绝经对血液单核细胞基因表达谱的影响:基因芯片初步研究

绝经是女性一生中很重要的生理现象之一,它能增加一系列复杂免疫、神经退化、新陈代谢和心血管方面的疾病。血液单核细胞能分化成各种各样的细胞,这些细胞在组织形态发生和免疫应答方面起着很重要的作用。本研究中采用了包含大约14,500个基因探针的Affymetrix Human U133A基因芯片来研究健康的绝经前和绝经后女性外周血液单核细胞中的基因表达谱。样本之间的对比分析表明有20个基因上调,20个基因下调。其中的28个基因根据它们的生物过程如细胞繁殖、免疫应答、细胞代谢等等被分成了6个主要的GO类别;剩下的12个基因其生物学功能还没有被鉴定。研究结果支持了我们的假设:血液单核细胞的功能状态确实受到绝经的影响,而且由此带来的改变可能是由全基因组范围的基因表达谱而决定的。本研究中鉴定的一些差异表达基因有可能作为以后研究与绝经相关的系统免疫、神经退化和心血管疾病的候选基因研究。此工作是这个研究方向的第一次尝试,为将来的进一步研究奠定了基础。     

Early-stage litter decomposition and its influencing factors in the wetland of the Sanjiang Plain, China

Using the litter bag technique, the decomposition rates and their influencing factors were studied by investigating three wetland macrophytes, Calamagrostics angustifolia, Carex meyeriana and Carex lasiocapa, in Sanjiang Plain, Northeast China. It was revealed that C. lasiocapa lost 28.91% of its weight, C. angustifolia lost 31.98% and C. meyeriana lost 32.99% after 164 days. Another finding was that the amount of organic carbon in the litter of C. angustifolia and C. lasiocapa fluctuated, but continuously decreased in that of C. meyeriana. However, all the three types of litter released organic carbon. Nitrogen was released substantially from the litter of both C. angustifolia and C. meyeriana, but accumulated in the litter of C. lasiocapa. Phosphorus concentrations in all the three types of litter apparently decreased first and then slightly increased. Overall, P release was observed in all the three types of litter. The C/N and C/P ratios varied significantly in the decomposition process. The decomposition rates and nutrient content variations were simultaneously influenced by the quality of the litter as well as the environmental factors in the Sanjian Plain, but they were more strongly affected by the quality of the litter.     

Phylogeny and biogeography of Chinese and Australasian Polystichum ferns as inferred from chloroplast trnL-F and rps4-trnS sequence data

Polystichum, one of the largest genera of ferns, occurs worldwide with the greatest diversity in southwest China and adjacent regions. Although there have been studies of Chinese Polystichum on its traditional classification, geographic distributions, and even a few on its molecular systematics, its relationships to other species outside China remain little known. Here, we investigated the phylogeny and biogeography of the Polystichum species from China and Australasia. The evolutionary relationships among 42 Polystichum species found in China (29 taxa) and Australasia (13 taxa) were inferred from phylogenetic analyses of two chloroplast DNA sequence data sets: rps4-trnS and trnL-F intergenic spacers. The divergence time between Chinese and Australasian Polystichum was estimated. The results indicated that the Australasian species comprise a monophyletic group that is nested within the Chinese diversity, and that the New Zealand species are likewise a monophyletic group nested within the Australasian species. The divergence time estimates suggested that Chinese Polystichum migrated into Australasia from around 40 Ma ago, and from there to New Zealand from about 14 Ma. The diversification of the New Zealand Polystichum species began about 10 Ma. These results indicated that Polystichum probably originated in eastern Asia and migrated into Australasia: first into Australia and then into New Zealand.     

An evaluation of the vitamin D3 content in fish: Is the vitamin D content adequate to satisfy the dietary requirement for vitamin D?

It has been suggested that the major source of vitamin D should come from dietary sources and not sun exposure. However, the major fortified dietary source of vitamin D is milk which often does not contain at least 80% of what is stated on the label. Fish has been touted as an excellent source of vitamin D especially oily fish including salmon and mackerel. Little is known about the effect of various cooking conditions on the vitamin D content in fish. We initiated a study and evaluated the vitamin D content in several species of fish and also evaluated the effect of baking and frying on the vitamin D content. Surprisingly, farmed salmon had approximately 25% of the vitamin D content as wild salmon had. The vitamin D content in fish varied widely even within species. These data suggest that the tables that list the vitamin D content are out-of-date and need to be re-evaluated.     

