CCR2-64Ⅰ在中国南方14个少数民族群体中的分布

为调查HIV-1感染相关等位基因CCR2-64Ⅰ在我国南方14个少数民族群体的频率和多态性分布, 从上述人群外周血中抽提基因组DNA, 采用PCR和PCR-RFLP等方法进行基因分型。在791例调查对象中, 636例是野生纯合子基因型, 104例为杂合子基因型, 51例为突变纯合子基因型。上述各群体等位基因型的分布符合Hardy-Weinberg平衡。14个民族群体的平均突变基因频率为13.6％, 等位基因频率范围分布在1.6％~30.3％之间, 14个民族群体之间突变基因频率具有显著差异(P<0.05)。广西壮族群体CCR2-64Ⅰ突变基因频率最低, 为1.6％, 云南的六库傈僳族频率最高, 为30.3％。12个群体的突变基因频率均低于中国汉族健康群体, 南方3个少数民族群体基因突变频率显著低于西南11个少数民族群体, 该突变基因在艾滋病发病过程中的影响值得进一步深入研究。     

MLT抑制E2诱发的大鼠垂体PRL瘤的增生涉及雌激素受体的作用

目的：探讨褪黑素（melatonin,MLT）抑制17-β-雌二醇（17-β-estradiol,E2）诱发垂体催乳素（prolactin,PRL）瘤的增生及其机制的初始阶段,MLT对雌激素受体的作用。方法：在体实验采用每日定时给各组SD大鼠分别皮下注射不同浓度的MLT,建立MLT抑制E2诱发的垂体PRL瘤增生的动物模型。离体实验采用原代培养细胞原位杂交方法,探讨垂体PRL瘤细胞MLT受体的表达、MLT对雌激素受体（ER）表达的作用;应用电泳迁移率改变（EMSA）的方法,观察MLT对雌激素受体（ER）与雌激素反应元件（ERE）结合的效应。结果：每只大鼠每日定时皮下注射0．25或0．50mg MLT能显著抑制E2诱发的垂体PRL瘤的增生（分别P〈0．01、P〈0．05）;F4诱发的垂体PRL瘤细胞内有MLT受体MLT1a和MLT1b;给0．25mg／day／rat MLT组的大鼠垂体PRL瘤细胞内ER的表达显著减少（P〈0．01）、ER与ERE的结合量显著降低（P〈0．01）。结论：一定剂量的MLT能显著抑制E2诱发的SD大鼠垂体PRL瘤的增生,其作用机制之一可能与MLT抑制ER的表达、及其部分阻断ER与ERE的结合有关。     

Functional polymorphisms of matrix metallopeptidase-9 and risk of coronary artery disease in a Chinese population

Matrix metallopeptidase-9 (MMP-9) plays a pivotal role in vascular remodeling and development of atherosclerotic lesion. The potentially functional MMP-9 polymorphisms may contribute to the susceptibility of coronary artery disease (CAD). A case–control study composed of 762 CAD cases and 555 CAD-free controls was conducted in a Chinese population to investigate the association between the MMP-9 ?1562 C>T, R279Q, P574R and R668Q polymorphisms and CAD risk. It was found that the variant genotypes of R279Q, P574R and R668Q were associated with a non-significant decreased risk of CAD when compared with their wild-type genotypes, respectively, Furthermore, compared with those without any variant genotypes for these four nonsynonymouse loci, individuals carrying all four variant genotypes (?1562 CT/TT, 279 RQ/QQ, 574 PR/RR and 668 RQ/QQ) had a 51% decreased risk of CAD (adjusted OR = 0.49; 95% CI = 0.26–0.95, P = 0.033). Although no significant main effects were observed for MMP-9 ?1562 C>T locus on CAD risk, variant genotypes of ?1562 C>T were associated with a 2.53 increased risk of CAD in subjects with diabetes mellitus (DM) (95% CI = 1.18–5.45, P = 0.018). In CAD cases, variant genotypes of ?1562 C>T were associated with a significantly increased risk of MI (adjusted OR, 1.48, 95% CI, 1.01–2.20, P = 0.048). These findings suggest that MMP-9 R279Q, P574R and R668Q may have combined effect in the occurrence of CAD and ?1562 CT/TT genotypes may contribute to CAD in diabetics and MI in CAD patients.     

