Low preoperative selenium is associated with post-operative atrial fibrillation in patients having intermediate-risk coronary artery surgery.

BACKGROUND/OBJECTIVES: Post-operative atrial fibrillation (POAF) is a frequent complication of cardiac surgery. Oxidative stress and reduced antioxidant function have major roles in its development. Selenium is a key to normal antioxidant function, and levels are often low before cardiac surgery. This study investigated whether low preoperative selenium levels were associated with POAF in cardiac surgical patients. SUBJECTS/METHODS: Using the Society of Thoracic Surgeons (STS) Mortality risk score, 50 patients having primary coronary artery bypass grafts (CABG) surgery were divided into two groups: (i) low-risk group (STS ⩽0.5%; n=26) and (ii) intermediate-risk group (STS ⩾2.0%; n=24). Plasma levels of selenium, glutathione peroxidase (GPx) and malondialdehyde (MDA) were measured in all patients at anaesthetic induction, after aortic cross-clamp removal, 3 h post cardiopulmonary bypass and on post-operative days 1 and 5. Multiple logistic regression was used to assess whether selenium levels were associated with POAF development. RESULTS: Seventeen patients developed POAF (14 patients in the intermediate-risk group and 3 patients in the low-risk group). Preoperative selenium was lower in patients who developed POAF compared with those with normal sinus rhythm (0.73±0.16 vs 0.89±0.13 µmol/l, P=0.005), and this was independently associated with POAF (PR 0.32; 95% confidence credible interval (95%cI) 0.06-0.85, P=0.016). Regardless of POAF, preoperative selenium was lower in the intermediate-risk patients than in the low-risk patients (0.77±0.15 vs 0.89±0.14 µmol/l; P=0.004). CONCLUSIONS: Intermediate-risk patients with low preoperative selenium levels may be at a greater risk of developing POAF following CABG. This raises the question of whether selenium supplementation in select cardiac surgical patients may reduce their POAF risk.

Protocol guided bleeding management improves cardiac surgery patient outcomes.

BACKGROUND AND OBJECTIVES: Excessive bleeding is a risk associated with cardiac surgery. Treatment invariably requires transfusion of blood products; however, the transfusion itself may contribute to postoperative sequelae. Our objective was to analyse a quality initiative designed to provide an evidenced-based approach to bleeding management. MATERIALS AND METHODS: A retrospective analysis compared blood product transfusion and patient outcomes 15 months before and after implementation of a bleeding management protocol. The protocol incorporated point-of-care coagulation testing (POCCT) with ROTEM and Multiplate to diagnose the cause of bleeding and monitor treatment. RESULTS: Use of the protocol led to decreases in the incidence of transfusion of PRBCs (47·3% vs. 32·4%; P < 0·0001), FFP (26·9% vs. 7·3%; P < 0·0001) and platelets (36·1% vs. 13·5%; P < 0·0001). During the intra-operative period, the percentage of patients receiving cryoprecipitate increased (2·7% vs. 5·1%; P = 0·002), as did the number of units transfused (248 vs. 692; P < 0·0001). The proportion of patients who received tranexamic acid increased (13·7% to 68·2%; P < 0·0001). There were reductions in re-exploration for bleeding (5·6% vs. 3·4; P = 0·01), superficial chest wound (3·3% vs. 1·4%; P = 0·002), leg wound infection (4·6% vs. 2·0%; P < 0·0001) and a 12% reduction in mean length of stay from operation to discharge (95%: 9-16%, P < 0·0001). Acquisition cost of blood products decreased by $1 029 118 in the 15-month period with the protocol. CONCLUSIONS: The implementation of a bleeding management protocol supported by POCCT in a cardiac surgery programme was associated with significant reductions in the transfusion of allogeneic blood products, improved outcomes and reduced cost.

Prevalence of Cysticercus bovis in Australian cattle.

OBJECTIVE: The first national abattoir survey of Cysticercus bovis ('beef measles') in cattle was conducted in February 2008. METHODS: During the data collection period, 493,316 cattle were subjected to standard postmortem procedures, including incision of the masseter and heart muscles. On-site veterinarians were asked to record the location of any C. bovis cysts, as well as the National Livestock Identification System ear tag numbers of infected animals. Veterinarians were asked to submit samples for laboratory confirmation by histology and polymerase chain reaction testing. RESULTS: Of the 23 samples submitted, none was positive for C. bovis by either diagnostic method. CONCLUSIONS: Occasional, isolated diagnoses of beef measles are still made in most states of Australia, but since the last regional surveys were conducted 30 years ago, when the estimated prevalence was 50 to 200 per 100,000 cattle slaughtered, the parasite has become extremely rare.

Simultaneous binding of PtdIns(4,5)P2 and clathrin by AP180 in the nucleation of clathrin lattices on membranes.