Can mobile sandy land be vegetated in the cold and dry Tibetan Plateau in China?

Desertification has taken place in the overgrazed grassland of the Tibetan Plateau,China,and the area of mobile sandy land has increased in recent decades.The challenging problem about desertification control is how to restore the vegetation of mobile sandy lands caused by severe desertification.Sand drifting is now regarded as the limiting factor of vegetation restoration in such lands.The initial phase of vegetation restoration is plant colonization,but it is often aborted due to sand drifting,and then vegetation restoration fails to proceed.For the sake of revegetation,the first step is to stop sand drifting to ensure plant colonization.In the northeastern Tibetan Plateau,China,feasible approaches have been found through long-term experiments,and the vegetation is being restored satisfactorily with these approaches in experimental sandy lands.The approaches comprise three types:enclosure,mechanical barriers and biological barriers.Different sandy lands require dissimilar combinations of these approaches.Enclosures may be adequate to revegetate inter-dunes or degraded grassland even in cold regions like the Tibetan Plateau,China,but it is deficient for revegetation of the shifting sand dunes unless mechanical and biological barriers are established simultaneously.     

Assimilation of Endogenous Nicotinamide Riboside Is Essential for Calorie Restriction-mediated Life Span Extension in Saccharomyces cerevisiae

NAD⁺ (nicotinamide adenine dinucleotide) is an essential cofactor involved in various biological processes including calorie restriction-mediated life span extension. Administration of nicotinamide riboside (NmR) has been shown to ameliorate deficiencies related to aberrant NAD⁺ metabolism in both yeast and mammalian cells. However, the biological role of endogenous NmR remains unclear. Here we demonstrate that salvaging endogenous NmR is an integral part of NAD⁺ metabolism. A balanced NmR salvage cycle is essential for calorie restriction-induced life span extension and stress resistance in yeast. Our results also suggest that partitioning of the pyridine nucleotide flux between the classical salvage cycle and the NmR salvage branch might be modulated by the NAD⁺-dependent Sir2 deacetylase. Furthermore, two novel deamidation steps leading to nicotinic acid mononucleotide and nicotinic acid riboside production are also uncovered that further underscore the complexity and flexibility of NAD⁺ metabolism. In addition, utilization of extracellular nicotinamide mononucleotide requires prior conversion to NmR mediated by a periplasmic phosphatase Pho5. Conversion to NmR may thus represent a strategy for the transport and assimilation of large nonpermeable NAD⁺ precursors. Together, our studies provide a molecular basis for how NAD⁺ homeostasis factors confer metabolic flexibility.The pyridine nucleotide NAD⁺ and its reduced form NADH are primary redox carriers involved in metabolism. In addition to serving as a coenzyme in redox reactions, NAD⁺ also acts as a cosubstrate in protein modification reactions including deacetylation and ADP-ribosylation (1, 2). NAD⁺ also plays an important role in calorie restriction (CR)²-mediated life span extension via regulating NAD⁺-dependent longevity factors (3, 4). CR is the most effective regimen known to extend life span in various species (5, 6). CR also ameliorates many age-related diseases such as cancer and diabetes (5). The Sir2 family proteins are NAD⁺-dependent protein deacetylases, which have been shown to play important roles in several CR models in yeast (3, 7) and higher eukaryotes (8, 9). By coupling the cleavage of NAD⁺ and deacetylation of target proteins, the Sir2 family proteins serve as a molecular link relaying the cellular energy