Alternative pathway of complement: recruitment of precursor properdin by the labile C3/C5 convertase and the potentiation of the pathway

In this study the physiological role of properdin and the differential subunit composition of the solid phase enzymes of the pathway have been explored. Cell-bound C3 and C5 convertase differ in their C3b requirement. Apparently one molecule of C3b is sufficient to allow formation of C3 convertase (C3b,B), whereas two or more are required for generation of C5 convertase (C3bn,B). This conclusion was drawn from results indicating the critical role of the spacial distribution of C3b molecules on the cell surface in enzyme formation. While the C3/C5 convertase is fully capable of acting on C5 and thereby initiating the assembly of the cytolytic membrane attack complex, it is exceedingly labile and vulnerable to destruction by the C3b inactivator. It is the apparent role of properdin to confer a degree of stability upon the labile enzyme and to protect its C3 convertase function against enzymatic destruction. To achieve these effects, precursor properdin (pre-P) is recruited in a binding-activation reaction by the labile C3/C5 convertase. Multiple C3b molecules appear to be needed for the formation of properdin-activating principle. Three modes of regulation have been described, which involve spontaneous dissociation enzymatic degradation by C3b inactivator and disassembly by beta1H. The functional differences of pre-P and activated properdin (P) were delineated, pre-P displaying a weak affinity for C3b and P the capacity of strong interaction, P generating a soluble C3 convertase in serum and pre-P being unable to do so. Because of the profound differences between native pre-P and the laboratory product P, the question was raised as to whether soluble P represents an unphysiological form of the protein. On the basis of this and other studies, the conclusion was reached that in vitro properdin recruitment constitutes the terminal event of the properdin pathway, and that properdin augments the function of C3/C5 convertase without changing its substrate specificity.     

Simulation of the AII amacrine cell of mammalian retina: functional consequences of electrical coupling and regenerative membrane properties

The AII amacrine cell of mammalian retina collects signals from several hundred rods and is hypothesized to transmit quantal "single-photon" signals at scotopic (starlight) intensities. One problem for this theory is that the quantal signal from one rod when summed with noise from neighboring rods would be lost if some mechanism did not exist for removing the noise. Several features of the AII might together accomplish such a noise removal operation: The AII is interconnected into a syncytial network by gap junctions, suggesting a noise-averaging function, and a quantal signal from one rod appears in five AII cells due to anatomical divergence. Furthermore, the AII contains voltage-gated Na+ and K+ channels and fires slow action potentials in vitro, suggesting that it could selectively amplify quantal photon signals embedded in uncorrelated noise. To test this hypothesis, we simulated a square array of AII somas (Rm = 25,000 Ohm-cm2) interconnected by gap junctions using a compartmental model. Simulated noisy inputs to the AII produced noise (3.5 mV) uncorrelated between adjacent cells, and a gap junction conductance of 200 pS reduced the noise by a factor of 2.5, consistent with theory. Voltage-gated Na+ and K+ channels (Na+: 4 nS, K+: 0.4 nS) produced slow action potentials similar to those found in vitro in the presence of noise. For a narrow range of Na+ and coupling conductance, quantal photon events (approximately 5-10 mV) were amplified nonlinearly by subthreshold regenerative events in the presence of noise. A lower coupling conductance produced spurious action potentials, and a greater conductance reduced amplification. Since the presence of noise in the weakly coupled circuit readily initiates action potentials that tend to spread throughout the AII network, we speculate that this tendency might be controlled in a negative feedback loop by up-modulating coupling or other synaptic conductances in response to spiking activity.     

EEA1, an early endosome-associated protein. EEA1 is a conserved alpha-helical peripheral membrane protein flanked by cysteine "fingers" and contains a calmodulin-binding IQ motif

