High-throughput assay for detection of soybean lipoxygenase-1

A high-throughput assay was developed to detect soybean lipoxygenase 1 (LOX-1) using a multilabel plate reader. The assay was also adapted to a single cell fluorometer. Fluorescein is degraded by linoleic hydroperoxide produced from soybean lipoxygenase and linoleic acid. The decrease in fluorescence is measured over time, and the area-under-the-curve (AUC) is used to quantify the LOX-1 content of soybean extract. A dose-dependent response is seen with varied dilutions of pure LOX enzyme or soybean extracts. Percent recovery was between 97% and 108%, and relative standard deviation was 4.3%. Advantages of the assay include the reduced preparation time of samples and reduced use of reagents in the high-throughput assay. Multiple samples can be measured in a single run with a multilabel plate reader.     

Effects of xylanase treatments on gelling and water-uptaking properties of psyllium

The effects of a commercial food-grade xylanase on the physicochemical properties of psyllium were evaluated. The enzymatic reactions were conducted in solid state at ambient temperature. The enzyme-treated psyllium preparations were analyzed and compared with the starting psyllium for their water-uptake properties, gelling capacities, soluble and insoluble fiber contents, and surface structures. The solid-state xylanase treatment significantly reduced both water-uptake and gelling capacities of psyllium (p < 0.01), with a slight decrease of soluble fiber content, whereas no effect on insoluble fiber content was observed. The xylanase treatment also resulted in a smoother surface structure of psyllium particles. In addition, no special equipment and operation were required to conduct the enzymatic reaction, which generated no waste. These data indicated a potential to improve the physicochemical properties of psyllium by use of the solid-state xylanase reactions to promote the utilization of psyllium fiber in functional foods for promoting human health.     

Disruption of the epithelial apical-junctional complex by Helicobacter pylori CagA

Helicobacter pylori translocates the protein CagA into gastric epithelial cells and has been linked to peptic ulcer disease and gastric carcinoma. We show that injected CagA associates with the epithelial tight-junction scaffolding protein ZO-1 and the transmembrane protein junctional adhesion molecule, causing an ectopic assembly of tight-junction components at sites of bacterial attachment, and altering the composition and function of the apical-junctional complex. Long-term CagA delivery to polarized epithelia caused a disruption of the epithelial barrier function and dysplastic alterations in epithelial cell morphology. CagA appears to target H. pylori to host cell intercellular junctions and to disrupt junction-mediated functions.     

Essential role of Fkbp6 in male fertility and homologous chromosome pairing in meiosis

Meiosis is a critical stage of gametogenesis in which alignment and synapsis of chromosomal pairs occur, allowing for the recombination of maternal and paternal genomes. Here we show that FK506 binding protein (Fkbp6) localizes to meiotic chromosome cores and regions of homologous chromosome synapsis. Targeted inactivation of Fkbp6 in mice results in aspermic males and the absence of normal pachytene spermatocytes. Moreover, we identified the deletion of Fkbp6 exon 8 as the causative mutation in spontaneously male sterile as/as mutant rats. Loss of Fkbp6 results in abnormal pairing and misalignments between homologous chromosomes, nonhomologous partner switches, and autosynapsis of X chromosome cores in meiotic spermatocytes. Fertility and meiosis are normal in Fkbp6 mutant females. Thus, Fkbp6 is a component of the synaptonemal complex essential for sex-specific fertility and for the fidelity of homologous chromosome pairing in meiosis.     

Understanding agri-food networks as social relations

Actor network theory and supply chainmanagement theory provide suggestive researchdirections for understanding regional agri-foodnetworks. These theories claim that relationshipsbased upon trust and cooperation are critical to thestrength and vitality of the network. This means thatexploring and detailing these relationships among thesuppliers, producers, workers, processors, brokers,wholesalers, and retailers within specific regionalgeographies of these networks are critical forfurthering cooperation and trust. Key areas ofcooperation include resource sharing andapprenticeship programs. Employing food networks as akey unit of contextual analysis will deepen ourunderstanding of how to enhance their resiliency andvibrancy. Important questions can be raised about thedifference gender makes for farmers, brokers,entrepreneurs, and workers in local food networks.     

