Role of the mitochondrial pathway in serum deprivation-induced apoptosis of rat endplate cells

The apoptosis of cartilage endplates (CEPs), acting as an initiating factor, plays a vital role in the pathogenesis of intervertebral disc degenerative diseases, the underlying molecular mechanism of the apoptotic process in CEPs is still not clear. The present study aimed to investigate the mechanism of CEP cell apoptosis. We found that low levels of fetal bovine serum (FBS) can induce cell apoptosis. Serum deprivation led to high expression levels of caspase-9, caspase-3, PARP, cytochrome-c and Bax. Flow cytometric analysis showed that inhibition of the intrinsic pathway by a caspase-9 inhibitor (z-LEHD-fmk) significantly suppressed serum deprivation-induced apoptosis. However, a caspase-8 inhibitor (z-IETD-fmk) did not reduce apoptotic cell death. These data suggest that serum deprivation induces apoptosis in rat CEP cells via the activation of the intrinsic apoptotic pathway. The efficacy of a caspase-9 inhibitor in attenuating or preventing apoptosis of serum deprivation-induced disc cell apoptosis suggests that targeting the intrinsic apoptotic pathway may be used as a potential therapy for the treatment of disc degeneration.     

谱系年代研究进展

谱系年代学是结合化石记录和分子钟方法计算"生命树"(Tree of Life)上各分歧点时间的一个新兴交叉学术领域.由于化石记录的不完整性,各类生物的起源年代和支系分化时间的确定可借助于部分化石记录和通过计算现生生物类群之间的遗传距离转换得出的相对分歧时间相结合的办法进行讨论.化石记录可代表部分生物类群起源时间的保守估计值,而分子钟方法可为那些不易保存为化石的生物类群与其姊妹群的分化时间提供依据.如果使用得当的话,两者可进行相互验证.在分子谱系年代分析中,正确使用化石校准方案是获得准确分歧时间的关键,这需要:1)正确确定化石物种在谱系树上的位置;2)正确解释化石记录所代表的时间含义(最小值、确定值、最大值及其标准差).分子钟估算分歧时间的技术在不断改进,目前常用的分子分歧时间估算法(宽松分子钟法,如贝叶斯法、补偿性似然法等)包容分子演化速率在谱系间和随时间的变化.随着谱系年代研究的不断深入,长期困扰人们的化石记录时间与分子钟计算结果悬殊的问题正在逐渐趋于和谐并得到正确诠释.文中还讨论了有关动物起源与早期分化时间以及早期陆生节肢动物的谱系年代学研究进展.我们强调,化石记录和分子钟分析可以优势互补,两者的整合无疑将提高生物演化历史时间格架的准确度和精度,以利更好地将生命演化事件置于地球系统科学及地球环境演化史之中.     

细菌素Paracin 1.7的纯化与性质研究

摘要：【目的】分离纯化（Lactobacillus paracasei）HD1.7所产生的细菌素并分析其特性。【方法】细菌素Paracin 1.7的纯化采用色谱技术,其分子量检测采用十二烷基磺酸钠－聚丙烯酰胺凝胶电泳（SDS-PAGE）,利用琼脂扩散法测定细菌素活力。【结果】Paracin 1.7分离于我国传统发酵食品酸菜发酵液中,其产生菌为副干酪乳杆菌。 Paracin 1.7可以抑制其它微生物的生长,为细菌素。该菌在稳定期可产生大量Paracin 1.7。经过阳离子交换层析、凝胶过滤层析以及高效液相色谱（HPLC）,对该细菌素进行了初步纯化,并经Tricine-SDS-PAGE检测其分子量大约为11 kDa。Paracin 1.7抑菌谱较广,其抑菌范围包括Proteus, Bacillus, Enterobacter, Staphylococcus, Escherichia, Lactobacillus, Microccus, Pseudomonas, Salmonella, Saccharomyces,其中有些为食品源致病菌。该细菌素在酸性及高温下稳定,对几种蛋白质酶敏感。该细菌素对敏感菌株的作用方式为抑菌。在4oC保存4个月后,Paracin 1.7的抑菌活性保持稳定。【结论】基于细菌素Paracin 1.7的性质,该细菌素可用作食品防腐剂。     

Effectively reducing sensory eye dominance with a push-pull perceptual learning protocol

Much knowledge of sensory cortical plasticity is gleaned from perceptual learning studies that improve visual performance [1-7]. Although the improvements are likely caused by modifications of excitatory and inhibitory neural networks, most studies were not primarily designed to differentiate their relative contributions. Here we designed a novel push-pull training protocol to reduce sensory eye dominance (SED), a condition that is mainly caused by unbalanced interocular inhibition [8-10]. During the training, an attention cue presented to the weak eye precedes the binocular competitive stimulation. The cue stimulates the weak eye (push) while causing interocular inhibition of the strong eye (pull). We found that this push-pull protocol reduces SED (shifts the balance toward the weak eye) and improves stereopsis more so than the push-only protocol, which solely stimulates the weak eye without inhibiting the strong eye. The stronger learning effect with the push-pull training than the push-only training underscores the crucial involvement of a putative inhibitory mechanism in sensory plasticity. The design principle of the push-pull protocol can potentially lend itself as an effective, noninvasive treatment of amblyopia.     