state to the machinery of life span regulation. Mammalian Sir2 family proteins (SIRT1–7) have also been implicated in stress response, cell survival, and insulin and fat metabolism (8–10), supporting a role for SIRT proteins in age-related metabolic diseases and perhaps human aging.In eukaryotes, NAD⁺ is generated by de novo synthesis and by salvaging various intermediary precursors (see Fig. 1A). In yeast, the de novo pathway is mediated by Bna1–5 and Qpt1 (Bna6), which produces nicotinic acid mononucleotide (NaMN) from tryptophan (11). Because the de novo pathway requires molecular oxygen as a substrate, cells grown under anaerobic growth conditions would rely on exogenous NAD⁺ precursors for the nicotinamide (Nam) moiety (11). Yeast cells also salvage Nam from NAD⁺ consuming reactions or nicotinic acid (NA) from environment via Tna1, Pnc1, and Npt1, leading to NaMN production. NaMN is then converted to NAD⁺ via Nma1/2 and Qns1 (see Fig. 1A). Nma1/2 are adenylyltransferases with dual specificity toward NMN and NaMN (12, 13), and Qns1 is a glutamine-dependent NAD⁺ synthetase. Recent studies also showed that supplementing nicotinamide riboside (NmR) and nicotinic acid riboside (NaR) to growth medium rescued the lethality of NAD⁺ auxotrophic mutants (14–16). Assimilations of exogenous NmR and NaR are mainly mediated by a conserved NmR kinase (Nrk1) and three nucleosidases (Urh1, Pnp1, and Meu1). Nrk1 phosphorylates NmR and NaR to produce nicotinamide mononucleotide (NMN) and NaMN, respectively (14, 16). Urh1, Pnp1, and Meu1 catabolize NmR and NaR to generate Nam and NA (15, 16).Open in a separate windowFIGURE 1.Nicotinamide riboside (NmR) is an endogenous metabolite in yeast. A, the current model of the NAD⁺ biosynthesis pathways. Extracellular NmR enters the salvage cycle through Nrk1, Urh1, Pnp1, and Meu1. B, NAD⁺ prototrophic cells release metabolites into growth medium to cross-feed NAD⁺ auxotrophic cells (the npt1Δqpt1Δ and qns1Δ mutants). Micro-colonies of the NAD⁺ auxotrophic mutants become visible after 2-day incubation at 30 °C, which show “gradient” growth patterns descending from the side adjacent to WT. C, Nrk1 is required for NAD⁺ auxotrophic cells to utilize NmR. Anaerobic growth conditions (−O₂) are utilized to block de novo NAD⁺ biosynthesis in the npt1Δ and npt1Δnrk1Δ mutants. D, Nrk1 is required to utilize cross-feeding metabolites. E, cross-feeding activity is modulated by factors in NmR metabolism. Cells defective in NmR utilization (left panel) or transport (middle panel) show increased cross-feeding in spot assays. Overexpressing Nrk1 decreases cross-feeding activity (right panel). The results show growth of the npt1Δqpt1Δ recipient (plated on YPD at a density of ∼9000 cells/cm²) supported by feeder cells (∼2 × 10⁴ cells spotted directly onto the recipient lawn). oe, overexpression.NmR supplementation has recently been shown to be a promising strategy for prevention and treatment of certain diseases (17). For example, NmR protected neurons from axonal degeneration via functioning as a NAD⁺ precursor (18, 19). Given that several NmR assimilating enzymes and NmR transporters have been characterized and many are conserved from fungi to mammals (14, 15, 20–22), NmR has been speculated to be an endogenous NAD⁺ precursor (17, 23). Here, we provided direct evidence for endogenous NmR as an integral part of NAD⁺ metabolism in yeast. We also determined the biological significance of salvaging endogenous NmR and studied its role in CR-induced life span extension. Moreover, we demonstrated that the NmR salvage machinery was also required for utilizing exogenous NMN, which has recently been shown to increase NAD⁺ levels in mammalian cells (24). Finally, we discussed the role of Sir2 in modulating the flux of pyridine nucleotides between alternate routes.     

