from the perspective of rebuilding myocardial tissue

The potential for bone marrow-derived progenitor cells (BMDPC) to regenerate myocardial tissue following infarction depends on homing, migration, nourishment, and spatially appropriate growth of BMDPC. Requisite to these objectives is the expression of adhesion molecules (ICAM-1) and chemoattractant cytokines (MCP-1), matrix metalloproteinase (MMP) activity, a neovasculature, and fibrillar collagen scaffolding. We found that enhanced ICAM-l and MCP-1, as well as MMP activity on day 3 and 7 postMI, are present to facilitate the homing, chemotaxis, and migration of circulating cells into the infarct site. The vascular network formed at the infarct site contains a ratio of conduit to exchange vessels that is greater than that for control tissue and therefore its ability to nourish BMDPC for their growth appears to be tenuous. These findings, together with the dense formation of a fibrillar collagen scar beyond week 2, suggest the optimal time to rebuild myocardium from BMDPC resides within 2 week postMI.     

Mechanism of Crosstalk between the LSD1 Demethylase and HDAC1 Deacetylase in the CoREST Complex

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HIV-1蛋白酶的表达、纯化及其抑制剂体外筛选方法的建立

从HIV-1 ⅢB病毒RNA经RT-PCR得到HIV-1蛋白酶编码序列,克隆到pet28a质粒中构建HIV-1蛋白酶表达载体.阳性克隆转染E.coli BL21 DE3,经IPTG诱导,蛋白酶以包涵体的形式表达,表达量占菌体总蛋白量的40%.包涵体经Triton X-100洗涤后溶解于8M尿素,溶解后的蛋白溶液经sephacyl s-200 H.R分子筛柱纯化后纯度达到90%以上,收集蛋白酶峰稀释复性并通过超滤进行浓缩.经检测,纯化的蛋白酶具有较高的活性.用荧光标记的蛋白酶底物检测不同浓度indinavir对蛋白酶活性的影响,表明该方法可以用于蛋白酶抑制剂的筛选.     

大鼠脑内caveolin-1蛋白的表达及其在分辨学习中的作用

Caveolin-1（Cav—1）蛋白作为细胞质膜结构小窝（caveolae）的标志蛋白,在胆固醇运输、膜组装、信号转导和细胞转化过程中扮演重要的角色。为了探讨Cav-1蛋白在中枢神经系统可塑性及学习记忆中的作用,本文以Sprague—Dawley大鼠为实验对象,利用蛋白质免疫印迹杂交方法观察了Car-1蛋白在不同年龄大鼠脑内表达的特征,并研究了Y-迷宫训练前后Cav-1蛋白表达的变化。结果表明：（1）大鼠不同脑区Cav-1蛋白表达的年龄特征不同。海马内的表达属青年鼠最高,其次是老年鼠和幼年鼠;皮层内的表达属幼年鼠最高,其次是老年鼠,青年鼠最低;小脑内的表达无明显年龄差异。（2）Y-迷宫训练引起青年鼠海马和前额叶皮层内Cav-1蛋白的表达显著增加。结果提示,Cav-1蛋白与动物脑发育和学习记忆有密切关系,可能参与中枢可塑性的调节。