The Recombination Protein RAD52 Cooperates with the Excision Repair Protein OGG1 for the Repair of Oxidative Lesions in Mammalian Cells

Oxidized bases are common types of DNA modifications. Their accumulation in the genome is linked to aging and degenerative diseases. These modifications are commonly repaired by the base excision repair (BER) pathway. Oxoguanine DNA glycosylase (OGG1) initiates BER of oxidized purine bases. A small number of protein interactions have been identified for OGG1, while very few appear to have functional consequences. We report here that OGG1 interacts with the recombination protein RAD52 in vitro and in vivo. This interaction has reciprocal functional consequences as OGG1 inhibits RAD52 catalytic activities and RAD52 stimulates OGG1 incision activity, likely increasing its turnover rate. RAD52 colocalizes with OGG1 after oxidative stress to cultured cells, but not after the direct induction of double-strand breaks by ionizing radiation. Human cells depleted of RAD52 via small interfering RNA knockdown, and mouse cells lacking the protein via gene knockout showed increased sensitivity to oxidative stress. Moreover, cells depleted of RAD52 show higher accumulation of oxidized bases in their genome than cells with normal levels of RAD52. Our results indicate that RAD52 cooperates with OGG1 to repair oxidative DNA damage and enhances the cellular resistance to oxidative stress. Our observations suggest a coordinated action between these proteins that may be relevant when oxidative lesions positioned close to strand breaks impose a hindrance to RAD52 catalytic activities.Oxidative DNA damage is generated at high levels in mammalian cells, even in cells not exposed to exogenous sources of reactive oxygen species. Several kinds of DNA modifications are formed upon oxidative stress (8). The most prevalent modifications, quantitatively, are single-strand breaks and oxidized bases. Clustered DNA damage, when two or more modifications are closely positioned in opposite strands, is detectable after gamma irradiation and has recently been shown to be generated by normal oxidative metabolism (3, 35). One unique aspect of such clustered lesions is that they can be converted into double-strand breaks (DSB) if a DNA glycosylase removes the two opposite bases and an apurinic/apyrimidinic (AP)-endonuclease cleaves the resulting abasic sites. Thus, although quantitatively minor, DSB are possible outcomes of oxidative DNA damage.Oxidized DNA bases are repaired primarily by the base excision repair pathway (BER) (22, 39). BER is initiated by a lesion-specific DNA N-glycosylase that recognizes and excises the damaged base. Eight-hydroxyguanine (8-oxoG) is one of the most abundant oxidized bases detected in cellular DNA. This adduct is easily bypassed by replicative polymerases; however, it can direct the misincorporation of adenine opposite 8-oxoG, thus leading to G·C-to-T·A transversion mutations (31). 8-oxoG accumulation has been causally associated with carcinogenesis and aging in several experimental models (1, 12). In eukaryotes, oxoguanine DNA glycosylase (OGG1) is the major 8-oxoG DNA glycosylase. OGG1 possesses an associated AP-lyase activity, such that it removes 8-oxoG and cleaves the DNA backbone. Human cells express two distinct OGG1 isoforms, α and β, which share the first 316 amino acids but differ significantly in their C termini (25). While OGG1-α is a bone fide DNA glycosylase (5) and localizes both to nuclei and mitochondria, OGG1-β localizes exclusively to mitochondria. We recently showed that the recombinant OGG1-β protein has no DNA glycosylase activity (13). The high degree of conservation of repair pathways for 8-oxoG, from bacteria to humans, along with epidemiological data correlating OGG1 polymorphisms and activity with predisposition to some cancers (11, 27, 33) attest to the biological importance of the repair of 8-oxoGs and other oxidative DNA lesions.Until recently, distinct classes of DNA lesions were believed to be metabolized by different and independent repair pathways. However, experimental evidence indicates that these pathways can interact and that there is a considerable degree of overlap in their substrate specificity and in the proteins that participate in each pathway. Experiments using yeast strains lacking one or more distinct DNA repair genes suggest that DSB repair pathways may play a role in repair of oxidative DNA damage. Swanson et al. showed that while yeast cells lacking ntg1 and ntg2 (homologues of Escherichia coli endonuclease III, a DNA glycosylase specific for pyrimidine lesions formed by oxidation) and apn1 (the major yeast abasic site endonuclease) are not overtly sensitive to oxidative stress, the additional disruption of the rad52 gene significantly increases sensitivity to H₂O₂ and menadione (36). Similarly, yeast cells expressing decreased levels of frataxin, which leads to elevated oxidative stress, show accumulation of oxidative damage in nuclear DNA only in a rad52 mutant background (18). RAD52 is a member of the RAD51 epistatic group. These proteins are believed to be involved in the early steps of homologous recombination, contributing to homology search and strand invasion; disruption of the corresponding genes renders cells deficient in DSB repair and hyper-recombinogenic (19).These results suggested a possible role for RAD52 in the repair of oxidative DNA damage. Moreover, an in vitro screening of protein partners that interact physically with OGG1-β performed in our lab (unpublished data) showed that human RAD52 strongly interacted with this glycosylase, again suggesting a possible function for RAD52 in the oxidative DNA damage response. Thus, we investigated whether RAD52 plays a role in the repair of oxidative DNA damage in human cells. We show here that human RAD52 physically interacts with both OGG1-α and -β, in vitro and in cell extracts. We also show that OGG1-α and -β inhibit RAD52 enzymatic activities. Conversely, RAD52 stimulates OGG1-α 8-oxoG incision activity. RAD52 colocalizes with OGG1-α in cells, and this colocalization increases after oxidative stress. Moreover, lower RAD52 expression, via gene knockdown (KD) or disruption of the RAD52 gene, render cells sensitive to oxidative stress. Based on our results, we discuss a model in which OGG1 and RAD52 cooperate to repair 8-oxoG lesions.     

