Naturally derived micelles for rapid in vitro screening of potential cholesterol-lowering bioactives

A high plasma cholesterol level, especially low-density lipoprotein cholesterol, indicates increased risk of cardiovascular diseases. Plasma cholesterol levels are influenced by diet and cholesterol biosynthesis, uptake, and secretion. Cholesterol uptake involves solubilization into complex phospholipid spherical bodies termed micelles that facilitate the transport of lipids through the gut brush border membrane into enterocytes. In vitro assays reported to date to determine potential cholesterol-lowering effects of various compounds require artificial micelle preparations that are elaborate and time-consuming to prepare. The aims of this study were to compare the efficacy of artificially prepared micelles with naturally derived micelles from pig's bile and to test their ability to assess potential inhibitors of cholesterol uptake. The suitability of pig's bile-derived micelles was tested both at the level of the micelle and at cellular uptake using cultured Caco-2 cells. Known cholesterol uptake inhibitors at the micelle (green tea catechins) and at the Caco-2 cell (beta-lactoglobulin-derived peptide, IIAEK) were used as reference inhibitory compounds. It was concluded that pig's bile was a rapid, reproducible, convenient, and cost-effective source of micelles for cholesterol micelle solubility and cellular uptake assay systems and is suitable for screening purposes focused on identifying potential cholesterol-lowering agents.     

Enhancing Embryo Yield in Superovulated Holstein Heifers by Immunization Against Inhibin

Eight heifers, aged 16–17 months and showing normal oestrous cycles, were immunized against a recombinant porcine inhibin α subunit immunogen, together with another 10 heifers of the same age as controls and treated with placebo immunogen. Primary (1 mg immunogen) and two booster (0.5 mg immunogen each) immunizations were administered at 28‐day intervals. Ten days after the second booster immunization, both groups of heifers underwent a superovulation treatment. Each animal was given an intravaginal progesterone releasing sponge, which was withdrawn 7 days following an i.m. injection of 0.5 mg cloprostenol. Heifers were treated with FSH for 4 days and artificially inseminated after oestrus occurred. The embryos were flushed and evaluated 7 days after insemination. Immunization significantly (p < 0.01) increased blood antibody titres against recombinant porcine inhibin α subunit, from pre‐immunizaion and control values of approximately 0.06 of ELISA 450 nm reading to 0.6 to 0.7 after two or three immunizations. The immunized heifers produced on average 15.8 ± 2.8 embryos, significantly (p < 0.05) higher than the yield of 8.3 ± 1.5 in the controls. The number of transferable embryos were non‐significantly higher in immunized than in control heifers (9.6 ± 3.1 vs 5.8 ± 1.6, p > 0.05). The peak plasma oestradiol concentrations were significantly higher in immunized than in control heifers, both immediately after FSH treatment and 20 days thereafter. Plasma P4 concentrations after superovulation were in the range of 20 ng / ml in the immunized heifers, significantly (p < 0.05) higher than the values approximately 15 ng / ml in control heifers. These results indicated that prior immunization against inhibin α subunit stimulated production of antibodies against inhibin, which enhanced follicular developmental response to superovulation and lead to higher yield of total and transferable embryos. Therefore immunization combined with the conventional superovulatory gonadotrophin treatment, can be a simple and efficient method to produce low cost bovine embryos.     

Exogenous DNA Uptake of Boar Spermatozoa by a Magnetic Nanoparticle Vector System

The sperm‐mediated gene transfer method is applicable to transgenesis in many species that use spermatozoa for reproduction recently, which has been shown various results. In the current study, we show that transgenic porcine embryos can be efficiently produced by employing a simple transfection method that uses magnetic nanoparticles (MNPs). The complexes formed between plasmid DNA and MNPs were bounded on ejaculated boar spermatozoa at a higher efficiency compared to methods using DNA alone or lipofection. Using confocal microscopy, rhodamine fluorophore‐labelled MNPs were detected on external surfaces of the spermatozoa membrane, which were bounded on zona pellucida of in vitro maturated oocyte during in vitro fertilization. Electron microscopy revealed that clusters of MNPs were detected in inside of plasma membrane and nucleus of the spermatozoa head. Additionally, we found that magnetofected boar spermatozoa could be fertilized with oocytes in vitro and that the resulting gene of green fluorescent protein was detected in fertilized eggs by genomic PCR analysis. Taken together, these results suggest that MNPs can be used to efficiently introduce a transgene into embryo via spermatozoa.     

