Isoproterenol evokes extracellular Ca2+ spikes due to secretory events in salivary gland cells

Secretory cells should in principle export substantial amounts of calcium via exocytosis since Ca2+ is sequestered in secretory granules. Based on a new technique for measurements of the extracellular calcium concentration in the vicinity of the cell membrane and on the droplet technique, we have monitored the rate of calcium extrusion from salivary gland acinar cells. Isoproterenol (ISP), a beta-adrenergic agonist and powerful secretogogue, evoked no change in the cytosolic free Ca2+ concentration ([Ca2+]i) but induced vigorous extracellular Ca2+ concentration ([Ca2+]i) spiking. The absence of [Ca2+]i elevation and the pulsatile nature of the changes in [Ca2+]i indicate that these spikes are most likely due to calcium release from secretory granules. The cholinergic agonist acetylcholine (ACh), which induces moderate secretion, evoked a marked rise in [Ca2+]i and a smooth rise in [Ca2+]i, most likely induced by plasma membrane calcium pumps, on which shortlasting [Ca2+]i spikes were superimposed. The rate of ISP-induced calcium efflux was very substantial. The calculated calcium loss during the first 100 s of supramaximal stimulation corresponded to a reduction of the total cellular calcium concentration of approximately 0.4 mM. We conclude that in salivary glands, calcium release via exocytosis is one of the main mechanisms extruding calcium from cells to the extracellular milieu.     

Influence of the major histocompatibility complex on age at onset of chronic lymphoid leukaemia

Goodpasture syndrome is an often fatal autoimmune disease associated with glomerulonephritis and/or pulmonary hemorrhage. The clinical manifestations of this disease correlate well with the presence of circulating antiglomerular basement membrane (GBM) autoantibodies. The primary target antigen in glomerular and alveolar basement membranes is thought to be the alpha 3 chain of type IV collagen. Nearly all that is known about anti-GBM antibodies in humans comes from work on unbound circulating antibody. We recently had the unique and rare opportunity to obtain early postmortem antibody and tissues from a patient who died with catastrophic Goodpasture syndrome. The specificity of circulating, kidney-bound and lung-bound autoantibodies from this patient was evaluated against a variety of purified basement membrane constituents. The results indicate that the primary target for the circulating and tissue-bound autoantibodies is the NC1 domain of the alpha 3(IV) chain of type IV collagen. Additionally, all the antibodies recognize a cryptic epitope/s on the alpha 3(IV)NC1 hexamer. Furthermore, tissue-bound and circulating antibodies compete with one another for overlapping epitopes on the antigen. These findings demonstrate that circulating autoantibodies in Goodpasture syndrome are highly representative of those bound to organ tissues, strengthening the notion that pathogenic autoantibodies are targeted to the alpha 3(IV)NC1 collagen, and that previous reports of findings in the circulation may be applicable to tissue injury.     

Evidence for Remote Effects of Dreissenid Mussels on the Amphipod Diporeia: Analysis of Lake Ontario Benthic Surveys, 1972–2003

The status of invasive dreissenid mussels (Dreissena polymorpha and D. bugensis) and native amphipods (Diporeia spp.) in Lake Ontario was assessed in 2003 and compared with historical data. D. polymorpha (zebra mussels) were rarely observed in 2003, having been displaced by D. bugensis (quagga mussels). D. bugensis expanded its depth range from 38 m depth in 1995 to 174 m in 2003 and this dreissenid reached densities averaging 8,000/m² at all sites < 90 m. During the same time period, Diporeia populations almost completely disappeared from 0–90 m depth, continuing a declining trend from 1994–1997 reported in previous studies. The average density of Diporeia in the 30–90 m depth interval decreased from 1,380/m² to 63/m² between 1997 and 2003. Prior to 2003, areas deeper than 90 m represented a refuge for Diporeia, but even these deep populations decreased, with densities declining from 2,181/m2 in 1999 to 545/m2 in 2003. Two common hypotheses for the decline of Diporeia in the Great Lakes are food limitation and a toxin/pathogen associated with dreissenid pseudofeces. The Diporeia decline in deep waters preceded the expansion of D. bugensis to these depths, and suggests that shallow dreissenid populations remotely influence profundal habitats. This pattern of decline is consistent with mechanisms that act from some distance including nearshore dreissenid grazing and downslope transport of pseudofeces.     

Detection of optic disc changes with Glaucoma-Scope probability maps

Before use of cardiovascular surgical techniques and procedures in humans, many experiments, e.g., hypothermic circulatory arrest and cardiopulmonary bypass using the heart-lung machine, have been performed in the dog. As a consequence experimental canine cardiovascular surgery is highly developed. This has not resulted in the routine performance of open heart surgery in veterinary medicine, probably because of the high costs. Cardiovascular surgery in the dog is generally limited to interventions not depending on hypothermic circulatory arrest or cardiopulmonary bypass. The clinical cardiovascular surgery in dogs can be divided into routine and more specialized interventions. The first category includes correction of peritoneopericardial diaphragmatic hernia, pericardial fenestration in dogs with pericardial effusion, treatment of persistent right aortic arch, and patent ductus closure. The specialized interventions include dilation of pulmonic and aortic stenoses and pacemaker implantation. The diagnosis and surgical treatment of such diseases is described. New developments in cardiovascular surgical treatment that can be expected include catheter techniques for occlusion of shunts and dilations using balloons, because the financial costs of these procedures are not prohibitive.     