千岛湖浮游甲壳动物垂直分布与昼夜垂直移动

2004年6月、9月、12月及2005年3月于千岛湖温馨岛站点(29°38′10.5″N,119°01′54.1″E)进行浮游甲壳动物垂直分布分层采样,分析千岛湖浮游甲壳动物的垂直分布特征及优势种昼夜垂直移动状况。结果显示,千岛湖浮游甲壳动物主要分布在10～21 m水层,不同季节中心水层分布深度12月9月6月3月;浮游动物昼夜垂直移动春(3月)、夏(6月)两季幅度较大,秋(9月)、冬(12月)季节幅度较小;浮游甲壳动物优势种在各个水层都有分布,密集区大都集中在中上水层(10～21 m),不同种类昼夜垂直移动幅度有所不同;昼夜垂直移动显著的种类有特异荡镖水蚤(Neutrodiaptanus incongruens)、近邻剑水蚤(Cyclops vicinus)、球状许水蚤(Schmackeria forbesi)、透明溞(Daphnia hyaline)、长额象鼻溞(Bosmina longirostris),不显著的种类为右突新镖水蚤(Neodiaptomus schmackeri)、短尾秀体溞(Diaphanosoma brachyurum)。光照、温度、饵料是影响千岛湖浮游甲壳动物垂直分布及昼夜垂直移动的主要因素。     

微生物降解苯甲酸的研究进展

苯甲酸在工业中的广泛应用使其成为环境中的常见污染物,对微生物好氧降解苯甲酸的邻位途径、间位途径、龙胆酸途径和原儿茶酸途径及厌氧降解途径等进行总结,并对苯甲酸降解过程中发挥重要作用的苯甲酸双加氧酶的种类、不同组分及苯甲酸降解基因和调控基因的基因簇进行介绍,同时展望微生物降解污染物的发展方向。     

Contribution of arbuscular mycorrhizal fungi of sedges to soil aggregation along an altitudinal alpine grassland gradient on the Tibetan Plateau

The diversity of arbuscular mycorrhizal fungi (AMF) in sedges on the Tibetan Plateau remains largely unexplored, and their contribution to soil aggregation can be important in understanding the ecological function of AMF in alpine ecosystems. Roots of Kobresia pygmaea C.B. Clarke and Carex pseudofoetida Kük. in alpine Kobresia pastures along an elevational transect (4149–5033 m) on Mount Mila were analysed for AMF diversity. A structural equation model was built to explore the contribution of biotic factors to soil aggregation. Sedges harboured abundant AMF communities covering seven families and some operational taxonomic units are habitat specific. The two plant species hosted similar AMF communities at most altitudes. The relative abundance of the two sedges contributed largely to soil macroaggregates, followed by extraradical mycorrhizal hyphae (EMH) and total glomalin‐related soil protein (T‐GRSP). The influence of plant richness was mainly due to its indirect influence on T‐GRSP and EMH. There was a strong positive correlation between GRSP and soil total carbon and nitrogen. Our results indicate that mycorrhization might not be a major trait leading to niche differentiation of the two co‐occurring sedge species. However, AMF contribute to soil aggregation and thus may have the potential to greatly influence C and N cycling in alpine grasslands.     

东方田鼠胚胎成纤维细胞永生化细胞系的建立

目的建立东方田鼠胚胎成纤维永生化细胞系,为全面研究东方田鼠抗日本血吸虫机制以及开展不同动物成纤维细胞间比较研究奠定基础和提供细胞实验材料。方法运用脂质体介导的基因转染法将pSV3neo质粒导入第3代东方田鼠胚胎成纤维细胞,经G418筛选抗性克隆并扩大培养,建立永生化细胞系;用PCR检测细胞株中SV40T基因的整合,RT-PCR鉴定SV40T基因在转染细胞中的表达;绘制东方田鼠胚胎成纤维永生化细胞生长曲线。结果阳性细胞克隆已扩大培养并稳定传代50代,经鉴定SV40T抗原已整合到东方田鼠胚胎成纤维细胞中且稳定表达。结论成功建立东方田鼠胚胎成纤维永生化细胞系。