Pattern of Expression and Substrate Specificity of Chloroplast Ferredoxins from Chlamydomonas reinhardtii

Ferredoxin (Fd) is the major iron-containing protein in photosynthetic organisms and is central to reductive metabolism in the chloroplast. The Chlamydomonas reinhardtii genome encodes six plant type [Fe₂S₂] ferredoxins, products of PETF, FDX2–FDX6. We performed the functional analysis of these ferredoxins by localizing Fd, Fdx2, Fdx3, and Fdx6 to the chloroplast by using isoform-specific antibodies and monitoring the pattern of gene expression by iron and copper nutrition, nitrogen source, and hydrogen peroxide stress. In addition, we also measured the midpoint redox potentials of Fd and Fdx2 and determined the kinetic parameters of their reactions with several ferredoxin-interacting proteins, namely nitrite reductase, Fd:NADP⁺ oxidoreductase, and Fd:thioredoxin reductase. We found that each of the FDX genes is differently regulated in response to changes in nutrient supply. Moreover, we show that Fdx2 (E_m = −321 mV), whose expression is regulated by nitrate, is a more efficient electron donor to nitrite reductase relative to Fd. Overall, the results suggest that each ferredoxin isoform has substrate specificity and that the presence of multiple ferredoxin isoforms allows for the allocation of reducing power to specific metabolic pathways in the chloroplast under various growth conditions.Ferredoxins are small (∼11,000-kDa), soluble, iron-sulfur cluster-containing proteins with strongly negative redox potentials (−350 to −450 mV) that function as electron donors at reductive steps in various metabolic pathways (1–3). In photosynthetic organisms, the well studied ferredoxin (Fd⁴; the product of the PETF gene) is the most abundant iron-containing protein in the chloroplast and is central to the distribution of photosynthetically derived reductive power (4).The most well known Fd-dependent reaction is the transfer of electrons from photosystem I (PSI) to NADPH, catalyzed by Fd:NADP⁺ oxidoreductase (FNR). The NADPH produced by this reaction donates electrons to the only reductant-requiring step in the Calvin cycle and other steps in anabolic pathways that require NADPH as reductant. In addition, reduced Fd directly donates electrons to other metabolic pathways by interacting with various enzymes in the chloroplast. This includes Fd:thioredoxin reductase (FTR), which converts a light-driven electron signal into a thiol signal that is transmitted to thioredoxins (TRXs) present in the plastid as different types (or different isoforms). Once reduced, TRXs interact with specific disulfide bonds on target enzymes, modulating their activities (5). Other Fd targets include hydrogenase, which is responsible for hydrogen production in anaerobic conditions in green algae; glutamine-oxoglutarate amidotransferase in amino acid synthesis; nitrite and sulfite reductases in nitrate and sulfate assimilation, respectively; stearoyl-ACP Δ9-desaturase in fatty acid desaturation; and phycocyanobilin:Fd oxidoreductase in synthesis of phytochromobilin (6). Fd also functions in non-photosynthetic cells. Here, FNR catalyzes the reduction of Fd by NADPH produced in the oxidative pentose phosphate pathway, enabling Fd-dependent metabolism to occur in the dark (7, 8).The single-celled green alga, Chlamydomonas reinhardtii is an excellent reference organism for studying both metabolic adaptation to nutrient stress and photosynthesis (9–13). The Chlamydomonas genome encodes six highly related plant type ferredoxin genes (9). Until recently, only the major photosynthetic ferredoxin, Fd (encoded by PETF), which mediates electron transfer between PSI and FNR, had been characterized in detail (14).Many land plants are known to have multiple ferredoxins. Typically, they are differently localized on the basis of their function. Photosynthetic ferredoxins reduce NADP⁺ at a faster rate and are localized to the leaves, whereas non-photosynthetic ferredoxins are more efficiently reduced by NADPH and are localized to the roots. Arabidopsis thaliana has a total of six ferredoxin isoforms (15). Of these, two are photosynthetic and localized in the leaves. The most abundant, AtFd2, is involved in linear electron flow, and the less abundant (5% of the ferredoxin pool), AtFd1, has been implicated in cyclic electron flow (16). There is one non-photosynthetic ferredoxin located in the roots, AtFd3, which is nitrate-inducible. This protein has higher electron transfer activity with sulfite reductase in in vitro assays compared with other Arabidopsis ferredoxin isoforms, suggesting in vivo function of AtFd3 in nitrate and sulfate assimilation (15, 17). In addition, there is one evolutionarily distant ferredoxin, AtFd4, of unknown function with a more positive redox potential present in the leaves and two other proteins which are “ferredoxin-like” and uncharacterized (15). Zea mays has four ferredoxin isoforms, two photosynthetic and two non-photosynthetic (18). One of the non-photosynthetic isoforms is specifically induced by nitrite, suggestive of a role in nitrate metabolism (19). A cyanobacterium, Anabaena 7120, has two ferredoxins, vegetative and heterocyst type (by analogy to leaf and root types, respectively). The heterocyst type is present only in cells that have differentiated into nitrogen-fixing cells, indicating that this form may serve to transfer electrons to nitrogenase (20).We hypothesize that the presence of as many as six ferredoxin isoforms in a single-celled organism like C. reinhardtii allows for the differential regulation of each isoform and therefore the prioritization of reducing power toward certain metabolic pathways under changing environmental conditions. To test this hypothesis, expression of the genes (PETF and FDX2–FDX6) encoding the six ferredoxin isoforms in Chlamydomonas reinhardtii was monitored under various conditions in which well characterized ferredoxin-dependent enzymes are known to be expressed. In addition, we also analyzed the interaction of Fd and Fdx2 with several ferredoxin-interacting proteins, such as NiR, FNR, and FTR, and determined the kinetic parameters of the corresponding reactions.We found that each of the FDX genes is indeed differently regulated in response to changes in nutrient supply. In the case of FDX2 whose product is most similar to classical Fd, we suggest that it has specificity for nitrite reductase based on its pattern of expression and activity with nitrite reductase.     

