Gene expression changes in male accessory glands during ageing are accompanied by reproductive decline in Drosophila melanogaster

Senescence is accompanied by loss of reproductive functions. Here, we studied reproductive ageing in Drosophila melanogaster males and asked whether the expected decline in male reproductive success is due to diminished functionality of the male accessory gland (AG). The male AG produces the majority of seminal fluid proteins (SFPs) transferred to the female at mating. SFPs induce female postmating changes and are key to male reproductive success. We measured age‐dependent gene expression changes for five representative SFP genes in males from four different age groups ranging from 1 to 6 weeks after eclosion. Simultaneously, we also measured male reproductive success in postmating traits mediated by transfer of these five SFPs. We found a decreased in male SFP gene expression with advancing age and an accompanying decline in male postmating success. Hence, male reproductive senescence is associated with a decline in functionality of the male AG. While overall individual SFP genes decreased in expression, our results point towards the idea that the composition of an ejaculate might change with male age as the rate of change was variable for those five genes.     

An improved algorithm for the detection of genomic variation using short oligonucleotide expression microarrays

High‐throughput microarray experiments often generate far more biological information than is required to test the experimental hypotheses. Many microarray analyses are considered finished after differential expression and additional analyses are typically not performed, leaving untapped biological information left undiscovered. This is especially true if the microarray experiment is from an ecological study of multiple populations. Comparisons across populations may also contain important genomic polymorphisms, and a subset of these polymorphisms may be identified with microarrays using techniques for the detection of single feature polymorphisms (SFP). SFPs are differences in microarray probe level intensities caused by genetic polymorphisms such as single‐nucleotide polymorphisms and small insertions/deletions and not expression differences. In this study, we provide a new algorithm for the detection of SFPs, evaluate the algorithm using existing data from two publicly available Affymetrix Barley (Hordeum vulgare) microarray data sets and compare them to two previously published SFP detection algorithms. Results show that our algorithm provides more consistent and sensitive calling of SFPs with a lower false discovery rate. Simultaneous analysis of SFPs and differential expression is a low‐cost method for the enhanced analysis of microarray data, enabling additional biological inferences to be made.     

A review of basin morphology and pool hydrology of isolated ponded wetlands: implications for seasonal forest pools of the northeastern United States

Seasonal forest pools (SFPs) are geographically- and hydrologically-isolated ponded wetlands, in that they are topographically isolated from other surface waters. SFPs occur commonly throughout the temperate forests of the eastern United States and adjacent Canada. SFPs are ephemeral in occurrence, typically drying annually. The regular drying of SFPs excludes fish from these habitats, and as a result, they are the preferred breeding habitat of some amphibians, notably ambystomid (‘mole’) salamanders and wood frogs (Rana sylvatica Le Conte). The pools also support a rich and diverse invertebrate fauna. The duration of the wet phase, or hydroperiod of SFPs, has been repeatedly shown to be the dominant influence on the composition and fitness of the faunal community of the pools. Despite the importance of SFP hydrology, it is a poorly studied subject. This paper reviews the limited state-of-knowledge of seasonal forest pool hydrology and associated basin morphology. The review discusses findings from studies of other isolated ponded wetlands that could be applicable to our understanding of the hydrology of SFPs.     

Hypoglycemic and hypolipidemic effects and antioxidant activity of fruit extracts from Lycium barbarum

