Isolation and characterization of a cone snail protease with homology to CRISP proteins of the pathogenesis-related protein superfamily

The pathogenesis-related (PR) protein superfamily is widely distributed in the animal, plant, and fungal kingdoms and is implicated in human brain tumor growth and plant pathogenesis. The precise biological activity of PR proteins, however, has remained elusive. Here we report the characterization, cloning and structural homology modeling of Tex31 from the venom duct of Conus textile. Tex31 was isolated to >95% purity by activity-guided fractionation using a para-nitroanilide substrate based on the putative cleavage site residues found in the propeptide precursor of conotoxin TxVIA. Tex31 requires four residues including a leucine N-terminal of the cleavage site for efficient substrate processing. The sequence of Tex31 was determined using two degenerate PCR primers designed from N-terminal and tryptic digest Edman sequences. A BLAST search revealed that Tex31 was a member of the PR protein superfamily and most closely related to the CRISP family of mammalian proteins that have a cysteine-rich C-terminal tail. A homology model constructed from two PR proteins revealed that the likely catalytic residues in Tex31 fall within a structurally conserved domain found in PR proteins. Thus, it is possible that other PR proteins may also be substrate-specific proteases.

Muscarinic and nicotinic ACh receptor activation differentially mobilize Ca2+ in rat intracardiac ganglion neurons

The origin of intracellular Ca2+ concentration ([Ca2+](i)) transients stimulated by nicotinic ( nAChR) and muscarinic ( mAChR) receptor activation was investigated in fura-2-loaded neonatal rat intracardiac neurons. ACh evoked [Ca2+](i) increases that were reduced to similar to 60% of control in the presence of either atropine ( 1 muM) or mecamylamine ( 3 muM) and to < 20% in the presence of both antagonists. Removal of external Ca2+ reduced ACh-induced responses to 58% of control, which was unchanged in the presence of mecamylamine but reduced to 5% of control by atropine. The nAChR-induced [Ca2+](i) response was reduced to 50% by 10 μM ryanodine, whereas the mAChR-induced response was unaffected by ryanodine, suggesting that Ca2+ release from ryanodine-sensitive Ca2+ stores may only contribute to the nAChR-induced [Ca2+](i) responses. Perforated-patch whole cell recording at - 60 mV shows that the rise in [Ca2+](i) is concomitant with slow outward currents on mAChR activation and with rapid inward currents after nAChR activation. In conclusion, different signaling pathways mediate the rise in [Ca2+](i) and membrane currents evoked by ACh binding to nicotinic and muscarinic receptors in rat intracardiac neurons.

Identification of slow and fast-acting toxins in a highly ciguatoxic barracuda (Sphyraena barracuda) by HPLC/MS and radiolabelled ligand binding

A barracuda implicated in ciguatera fish poisoning in Guadeloupe was estimated to have an overall flesh toxicity of 15 MUg/g using mouse bioassay. A lipid soluble extract was separated into two toxic fractions, FrA and FrB, on a LH20 Sephadex column eluted with dichloromethane/methanol (1:1). When intraperitoneal injected into mice, FrA provoked symptoms characteristic of slow-acting ciguatoxins, whereas FrB produced symptoms indicative of fast-acting toxins (FAT). High performance liquid chromatography/mass spectrometry/radio-ligand binding (HPLC/MS/RLB) analysis confirmed the two fractions were distinct, because only a weak overlap of some compounds was observed. HPLC/MS/RLB analysis revealed C-CTX-1 as the potent toxin present in FrA, and two coeluting active compounds at m/z 809.43 and 857.42 in FrB, all displaying the characteristic pattern of ion formation for hydroxy-polyethers. Other C-CTX congeners and putative hydroxy-polyether-like compounds were detected in both fractions, however, the RLB found them inactive. C-CTX-1 accounted for >90% of total toxicity in this barracuda and was confirmed to be a competitive inhibitor of brevetoxin binding to voltage-sensitive sodium channels (VSSCs) with a potency two-times lower than P-CTX-1. However, FAT active on VSSCs and

Clinical aspects of gestational trophoblastic disease: A review based partly on 25-year experience of a statewide registry