Fast fracture of rubber-toughened thermoplastics used for the shells of motorcycle helmets

The effects of outdoor ageing and of accelerated UV exposure on the grade of acrylonitrile butadiene styrene copolymer (ABS) used by UK manufacturers of motorcycle helmets were compared. Although the surface layer becomes embrittled the helmets still survive an impact test after conditioning at -20° C because the cracks are arrested before they penetrate the full thickness of the shell. A 4 mm thick rubber-toughened polycarbonate, used in helmets from continental suppliers, shows a transition from tough to brittle crack propagation behaviour at -15° C. For 4 mm thick ABS this transition had not occurred at the lowest test temperature of -20° C, but if 6 mm sheet is tested the transition is at -15° C. Temperature increases in the 40 sec delay between conditioning a helmet at -20° C and impact testing are shown to be significant.     

The Role of Rigid Residues in Modulating TEM-1 β-Lactamase Function and Thermostability

The relationship between protein motions (i.e., dynamics) and enzymatic function has begun to be explored in β-lactamases as a way to advance our understanding of these proteins. In a recent study, we analyzed the dynamic profiles of TEM-1 (a ubiquitous class A β-lactamase) and several ancestrally reconstructed homologues. A chief finding of this work was that rigid residues that were allosterically coupled to the active site appeared to have profound effects on enzyme function, even when separated from the active site by many angstroms. In the present work, our aim was to further explore the implications of protein dynamics on β-lactamase function by altering the dynamic profile of TEM-1 using computational protein design methods. The Rosetta software suite was used to mutate amino acids surrounding either rigid residues that are highly coupled to the active site or to flexible residues with no apparent communication with the active site. Experimental characterization of ten designed proteins indicated that alteration of residues surrounding rigid, highly coupled residues, substantially affected both enzymatic activity and stability; in contrast, native-like activities and stabilities were maintained when flexible, uncoupled residues, were targeted. Our results provide additional insight into the structure-function relationship present in the TEM family of β-lactamases. Furthermore, the integration of computational protein design methods with analyses of protein dynamics represents a general approach that could be used to extend our understanding of the relationship between dynamics and function in other enzyme classes.     

Solidification and crystallization properties of CaO-SiO2-Na2O based mold fluxes

Crystallization properties play an important role in keeping a smooth running of continuous casting process and high sur-face quality of cast strands. To reduce fluorine pollution in slag, a new type of CaO-SiO2-Na2O (CSN) based mold flux was studied. The solidification and crystallization properties, including crystallization temperature, crystallization ratio and solidification minera-graphy, were measured, which were compared with the CaO-SiO2-CaF2 (GF) mold flux. The results show that the crystallization performance is equal to the high fluoride mold powder and CSN can be used for peritectic steel grades sensitive to longitudinal cracking in continuous casting.     

The use of electrochemical noise methods (ENM) to study thick, high impedance coatings

Thick, high impedance organic coatings are those class of coatings used to provide corrosion protection to naval vessels, pipelines, gasoline storage tanks, and other large structures such as bridges and plant structures. These coatings, especially the newest generations now being used in practice, can provide exceptional protection and lifetime of performance such that properly and accurately assessing and differentiating among competing coatings is a very difficult task. The standard protocol of salt fog testing (ASTM B117), immersion testing, and outdoor exposure in a corrosive environment with subjective evaluation of a coating's performance durings and after testing, does not adequatcly rank and predict coating lifetimes for new coating systems, especially for the environmentally compliant coating systems such as powder coatings (especially the thick, fusion bonded epoxy (FBE) coatings used for pipelines), two component epoxy and urethane coatings and waterborne coatings. New, objective test methods are desperately needed by users and manufacturers of coatings. A relatively new electrochemical test procedure, electrochemical noise methods (ENM), as developed by Skerry and Eden, has been shown in our laboratory to be very successful in the ranking and prediction of relative coating performance. We have used the method successfully on naval ship coatings, several pipeline coatings and other related systems, and Skerry has used them successfully on industrial maintenance coatings. We have used these methods in conjunction with electrochemical impedance spectroscopy, d.c. resistance measurements and cyclic salt fog testing of the Prohesion^TM type. In our studies of pipeline coatings, we needed to investigate thermal effects because of their extended range of use temperature. In these studies, we have discovered that electrochemical methods can be used for an in situ measurement of the T_g of coatings in electrolyte immersion. Further, the ‘plasticizing’ effect of aqueous electrolyte absorption as well as its relative irreversibility has been shown. For all coatings studied, ENM provided useful, objective, numerical data which rapidly ranks coatings and provides useful information on the relative lifetime prediction of coatings which may provide up to 30 years of service.