The hypoglycemic and hypolipidemic effects of Lycium barbarum fruit water decoction, crude polysaccharide extracts (crude LBP), and purified polysaccharide fractions (LBP-X) in alloxan-induced diabetic or hyperlipidemic rabbits were investigated through designed sequential trials and by measuring blood glucose and serum lipid parameters. Total antioxidant capacity was also assessed using trolox equivalent antioxidant capacity (TEAC) and oxygen radical absorbance capacity (ORAC) assay. It was found that the three Lycium barbarum fruit extracts/fractions could significantly reduce blood glucose levels and serum total cholesterol (TC) and triglyceride (TG) concentrations and at same time markedly increase high density lipoprotein cholesterol (HDL-c) levels after 10 days treatment in tested rabbits, indicating that there were substantial hypoglycemic and hypolipidemic effects. Hypoglycemic effect of LBP-X was more significant than those of water decoction and crude LBP, but its hypolipidemic effect seemed to be weaker. Total antioxidant capacity assay showed that all three Lycium barbarum extracts/fractions possessed antioxidant activity. However, water and methanolc fruit extracts and crude polysaccharide extracts exhibited stronger antioxidant activity than purified polysaccharide fractions because crude extracts were identified to be rich in antioxidants (e.g., carotenoids, riboflavin, ascorbic acid, thiamine, nicotinic acid). Lycium barbarum polysaccharides (glycocojugates), containing several monosaccharides and 17 amino acids, were major bioactive constituents of hypoglycemic effect. Both polysaccharides and vitamin antioxidants from Lycium barbarum fruits were possible active principles of hypolipidemic effect.     

Hypolipidemic Effect of Fenugreek Seeds Is Mediated Through Inhibition of Fat Accumulation and Upregulation of LDL Receptor

Fenugreek (Trigonella foenum‐graecum) seeds, used as a condiment, are documented for health benefits including amelioration of abnormalities in lipid homeostasis due to its hypolipidemic properties. However, molecular mechanisms underlying the hypolipidemic effect of fenugreek seeds remain obscure. In this study, hypolipidemic effect of a novel thermostable extract of fenugreek seeds (TEFS) was evaluated in vitro by employing differentiating and differentiated 3T3‐L1 cells, and HepG2 cells cultured in normal or sterol‐enriched conditions. Hypolipidemic effect was studied by quantifying decrease in accumulation of fat or by western blot analysis of adipogenic and lipogenic factors. At molecular level, TEFS inhibited accumulation of fat in differentiating and differentiated 3T3‐L1 cells via decreased expression of adipogenic factors such as peroxisome proliferators activated‐receptor‐γ (PPAR‐γ), sterol regulatory element‐binding protein‐1 (SREBP‐1), and CAAT element‐binding proteins‐α (c/EBP‐α). We also show that following TEFS treatment, cellular triglycerides (TGs), and cholesterol concentrations decreased significantly (P < 0.05) in HepG2 cells via reduced expression of SREBP‐1, at mRNA as well as protein level. Under sterol enriched condition, TEFS upregulated low‐density lipoprotein receptor (LDLR) expression resulting in enhanced LDL uptake. Treating fat supplement fed C57BL6/J mice with TEFS for 15 days resulted in decrease of serum TG, LDL‐cholesterol (LDLc), and body weight in a dose‐ and time‐dependent manner (P < 0.05). Results indicate that hypolipidemic effect of TEFS is due to inhibition of fat accumulation and upregulation of LDLR. Taken together, the study suggests that TEFS may have potential application in the management of dyslipidemia and its associated metabolic disorders.     

Protective effects of cynaroside on oxidative stress in retinal pigment epithelial cells

Cynaroside is a flavonoid compound proved to possess antioxidant activity, but its protective effect on age‐related macular degeneration still remains unclear. In this study, the protective effects of cynaroside on oxidative stress and apoptosis in retinal pigment epithelial (RPE) cells induced by hydrogen peroxide (H₂O₂) were investigated. Results showed that cynaroside effectively attenuated the decrease of cell activity induced by H₂O₂. The total reactive oxygen species can be remitted by decreasing malondialdehyde level, as well as increasing glutathione level, and superoxide dismutase and catalase activities. In addition, Western blot analysis indicated that cynaroside protected ARPE‐19 cells from apoptosis through downregulation of caspase‐3 protein activation which was controlled by the upstream proteins Bcl‐2 and Bax. It was finally proved that cynaroside could enhance the antioxidant and antiapoptotic ability in ARPE‐19 cells by promoting the expression of p‐Akt.     