Adaptor protein 180 (AP180) and its homolog, clathrin assembly lymphoid myeloid leukemia protein (CALM), are closely related proteins that play important roles in clathrin-mediated endocytosis. Here, we present the structure of the NH2-terminal domain of CALM bound to phosphatidylinositol-4,5- bisphosphate [PtdIns(4,5)P2] via a lysine-rich motif. This motif is found in other proteins predicted to have domains of similar structure (for example, Huntingtin interacting protein 1). The structure is in part similar to the epsin NH2-terminal (ENTH) domain, but epsin lacks the PtdIns(4,5)P2-binding site. Because AP180 could bind to PtdIns(4,5)P2 and clathrin simultaneously, it may serve to tether clathrin to the membrane. This was shown by using purified components and a budding assay on preformed lipid monolayers. In the presence of AP180, clathrin lattices formed on the monolayer. When AP2 was also present, coated pits were formed.

The structure and function of the beta 2-adaptin appendage domain.

The heterotetrameric AP2 adaptor (alpha, beta 2, mu 2 and sigma 2 subunits) plays a central role in clathrin-mediated endocytosis. We present the protein recruitment function and 1.7 A resolution structure of its beta 2-appendage domain to complement those previously determined for the mu 2 subunit and alpha appendage. Using structure-directed mutagenesis, we demonstrate the ability of the beta 2 appendage alone to bind directly to clathrin and the accessory proteins AP180, epsin and eps15 at the same site. Clathrin polymerization is promoted by binding of clathrin simultaneously to the beta 2-appendage site and to a second site on the adjacent beta 2 hinge. This results in the displacement of the other ligands from the beta 2 appendage. Thus clathrin binding to an AP2-accessory protein complex would cause the controlled release of accessory proteins at sites of vesicle formation.

Clathrin coat construction in endocytosis.

Electron cryomicroscopy of the clathrin coat and X-ray crystallography of parts of the clathrin heavy chain combine to give a detailed picture of the clathrin molecule, assembled as a cage. Recently determined domain structures of other components of the endocytic machinery, particularly the mu2 subunit and the alpha-appendage domain of the AP2 adaptor complex, provide important information on the sequence of recognition events involved in the dynamic process of clathrin coat assembly.

Clathrin: anatomy of a coat protein.

Clathrin is a vesicle coat protein involved in the assembly of membrane and cargo into transport vesicles at the plasma membrane and on certain intracellular organelles. Recently, crystal structures of two separate parts of the clathrin heavy chain, a fragment of the proximal leg and the N-terminal domain, have been analysed, providing the first high-resolution data for a vesicle coat protein. Viewing these structures in the context of a hexagonal barrel coat, recently determined to 21 A by cryo-electron microscopy, provides new insights into the assembly of clathrin coats.

Functional organization of clathrin in coats: combining electron cryomicroscopy and X-ray crystallography.

The sorting of specific proteins into clathrin-coated pits and the mechanics of membrane invagination are determined by assembly of the clathrin lattice. Recent structures of a six-fold barrel clathrin coat at 21 A resolution by electron cryomicroscopy and of the clathrin terminal domain and linker at 2.6 A by X-ray crystallography together show how domains of clathrin interact and orient within the coat and reveal the strongly puckered shape and conformational variability of individual triskelions. The beta propeller of the terminal domain faces the membrane so that recognition segments from adaptor proteins can extend along its lateral grooves. Clathrin legs adapt to different coat environments in the barrel by flexing along a segment at the knee that is free of contacts with other molecules.

Clathrin coats at 21 A resolution: a cellular assembly designed to recycle multiple membrane receptors.

We present a map at 21 A resolution of clathrin assembled into cages with the endocytic adaptor complex, AP-2. The map was obtained by cryo-electron microscopy and single-particle reconstruction. It reveals details of the packing of entire clathrin molecules as they interact to form a cage with two nested polyhedral layers. The proximal domains of each triskelion leg depart from a cage vertex in a skewed orientation, forming a slightly twisted bundle with three other leg domains. Thus, each triskelion contributes to two connecting edges of the polyhedral cage. The clathrin heavy chains continue inwards under the vertices with local 3-fold symmetry, the terminal domains contributing to 'hook-like' features which form an intermediate network making possible contacts with the surface presented by the inner adaptor shell. A node of density projecting inwards from the vertex may correspond to the C-termini of clathrin heavy chains which form a protrusion on free triskelions at the vertex. The inter-subunit interactions visible in this map provide a structural basis for considering the assembly of clathrin coats on a membrane and show the contacts which will need to be disrupted during disassembly.

The non-refluxing gastrostomy: an evaluation.

