Identification of in vitro phosphorylation sites in the growth cone protein SCG10. Effect Of phosphorylation site mutants on microtubule-destabilizing activity

SCG10 is a neuron-specific, membrane-associated protein that is highly concentrated in growth cones of developing neurons. Previous studies have suggested that it is a regulator of microtubule dynamics and that it may influence microtubule polymerization in growth cones. Here, we demonstrate that in vivo, SCG10 exists in both phosphorylated and unphosphorylated forms. By two-dimensional gel electrophoresis, two phosphoisoforms were detected in neonatal rat brain. Using in vitro phosphorylated recombinant protein, four phosphorylation sites were identified in the SCG10 sequence. Ser-50 and Ser-97 were the target sites for protein kinase A, Ser-62 and Ser-73 for mitogen-activated protein kinase and Ser-73 for cyclin-dependent kinase. We also show that overexpression of SCG10 induces a disruption of the microtubule network in COS-7 cells. By expressing different phosphorylation site mutants, we have dissected the roles of the individual phosphorylation sites in regulating its microtubule-destabilizing activity. We show that nonphosphorylatable mutants have increased activity, whereas mutants in which phosphorylation is mimicked by serine-to-aspartate substitutions have decreased activity. These data suggest that the microtubule-destabilizing activity of SCG10 is regulated by phosphorylation, and that SCG10 may link signal transduction of growth or guidance cues involving serine/threonine protein kinases to alterations of microtubule dynamics in the growth cone.     

Magnetic resonance of the encephalon in 17 patients with ocular Beh?et''s disease

OBJECTIVE: To evaluate the relationships between patient and physician pretreatment expectations of pain relief and subsequent pain relief reported by chronic pain patients immediately after treatment. DESIGN: Prospective study of consecutive patients undergoing a procedure in a pain clinic for treatment of chronic pain. Patients rated their current pain level and their expectation of pain relief immediately prior to undergoing a procedure (e.g., intravenous drug infusion, nerve block) for the treatment of chronic pain. Simultaneously and independently, the treating physician completed a similar questionnaire. At completion of the procedure, patients rated their current pain level and degree of pain relief. SETTING: University of Washington Multidisciplinary Pain Center procedure suite. PATIENTS: Forty-six consecutive chronic pain patients. INTERVENTION: Intravenous drug infusions and nerve blocks. OUTCOME MEASURES: Current pain and pain relief ratings. RESULTS: Patients' pain relief expectation ratings were not correlated significantly with their postprocedure pain relief ratings or pre-post procedure changes in pain ratings. However, a statistically significant correlation was found between physician expectations of pain relief and patient pain relief ratings and patient pre-post procedure changes in pain. CONCLUSIONS: The results of this study suggest that physicians are better predictors than are patients of patients responses to these procedures and/or that physicians may somehow subtly communicate their expectations to patients during the procedure, and these expectations then influence patient response. Patient pretreatment expectations may not always play a significant role in nonspecific treatment effects.     

Snowboard traumatology: an epidemiological study

OBJECTIVE: To estimate the frequency of perioperative morbidities in patients who underwent anesthesia and a surgical procedure with no preoperative laboratory testing. MATERIAL AND METHODS: We conducted an electronic database search of medical records of 56,119 patients who underwent surgical or diagnostic procedures and anesthesia at Mayo Clinic Rochester in 1994 and found 5,120 who had no laboratory tests done within 90 days before the procedure. From this group, we randomly selected 1,044 patients (87 from each month) to document the absence of preoperative tests, the presence of preexisting disease (by organ system), the type of anesthetic agent, and the outcomes and tests intraoperatively and postoperatively. RESULTS: The 1,044 patients ranged in age from 0 to 95 years (median age, 21). No deaths or major perioperative morbidities occurred (0.0%; exact 95% confidence interval, 0.00 to 0.35%). Although 10 patients underwent blood typing and screening for antibodies immediately preoperatively, no blood transfusions were necessary. Intraoperatively, 17 laboratory tests and 1 electrocardiogram were obtained, and 3 results were abnormal. Postoperatively, 42 blood tests and 2 electrocardiographic procedures were performed. Five of the 42 blood tests showed abnormal results (hemoglobin levels in 3, serum sodium in 1, and arterial blood gases in 1). One electrocardiogram showed normal findings, and the other revealed normal results except for premature ventricular contractions. No laboratory test done intraoperatively or postoperatively was found to change surgical or medical management substantially. One patient who had unanticipated blood loss during an outpatient procedure was admitted to the hospital for observation. CONCLUSION: All 1,044 patients, 97% of whom were relatively healthy, with no recent laboratory testing safely underwent anesthesia and an operation. We conclude that patients who have been assessed by history and physical examination and determined to have no preoperative indication for laboratory tests can safely undergo anesthesia and operation with tests obtained only as indicated intraoperatively and post-operatively. Current anesthetic and medical practices rapidly identify perioperative indications for laboratory evaluation as they arise.     

Inorganic–organic hybrid materials with zirconium oxoclusters as protective coatings on aluminium alloys

Inorganic–organic hybrid materials are attracting a strong scientific interest mainly for their outstanding inherent mechanical and thermal properties, which can be traced back to the intimate coupling of both inorganic and organic components. By carefully choosing the experimental parameters used for their synthesis, chemically and thermally stable acrylate-based hybrid material embedding the zirconium oxocluster Zr₄O₄(OMc)₁₂, where OMcCH₂C(CH₃)C(O)O, can be deposited as UV-cured films on aluminium alloys.