Design of a Potent d-Peptide HIV-1 Entry Inhibitor with a Strong Barrier to Resistance

The HIV gp41 N-trimer pocket region is an ideal viral target because it is extracellular, highly conserved, and essential for viral entry. Here, we report on the design of a pocket-specific d-peptide, PIE12-trimer, that is extraordinarily elusive to resistance and characterize its inhibitory and structural properties. d-Peptides (peptides composed of d-amino acids) are promising therapeutic agents due to their insensitivity to protease degradation. PIE12-trimer was designed using structure-guided mirror-image phage display and linker optimization and is the first d-peptide HIV entry inhibitor with the breadth and potency required for clinical use. PIE12-trimer has an ultrahigh affinity for the gp41 pocket, providing it with a reserve of binding energy (resistance capacitor) that yields a dramatically improved resistance profile compared to those of other fusion inhibitors. These results demonstrate that the gp41 pocket is an ideal drug target and establish PIE12-trimer as a leading anti-HIV antiviral candidate.The HIV envelope protein (Env) mediates viral entry into cells (11). Env is cleaved into surface (gp120) and transmembrane (gp41) subunits that remain noncovalently associated to form trimeric spikes on the virion surface (16). gp120 recognizes target cells by interacting with cellular receptors, while gp41 mediates membrane fusion. Peptides derived from heptad repeats near the N and C termini of the gp41 ectodomain (N and C peptides) interact in solution to form a six-helix bundle, representing the postfusion structure (3, 55, 56). In this structure, N peptides form a central trimeric coiled coil (N trimer), creating grooves into which C peptides bind. This structure, in conjunction with the dominant-negative inhibitory properties of exogenous N and C peptides, suggests a mechanism for Env-mediated entry (10, 22, 58-60).During entry, gp41 forms an extended prehairpin intermediate that leaves the exposed N-trimer region vulnerable to inhibition for several minutes (18, 35). This intermediate ultimately collapses as the C-peptide regions bind to the N-trimer grooves to form a trimer of hairpins (six-helix bundle), juxtaposing viral and cellular membranes and inducing fusion. Enfuvirtide (Fuzeon), the only clinically approved HIV fusion inhibitor, is a C peptide that binds to part of the N-trimer groove and prevents six-helix bundle formation in a dominant-negative manner (61). Enfuvirtide is active in patients with multidrug resistance to other classes of inhibitors and is a life-prolonging option for these patients (30, 31). However, enfuvirtide use is restricted to salvage therapy due to several limitations, including (i) high dosing requirements (90 mg, twice-daily injections), (ii) high cost (∼$30,000/year/patient in the United States), and (iii) the rapid emergence of resistant strains (21, 47).A deep hydrophobic pocket at the base of the N-trimer groove is an especially attractive inhibitory target because of its high degree of conservation (3, 12, 48), poor tolerance to substitution (4, 34), and critical role in membrane fusion (2). Indeed, this region is conserved at both the amino acid level (for gp41 function in membrane fusion) and the nucleotide level (for the structured RNA region of the Rev-responsive element). Enfuvirtide binds to the N-trimer groove just N terminal to the pocket and is significantly more susceptible to resistance mutations than 2nd-generation C-peptide inhibitors, such as T-1249, that also bind to the pocket (8, 13, 29, 44, 46, 47, 58).Peptide design, molecular modeling, and small-molecule screening have produced a diverse set of compounds that interact with the gp41 pocket and inhibit HIV-1 entry with modest potency, but often with significant cytotoxicity (7, 14, 15, 17, 23, 24, 26, 34, 51, 54). The first direct evidence that pocket-specific binders are sufficient to inhibit HIV entry came with the discovery of protease-resistant d-peptides identified using mirror-image phage display (12). In this technique, a phage library is screened against a mirror-image version of the target protein (synthesized using d-amino acids) (50). By symmetry, mirror images (d-peptides) of the discovered sequences will bind to the natural l-peptide target. As the mirror images of naturally occurring l-peptides, d-peptides cannot be digested by natural proteases. Protease resistance provides d-peptides theoretical treatment advantages of extended survival in the body and possible oral bioavailability (41, 42, 49).These 1st-generation d-peptide entry inhibitors possess potency against a laboratory-adapted isolate (HXB2) at low to mid-μM concentrations (12). We previously reported an affinity-matured 2nd-generation d-peptide called PIE7, pocket-specific inhibitor of entry 7 (57). A trimeric version of PIE7 is the first high-affinity pocket-specific HIV-1 inhibitor and has potency against X4-tropic (HXB2) and R5-tropic (BaL) strains at sub-nM concentrations. However, significant further optimization is required to create a robust clinical candidate for two reasons. First, this d-peptide is much less potent (requiring high nM concentrations) against JRFL, a primary R5-tropic strain. Therefore, improved PIE potency is necessary to combat diverse primary strains. Second, by improving the affinity of our inhibitors for the pocket target, we hope to provide a reserve of binding energy that will delay the emergence of drug resistance, as described below.We and others have reported a potency plateau for some gp41-based fusion inhibitors that is likely imposed by the transient exposure of the prehairpin intermediate (9, 27, 53, 57). For very high-affinity inhibitors, association kinetics (rather than affinity) limits potency so that two inhibitors with significantly different affinities for the prehairpin intermediate can have similar antiviral potencies. We proposed that overengineering our d-peptides with substantial affinity beyond this potency plateau would provide a reserve of binding energy that would combat affinity-disrupting resistance mutations (57). Such a resistance capacitor should also prevent the stepwise accumulation of subtle resistance mutations in Env by eliminating the selective advantage that such mutants would otherwise confer.Here, we report on the design and characterization of a 3rd-generation pocket-specific d-peptide, PIE12-trimer, with ∼100,000-fold improved target binding compared to that of the best previous d-peptide, significantly broadened inhibitory potency, and an enhanced resistance capacitor that provides a strong barrier to viral resistance. We achieved this increased potency via structure-guided phage display and crosslinker optimization. PIE12-trimer has a dramatically improved resistance profile compared to the profiles of earlier d-peptides, as well as those of enfuvirtide and T-1249. These results validate the resistance capacitor hypothesis and establish PIE12-trimer as a leading anti-HIV therapeutic candidate.     