Brain mitochondria from rats treated with sulforaphane are resistant to redox-regulated permeability transition

Oxidative stress promotes Ca²⁺-dependent opening of the mitochondrial inner membrane permeability transition pore (PTP), causing bioenergetic failure and subsequent cell death in many paradigms, including those related to acute brain injury. One approach to pre-conditioning against oxidative stress is pharmacologic activation of the Nrf2/ARE pathway of antioxidant gene expression by agents such as sulforaphane (SFP). This study tested the hypothesis that administration of SFP to normal rats increases resistance of isolated brain mitochondria to redox-sensitive PTP opening. SFP or DMSO vehicle was administered intraperitoneally to adult male rats at 10 mg/kg 40 h prior to isolation of non-synaptic brain mitochondria. Mitochondria were suspended in medium containing a respiratory substrate and were exposed to an addition of Ca²⁺ below the threshold for PTP opening. Subsequent addition of tert-butyl hydroperoxide (tBOOH) resulted in a cyclosporin A-inhibitable release of accumulated Ca²⁺ into the medium, as monitored by an increase in fluorescence of Calcium Green 5N within the medium, and was preceded by a decrease in the autofluorescence of mitochondrial NAD(P)H. SFP treatment significantly reduced the rate of tBOOH-induced Ca²⁺ release but did not affect NAD(P)H oxidation or inhibit PTP opening induced by the addition of phenylarsine oxide, a direct sulfhydryl oxidizing agent. SFP treatment had no effect on respiration by brain mitochondria and had no effect on PTP opening or respiration when added directly to isolated mitochondria. We conclude that SFP confers resistance of brain mitochondria to redox-regulated PTP opening, which could contribute to neuroprotection observed with SFP.     

The Drosophila melanogaster seminal fluid protease "seminase" regulates proteolytic and post-mating reproductive processes

Proteases and protease inhibitors have been identified in the ejaculates of animal taxa ranging from invertebrates to mammals and form a major protein class among Drosophila melanogaster seminal fluid proteins (SFPs). Other than a single protease cascade in mammals that regulates seminal clot liquefaction, no proteolytic cascades (i.e. pathways with at least two proteases acting in sequence) have been identified in seminal fluids. In Drosophila, SFPs are transferred to females during mating and, together with sperm, are necessary for the many post-mating responses elicited in females. Though several SFPs are proteolytically cleaved either during or after mating, virtually nothing is known about the proteases involved in these cleavage events or the physiological consequences of proteolytic activity in the seminal fluid on the female. Here, we present evidence that a protease cascade acts in the seminal fluid of Drosophila during and after mating. Using RNAi to knock down expression of the SFP CG10586, a predicted serine protease, we show that it acts upstream of the SFP CG11864, a predicted astacin protease, to process SFPs involved in ovulation and sperm entry into storage. We also show that knockdown of CG10586 leads to lower levels of egg laying, higher rates of sexual receptivity to subsequent males, and abnormal sperm usage patterns, processes that are independent of CG11864. The long-term phenotypes of females mated to CG10586 knockdown males are similar to those of females that fail to store sex peptide, an important elicitor of long-term post-mating responses, and indicate a role for CG10586 in regulating sex peptide. These results point to an important role for proteolysis among insect SFPs and suggest that protease cascades may be a mechanism for precise temporal regulation of multiple post-mating responses in females.     

Antidyslipidemic action of fenofibrate in dyslipidemic-diabetic hamster model

Fenofibrate is the ligand for PPARalpha subtype that mediates the action of its agonists' in lipid metabolism. How fibrate exerts hypolipidemic effect? The mechanism is studied in a newly developed high-fat fructose enriched diet induced dyslipidemia-diabetic hamster model. Fenofibrate lowered the basal plasma lipids like TC, TG, PL, FFA, glycerol, VLDL, and LDL, but HDL was increased. The activity of lipoprotein lipase in liver, adipose tissue, and small intestine was upregulated. However, that of triglyceride lipase was downregulated in liver. It has also improved the insulin secretion and plasma glucose lowering, caused by impairment in insulin secretion due to high-fat load. The drug was found effective in reducing body weight and diet due to rise in leptin level. Fenofibrate also enhanced the fecal excretion of total lipids, cholic acid, and deoxycholic acid probably by the activation of 7alpha cholesterol hydroxylase enzyme. Thus, causing broad-spectrum lipid lowering along with inhibition of hepatic lipid biosynthesis and maintaining lipid-glucose homeostasis.     