GSK-3 Phosphorylates ��-Catenin and Negatively Regulates Its Stability via Ubiquitination/Proteosome-mediated Proteolysis

δ-Catenin was first identified because of its interaction with presenilin-1, and its aberrant expression has been reported in various human tumors and in patients with Cri-du-Chat syndrome, a form of mental retardation. However, the mechanism whereby δ-catenin is regulated in cells has not been fully elucidated. We investigated the possibility that glycogen-synthase kinase-3 (GSK-3) phosphorylates δ-catenin and thus affects its stability. Initially, we found that the level of δ-catenin was greater and the half-life of δ-catenin was longer in GSK-3β^−/− fibroblasts than those in GSK-3β^+/+ fibroblasts. Furthermore, four different approaches designed to specifically inhibit GSK-3 activity, i.e. GSK-3-specific chemical inhibitors, Wnt-3a conditioned media, small interfering RNAs, and GSK-3α and -3β kinase dead constructs, consistently showed that the levels of endogenous δ-catenin in CWR22Rv-1 prostate carcinoma cells and primary cortical neurons were increased by inhibiting GSK-3 activity. In addition, it was found that both GSK-3α and -3β interact with and phosphorylate δ-catenin. The phosphorylation of ΔC207-δ-catenin (lacking 207 C-terminal residues) and T1078A δ-catenin by GSK-3 was noticeably reduced compared with that of wild type δ-catenin, and the data from liquid chromatography-tandem mass spectrometry analyses suggest that the Thr¹⁰⁷⁸ residue of δ-catenin is one of the GSK-3 phosphorylation sites. Treatment with MG132 or ALLN, specific inhibitors of proteosome-dependent proteolysis, increased δ-catenin levels and caused an accumulation of ubiquitinated δ-catenin. It was also found that GSK-3 triggers the ubiquitination of δ-catenin. These results suggest that GSK-3 interacts with and phosphorylates δ-catenin and thereby negatively affects its stability by enabling its ubiquitination/proteosome-mediated proteolysis.δ-Catenin was first identified as a molecule that interacts with presenilin-1 (PS-1)² by yeast two-hybrid assay (1) and was found to belong to the p120-catenin subfamily of armadillo proteins, which characteristically contain 10 Arm repeats (2). In addition to its interaction with PS-1 and its abundant expression in brain (3, 4), several lines of evidence indicate that δ-catenin may play a pivotal role in cognitive function. First, the hemizygous loss of δ-catenin is known to be closely correlated with Cri-du-Chat syndrome, a severe form of mental retardation in humans (5). Second, severe learning deficits and abnormal synaptic plasticity were found in δ-catenin-deficient mice (6). Moreover, in δ-catenin^−/− mice, paired pulse facilitation (a form of short term plasticity) was found to be reduced, and long term potentiation, which is related to the forming and storage mechanisms of memory, was deficient (7, 8). Third, δ-catenin interacting molecules, such as PSs (1, 9), cadherins (10), S-SCAM (2), and PSD-95 (11), have been shown to play important roles in modulating synaptic plasticity. However, even though the maintenance of an adequate δ-catenin level