Type A botulinum toxin disorganizes quantal acetylcholine release and inhibits energy metabolism

The physiological, morphological and biochemical effects of type A Botulinum toxin (BoTX) were analysed in the electric organ of Torpedo, a modified neuromuscular system. The quantal content of the postsynaptic potential, or electroplaque potential (EPP), was reduced by BoTX but the quantum size remained unchanged till complete failure of the neurally evoked transmission. BoTX also suppressed the occurrence of spontaneous electroplaque potentials (MEPPs) of a quantal size but potentials of a smaller amplitude still kept on occurring in the intoxicated synapses. BoTX inhibited the evoked release of acetylcholine (ACh; biochemically measured) but the rate of spontaneous ACh release transiently increased during the period when evoked release went down. On the other hand, there were no significant change of ACh content, of ACh turnover, of ACh repartition in the vesicular and free compartments, or in the number of synaptic vesicles. Surprisingly, the amount of ATP was reduced to 50% in BoTX treated tissue at the time of transmission failure; also the level of creatine phosphate (CrP) was lowered to less than 20% and the rate of activity of creatine kinase was reduced. It was concluded that, electrophysiologically, BoTX affects synaptic transmission in a very similar way in the electric organ and in the neuromuscular junctions. On the other hand, the shortage of ATP supply found in the present study may play a role in the pathophysiology of intoxication and should be taken into account in investigations designed to see whether BoTX affects various phosphorylations in cholinergic nerve terminals

Association between low empathy and high burnout among primary care physicians and nurses in Lleida, Spain

Altres ajuts: Sociedad Espanola de Medicina Familiar y Comunitaria ~ [3000 e]Background: Burnout is a growing problem among healthcare professionals and may be mitigated and even prevented by measures designed to promote empathy and resilience. Objectives: We studied the association between burnout and empathy in primary care practitioners in Lleida, Spain and investigated possible differences according to age, sex, profession, and place of practice (urban versus rural). Methods: All general practitioners (GPs) and family nurses in the health district of Lleida (population 366 000) were asked by email to anonymously complete the Maslach Burnout Inventory (MBI) and the Jefferson Scale of Physician Empathy (JSPE) between May and July 2014. Tool consistency was evaluated by Cronbach's α, the association between empathy and burnout by Spearman's correlation coefficient, and the association between burnout and empathy and sociodemographic variables by the χ 2 test. Results: One hundred and thirty-six GPs and 131 nurses (52.7% response rate) from six urban and 16 rural practices participated (78.3% women); 33.3% of respondents had low empathy, while 3.7% had high burnout. The MBI and JSPE were correlated (P <.001) and low burnout was associated with high empathy (P <.05). Age and sex had no influence on burnout or empathy. Conclusion: Although burnout was relatively uncommon in our sample, it was associated with low levels of empathy. This finding and our observation of lower empathy levels in rural settings require further investigation

Drp1 Mediates Caspase-Independent Type III Cell Death in Normal and Leukemic Cells▿ †

Ligation of CD47 triggers caspase-independent programmed cell death (PCD) in normal and leukemic cells. Here, we characterize the morphological and biochemical features of this type of death and show that it displays the hallmarks of type III PCD. A molecular and biochemical approach has led us to identify a key mediator of this type of death, dynamin-related protein 1 (Drp1). CD47 ligation induces Drp1 translocation from cytosol to mitochondria, a process controlled by chymotrypsin-like serine proteases. Once in mitochondria, Drp1 provokes an impairment of the mitochondrial electron transport chain, which results in dissipation of mitochondrial transmembrane potential, reactive oxygen species generation, and a drop in ATP levels. Surprisingly, neither the activation of the most representative proapoptotic members of the Bcl-2 family, such as Bax or Bak, nor the release of apoptogenic proteins AIF (apoptosis-inducing factor), cytochrome c, endonuclease G (EndoG), Omi/HtrA2, or Smac/DIABLO from mitochondria to cytosol is observed. Responsiveness of cells to CD47 ligation increases following Drp1 overexpression, while Drp1 downregulation confers resistance to CD47-mediated death. Importantly, in B-cell chronic lymphocytic leukemia cells, mRNA levels of Drp1 strongly correlate with death sensitivity. Thus, this previously unknown mechanism controlling caspase-independent type III PCD may provide the basis for novel therapeutic approaches to overcome apoptotic avoidance in malignant cells

Caspase-independent type III programmed cell death in chronic lymphocytic leukemia: the key role of the F-actin cytoskeleton

Stimulation of the CD47 membrane receptor activates an original mechanism of cell death in chronic lymphocytic leukemia (CLL) cells that is caspase-independent. This study provides an exhaustive characterization of the CD47 anti-tumor signaling in CLL that may help to define new targets for further therapeutic strategies

Production of an anti-Aβ antibody fragment in Pichia pastoris and in vitro and in vivo validation of its therapeutic effect

ScFv-h3D6 has been shown as an efficient therapy in the 3xTg-AD mouse model of Alzheimer's Disease. Because one of the major bottlenecks for the therapeutic uses of proteins produced in Escherichia coli is their potential contamination with endotoxins, LPS were extensively removed by a rather low-efficient, expensive, and time-consuming purification step. In addition, disulfide scrambling is favored in the reducing bacterial cytoplasm albeit the use of reductase deficient strains. To overcome these hurdles, as well as to improve the yield, the yeast Pichia pastoris, an endotoxin-free host system for recombinant protein production, has been used to produce scFv-h3D6, both in flask and in a fed-batch bioreactor. Comparison of the thermal stability of the obtained protein with that from E. coli showed no differences. Opposite to the case of the protein obtained from E. coli, no disulfide scrambled conformations or LPS traces were detected in that produced in P. pastoris. Cytotoxicity assays in SH-SY5Y neuroblastoma cell-cultures demonstrated that proteins from both expression systems were similarly efficient in precluding Aβ-induced toxicity. Finally, the 3xTg-AD mouse model was used to test the therapeutic effect of both proteins. Quantification of Aβ levels from cortex and hippocampus protein extracts by ELISA, and Aβ-immunohistochemistry, showed that both proteins reduced Aβ burden. This work demonstrates that scFv-h3D6 obtained from P. pastoris shows the same benefits as those already known for that obtained from E. coli, with multiple advantages in terms of recombinant production and safety