Duration and dose-dependency of female sexual receptivity responses to seminal fluid proteins in Aedes albopictus and Ae. aegypti mosquitoes

Male mosquitoes transfer seminal fluid proteins (hereafter ‘SFPs’) during mating. These proteins can have profound effects on female behavior in the yellow fever mosquito Aedes aegypti and the Asian tiger mosquito Aedes albopictus. SFPs are thought to be responsible for female refractoriness to mating in both species. However, only limited information is available about the duration of induced refractoriness or the quantity of SFPs required to be effective in Ae. albopictus. Here, we tested the duration of the effect of SFPs on female refractory behavior for both Aedes species. Additionally, we determined the lowest SFP dose required to induce female refractory behavior in Ae. aegypti. Virgin females were injected intra-thoracically with doses ranging from 0.25 to 0.008 equivalents of one male’s SFP amount. Our results demonstrate high sensitivity of female Ae. aegypti and Ae. albopictus to SFPs of their own species, with the majority of females becoming refractory at doses ? 0.031 male-equivalents after injection into the hemocoel. This effect was long-lasting in both species; none of the injected females were inseminated when presented with males of their own species 30 to 34 days post-injection, whereas most saline-injected control females mated at this time point. These results will aid future work to characterize individual SFPs involved in post-mating refractoriness in these two species. Moreover, they show that as is the situation in the mosquito Anopheles gambiae, and unlike Drosophila melanogaster, sperm are not required for the maintenance of a sexual refractoriness response in Ae. aegypti and Ae. albopictus.     

疏花水柏枝内生真菌QY-1的抗氧化活性分析

以从疏花水柏枝(Myricaria laxiflora)中分离的1株内生真菌QY-1为研究对象,为分析其体内及体外的抗氧化活性,对其发酵液及其提取物的活性进行了综合评价。对nr DNA内转录间隔区进行测序鉴定。分别采用总抗氧化力试剂盒、自由基清除试剂盒和还原力测定试剂盒评价发酵液及粗提物的体外抗氧化能力活性;对发酵液提取物及其主成分进行了大肠埃希菌、人神经母瘤细胞SH-SY5Y的体内抗氧化损伤保护作用分析。结果表明QY-1是1株球毛壳菌(Chaetomium globosum),其粗提物具有很高的抗氧化活性,总抗氧化活力、自由基清除率及总还原力均接近维生素C的50%,远高于国际报道水平。且其发酵液提取物有促进细胞增殖和抗氧化保护作用,显著提高大肠埃希菌细胞和神经细胞在H2O2胁迫下的存活率。其粗提物可保护神经细胞膜的完整性,有效减低乳酸脱氢酶(LDH)的渗漏率。从粗提物中分离到一种主成分SF2,可显著抑制凋亡蛋白Caspase 3和Caspase 9的活性。结果表明内生真菌球毛壳菌QY-1不管在体内还是体外都具备较强的抗氧化能力,是一种具有潜在开发价值的天然抗氧化剂资源。     

Lipid lowering and antioxidant effects of Amomum subulatum seeds (Family Zingiberaceae) in cholesterol fed rabbits