带有干扰和非线性指标的脉冲时滞SI模型的渐近性质(英文)

研究了一个带有干扰和非线性指标的脉冲时滞SI模型,通过引入三个引理,获得该疾病最终灭绝和保持持久的充分性条件。结果表明,时滞因素对该模型的全局吸引性和持久性都有影响。此外。如果脉冲免疫接种率的最大值和最小值之比大于某一阈值,则该疾病最终灭绝,本文的主要特点是将多时滞和变系数引入脉冲SI模型,数值模拟表明,该系统具有复杂的动力学行为,包括周期解和周期振荡.     

Flexible interwoven termini determine the thermal stability of thermosomes

Group II chaperonins, which assemble as double-ring complexes, assist in the refolding of nascent peptides or denatured proteins in an ATP-dependent manner. The molecular mechanism of group II chaperonin assembly and thermal stability is yet to be elucidated. Here, we selected the group II chaperonins (cpn-α and cpn-β), also called thermosomes, from Acidianus tengchongensis and investigated their assembly and thermal stability. We found that the binding of ATP or its analogs contributed to the successful assembly of thermosomes and enhanced their thermal stabilities. Cpn-β is more thermally stable than cpn-α, while the thermal stability of the hetero thermosome cpn-αβ is intermediate. Cryo-electron microscopy reconstructions of cpn-α and cpn-β revealed the interwoven densities of their non-conserved fl exible N/C-termini around the equatorial planes. The deletion or swapping of their termini and pH-dependent thermal stability assays revealed the key role of the termini electrostatic interactions in the assembly and thermal stability of the thermosomes.     