A gastrostomy is often essential to deliver adequate and safe nutrition. Various types are now available such that the technique can be tailored to the specific needs of the patient. This paper explores the non-refluxing gastrostomy for long-term intermittent gastrostomy feeding, avoiding the need for a permanent indwelling appliance. A full-thickness vascularized flap based on the right gastroepiploic vessels is raised from the greater curve of the stomach. The proximal half of the tubularized flap is buried in a submucosal tunnel and the free distal end is brought to the skin surface as a catheterizable stoma. Fifteen children with varied mental and physical disabilities formed the cohort of the study. There were 3 stomal stenoses and 3 mild mucosal eversions requiring minor surgical adjustments. One child had a wound dehiscence 10 days postoperatively. Once the stoma had healed, the majority fed by intermittent catheterization and bolus feeds at conventional feed times during the day. Intermittent catheterization was painless and easy and was well accepted by caregivers and patients. Perhaps the most important advantages were the increased patient and caregiver confidence and independence, as well as the reduction in anxiety and hospital attendance.

A chimera of the cytoplasmic tail of the mannose 6-phosphate/IGF-II receptor and lysozyme localizes to the TGN rather than prelysosomes where the bulk of the endogenous receptor is found.

We fused the cytoplasmic and transmembrane domains of the bovine mannose 6-phosphate/IGF-II receptor (MPR) to lysozyme, a monomeric secretory protein thought to be devoid of sorting information. When the resulting chimera (lys/MPR) was transiently expressed in COS cells or stably expressed in CV1 cells, it had a predominantly intracellular distribution in the trans-Golgi region, with less than 10% present on the surface. In contrast, a similar chimera containing the transmembrane and cytoplasmic domains of the low density lipoprotein receptor (lys/LDLR) was localized to the plasma membrane, even though it endocytoses efficiently. Exchanging domains between the lys/MPR and lys/LDLR chimeras indicated that the MPR cytoplasmic domain contains the information necessary to specify the intracellular localization of the chimeric molecule. This signal must be located in the membrane-proximal third of the tail, as deletion of the last 120 residues of the 163 residue tail has no obvious effect on the distribution of lys/MPR. However, the recycling of the lys/MPR does not completely mimic that of the intact endogenous MPR, as immunofluorescence labelling shows that they are predominantly in different locations, indicating a role for the lumenal domain of the MPR in determining the steady-state distribution of the MPR itself.

Cloning of Drosophila beta-adaptin and its localization on expression in mammalian cells.

A Drosophila cDNA (BAD1) encoding a structural and assembly-competent homologue of the mammalian coated pit beta-adaptins (beta and beta') has been cloned and sequenced. In its amino-terminal region (residues 1-575), the BAD1 sequence appears intermediate between that of the mammalian beta-adaptin and a predicted sequence, from cDNA 105a, which appears to code for a version of beta'-adaptin. To test its functional characteristics, a 'myc'-tagged version of BAD1 was expressed in Cos cells. The BAD1 protein was detected most clearly in plasma membrane coated pits, where it colocalized with alpha-adaptin, although other coated pits were noted which apparently did not contain alpha-adaptin. However, these are probably gamma-adaptin containing pits, as BAD1 was also found colocalized with gamma-adaptin in Golgi coated pits in which, typically, alpha-adaptin is absent. Immunoprecipitation experiments confirmed that the BAD1 protein was present in both types of adaptor complex, unlike beta-adaptin which complexes with alpha-adaptin and beta'-adaptin which partners gamma-adaptin exclusively. In spite of this, BAD1 expression does not appear to mix alpha-adaptin and gamma-adaptin distribution amongst all the coated pits: thus the location of these adaptor complexes in mammalian cells does not depend on the differences between beta subunits but rather on membrane-specific interactions of other adaptor polypeptides. The differential interaction of beta with alpha-adaptin and beta' with gamma-adaptin in mammalian cells is likely to depend on the few non-conservative differences between their respective sequences and BAD1. Four of these (one with respect to beta and three versus 105a) are clustered in a particular region (residues 155 to 305), which may therefore represent a domain that influences the choice of partner adaptin.

Comparison of a liquid and a powder insecticidal dressing to aid healing and prevent flystrike of mulesing wounds in lambs.

A proprietary insecticidal mulesing powder containing diazinon and an experimental liquid dressing based on eucalyptus oil, naphthalene, cresylic acid and chlorfenvinphos in a carrier of liquid hydrocarbons and petroleum oil were compared for their ability to promote wound healing and reduce the incidence of fly strike in freshly mulesed lambs. Throughout the trial period of 4 weeks, fly trapping confirmed the presence of Lucilia cuprina in the paddock where the ewes and lambs were grazing At inspection one month after mulesing, no deaths had occurred since mulesing and no lambs showed evidence of cutaneous myiasis, although a number of their dams (with 8 months wool) were struck. At 4 weeks after mulesing, the wounds of the lambs treated with the experimental liquid dressing showed better healing than those treated with the powder dressing. It was concluded that both mulesing preparations were effective in inhibiting flystrike, but the liquid dressing promoted faster wound healing than did the powder.