In the present study, hypolipidemic activity of fraction (50:50; CHCl₃:CH₃OH) of Amomum subulatum (Zingiberaceae) seeds was evaluated in cholesterol-fed rabbits. Hyperlipidemia induced by feeding atherogenic diet for 120 days resulted in a significant increase in serum total cholesterol, phospholipid and triglyceride levels when compared with control group. The levels of LDL and VLDL-cholesterol were increased significantly, but the HDL-cholesterol ratio was decreased. The changes in the antioxidant parameters were accompanied by an increase in lipid peroxidation and reduction in glutathione (GSH) and catalase activity. The level of lipid peroxidation was reduced whereas GSH content and catalase activity were elevated after the treatment with A. subulatum fraction at the dose level of 100 mg/kg.b.wt/day. A significant reduction was observed in total cholesterol, triglyceride, phospholipid, LDL and VLDL cholesterol where as HDL-cholesterol ratio was increased after administration of A. subulatum. The results of the present study indicate that fraction of A. subulatum possesses lipid-lowering and antioxidant activity and could be beneficial in the treatment of hyperlipidemia.     

The antioxidant characteristics of medicinal plant extracts from Western Siberia

An antioxidant activity of the water-alcohol extracts of leaves of ten herbs from Western Siberia was studied. In vivo the capability of extracts to protect cells of Escherichia coli against the bacteriostatic action of H ₂O₂ and the influence of the extracts on the expression of the antioxidant gene katG coding catalase-hydroperoxidase I were investigated. In vitro the radical-binding activity with DPhPG^· (1,1-diphenyl-2-picrylhydrazyl radical), the chelating capability with ferrozine, and total composition of flavonoids and tannins were determined. The extracts of Filipendula stepposa and Limonium gmelinii were characterized by the highest antioxidant activity. According to data, the test extracts could have an antioxidant effect on bacteria in different ways at once including the direct inhibition of ROS (reactive oxygen species), iron ion chelation and antioxidant gene induction.     

One of two tandem Arabidopsis genes homologous to monosaccharide transporters is senescence-associated

A gene designated SFP1, which is similar to major facilitator superfamily monosaccharide transporters, is induced during leaf senescence. Genomic sequence analysis identified a second highly similar and closely linked gene, SFP2, suggesting that SFP1 and SFP2 may have arisen through a recent duplication event. However, RNA gel-blot analyses and histochemical localization of a reporter gene activity in transgenic plants show that SFP1 and SFP2 are differentially regulated and that only SFP1 is induced during leaf senescence. The increase in SFP1 gene expression during leaf senescence is paralleled by an accumulation of monosaccharides. Possible roles for SFP1 in sugar transport during leaf senescence are discussed.     

Sulforaphane inhibits mitochondrial permeability transition and oxidative stress

Exposure of mitochondria to oxidative stress and elevated Ca²⁺ promotes opening of the mitochondrial permeability transition pore (PTP), resulting in membrane depolarization, uncoupling of oxidative phosphorylation, and potentially cell death. This study tested the hypothesis that treatment of rats with sulforaphane (SFP), an activator of the Nrf2 pathway of antioxidant gene expression, increases the resistance of liver mitochondria to redox-regulated PTP opening and elevates mitochondrial levels of antioxidants. Rats were injected with SFP or drug vehicle and liver mitochondria were isolated 40 h later. Respiring mitochondria actively accumulated added Ca²⁺, which was then released through PTP opening induced by agents that either cause an oxidized shift in the mitochondrial redox state or directly oxidize protein thiol groups. SFP treatment of rats inhibited the rate of pro-oxidant-induced mitochondrial Ca²⁺ release and increased expression of the glutathione peroxidase/reductase system, thioredoxin, and malic enzyme. These results are the first to demonstrate that SFP treatment of animals increases liver mitochondrial antioxidant defenses and inhibits redox-sensitive PTP opening. This novel form of preconditioning could protect against a variety of pathologies that include oxidative stress and mitochondrial dysfunction in their etiologies.     