Early endosomes are cellular compartments receiving endocytosed material and sorting them for vesicular transport to late endosomes and lysosomes or for recycling to the plasma membrane. We have cloned a human cDNA encoding an evolutionarily conserved 180-kDa protein on early endosomes named EEA1 (Early Endosome Antigen1). EEA1 is associated with early endosomes since it co-localizes by immunofluorescence with the transferrin receptor and with Rab5 but not with Rab7. Immunoelectron microscopy shows that it is associated with tubulovesicular early endosomes containing internalized bovine serum albumin-gold. EEA1 is a hydrophilic peripheral membrane protein present in cytosol and membrane fractions. It partitions in the aqueous phase after Triton X-114 solubilization and is extracted from membranes by 0.3 M NaCl. It is a predominantly alpha-helical protein sharing 17-20% sequence identity with the myosins and contains a calmodulin-binding IQ motif. It is flanked by metal-binding, cysteine "finger" motifs. The COOH-terminal fingers, Cys-X2-Cys-X12-Cys-X2-Cys and Cys-X2-Cys-X16-Cys-X2-Cys, are present within a region that is strikingly homologous with Saccharomyces cerevisiae FAB1 protein required for endocytosis and with Caenorhabditis elegans ZK632. These fingers also show limited conservation with S. cerevisiae VAC1, Vps11, and Vps18p proteins implicated in vacuolar transport. We propose that EEA1 is required for vesicular transport of proteins through early endosomes and that its finger motifs are required for this activity.     

Kinetics of nonenzymatic glycation of ribonuclease A leading to advanced glycation end products. Paradoxical inhibition by ribose leads to facile isolation of protein intermediate for rapid post-Amadori studies

Nonenzymatic glycation (Maillard reaction) of long-lived proteins is a major contributor to the pathology of diabetes and possibly aging and Alzheimer's disease. We report here kinetic studies of the glycation of the model protein ribonuclease A by glucose and ribose leading to the formation of antigenic advanced glycation end products ("AGEs"), detectable by AGE-specific polyclonal antibodies, and pentosidine, an acid-stable fluorescent AGE. As anticipated, the kinetics of glycation by ribose were considerably faster than by glucose, and the rate of AGE formation initially increased with increasing sugar concentrations. However, ribose above 0.15 M appeared to paradoxically slow the kinetics of AGE formation, suggesting ribose inhibits the conversion of "early" Amadori rearrangement products to "late" AGEs and thus favors the accumulation of reactive Amadori intermediates. The facile isolation of such protein intermediates was achieved by an "interrupted glycation" protocol which free and reversibly bound (Schiff base) ribose was removed following a short (24h) initial incubation of 0.5 M ribose at 37 degrees C. The kinetics of buildup of the Amadori intermediates and the kinetics of their post-Amadori conversion to antigenic AGEs were independently studied. A rapid and reversible inhibition of the post-Amadori kinetics by free ribose was verified by direct re-addition of ribose to the isolated, sugar-free intermediate. The pH dependence of the kinetics of antigenic AGE formation from such intermediates was measured and exhibited an unusual bell-shaped profile over the pH range of 5.0-9.5 with a maximum near pH 8.0. Aminoguanidine, a pharmacological AGE inhibitor, was found to moderately or weakly inhibit antigenic AGE formation in such post- Amadori steps. The isolation of the glycated ribonuclease intermediate thus simplifies kinetic and mechanistic studies of AGE formation, permits AGE studies in the absence of complications arising from free or Schiff base bound sugar, and provides a novel methodology for evaluating the mechanism and efficacy of therapeutic agents that may inhibit AGE formation.     

What is the psychiatrist's role in drugs and alcohol?

OBJECTIVE: This article describes a consensus view of the role of psychiatrists in respect of alcohol and other drug (AOD) problems, in response to the view expressed by Wodak [1]. METHOD: The data were selected on the basis of the knowledge and experience of the authors. RESULTS: Psychiatrists have made major contributions in the primary, secondary and tertiary prevention of AOD problems over many years in Australia and New Zealand. In recent years there has been an explosion of new knowledge in the AOD area and a shift from mental health to primary and public health care for these patients. Substance use disorders (SUD) are highly prevalent in all areas of psychiatric practice, requiring treatment in their own right as well as complicating the treatment of coexisting psychiatric illness. CONCLUSION: It is argued that psychiatrists have important roles in harm reduction, prevention and policy development; brief and early intervention in SUD in liaison and child psychiatry; and systematic treatment for those with dependence and other psychiatric comorbidity. A research and collaborative approach to AOD services and patients should be encouraged, rather than engaging in divisive debate over "ownership' of this area of clinical practice.     

