植物抗冷分子生物学研究进展（综述）

温度是植物生长的必要条件,然而低温却是限制作物生产的重要因素,为此,各国政府及研究部门一直都把植物低温适应性问题作为一个重要的研究课题．按照低温的不同程度,植物的低温伤害可分为冷害（ｃｈｉｌｌｉｎｇ ｉｎｊｕｒｙ;零上低温对植物的伤害）和冻害（ｆｒｅｅｚｉｎｇ ｉｎｊｕｒｙ;零下低温对植物的伤害）两大类。早期关于植物冷害机理和抗冷机理的研究,是从比较冷敏感（ｃｈｉｌｌｉｎｇ－ｓｅｎｓｉｔｉｖｅ）植物和抗冷（ｃｈｉｌｌｉｎｇ－ｉｎｓｅｎｓｉｔｉｖｅ）植物或比较未经冷驯化的植物（ｎｏｎ－ｃｏｌｄ－ａｃ…     

Pregnancy induced hypertension: a role for peroxidation in microvillus plasma membranes

It has been recently hypothesized that in PIH a placental oxidant-antioxidant imbalance might cause the release of lipoperoxidation products into the circulation, with subsequent damage of endothelial cell membranes. In this hypothesis the endothelial cell and further increase in circulating lipoperoxide levels, which are by themselves able to induce smooth muscle constriction and increased pressor responsiveness to angiotensin II. In order to investigate this issue, we studied the basal content of lipid peroxides in terms of malondialdehyde (MDA) in the syncytiotrophoblast plasma membranes (SPM) from PIH women. Moreover, we investigated the susceptibility to peroxidation of SPM using anin vitro oxidative stress as a tool to verify the predisposition to thein vivo development of peroxidation products. The fatty acid composition of the membranes was also analyzed. Microvillus membrane lipoperoxide concentrations were significantly increased in PIH women (62.8±7.6 ng MDA/mg prot) compared with healthy pregnant subjects (37.6±4.8 ng MDA/mg prot; p<0.01).The formation of TBARS under the action of phenylhydrazine was significantly greater in PIH women (90.3±7.4 mmol MDA/mol cholesterol) than in normal pregnant subjects (68.6±6.4 mmol MDA/mol cholesterol; p<0.01). In PIH microvillus membrane we also observed a significant increase of the content of polyunsaturated arachidonic acid.The increased susceptibility to oxidative stress of SPMs from PIH women might be due either to reduced antioxidant systems or to an abnormality of the lipid composition of the membrane. The present work also demonstrated in PIH a reduction in the SPM content of saturated fatty acids with an increase in polyunsaturated fatty acids, which are the major substrate for peroxidation. On the other hand, the higher lipoperoxidation may be due to the observed increased susceptibility to peroxidative stress, to a primary reduction in placental perfusion with tissue hypoxia or to both factors, which can potentiate each other.     

Diverse effects of essential (n−6 andn−3) fatty acids on cultured cells

Fatty acids (FAs) have long been recognized for their nutritional value in the absence of glucose, and as necessary components of cell membranes. However, FAs have other effects on cells that may be less familiar. Polyunsaturated FAs of dietary origin (n–6 andn–3) cannot be synthesized by mammals, and are termed essential because they are required for the optimal biologic function of specialized cells and tissues. However, they do not appear to be necessary for normal growth and metabolism of a variety of cells in culture. The essential fatty acids (EFAs) have received increased attention in recent years due to their presumed involvement in cardiovascular disorders and in cancers of the breast, pancreas, colon and prostate. Manyin vitro systems have emerged which either examine the role of EFAs in human disease directly, or utilize EFAs to mimic thein vivo cellular environment. The effects of EFAs on cells are both direct and indirect. As components of membrane phospholipids, and due to their varying structural and physical properties, EFAs can alter membrane fluidity, at least in the local environment, and affect any process that is mediated via the membrane. EFAs containing 20 carbons and at least three double bonds can be enzymatically converted to eicosanoid hormones, which play important roles in a variety of physiological and pathological processes. Alternatively, EFAs released into cells from phospholipids can act as second messengers that activate protein kinase C. Furthermore, susceptibility to oxidative damage increases with the degree of unsaturation, a complication that merits consideration because lipid peroxidation can lead to a variety of substances with toxic and mutagenic properties. The effects of EFAs on cultured cells are illustrated using the responses of normal and tumor human mammary epithelial cells. A thorough evaluation of EFA effects on commercially important cells could be used to advantage in the biotechnology industry by identifying EFA supplements that lead to improved cell growth and/or productivity.Abbreviations AA arachidonic acid (20 carbons: 4 double bonds,n–6) - BHA butylated hydroxyanisole - BHT butylated hydroxytoluene - cAMP cyclic adenosine monophosphate - CHO Chinese hamster ovary - DAG diacylglycerol - DGLNA dihomo--linolenic acid (203,n–6) - DHA docosahexaenoic acid (226,n–3) - EFA essential fatty acid - EGF epidermal growth factor - EGFR epidermal growth factor receptor - EPA eicosapentaenoic acid (205,n–3) - FA fatty acid - FBS fetal bovine serum - GLNA -linolenic acid (183,n–6) - LA linoleic acid (182,n–6) - LNA -linolenic acid (183,n–3) - LT leukotriene - MDA malondialdehyde - NAD nicotinamide adenine dinucleotide - NDGA nordihydroguaiaretic acid - OA oleic acid (181,n–9) - PG prostaglandin - PKC protein kinase C - PUFA polyunsaturated fatty acid - SFM serum-free medium - TX thromboxane     

