Fibrinolysis: strategies to enhance the treatment of acute ischemic stroke.

Stroke is a major cause of disability worldwide, and is the second leading cause of death after ischemic heart disease. Until recently, tissue-type plasminogen activator (t-PA) was the only treatment for acute ischemic stroke. If administered within 4.5 h of symptom onset, t-PA improves the outcome in stroke patients. Mechanical thrombectomy is now the preferred treatment for patients with acute ischemic stroke resulting from a large-artery occlusion in the anterior circulation. However, the widespread use of mechanical thrombectomy is limited by two factors. First, only ⁓ 10% of patients with acute ischemic stroke have a proximal large-artery occlusion in the anterior circulation and present early enough to undergo mechanical thrombectomy within 6 h; an additional 9-10% of patients presenting within the 6-24-h time window may also qualify for the procedure. Second, not all stroke centers have the resources or expertise to perform mechanical thrombectomy. Nonetheless, patients who present to hospitals where thrombectomy is not an option can receive intravenous t-PA, and those with qualifying anterior circulation strokes can then be transferred to tertiary stroke centers where thrombectomy is available. Therefore, despite the advances afforded by mechanical thrombectomy, there remains a need for treatments that improve the efficacy and safety of thrombolytic therapy. In this review, we discuss: (i) current treatment options for acute ischemic stroke; (ii) the mechanism of action of fibrinolytic agents; and (iii) potential strategies to manipulate the fibrinolytic system to promote endogenous fibrinolysis or to enhance the efficacy of fibrinolytic therapy.

Robust anti-obesity and metabolic effects of a dual GLP-1/glucagon receptor peptide agonist in rodents and non-human primates.

AIMS: To characterize the pharmacology of MEDI0382, a peptide dual agonist of glucagon-like peptide-1 (GLP-1) and glucagon receptors. MATERIALS AND METHODS: MEDI0382 was evaluated in vitro for its ability to stimulate cAMP accumulation in cell lines expressing transfected recombinant or endogenous GLP-1 or glucagon receptors, to potentiate glucose-stimulated insulin secretion (GSIS) in pancreatic ß-cell lines and stimulate hepatic glucose output (HGO) by primary hepatocytes. The ability of MEDI0382 to reduce body weight and improve energy balance (i.e. food intake and energy expenditure), as well as control blood glucose, was evaluated in mouse models of obesity and healthy cynomolgus monkeys following single and repeated daily subcutaneous administration for up to 2 months. RESULTS: MEDI0382 potently activated rodent, cynomolgus and human GLP-1 and glucagon receptors and exhibited a fivefold bias for activation of GLP-1 receptor versus the glucagon receptor. MEDI0382 produced superior weight loss and comparable glucose lowering to the GLP-1 peptide analogue liraglutide when administered daily at comparable doses in DIO mice. The additional fat mass reduction elicited by MEDI0382 probably results from a glucagon receptor-mediated increase in energy expenditure, whereas food intake suppression results from activation of the GLP-1 receptor. Notably, the significant weight loss elicited by MEDI0382 in DIO mice was recapitulated in cynomolgus monkeys. CONCLUSIONS: Repeated administration of MEDI0382 elicits profound weight loss in DIO mice and non-human primates, produces robust glucose control and reduces hepatic fat content and fasting insulin and glucose levels. The balance of activities at the GLP-1 and glucagon receptors is considered to be optimal for achieving weight and glucose control in overweight or obese Type 2 diabetic patients.

Risk management in clinical practice. Part 11. Oral surgery.

Oral surgery is often an unpleasant experience for a patient and if managed inadequately can be a cause for complaint or a claim in negligence. A practitioner can reduce their risk of complaints, claims or even regulatory body investigations by following some straightforward risk management strategies. Effective communication skills deployed throughout the interaction with the patient, especially during the consent process, are a pre-requisite, as is a proper understanding of the law on consent. An honest reflection by the practitioner on their competence to carry out a procedure, considering their skills, the equipment and support available will result in fewer medico-legal cases. In this article, each stage of the patient's journey is discussed and risk management advice offered for a range of procedures that are regularly encountered in general dental practice.

Mg2+-dependent compaction and folding of yeast tRNAPhe and the catalytic domain of the B. subtilis RNase P RNA determined by small-angle X-ray scattering.