Purification and Biological Activities of Enzymatically Degraded Sargassum fusiforme Polysaccharides

Enzymatic hydrolysate of the crude polysaccharide (SFP) extracted from Sargassum fusiforme was purified by column DEAE-52 and Sephadex G-100 to yield four components, namely, ESFP1, ESFP2, ESFP3 and ESFP4. These components were characterized by chemical composition assay, GC/MS, HPGPC, UV and FT-IR techniques. The in vitro antioxidant activities of the four purified fractions were investigated by measuring their radical scavenging activity and reducing power. The results suggested that all the four components possess good antioxidant activities. Among them, ESFP1 was found to possess the strongest 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical and hydroxyl radical-scavenging activity, and the greatest ferric reducing power. The immunomodulatory effect of these four polysaccharides was demonstrated by their ability to promote proliferation, and to enhance both phagocytic activity and NO release in a macrophage RAW264.7 model. The results revealed that the bioactivities of the polysaccharides are related to their molecular weight, and the uronic acid and sulfate contents.     

Free radical and reactive oxygen species scavenging activities of the extracts from seahorse, Hippocampus kuda Bleeler

Seahorse, Hippocampus kuda (SH) a marine teleost fish, is well known not only for its special medicinal composition and used as one of the most famous and expensive materials of traditional Chinese medicine. It was extracted with water (SHW), methanol (SHM), and ethanol (SHE), respectively and evaluated by various antioxidant assays. The including reducing power, total antioxidant, DPPH radical scavenging, hydroxyl radical scavenging, superoxide anion radical scavenging, alkyl radical scavenging, and protective effect on DNA damage caused by hydroxyl radicals generated. Further, the ROS level was detected using a fluorescence probe, 2′,7′-dichlorofluorescin diacetate (DCFH-DA), which could be converted to highly fluorescent dichlorofluorescein (DCF) with the presence of intracellular ROS on mouse macrophages, RAW264.7 cell and inhibited myeloperoxidase (MPO) activity in human myeloid, HL60 cells, respectively. Those various antioxidant activities were compared to standard antioxidants such as α-tocopherol. Among SHM exhibited the highest antioxidant activity in linoleic acid system, effective reducing power, DPPH radical scavenging, hydroxyl radical scavenging, superoxide radical scavenging, alkyl radical scavenging, inhibitory intracellular ROS, and inhibited MPO activity. Furthermore, MTT assay showed no cytotoxicity on mouse macrophages cell (RAW264.7) and human cell lines (MRC-5, HL60, U937). This antioxidant property depends on concentration and increasing with increased amount of extracts. The results obtained in the present study indicated that the see horse (Hippocampus kuda Bleeker) is a potential source of natural antioxidant.     

Taiwanofungus camphoratus Activates Peroxisome Proliferator-Activated Receptors and Induces Hypotriglyceride in Hypercholesterolemic Rats

Taiwanofungus camphoratus (T. camphoratus), a fungus and a Taiwan-specific, well-known traditional Chinese medicine, has long been used to treat diarrhea, hypertension, itchy skin, and liver cancer. To gain a large amount of T. camphoratus, several culture techniques have been developed, including solid-state culture and liquid-state fermentation. Peroxisome proliferator-activated receptor gamma (PPARγ) has been described as a hypoglycemic agent that increases insulin sensitivity in peripheral tissues and results in reduced blood glucose, insulin, and triglyceride levels in insulin-resistant animals and in type-2 (non-insulin-dependent) diabetic patients. In this study, we investigate the possibility that T. camphoratus might activate PPARγ in vitro and hypolipidemic activity in vivo. The results show that an aqueous extract of the wild fruiting bodies of T. camphoratus was able to increase the PPARγ activity in cells transfected with the PPARγ expression plasmid and the AOx-TK reporter plasmid. Based on the cell experiment, we examined the hypolipidemic effect of wild fruiting bodies (WFT) and a solid-state culture (SST) of T. camphoratus on SD rats fed on a high-cholesterol (HC) diet. The results show that WFT significantly decreased the serum triglyceride level, but could not affect the cholesterol level. SST only slightly decreased the serum triglyceride level. In addition, both WFT and SST significantly decreased the serum alanine transaminase (ALT) level and protected against the liver damage induced by the HC diet from the results of a histological examination. These results suggest that T. camphoratus might contain PPARγ ligands and result in a hypotriglyceridemic effect, and that it also exhibits a liver protective activity.