In particular, the molar ratios between the oxocluster and the monomer, the polymerisation time, the amount of photo-initiator and the deposition conditions, by using an home-made spray-coating equipment, were optimised in order to obtain the best performing layers in terms of transparency and hardness to coat aluminium alloy (AA1050, AA6060 and AA2024) sheets. Furthermore, it was also evaluated whether the hybrid coatings behave as barrier to corrosion.

Several coated samples were prepared and characterised. Environmental scanning electronic microscopy (ESEM) and scratch test were used to investigate the morphology of the films and to evaluate their scratch resistance, respectively. Electrochemical impedance spectroscopy (EIS) was performed in order to evaluate if the coatings actually protect the metallic substrate from corrosion.

In order to measure shear storage modulus (G′) and loss modulus (G″) of the materials used for coatings, bulk samples were also obtained by UV-curing of the precursors solution. Dynamical mechanical thermal analysis (DMTA) was performed in shear mode on cured disks of both the hybrid materials and pristine polymer for comparison. The values of T_g were read off as the temperatures of peak of loss modulus. The length and mass of all the samples were measured before and after the DMTA analysis, so that the shrinkage of the materials in that temperature range was exactly evaluated.     

Effect of Bonder at Skin/Core Interface on the Mechanical Performances of Sandwich Structures Used in Marine Industry

The present work is aimed to extend the knowledge of mechanical properties of sandwich structures used for marine applications focusing on the possibility to increase the performances of such structures by adding a bonder at the skin/core interface. Therefore, three sandwich structures that are utilised in different structural components of a yacht were realised by manual lay-up. The mechanical characterisation was performed by flatwise compressive, edgewise compressive and three point flexural tests. The tests execution has allowed both to determine the mechanical performances and to understand the fracture mechanisms that take place when the bonder is added in the stacking sequence of the samples.     

Non-pinched, minimum energy distillation designs

Non-pinched, minimum energy solutions are important class of distillation designs that offer the potential advantage of a better trade-off between capital investment and operating costs. In this paper, two important tasks associated with non-pinched distillation designs are studied. Thus the novel contributions of this work to the literature are

(1) A comprehensive methodology for finding non-pinched minimum energy designs.

(2) Understanding of the reasons for the existence of non-pinched distillation designs.

It is shown that the recent shortest stripping line distance approach of Lucia et al. [Lucia, A., Amale, A. and Taylor, R., 2007, Distillation pinch points and more. Comput Chem Eng, available on-line] is capable of systematically and reliably finding non-pinched, minimum energy distillation designs. In addition, we provide an understanding of the reasons behind the existence of non-pinched designs, which include trajectories that follow unstable branches of a pinch point curve in azeotropic systems, the inherent looping structure of trajectories in hydrocarbon separations, and the presence of ancillary constraints in multi-unit processes like extraction/distillation. Several distillation examples are studied and many numerical results and geometric illustrations are presented that show the shortest stripping line distance methodology is indeed a powerful and systematic tool for computing non-pinched, minimum energy designs and that support the underlying reason we provide for the existence of non-pinched designs.     

Dopamine D2 receptor signaling via the arachidonic acid cascade: modulation by cAMP-dependent protein kinase A and prostaglandin E2

Recent studies have shown that, in Chinese hamster ovary cells transfected with D2-receptor cDNA, CHO(D2) cells, D2 agonists are potent in enhancing the release of [3H]arachidonic acid (AA) induced by stimulation of constitutive purinergic receptors or by application of Ca2+ ionophores. This facilitatory action is further amplified by the concomitant activation of D1 receptors, which per se have no effect on evoked [3H]AA release. Here, we review a series of experiments aimed at examining the molecular mechanism of this synergistic interaction. The results show that, in CHO(D2) cells: (a) application of 8-Br-cAMP or stimulation of constitutive prostaglandin (PG)E2 receptors augment the AA response produced by D2 agonists; (b) in CHO(D2) cells transfected with human beta 2-receptor cDNA, the beta-agonist, isoproterenol, produces a similar effect; (c) the potentiation of [3H]AA release produced by PGE2 and 8-Br-cAMP is prevented by overexpressing either a protein inhibitor of cAMP-dependent protein kinase (PKA) or a mutated form of pKA regulatory subunit incapable of binding cAMP; (d) mock-synergism is obtained in CHO(D2) cells overexpressing the catalytic subunit of PKA; (e) PGE2 is a major AA metabolite in stimulated CHO(D2) cells and its formation may contribute to the effect of D2 agonists on AA release. The results indicate that cAMP-induced activation of PKA represents a likely molecular basis for D1/D2 receptor synergism on AA release. They also suggest that additional membrane receptors, colocalized with D2 and positively linked to adenylyl cyclase, may exert a similar action. Furthermore, stimulation of PGE2 receptors by endogenously produced prostaglandin may participate in AA signaling at the D2 receptor, by providing a paracrine positive feedback loop.     

Novel diterpenoid diacylglycerols from marine molluscs: potent morphogens and protein kinase C activators

Five novel 1,2-sn-diacylglycerols with diterpenoid acyl moieties in the sn-1 position were isolated and characterized, together with the corresponding 1,3-sn-diacylglycerols, from three species of dorid nudibranchs molluscs. Their potent activity as morphogens in vivo in the Hydra tentacle regeneration assay and their parallel activity as activators of rat brain protein kinase C (PKC) in vitro are reported here. Our findings promote the use of these compounds as useful molecular probes for both in vivo and in vitro studies on the participation of PKC in cell development.