We apply synchrotron-based small-angle X-ray scattering to investigate the relationship between compaction, metal binding, and structure formation of two RNAs at 37 degrees C: the 76 nucleotide yeast tRNA(Phe) and the 255 nucleotide catalytic domain of the Bacillus subtilis RNase P RNA. For both RNAs, this method provides direct evidence for the population of a distinct folding intermediate. The relative compaction between the intermediate and the native state does not correlate with the size of the RNA but does correlate well with the amount of surface burial as quantified previously by the urea-dependent m-value. The total compaction process can be described in two major stages. Starting from a completely unfolded state (4-8 M urea, no Mg(2+)), the major amount of compaction occurs upon the dilution of the denaturant and the addition of micromolar amounts of Mg(2+) to form the intermediate. The native state forms in a single transition from the intermediate state upon cooperative binding of three to four Mg(2+) ions. The characterization of this intermediate by small-angle X-ray scattering lends strong support for the cooperative Mg(2+)-binding model to describe the stability of a tertiary RNA.

A cross-sectional survey of pain and pharmacological analgesia in Canadian neonatal intensive care units.

OBJECTIVE: To determine current practices for the use of analgesia term and preterm neonates cared for in Neonatal Intensive Care Units (NICUs). DESIGN: One-week survey of medical charts of current patients. SETTING: NICUs in Canada. PARTICIPANTS: A total of 14 of 38 invited NICUs participated. These units were not different on number of beds, admissions per year, or university affiliation from the nonparticipating units. MAIN OUTCOME MEASURES: Daily logs were kept of the frequency and type of procedures and analgesia administration for all ill neonates in each NICU during the study period. RESULTS: The sample consisted of 239 patients. A total of 2,134 invasive procedures were performed. Medication was given specifically 18 times for 17 invasive procedures (0.8%). For another 129 invasive procedures, the patient was receiving analgesia for reasons other than the procedure. Sixteen patients had surgery during the survey period, and another 14 had surgery prior to but within 4 days of the survey. Fifty-one patients received anaesthesia or analgesia specifically related to surgery (39 times), procedures (35 times), or other reasons (34 times), a total of 108 courses. Opioids were the most frequently used medications and were given for all reasons, by continuous infusion, intermittent bolus, or sometimes both methods for the same patient. CONCLUSIONS: Postoperative pain in neonates in Canadian NICUs appears to be consistently treated, primarily with opioid analgesics, but analgesia, opioid or nonopioid is rarely given for nonsurgical invasive procedures.

Monte Carlo modeling of small-angle scattering data from non-interacting homogeneous and heterogeneous particles in solution.

A Monte Carlo algorithm that rapidly generates the scattered intensity function for complex heterogeneous particles is described. The heterogeneous particles are built from any number and orientation of simple building blocks, which include ellipsoidal shells, hollow ellipsoidal cylinders, ellipsoidal helices, triangular prisms, rectangular prisms, and semi-ellipsoidal shells. Applications are discussed such as real proteins and their complexes, polysaccharides, void effects on I(q), Guinier range estimation, and calculation of Stuhrmann plots.

Solution structures of GroEL and its complex with rhodanese from small-angle neutron scattering.

BACKGROUND: Molecular chaperonins 60 are cylindrical oligomeric complexes which bind to unfolded proteins and assist in their folding. Studies to identify the location of the protein substrate have produced contradictory results: some suggest that the substrate-binding site is buried within the interior of the complex, whereas others indicate an external (polar) location. RESULTS: Small-angle neutron scattering (SANS) measurements were made on GroEL chaperonin and on a complex of GroEL with rhodanese. The radius of gyration and the molecular weight determined from SANS measurements of GroEL agree well with those from its crystal structure. The positions of residues which were unresolved in the crystal structure have been confirmed. In addition, through model fitting of the SANS data, conformational changes in solution have been assessed and the location of bound rhodanese has been determined. CONCLUSIONS: The overall structure of GroEL in solution is similar to the crystal structure. In GroEL the N-terminal and C-terminal residues are organized compactly near the equator of the cylinder and the apical domains are flared by about 5 degrees. The best fit of SANS data suggests the existence of an equilibrium between the complex and single rings and monomers. SANS data for the GroEL-rhodanese complex are consistent with a model wherein one rhodanese molecule binds across the opening to the chaperonin cavity, rather than within it.

Conformational changes in yeast phosphoglycerate kinase upon substrate binding.