Trans-endocytosis of CD80 and CD86: a molecular basis for the cell-extrinsic function of CTLA-4

Cytotoxic T lymphocyte antigen 4 (CTLA-4) is an essential negative regulator of T cell immune responses whose mechanism of action is the subject of debate. CTLA-4 shares two ligands (CD80 and CD86) with a stimulatory receptor, CD28. Here, we show that CTLA-4 can capture its ligands from opposing cells by a process of trans-endocytosis. After removal, these costimulatory ligands are degraded inside CTLA-4-expressing cells, resulting in impaired costimulation via CD28. Acquisition of CD86 from antigen-presenting cells is stimulated by T cell receptor engagement and observed in vitro and in vivo. These data reveal a mechanism of immune regulation in which CTLA-4 acts as an effector molecule to inhibit CD28 costimulation by the cell-extrinsic depletion of ligands, accounting for many of the known features of the CD28-CTLA-4 system.     

Molecular species of acylglycerols incorporating radiolabeled fatty acids from castor (Ricinus communis L.) microsomal incubations

Sixty-one molecular species of triacylglycerols (TAG) and diacylglycerols produced from castor microsomal incubations incorporating six different (14)C-labeled fatty acids have been identified and quantified. The preference for incorporation into TAG was in the order ricinoleate > oleate > linoleate > linolenate > stearate > palmitate. Ricinoleate was the major fatty acid incorporated, whereas stearate, linolenate, and palmitate were incorporated at low levels. Twenty-one molecular species of acylglycerols (HPLC peaks) in castor oil have also been assigned. The levels of TAG in castor oil are RRR (triricinolein) > RR-TAG > R-TAG > no R-TAG. The levels of the molecular species within the groups of RR-TAG, RL-TAG, and LL-TAG individually are ricinoleate > linoleate > oleate > linolenate, stearate, and palmitate. The results of the labeled fatty acid incorporation are consistent with ricinoleate being preferentially driven into TAG and oleate being converted to ricinoleate in castor oil biosynthesis.     

A comparison of progestin-based protocols to synchronize estrus in postpartum beef cows

Two progestin-based protocols for estrus synchronization in postpartum beef cows were compared following treatment administration on the basis of estrous response, interval to and synchrony of estrus, and pregnancy. Cows were assigned to one of the two treatment protocols by age, body condition score (BCS), and days postpartum (DPP). The MGA Select-treated cows (MGA Select; n = 109) were fed melengestrol acetate (MGA; 0.5mg x cow-1 x d(-1)) for 14 d, fed carrier for 8 d, GnRH (100 microg of Cystorelin) was injected i.m. 12 d after MGA withdrawal, and PG (25 mg of Lutalyse) was administered i.m. 7 d after GnRH. Cows assigned to the 7-11 Synch protocol (7-11 Synch; n = 111) were fed carrier for 15 d, fed MGA for 7 d, injected with PG on d 22 (d 7 of MGA), injected with GnRH on d 26, and injected with PG on d 33. Mean BCS (4.8 +/- 0.1, MGA Select; 4.7 +/- 0.1, 7-11 Synch) and DPP (40 +/- 1, MGA Select; 40 +/- 1, 7-11 Synch) did not differ between treatments. Blood samples were collected 8 d and 1 d before feeding of MGA or carrier to determine the pretreatment estrous cyclicity (progesterone > or = 1 ng/mL; 10/109 [9%], MGA Select; 12/111 [11%], 7-11 Synch), and again at PG on d 33 to evaluate treatment response (81/109 [74%], MGA Select; 84/111 (76%), 7-11 Synch). Serum concentrations of progesterone at PG on d 33 differed (P < 0.01) between treatments (3.3 +/- 0.3 ng/mL [MGA Select] vs. 1.7 +/- 0.1 ng/mL [7-11 Synch]). HeatWatch was used for 6 d after PG on d 33 to detect estrus, and AI was performed 12 h after the onset of estrus. Estrous response did not differ between treatments (100/109 [92%], MGA Select; 101/111 [91%], 7-11 Synch). Mean interval to estrus (65 +/- 2.7 h, MGA Select; 52 +/- 1.8 h, 7-11 Synch) and synchrony of estrus differed (P < 0.01) between treatments. Synchronized conception and pregnancy rates (61/100 [61%], 61/109 [56%], MGA Select; 71/101 [70%], 71/111 [64%], 7-11 Synch), and final pregnancy rates (94/109 [86%], MGA Select; 99/110 [90%], 7-11 Synch) did not differ between treatments. In summary, estrous response and fertility did not differ among cows assigned to the MGA Select or 7-11 Synch protocols. Synchrony of estrus, defined as the variance in the interval to estrus from PG, however, was improved following treatment with the 7-11 Synch protocol.