Cathepsin L1, the major protease involved in liver fluke (Fasciola hepatica) virulence.

The secretion and activation of the major cathepsin L1 cysteine protease involved in the virulence of the helminth pathogen Fasciola hepatica was investigated. Only the fully processed and active mature enzyme can be detected in medium in which adult F. hepatica are cultured. However, immunocytochemical studies revealed that the inactive procathepsin L1 is packaged in secretory vesicles of epithelial cells that line the parasite gut. These observations suggest that processing and activation of procathepsin L1 occurs following secretion from these cells into the acidic gut lumen. Expression of the 37-kDa procathepsin L1 in Pichia pastoris showed that an intermolecular processing event within a conserved GXNXFXD motif in the propeptide generates an active 30-kDa intermediate form. Further activation of the enzyme was initiated by decreasing the pH to 5.0 and involved the progressive processing of the 37 and 30-kDa forms to other intermediates and finally to a fully mature 24.5 kDa cathepsin L with an additional 1 or 2 amino acids. An active site mutant procathepsin L, constructed by replacing the Cys26 with Gly26, failed to autoprocess. However, [Gly26]procathepsin L was processed by exogenous wild-type cathepsin L to a mature enzyme plus 10 amino acids attached to the N terminus. This exogenous processing occurred without the formation of a 30-kDa intermediate form. The results indicate that activation of procathepsin L1 by removal of the propeptide can occur by different pathways, and that this takes place within the parasite gut where the protease functions in food digestion and from where it is liberated as an active enzyme for additional extracorporeal roles.

Fasciola hepatica cathepsin L-like proteases: biology, function and potential in the development of first generation liver fluke vaccines.

Fasciola hepatica secretes cathepsin L proteases that facilitate the penetration of the parasite through the tissues of its host, and also participate in functions such as feeding and immune evasion. The major proteases, cathepsin L1 (FheCL1) and cathepsin L2 (FheCL2) are members of a lineage that gave rise to the human cathepsin Ls, Ks and Ss, but while they exhibit similarities in their substrate specificities to these enzymes they differ in having a wider pH range for activity and an enhanced stability at neutral pH. There are presently 13 Fasciola cathepsin L cDNAs deposited in the public databases representing a gene family of at least seven distinct members, although the temporal and spatial expression of each of these members in the developmental stage of F. hepatica remains unclear. Immunolocalisation and in situ hybridisation studies, using antibody and DNA probes, respectively, show that the vast majority of cathepsin L gene expression is carried out in the epithelial cells lining the parasite gut. Within these cells the enzyme is packaged into secretory vesicles that release their contents into the gut lumen for the purpose of degrading ingested host tissue and blood. Liver flukes also express a novel multi-domain cystatin that may be involved in the regulation of cathepsin L activity. Vaccine trials in both sheep and cattle with purified native FheCL1 and FheCL2 have shown that these enzymes can induce protection, ranging from 33 to 79%, to experimental challenge with metacercariae of F. hepatica, and very potent anti-embryonation/hatch rate effects that would block parasite transmission. In this article we review the vaccine trials carried out over the past 8 years, the role of antibody and T cell responses in mediating protection and discuss the prospects of the cathepsin Ls in the development of first generation recombinant liver fluke vaccines. Author Keywords: Helminths; Trematodes; Parasites; Cathepsins; Proteases; Vaccines; Immunology; Biochemistry

Spectral simulation of laser ablated magnesium plasmas

We describe a collisional-radiative equilibrium model for predicting the optical emission spectrum of low-temperature magnesium plasmas, specifically those created by laser ablation. In the model, levels are populated by a balance of collisional and radiative rates. We include Stark widths of lines and trapping of radiation in the calculations. By use of this model we discuss various issues of importance in spectral analysis of laser ablated plasma plumes, such as the partial local thermodynamic equilibrium approximation, line trapping and time dependence.

Neural modelling, control and optimisation of an industrial grinding process

This paper describes the development of neural model-based control strategies for the optimisation of an industrial aluminium substrate disk grinding process. The grindstone removal rate varies considerably over a stone life and is a highly nonlinear function of process variables. Using historical grindstone performance data, a NARX-based neural network model is developed. This model is then used to implement a direct inverse controller and an internal model controller based on the process settings and previous removal rates. Preliminary plant investigations show that thickness defects can be reduced by 50% or more, compared to other schemes employed. (c) 2004 Elsevier Ltd. All rights reserved.