Small-angle neutron scattering (SANS) was used to measure the radius of gyration (Rg) of solutions of phosphoglycerate kinase (PGK) in a variety of substrate environments in D2O. The Rg of 24.0 A was measured for native PGK. A decrease in Rg was observed for the following: 23.7 A for PGK+sulphate; 23.5 A for PGK+ beta, gamma-bidentate Cr(H2O)4ATP (CrATP); 23.3 A for PGK + 3-phospho-D-glycerate (PGA)+CrATP; 22.9 A for PGK+CrATP+sulphate; 22.6 A for PGK+PGA+CrATP+sulphate. The statistical error was about +/- 0.3 A, which is less than systematic effects in this system. These results are consistent with catalysis by a hinge-bending motion of the enzyme. Since CrATP is not hydrolyzed, these results represent the conformational states of the bound substrates in the catalytically relevant ternary complex in the absence of product formation. The second virial coefficient is also measured for this system and this is consistent with that calculated from the protein volume only.

A heat and moisture exchanging filter in long-term ventilation.

We report our experience of using the Pall BB50T heat and moisture exchanging filter (HMEF) in critically ill patients undergoing long-term ventilation. Three hundred and sixty-four ventilated patients humidified with the Pall HMEF were studied prospectively. Thirty-five patients (mean APACHE II score 24.6, mean predicted mortality 50.9%) were ventilated for more than 10 days (mean 18.9), accounting for 661 patient-ventilated days. During this period two patients suffered tracheal tube occlusion. No other complications could be attributed to the filter. We conclude that the Pall BB50T heat and moisture exchanging filter is safe in patients undergoing long-term ventilation. However, its use demands a high degree of respiratory care.

Renal organic acid transport: uptake by rat kidney slices of a furan dicarboxylic acid which inhibits plasma protein binding of acidic ligands in uremia.

The furan dicarboxylic acid, 3-carboxy-4-methyl-5-propyl-2-furanpropanoic acid (5-propyl FPA), accumulates in uremic plasma and inhibits the binding of various drugs and marker ligands that are organic acids. 5-Propyl FPA is excreted unchanged in human urine and active tubular secretion is likely to be involved because of its high affinity for albumin. The uptake of 5-propyl FPA by rat kidney slices has been measured and compared with that of p-aminohippurate (PAH). The mean (+/- S.D.) slice/medium ratio for uptake of 5-propyl FPA (76 microM) was 22.7 +/- 2.6 (n = 11) and for PAH (75 microM) was 15.9 +/- 3.2 (n = 9) after incubation for 90 min at 25 degrees C. 5-Propyl FPA (149-829 microM) inhibited the uptake of PAH (77 microM) in a concentration-dependent manner, and likewise, PAH (150-830 microM) inhibited the uptake of 5-propyl FPA (77 microM). The mean apparent Km and Vmax values for the uptake of 5-propyl FPA were 194 +/- 125 microM and 55 +/- 28 nmol/g kidney/min, respectively, and 487 +/- 179 and 99 +/- 46 nmol/g kidney/min, respectively, for PAH. The kinetics of inhibition of uptake of PAH by 5-propyl FPA were mainly competitive. 5-Propyl FPA is thus likely to undergo active tubular secretion in a similar way to PAH, and this furan dicarboxylic acid, therefore, has the potential to inhibit the renal excretion of various drugs, drug conjugates and other endogenous organic acids.

Solution structure of phosphorylase kinase studied using small-angle X-ray and neutron scattering.

Small-angle X-ray and neutron scattering have been used to characterize the solution structure of rabbit skeletal phosphorylase kinase. The radius of gyration of the unactivated holoenzyme determined from neutron scattering is 94 A, and its maximum dimension is approximately 275-295 A. A planar model has been constructed that is in general agreement with the dimensions of the transmission electron microscope images of negatively stained phosphorylase kinase and that gives values for the radius of gyration, maximum linear dimension, and a pair distribution function for the structure that are consistent with the scattering data.

Neonatal screening for haemoglobin variants using filter paper-dried blood specimens.

Neonatal screening for haemoglobinopathies utilizing cord blood samples is well established, although it has a high miss rate and has the inherent problem of possible misdiagnosis from maternal contamination of the sample. The use of dried Guthrie card samples which are taken at six days of age avoids these problems and has the advantage of using an established system of sample collection. Controversy exists as to the method of choice for analysis of dried samples, this study of 2406 samples found that Iso-electric focusing (IEF) analysis of dried specimens gives excellent correlation when compared with cellulose acetate/citrate agar electrophoresis of liquid cord blood samples. The IEF results were clear and relatively simple to interpret even when the samples had been stored at room temperature for 4 weeks. The commercial enzyme linked immunosorbent assay (ELISA) screening test JOSHUA reliably determines the presence or absence of haemoglobin S in dried specimens. It could therefore be used as a relatively cheap and simple method for the confirmation of sickle cell trait in neonatal screening programmes based on dried specimens.