Effect of lupin (Lupinus angustifolius) supplementation on ovarian and pituitary activity in ewes.

In each of three experiments, thirty seasonally anoestrous Border Leicester ewes were fed on a maintenance ration of oaten chaff. Fifteen of them were given a supplement of 500 g lupin grain per head per day. The ewes were treated with 10 mg follicle stimulating hormone (Expt 1), 600 I.U. pregnant mare serum gonadotrophin (Expt 2) and either 150 or 300 micrograms gonadotrophin releasing hormone (Expt 3) to determine whether the ovaries and/or the anterior pituitary were capable of responding to the nutrient status of the animals and influencing ovulation rate. In each experiment, the number and size of corpora lutea and follicles in the lupin-supplemented and -unsupplemented groups were similar. It was concluded that the mechanism by which lupins increase the ovulation rate is probably neural and not a result of direct effect on either the pituitary or the ovaries.

An assessment of a composite sampling method for counting strongyle eggs in sheep faeces.

The conventional method for estimating the average strongyle egg count for a group of sheep was compared with a single count from a group composite faecal sample. Sixty-one groups of field samples were used. Composite samples were prepared in the laboratory by pooling equal amounts of faeces from individual samples. Data were logtransformed for analysis to meet the assumption of normality. There were no significant differences in the variances and overall mean counts obtained by the 2 methods. The regression line of log (composite count) on log (group arithmetic mean) did not differ significantly from the line of identity. When untransformed egg count data were categorised as low, moderate and high, the 2 methods were in agreement for 53 of the 61 groups. The mixing and counting process used for both methods (modified McMaster technique) gave highly repeatable results (repeatability = 0.94). The composite method was a quicker and valid alternative to the conventional method for monitoring helminthosis in sheep flocks.

Specificity of binding of clathrin adaptors to signals on the mannose-6-phosphate/insulin-like growth factor II receptor.

Adaptors mediate the interaction of clathrin with select groups of receptors. Two distinct types of adaptors, the HA-II adaptors (found in plasma membrane coated pits) and the HA-I adaptors (localized to Golgi coated pits) bind to the cytoplasmic portion of the 270 kd mannose 6-phosphate (M6P) receptor-a receptor which is concentrated in coated pits on both the plasma membrane and in the trans-Golgi network. Neither type of adaptor appears to compete with the other for binding, suggesting that each type recognizes a distinct site on the M6P receptor tail. Mutation of the two tyrosines in the tail essentially eliminates the interaction with the HA-II plasma membrane adaptor, which recognizes a 'tyrosine' signal on other endocytosed receptors (for example, the LDL receptor and the poly Ig receptor). In contrast, the wild type and the mutant M6P receptor tail (lacking tyrosines) are equally effective at binding HA-I adaptors. This suggests that there is an HA-I recognition signal in another region of the M6P receptor tail, C-terminal to the tyrosine residues, which remains intact in the mutant. This signal is presumably responsible for the concentration of the M6P receptor, with bound lysosomal enzymes, into coated pits which bud from the trans-Golgi network, thus mediating efficient transfer of these enzymes to lysosomes.

Characterization of coated-vesicle adaptors: their reassembly with clathrin and with recycling receptors.

Adaptors sort out those receptors that participate in assembly of coated pits from those that are excluded. Two distinct adaptor units have so far been identified: (1) adaptors restricted to plasma membrane coated pits (HA-II type, named according to their elution position during hydroxylapatite chromatography) and (2) adaptors restricted to Golgi region coated pits (HA-I type). Adaptors contain a heterodimer of two 100-kDa polypeptides, a beta-adaptin (possibly carrying an essentially common clathrin-binding domain) and a distinct alpha- or gamma-adaptin characteristic of the type of adaptor and its specific location. Each adaptor in constructed from four different polypeptides. Thus HA-II adaptors contain a beta-adaptin and an alpha-adaptin in combination with a 50-kDa protein and a 16-kDa polypeptide. The HA-I adaptors contain a beta-adaptin and a gamma-adaptin in combination with a 47-kDa protein and a 19-kDa polypeptide. Both types of adaptors and also a 180-kDa polypeptide will promote the assembly of clathrin to form coats, the size range of which appears to be relatively restricted compared to cages made from clathrin alone. The HA-II adaptors, characteristic of plasma membrane coated pits, bind to the cytoplasmic tail of the LDL receptor. They also assemble with the mannose 6-phosphate receptor in vitro in the absence of membrane. When clathrin is included, the adaptors promote the assembly of coats containing bound receptor.