Leucine aminopeptidase of the human blood flukes, Schistosoma mansoni and Schistosoma japonicum

An array of schistosome endoproteases involved in the digestion of host hemoglobin to absorbable peptides has been described, but the exoprotease responsible for catabolising these peptides to amino acids has yet to be identified. By searching the public databases we found that Schistosoma mansoni and Schistosoma japonicum express a gene encoding a member of the M17 family of leucine aminopeptidases (LAPs). A functional recombinant S. mansoni LAP produced in insect cells shared biochemical properties, including pH optimum for activity, substrate specificity and reliance on metal cations for activity, with the major aminopeptidase activity in soluble extracts of adult worms. The pH range in which the enzyme functions and the lack of a signal peptide indicate that the enzyme functions intracellularly. Immunolocalisation studies showed that the S. mansoni LAP is synthesised in the gastrodermal cells surrounding the gut lumen. Accordingly, we propose that peptides generated in the lumen of the schistosome gut are absorbed into the gastrodermal cells and are cleaved by LAP to free amino acids before being distributed to the internal tissues of the parasite. Since LAP was also localised to the surface tegument it may play an additional role in surface membrane re-modelling. (C) 2004 Australian Society for Parasitology Inc. Published by Elsevier Ltd. All rights reserved.

Del Fichero supremo a la Biblioteca de Babel: Salaverría, precursor de Borges

A juzgar por alguna declaración más bien negativa de Borges sobre la literatura española, se podría creer que esta no influyó demasiado en él. Sin embargo, existen indicios de que pudo haberse inspirado también en determinados escritores españoles coetáneos hoy casi olvidados. Uno de ellos pudo ser José María Salaverría, entre cuyos relatos destaca “El fichero supremo” (1926), del que se ha dicho que “anticipa algunas de las preocupaciones características de un tipo de relato que Jorge Luis Borges elevará años después a la máxima categoría estética”. De hecho, recuerda a “La biblioteca de Babel” (1941) borgiana por su planteamiento hasta el punto de que podría pensarse que el maestro argentino pudo tener presente, a la hora de escribir esa obra maestra, ese cuento de Salaverría, el cual se publicó por primera vez en Caras y Caretas, una revista porteña que Borges reconoció “devorar” en su juventud. Sin embargo, el interés mayor de la comparación entre “El fichero supremo” y “La biblioteca de Babel” no radica tanto en el carácter de posible fuente del primero como en el contraste entre sus formas de presentación narrativa: desde fuera y en tercera persona en Salaverría, en un marco realista; y desde dentro y en primera persona, prácticamente sin marco, en Borges. Este parece desarrollar, en el registro propio de la “imaginación razonada” descrito por él mismo, una virtualidad presente en el relato de Salaverría, cuya comparación con “La biblioteca de Babel” puede suscitar también alguna reflexión sobre el enigma de la identidad y el carácter de la voz enunciadora de la biblioteca universal de Babel. Al menos, esta parece haber hecho realidad en cierto modo, de forma sublime, el patético sueño divino del archivero imaginado por Salaverría.

Adjusting to Life After Esophagectomy: The Experience of Survivors and Carers

Following surgery for esophageal cancer, patients can experience complex physical, social, and emotional changes. Investigation of these challenges, particularly from the perspective of the patient and his or her carer, has been limited. The current study explored the emotional and cognitive experiences of esophageal cancer survivors and those of their carers, using focus groups conducted with members of a patient support group. Analysis of the patients’ data yielded three themes: coping with a death sentence, adjusting to and accepting an altered self, and the unique benefits of peer support. Analysis of the carers’ data also yielded three themes: the carer as buffer, representations of recovery and recurrence, and normalizing experiences through peer support. Esophageal cancer patients and their carers require holistic support in their efforts to adjust to the social, emotional, and physical consequences of esophagectomy. Peers could be an effective channel for the support of patients and carers.

Electron number density measurements in magnesium laser produced plumes

Interferometry has been used to investigate the spatio-temporal evolution of electron number density following 248 nm laser ablation of a magnesium target. Fringe shifts were measured as a function of laser power density for a circular spot obtained using a random phase plate. Line averaged electron number densities were obtained at delay times up to ∼100 ns after the laser pulse. Density profiles normal to the target surface were recorded for power densities on target in the range 125–300 MW cm−2.