奶山羊β-乳球蛋白基因5'、3'调控区的克隆和序列分析

目的:克隆奶山羊β-乳球蛋白(BLG)基因5'、3'调控区,并对其进行序列分析.方法:从徐淮奶山羊组织中提取基因组DNA,采用高保真长模板PCR法克隆得到奶山羊BLG基因4.2 kb的5'调控区和1.8 kb的3'调控区;将纯化后的PCR产物克隆到pGEM-T Easy载体中,经酶切鉴定和序列测定验证克隆的正确性.结果:部分序列分析表明,奶山羊BLG基因5'区与GenBank中登录的山羊和绵羊BLG基因5'区的同源性分别为100%和95%,3'区的同源性分别为99%和93%;克隆得到的奶山羊BLG基因调控区与山羊BLG伪基因显著不同,二者5'区和3'区的同源性分别为83%和88%.结论:克隆得到的奶山羊BLG基因调控区可用于构建乳腺特异性表达载体,用于外源基因在转基因动物乳腺中的高效表达.     

Cdk5 regulates accurate maturation of newborn granule cells in the adult hippocampus

Newborn granule cells become functionally integrated into the synaptic circuitry of the adult dentate gyrus after a morphological and electrophysiological maturation process. The molecular mechanisms by which immature neurons and the neurites extending from them find their appropriate position and target area remain largely unknown. Here we show that single-cell–specific knockdown of cyclin-dependent kinase 5 (cdk5) activity in newborn cells using a retrovirus-based strategy leads to aberrant growth of dendritic processes, which is associated with an altered migration pattern of newborn cells. Even though spine formation and maturation are reduced in cdk5-deficient cells, aberrant dendrites form ectopic synapses onto hilar neurons. These observations identify cdk5 to be critically involved in the maturation and dendrite extension of newborn neurons in the course of adult neurogenesis. The data presented here also suggest a mechanistic dissociation between accurate dendritic targeting and subsequent synapse formation.     

真正柑橘果树类植物基于AFLP分子标记的分类与进化研究

用AFLP分子标记技术对芸香科Rutaceae柑橘亚科Aurantioideae真正柑橘果树类(the True Citrus Fruit Trees Group) 6属植物的59个生物类型以及九里香Murraya exotica L.、蚝壳刺Severinia buxifolia Tenore和酒饼簕Atalantia buxifolia Oliv.进行了分析。从64对引物组合中筛选出15对分辨率好的引物对全部材料进行了扩增, 共获得312条带, 其中多态型带305条, 多态位点达97.8%。利用PAUP软件构建了全部材料基于AFLP数据的UPGMA距离树。结果表明, AFLP证据支持基于形态特征建立的真正柑橘果树群以及群内各属的划分, 只是金柑属Fortunella Swingle作为一个独立的分支嵌于柑橘属Citrus L.内。对于柑橘属, 我们的分子证据支持Swingle关于亚属的划分以及Scora等关于C. medica、C. grandis和C. reticulata是真正柑橘亚属subgen. Eucitrus三个基本种的观点。AFLP分子证据显示宜昌橙C. ichangensis Swingle位于真正柑橘亚属中, 与香橙C. junos Sieb. ex Tanaka的亲缘关系最近, 而与大翼橙亚属subgen. Papeda中马蜂柑C. hystrix DC.相隔甚远。另外, 莽山野橘C. mangshanensis S. W. He &; G. F. Liu在系统树上居于宽皮柑橘类的基部。最后, 对真正柑橘果树群6属植物间以及柑橘属各主要类型(种)的进化关系进行了详细的讨论。