is known to be critical for normal brain function, few studies have been undertaken to identify the factors that regulate δ-catenin stability in cells. We have previously demonstrated that PS-1 inhibits δ-catenin-induced cellular branching and promotes δ-catenin processing and turnover (12).Because of structural similarities among β-catenin, p120-catenin, and δ-catenin and to their shared binding partners (i.e. PS-1 (1, 9) and cadherins (10)), glycogen-synthase kinase-3 (GSK-3) drew our attention as a potential candidate effector of δ-catenin stability in cells. GSK-3 is a serine/threonine kinase and has two highly homologous forms, GSK-3α and GSK-3β, in mammals (13). Although GSK-3α and GSK-3β have similar structures, they differ in mass (GSK-3α (51 kDa) and GSK-3β (47 kDa) (13)) and to some extent in function (14). GSK-3 is a well established inhibitor of Wnt signaling. Moreover, it is known to phosphorylate β-catenin, which results in its degradation via ubiquitination/proteosome-dependent proteolysis (15). GSK-3 is ubiquitously distributed in the human body, but it is particularly abundant in brain (13), and it is interesting that δ-catenin is also abundant in the nervous system (4) and that GSK-3 participates in the progression of Alzheimer disease (16). The majority of GSK-3 substrates have the consensus sequence (Ser/Thr)-Xaa-Xaa-Xaa-(Ser/Thr) (17). Interestingly, we found that δ-catenin has several putative phosphorylation sites targeted by GSK-3, which suggests that δ-catenin can be regulated by GSK-3 in the same way as β-catenin.In this report, we demonstrate that both GSK-3α and -3β interact with and phosphorylate δ-catenin and that this leads to its subsequent ubiquitination and degradation via proteosome-dependent proteolysis. Our results strongly suggest that GSK-3 is a key regulator of δ-catenin stability in cells.     

Mus和Rattus属鼠类下颌骨形态特征测量分析系统建立

目的建立高精度低误差的《Mus和Rattus属鼠类下颌骨形态特征测量分析系统》。方法运用Microsoft Visual Basic6．0语言设计程序,自动测量、计算、分析下颌骨形态特征。实例运行评估其可操作性。结果本系统可提高测量精度28倍,减少计算误差40余倍。可适用于Mus和Rattus属的野生动物、实验动物的下颌骨形态特征测量分析。结论《Mm和Rottus属鼠类下颌骨形态特征测量分析系统》具有成为可操作的遗传检测方法。     

Curcumin-Induced DNA Damage and Inhibited DNA Repair Genes Expressions in Mouse–Rat Hybrid Retina Ganglion Cells (N18)

Curcumin is reported to be a potent inhibitor of the initiation and promotion of many cancer cells. We investigated to examine whether or not curcumin induce DNA damage in mouse–rat hybrid retina ganglion cell line N18 cells. The Comet assay showed that incubation of N18 cells with 10, 25 and 30 μM of curcumin led to a longer DNA migration smear (Comet tail). The DNA gel electrophoresis showed that 20 μM of curcumin for 24 and 48 h treatment induced DNA damage and fragments in N18 cells. The real time PCR analysis showed that 20 μM of curcumin for 48 h treatment decreased ATM, ATR, BRCA1, 14-3-3σ, DNA-PK and MGMT mRNA, and ATM and MGMT mRNA expression were inhibited in a time-dependent manner. Our results indicate that curcumin caused DNA damage and inhibited DNA repair genes which may be the factors for curcumin-inhibited cell growth. H.-F. Lu and J.-S. Yang are contributed equally to this study.