Inhibition of low-density lipoprotein oxidation and up-regulation of low-density lipoprotein receptor in HepG2 cells by tropical plant extracts

Twelve edible plant extracts rich in polyphenols were screened for their potential to inhibit oxidation of low-density lipoprotein (LDL) in vitro and to modulate LDL receptor (LDLr) activity in cultured HepG2 cells. The antioxidant activity (inhibition of LDL oxidation) was determined by measuring the formation of conjugated dienes (lag time) and thiobarbituric acid reagent substances (TBARS). Betel leaf (94%), cashew shoot (63%), Japanese mint (52%), semambu leaf (50%), palm frond (41%), sweet potato shoot, chilli fruit, papaya shoot, roselle calyx, and maman showed significantly increased lag time (>55 min, P < 0.05) and inhibition of TBARS formation (P < 0.05) compared to control. LDLr was significantly up-regulated (P < 0.05) by Japanese mint (67%), semambu (51%), cashew (50%), and noni (49%). Except for noni and betel leaf, most plant extracts studied demonstrated a positive association between antioxidant activity and the ability to up-regulate LDL receptor. Findings suggest that reported protective actions of plant polyphenols on lipoprotein metabolism might be exerted at different biochemical mechanisms.     

Effectiveness of controlled internal drug release device treatment to alleviate reproductive seasonality in anestrus lactating or dry Barki and Rahmani ewes during non‐breeding season

This study aimed to evaluate the effectiveness of hormonal treatments on ovarian activity and reproductive performance in Barki and Rahmani ewes during non‐breeding season. Forty‐eight multiparous ewes, 24 Barki and 24 Rahmani ewes were divided into two groups, 12 lactating and 12 dry ewes for each breed. Controlled internal drug release (CIDR) device was inserted in all ewes for 14 days in conjunction with intramuscular 500 IU equine chronic gonadotrophin (eCG) at day of CIDR removal. Data were analysed using PROC MIXED of SAS for repeated measures. Breed, physiological status and days were used as fixed effects and individual ewes as random effects. Barki ewes recorded higher (p < .05) total number of follicles, number of large follicles, serum estradiol concentration and estradiol: progesterone (E₂:P₄) ratio compared to Rahmani ewes. Lactating ewes recorded higher (p < .05) number of small follicles and lower concentration of total antioxidant capacity (TAC) compared to dry ewes. Number and diameter of large follicles recorded the highest (p < .05) values accompanied with disappearance of corpora lutea at day of mating. Serum progesterone concentration recorded lower (p < .05) value at day of mating and the highest (p < .05) value at day 35 after mating. CIDR‐eCG protocol induced 100% oestrous behaviour in both breeds, but Rahmani ewes recorded longer (p < .05) oestrous duration compared to Barki. Conception failure was higher (p < .05) in Barki compared to Rahmani ewes. In conclusion, CIDR‐eCG protocol was more potent in improving ovarian activity in Barki compared to Rahmani ewes, but this protocol seems to induce hormonal imbalance in Barki ewes that resulted in increasing conception failure compared to Rahmani ewes.     

防治危害板蓝的小菜蛾和菜青虫的药剂筛选

通过４种杀虫剂田间药效试验,筛选出农地乐对小菜蛾防效最好：功夫、农地乐、敌敌畏对菜青虫有良好防效。选用农地乐可防治小菜蛾兼治菜青虫。     

Response of Photosynthesis and Water Relations of Rice (Oryza sativa) to Elevated Atmospheric Carbon Dioxide in the Subhumid Zone of Sri Lanka

The objective of the present paper is to determine the response of the physiological parameters related to biomass production and plant water relations in a standard Sri Lankan rice (Oryza sativa) variety (BG‐300) to elevated CO₂ (i.e. 570 µmol/mol). During two seasons, rice crops were grown under three different experimental treatments; namely, at 570 µmol/mol (i.e. ‘elevated’) and 370 µmol/mol (‘ambient’) CO₂ within open top chambers, and at ambient CO₂ under open field conditions. Leaf net photosynthetic rate in the elevated treatment increased by 22–75 % in comparison to the ambient. However, the ratio between intercellular and ambient CO₂ concentrations remained constant across different CO₂ treatments and seasons. CO₂ enrichment decreased individual leaf stomatal conductance and transpiration rate per unit leaf area, and increased both leaf and canopy temperatures. However, the overall canopy stomatal conductance and daily total canopy transpiration rate of the elevated treatment were approximately the same as those achieved under ambient conditions. This was because of the significantly greater leaf area index and greater leaf–air vapour pressure deficit under CO₂ enrichment. The leaf chlorophyll content increased significantly under elevated CO₂; however, the efficiency (i.e. photochemical yield) of light energy capture by Photosystem II (i.e. F_v/F_m) in chlorophyll a did not show a significant and consistent variation with CO₂ enrichment.     