耐低温酵母Curvibasidium rogersii菌株在松萝样品中的首次分离鉴定及基于基因组分析的生物特性探究

【目的】对西藏松萝地衣来源的两株非常规酵母进行分离鉴定,并通过基因组序列分析探究其生物学特性和应用潜力。【方法】从西藏来源的松萝地衣样品内部分离得到2株耐低温酵母菌株,通过26S D1/D2和ITS序列比对分析以及生理生化实验进行菌种鉴定;通过全基因组序列分析和验证探究菌株的生物特性。【结果】两株酵母菌株经鉴定均为Curvibasidium rogersii,可以在10℃低温良好生长,在20℃生长最佳,25℃及以上温度生长缓慢或不生长。对其进行基因组测序和基因组挖掘,测序结果发现,其基因组注释出功能的部分与产油脂的低温酵母白冬孢酵母Leucosporidium creatinivorum具有最高相似性,尼罗红染色发现两株酵母都能够生产油脂,另外在基因组序列中还发现了可能参与木糖代谢的相关蛋白编码基因,实验证明两个酵母菌株可以利用木糖生长。【结论】首次分离鉴定了来自西藏松萝的酵母C. rogersii,为充分开发利用松萝和其他地衣来源微生物,以及利用可代谢木糖的新资源酵母生产微生物油脂提供了基础。     

Cell-penetrating peptide-conjugated lipid nanoparticles for siRNA delivery

Lipid nanoparticles (LNP) modified with cell-penetrating peptides (CPP) were prepared for the delivery of small interfering RNA (siRNA) into cells. Lipid derivatives of CPP derived from protamine were newly synthesized and used to prepare CPP-decorated LNP (CPP-LNP). Encapsulation of siRNA into CPP-LNP improved the stability of the siRNA in serum. Fluorescence-labeled siRNA formulated in CPP-LNP was efficiently internalized into B16F10 murine melanoma cells in a time-dependent manner, although that in LNP without CPP was hardly internalized into these cells. In cells transfected with siRNA in CPP-LNP, most of the siRNA was distributed in the cytoplasm of these cells and did not localize in the lysosomes. Analysis of the endocytotic pathway indicated that CPP-LNP were mainly internalized via macropinocytosis and heparan sulfate-mediated endocytosis. CPP-LNP encapsulating siRNA effectively induced RNA interference-mediated silencing of reporter genes in B16F10 cells expressing luciferase and in HT1080 human fibrosarcoma cells expressing enhanced green fluorescent protein. These data suggest that modification of LNP with the protamine-derived CPP was effective to facilitate internalization of siRNA in the cytoplasm and thereby to enhance gene silencing.     

A cyp19a1b‐gfp (aromatase B) transgenic zebrafish line that expresses GFP in radial glial cells

Aromatase is an enzyme that catalyzes the synthesis of estrogen in gonads and brain. Teleost fish express aromatase (AroB) strongly in the brain facilitating its detailed examination. To understand the function of AroB in the brain, we generated transgenic zebrafish that expresses green fluorescent protein (GFP) driven by the brain aromatase cyp19a1b promoter. GFP was found in the radial glial cells of transgenic larvae and adult fish that overlap with AroB immunoreactivity in the correct temporal and spatial pattern. GFP was also coexpressed with radial cell marker BLBP, but was not in neurons. In addition, GFP expression in the radial glial cells was stimulated by estrogen, same as endogenous AroB expression. Thus, this transgenic line faithfully mimics the regulation of AroB expression in radial glial cells. It provides a powerful tool to further characterize progenitor radial cells in adult and developing fish and to evaluate estrogenic activities of xenoestrogens and phytoestrogens. genesis 47:67–73, 2009. © 2008 Wiley‐Liss, Inc.     

Circulating tumor microemboli: Progress in molecular understanding and enrichment technologies

Circulating tumor cells (CTCs) and their clusters, also known as circulating tumor microemboli (CTM), have emerged as valuable tool that can provide mechanistic insights into the tumor heterogeneity, clonal evolution, and stochastic events within the metastatic cascade. However, recent investigations have hinted that CTM may not be mere aggregates of tumor cells but cells comprising CTM exhibit distinct phenotypic and molecular characteristics in comparison to single CTCs. Moreover, in many cases CTM demonstrated higher metastatic potential and resistance to apoptosis as compared to their single cell counterparts. Thus, their evaluation and enumeration may provide a new dimension to our understanding of cancer biology and metastatic cancer spread as well as offer novel theranostic biomarkers. Most of the existing technologies for isolation of hematogenous tumor cells largely favor single CTCs, hence there is a need to devise new approaches, or re-configure the existing ones, for specific and efficient CTM isolation. Here we review existing knowledge and insights on CTM biology. Furthermore, a critical commentary on current and emerging trends in CTM enrichment and characterization along with recently developed ex-vivo CTC expansion methodologies is presented with the aim to facilitate researchers to identify further avenues of research and development.     