Intron sequences are involved in the plastid- and light-dependent expression of the spinach PsaD gene

OBJECTIVE: To obtain preliminary information about the safety and efficacy of intravenous immune globulin (IVIG: Iveegam, Immuno AG, Vienna) in the treatment of polyarticular juvenile rheumatoid arthritis (poly-JRA) resistant to other forms therapy. METHODS: We used a multicentered, phase I/II blinded-withdrawal design with stratified entry. All patients began by receiving open infusions of IVIG at a dose between 1.5 and 2.0 g/kg/infusion (100 g maximum) bimonthly for the first 2 months, then monthly for up to 6 months. Beginning at Month 3, those who met the criteria for "clinically important improvement" were randomized to receive monthly infusions for 4 months of either placebo or IVIG in a double blind (DB) phase. Patients were permitted nonsteroidal antiinflammatory drugs, slow acting antirheumatic drugs, and low dose (< 10 mg/day) prednisone at constant doses. An "early escape" provision in the DB allowed those who showed "clinically important worsening" to again receive IVIG (if taking placebo) or a higher dose of IVIG (if taking the lower dose of IVIG). RESULTS: Efficacy. Twenty-five children entered the trial and 19 (76%) met the criteria for "clinically important improvement" during the open phase (OP) and entered the DB. Three patients completed the OP but failed to meet the criteria for response, and 3 patients dropped out of the OP, none of whom showed benefit from IVIG. Treatment effect sizes produced by IVIG were moderate to large for all variables in the OP. Patients who continued IVIG in the DB continued to show improvement over that achieved in the OP. Those given placebo showed a rapid loss of efficacy, suggesting IVIG has a limited duration of effect after discontinuation. Safety. No patient developed serious or unexpected adverse side effects in the open or DB phases, and none dropped out of the study due to toxicity or side effects. CONCLUSION: Substantial clinical improvement from IVIG is produced in about three-fourths of patients with poly-JRA during open administration, but the duration of the beneficial effect is short after discontinuation. Those with disease < 3 years' duration may be more likely to respond than those who have had their disease for > 5 years. Short term safety is excellent.     

Preliminary report of data collection in hospital quality assurance programs--a project of the Association of Hospitals of the Czech Republic

OBJECTIVES: This was the first attempt of the association representing all acute care hospitals in the Czech Republic to collect mutual data which might be used for quality assurance (QA) purposes and which might lead to the development of national standards of care which could be used for hospital accreditation. Data collected included information which was available universally and which could be measured; in addition, information was intended to be similar in each hospital. In most cases, the data collection systems were based on financial information and data had to be identified which might be used for QA purposes, rather than being able to design a system specific for QA purposes. DESIGN: Since the hospital payment system was established in 1992, hospitals have had to develop data collection systems to measure clinical activity; this current study was based on this data collection, adapted to QA purposes. SETTING: The Executive Committee of the Hospital Association agreed to a pilot study of hospitals in 1993; data were collected from approximately 40 hospitals, beginning in 1994. STUDY PARTICIPANTS: Hospitals were chosen based on their ability to collect data and participate in the program, and it was determined that there should be variability in the hospitals, in size, location and activities, but that the data collected should be generic. INTERVENTIONS: Raw data included 33 different items, most of which were irrelevant to QA. Using a computer program, various combinations of data were reviewed and evaluated to ascertain the most appropriate for QA purposes. MAIN OUTCOME MEASURES: Data were chosen for study which included (a) data from the largest departments in the individual hospitals; (b) length of stay for patients hospitalized in these departments; (c) number of occupied beds/physician in the department and (d) mortality/1000 admissions to the department. RESULTS: The combination of (1) a long length of stay; (2) a high occupied bed/doctor ratio; and (3) a high mortality rate/1000 admissions might be indicators of poor quality. Additional factors to consider include: the type of department-emergency, cancer, geriatric, etc.; the nature of the medical activity-acute, referral, primary care, etc.; whether or not "social" beds are included and, generally, comparability among departments. However, as a pilot study, certain indicators can be determined which then can be used for future study to determine quality of care. The ability to cooperate and collect seemingly comparable data indicates reason for optimism in the future; more detailed and accurate studies can be carried out which will enable assessment of the quality of care given in comparable situations in hospitals throughout the Czech Republic.