Interaction of 3-carboxy-4-methyl-5-propyl-2-furanpropanoic acid, an inhibitor of plasma protein binding in uraemia, with human albumin.

The furan dicarboxylic acid 3-carboxy-4-methyl-5-propyl-2-furanpropanoic acid (5-propyl FPA) accumulates in uraemic plasma and is a potent inhibitor of the binding of other anionic ligands to albumin. The interaction of 5-propyl FPA with human albumin has been investigated by equilibrium dialysis at 37 degrees and pH 7.4. Analysis of the binding data on the basis of a two-site model gave binding parameters of n1 = 0.6 and K1 = 4.8 x 10(6) M-1 for the primary binding site. 5-Propyl FPA binding was observed to decrease as the pH was raised from 6.4 to 8.3 which emphasizes the need for pH control of whole plasma or serum. Temperature, however, had little effect on binding as assessed by equilibrium dialysis at 10 degrees, 25 degrees and 37 degrees. The high affinity of 5-propyl FPA for albumin explains its retention in uraemic plasma, its potency as a binding inhibitor and points to active tubular secretion as the mechanism by which it is normally excreted by the kidney.

Low mortality associated with low volume pressure limited ventilation with permissive hypercapnia in severe adult respiratory distress syndrome.

Many animal studies have shown that high peak inspiratory pressures (PIP) during mechanical ventilation can induce acute lung injury with hyaline membranes. Since 1984 we have limited PIP in patients with ARDS by reducing tidal volume, allowing spontaneous breathing with SIMV and disregarding hypercapnia. Since 1987 50 patients with severe ARDS with a "lung injury score" greater than or equal to 2.5 and a mean PaO2/FiO2 ratio of 94 were managed in this manner. The mean maximum PaCO2 was 62 mmHg, the highest being 129 mmHg. The hospital mortality was significantly lower than that predicted by Apache II (16% vs. 39.6%, chi 2 = 11.64, p less than 0.001). Only one death was due to respiratory failure, caused by pneumocystis pneumonia. 10 patients had a "ventilator score" greater than 80, which has previously predicted 100% mortality from respiratory failure. Only 2 died, neither from respiratory failure. There was no significant difference in lung injury score, ventilator score, PaO2/FiO2 or maximum PaCO2 between survivors and non-survivors. We suggest that this ventilatory management may substantially reduce mortality in ARDS, particularly from respiratory failure.

Antibody-mediated pure neutrophil aplasia, recurrent myasthenia gravis and previous thymoma: case report and literature review.

We describe a case of neutrophil aplasia in a woman with recurrent myasthenia gravis and a past history of thymoma. Bone marrow showed virtually absent granulopoiesis but normal erythropoiesis and megakaryopoiesis. Bone marrow cultures showed no growth of granulocyte/mononuclear cell progenitors (CFU-GM). She was treated with immunosuppression including azathioprine, and her neutrophil count returned to normal. Serum before treatment, and also an IgG fraction thereof, inhibited CFU-GM growth both in autologous 'remission' marrow and in allogeneic marrow. She remains in complete remission 36 months after starting azathioprine. This association is extremely rare, and was formerly associated with a grim prognosis.

Structural relationships between the isoenzymes of human placental alkaline phosphatase: a serum factor converts M-PLAP to A- and B-PLAP.

A protein factor has been found in serum which converts the M form of placental alkaline phosphatase (PLAP) to the A and B forms. The identity of the conversion products has been confirmed by analysis of their dimers and polypeptides. Proteolysis is not implicated in this phenomenon. This report establishes microvillous M-PLAP as the precursor of the A and B forms.

Factors producing error in the total leucocyte count as measured on the Coulter Counter S Plus IVD.

Whole blood specimens, anticoagulated with potassium EDTA, which were seen to have particles interfering with the total leucocyte count were collected. This interference was detected, when the samples were processed through a Coulter Counter S Plus IVD, by R1 regional flags or backlighting of the leucocyte count. These specimens then had visual leucocyte counts performed using a counting chamber method and were examined microscopically, to ascertain possible causes of the interference. It was shown that the degree of error in the leucocyte count produced varies considerably but can be very high. Particles causing the error were found to include fibrin strands, platelet clumps, nucleated red cells and giant platelets.