Genotypic Variation in the Response of Rice (Oryza sativa L.) to Increased Atmospheric Carbon Dioxide and its Physiological Basis

Increasing atmospheric carbon dioxide concentration (C_a) is one aspect of global change that will have a significant impact on the productivity of agricultural crops. Crop yields have been shown to increase with increasing C_a. The magnitude of yield response to increased C_a could vary depending on genotypic and environmental factors. The objective of this study was to determine the genotypic variation of yield response and its physiological basis in rice (Oryza sativa L.) through an initial varietal screening using 16 genotypes. The genotypes were grown under two concentrations of C_a, i.e. 370 ± 28 (ambient) and 570 ± 42 (elevated) μmol mol⁻¹, in open top chambers under lowland field conditions at the Rice Research and Development Institute in Sri Lanka (7°50′N, 80°50′E) from May to August 2001 (yala season) and from November 2001 to March 2002 (maha). C_a within chambers was maintained around target concentrations by a computer‐based real‐time data acquisition and control system. There was significant variation between genotypes in the response of yield to elevated C_a, with absolute increases up to 530 g m⁻² in yala and 347 g m⁻² in maha. In relative terms, percentage yield increases from ambient to elevated C_a ranged from 4 % to 175 % in yala and from 3 % to 64 % in maha. Genotypic variation in yield showed significant positive correlations with light‐saturated net photosynthetic rate of the flag leaf during the grain‐filling stage. This indicated that increased assimilate supply and its genotypic variation contributed to the observed genotypic variation in yield response to elevated C_a. Furthermore, the capacity to develop a larger reproductive sink through increased panicle number per m² and increased number of grains per panicle contributed to greater yield at elevated C_a. There was a significant genotype × season interaction with genotypes responding differentially to increased C_a in the two seasons. This was mainly due to inter‐seasonal variation in incident radiation during the grain‐filling stage. Our results demonstrate the significant genotypic variation that exists within the rice germplasm, in the response to increased C_a of yield and its correlated physiological parameters. A subset of genotypes from screening trials such as the present study can be used for more in‐depth analysis of the influence of elevated C_a on processes responsible for yield determination in rice and for molecular studies to elucidate the genetic basis of the response to increased C_a. This could pave the way for breeding genotypes which are more productive in a future high CO₂ environment, provided that genotypes with greater flexibility in their physiology are selected to counter the genotype × environmental interaction.     

Semen Collection, Examination and Spermiogram in Ostriches

The level of fertility in the male ostrich exerts considerable influence on the efficiency of the fertilization procedure, and thus also on reproductive performance. The determination of the reproductive capacity is of particular interest with regard to the selection of single individuals for optimizing reproduction ratios. Although the breeding and raising of ostriches has become increasingly important in many countries, little research has been completed on reproductive parameters and factors that may possibly influence them. This study presents observations made concerning the quantity and quality of sperm as found in the spermatological testing of 411 ejaculate samples taken from male ostriches on two farms in Namibia. The semen volume varied between 0.1 and 1.5 ml (mean, 0.64 ml). Normal ejaculate colours ranged from white to ivory; the consistency ranged from thin creamy to viscous. The measured pH values lay between 6.4 and 8.0 (mean, 7.3). Microscopic investigations revealed sperm concentrations of 8.9–78.1 million/μl and individual sperm motility from 42 to 96% (mean, 78%). No mass motility was detectable in 42% of the ejaculates; weak mass motility was found in 46%, and clear mass movements were to be found in only 12% of samples. Regarding the morphology of the sperm, 5 to 26% were abnormal (mean, 17%) and 4 to 28% (mean, 20%) were dead. Seasonal patterns of sperm concentration and the influence of frequency of semen collection were investigated in a group of 56 healthy male ostriches. Peak sperm concentrations were found at the beginning of the breeding season in spring; the lowest values were found at the end of the breeding season in autumn. The highest quality ejaculate was obtained from those males whose semen was collected once a week. The results of this study provide fundamental data for the establishment of minimum quality requirements for ostrich sperm to be met by individuals receiving certification as breeding animals and for the selection of suitable males for use in artificial insemination.     

低温胁迫下外源CaCl2对玉米种子萌发及幼苗生长的影响

以杂C-546为试材,采用10、20、40、80、160 mmol/L外源CaCl_2浸种和叶面喷施不同浓度处理种子,比较分析低温胁迫下玉米种子发芽率、发芽势、发芽指数和幼苗叶片相对电导率、可溶性蛋白、Pn、Fv/Fm、POD和SOD活性等生理生化指标的变化。结果表明,在低温抑制玉米幼苗生长条件下,一定浓度CaCl_2(10~80 mmol/L)处理可降低玉米幼苗生长伤害程度。80 mmol/L CaCl_2浸种处理对玉米低温胁迫的缓解效果最佳,与对照比较,发芽率提高8%,发芽势提高24.6%,发芽指数提高33.2,相对电导率降低51.8%,可溶性蛋白含量增加76.3%,SOD活性增加35.7%。适宜浓度的CaCl_2在低温条件下可促进玉米种子的萌发,降低低温对玉米幼苗造成的伤害。