Achieving Molecular Selectivity in Imaging Using Multiphoton Raman Spectroscopy Techniques

In the case of most optical imaging methods, contrast is generated either by physical properties of the sample (Differential Image Contrast, Phase Contrast), or by fluorescent labels that are localized to a particular protein or organelle. Standard Raman and infrared methods for obtaining images are based upon the intrinsic vibrational properties of molecules, and thus obviate the need for attached fluorophores. Unfortunately, they have significant limitations for live-cell imaging. However, an active Raman method, called Coherent Anti-Stokes Raman Scattering (CARS), is well suited for microscopy, and provides a new means for imaging specific molecules. Vibrational imaging techniques, such as CARS, avoid problems associated with photobleaching and photo-induced toxicity often associated with the use of fluorescent labels with live cells. Because the laser configuration needed to implement CARS technology is similar to that used in other multiphoton microscopy methods, such as two-photon fluorescence and harmonic generation, it is possible to combine imaging modalities, thus generating simultaneous CARS and fluorescence images. A particularly powerful aspect of CARS microscopy is its ability to selectively image deuterated compounds, thus allowing the visualization of molecules, such as lipids, that are chemically indistinguishable from the native species.     

Effect of dietary fat or starch supply during gestation and/or lactation on the performance of sows, piglets' survival and on the performance of progeny after weaning

Two trials were carried out to compare the effects of fat or starch inclusion in sow's diet on sow and litter performance. In each trial, sows were assigned to one of two treatments. In trial 1, the sows were fed diets containing either soybean oil (5%, treatment GL5) or cornstarch (11.3%, GL0) from day 35 of gestation to weaning. Daily net energy and nutrient allowance were equalised during gestation. In trial 2, the same treatments were applied only after farrowing (treatments L5 and L0, respectively). Within each trial, a batch of piglets was studied until slaughter. In trial 1, adipose cell development and total lipid content were determined on some pigs at weaning (n = 6/treatment) and at slaughter in dorsal subcutaneous adipose tissue (n = 13/group at least) and in muscle (n = 46/group at least). Piglets' birth weight was not affected by treatment in trial 1. Survival rates at birth and after 24 h of life were higher in treatment GL5 (4.0% v. 7.5% stillborn piglets in GL0 treatment, P < 0.05; 8.7% v. 12.6% of piglets alive at 24 h of age died in treatment GL0, P = 0.06). Subsequently, overall survival rate until weaning was higher in treatment GL5 (81.4% v. 75.7% of total born piglets, P = 0.03), but litter size at weaning was not significantly affected (11.3). Litter growth rate before weaning was increased when a fat-enriched diet was provided during gestation and lactation (+140 g/day per litter; P < 0.01) and to a lower extent when provided only after farrowing (+90 g/day; P < 0.05). Energy supply through fat did not decrease the mobilisation of the sow's body reserve and backfat thickness loss was even higher with treatment GL5 (P < 0.05). After weaning, pigs' average daily gain, feed : gain ratio and carcass lean content were not affected by the energy source supplied before and/or after farrowing. At weaning, the number of adipose cells in the dorsal subcutaneous adipose tissue and in the Longissimus dorsi muscle was higher in the GL5 pigs. Muscle lipid content at weaning did not differ between treatments, but it was higher at slaughter, around 110 kg, in the GL5 pigs (3.46% v. 2.58%, P < 0.001).     

Opposing roles of cell death-inducing DFF45-like effector B and perilipin 2 in controlling hepatic VLDL lipidation

Regulation of hepatic very low density lipoprotein (VLDL) assembly and maturation is crucial in controlling lipid homeostasis and in the development of metabolic disorders, including obesity, hepatic steatosis, and insulin resistance. Cideb, a member of cell death-inducing DFF45-like effector (CIDE) protein family, has been previously shown to promote VLDL lipidation and maturation. However, the precise subcellular location of Cideb-mediated VLDL lipidation and the factors modulating its activity remain elusive. In addition to its localization to endoplasmic reticulum (ER) and lipid droplets (LD), we observed that Cideb was also localized to the Golgi apparatus. Mature and lipid-rich VLDL particles did not accumulate in the Golgi apparatus in Cideb(-/-) livers. Interestingly, we observed that hepatic perilipin 2/adipose differentiation-related protein (ADRP) levels were markedly increased in Cideb(-/-) mice. Liver-specific knockdown of perilipin 2 in Cideb(-/-) mice resulted in the reduced accumulation of hepatic triglycerides (TAG), increased VLDL-TAG secretion, and the accumulation of mature TAG-rich VLDL in the Golgi apparatus. These data reveal that Cideb and perilipin 2 play opposing roles in controlling VLDL lipidation and hepatic lipid homeostasis.