Background: Gestational trophoblastic disease is a fascinating group of pregnancy disorders characterised by abnormal proliferation of trophoblast, ranging from benign to malignant. Because the disease is uncommon, there is a need to formulate management with the assistance of collective information. Methodology: A review of available information from English written literature was undertaken especially data reported by registries around the world (Charing Cross Hospital in England, the North-western University and the New England area in the USA as well as our own experience in Queensland, Australia). Where possible, collated data from relevant studies were analysed to answer some of the questions posed in clinical practice, with reference to metastatic disease to liver and brain, twinning of molar gestation and coexisting fetus, and placental-site tumour. Results: We found that molar gestation can be classified according to its clinical presentation which influences the time taken to reach human chorionic gonadotropin (HCG) 'negativity' and the risk of persisting disease. Categorisation of risk is the basis for choice of chemotherapy to achieve good outcomes. Metastases to liver and brain remain problems in management; the development of 'new' metastases during chemotherapy is a very poor prognostic factor. In the variant of twinning with molar gestation and coexisting fetus, it is important to elucidate the fetal karyotype in planning management: a 69XXX fetus is not salvageable but a normal 46XX or 46XY fetus faces the prospect of early preterm delivery. The placental-site tumour is very rare; localised disease is curable by surgery; chemotherapy is less effective in disseminated disease. From collated worldwide data, the recurrence rate after one mole is 1.3% and after two or more is 20%. Reproductive outcome in subsequent pregnancies, even after multidrug chemotherapy, is not different from the general population. Because of the increased risk long-term of second tumours after multidrug chemotherapy a closer surveillance of these patients is necessary Conclusion: In general, the disease in its persisting or malignant form is 'a cancer model par excellence' because of an identifiable precursor condition, a reliable HCG marker, and sensitivity of the disease to cytotoxic drugs. With current management, retention of fertility is possible and normal reproductive outcome assured.

Expression of hNP22 is altered in the frontal cortex and hippocampus of the alcoholic human brain

Background: Human neuronal protein (hNP22) is a gene with elevated messenger RNA expression in the prefrontal cortex of the human alcoholic brain. hNP22 has high homology with a rat protein (rNP22). These proteins also share homology with a number of cytoskeleton-interacting proteins. Methods: A rabbit polyclonal antibody to an 18-amino acid epitope was produced for use in Western and immunohistochemical analysis. Samples from the human frontal and motor cortices were used for Western blots (n = 10), whereas a different group of frontal cortex and hippocampal samples were obtained for immunohistochemistry (n = 12). Results: The hNP22 antibody detected a single protein in both rat and human brain. Western blots revealed a significant increase in hNP22 protein levels in the frontal cortex but not the motor cortex of alcoholic cases. Immunohistochemical studies confirmed the increased hNP22 protein expression in all cortical layers. This is consistent with results previously obtained using Northern analysis. Immunohistochemical analysis also revealed a significant increase of hNP22 immunoreactivity in the CA3 and CA4 but not other regions of the hippocampus. Conclusions: It is possible that this protein may play a role in the morphological or plastic changes observed after chronic alcohol exposure and withdrawal, either as a cytoskeleton-interacting protein or as a signaling molecule.

A protective effect of early pregnancy factor on experimental autoimmune encephalomyelitis induced in Lewis rats by inoculation with myelin basic protein

Experimental antoimmune encephalomyelitis (EAE) is an organ-specific autoimmune disease characterised by inflammation and demyelination of the central nervous system and is the best available animal model of multiple sclerosis (MS). Since previous studies have shown that EAE is less severe or is delayed in onset during pregnancy and that administration of the pregnancy hormone early pregnancy factor (EPF) down-regulates EAE, experiments in the present study were designed to explore further the role of EPF in EAE. By using the rosette inhibition test, the standard bioassay for EPF and, by semi-quantitative RT-PCR techniques, we have now shown that inflammatory cells from the spinal cord of rats with EAE can produce and secrete EPF, with production being greatest during recovery from disease. Administration of EPF to rats with EAE resulted in a significant increase in the expression of IL-4 and IL-10 mRNA and a significant decrease in IFN-gamma mRNA expression in spinal cord inflammatory cells. Encephalitogenic MBP-specific T cell lines were prepared from popliteal lymph nodes of rats with EAE. Proliferation assays using these cells demonstrated the ability of exogenous EPF to down-regulate the responses of T lymphocytes to MBP. (C) 2003 Elsevier B.V. All rights reserved.

Neural reflex regulation of arterial pressure in pathophysiological conditions : interplay among the baroreflex, the cardiopulmonary reflexes and the chemoreflex

The maintenance of arterial pressure at levels adequate to perfuse the tissues is a basic requirement for the constancy of the internal environment and survival. The objective of the present review was to provide information about the basic reflex mechanisms that are responsible for the moment-to-moment regulation of the cardiovascular system. We demonstrate that this control is largely provided by the action of arterial and non-arterial reflexes that detect and correct changes in arterial pressure (baroreflex), blood volume or chemical composition (mechano- and chemosensitive cardiopulmonary reflexes), and changes in blood-gas composition (chemoreceptor reflex). The importance of the integration of these cardiovascular reflexes is well understood and it is clear that processing mainly occurs in the nucleus tractus solitarii, although the mechanism is poorly understood. There are several indications that the interactions of baroreflex, chemoreflex and Bezold-Jarisch reflex inputs, and the central nervous system control the activity of autonomic preganglionic neurons through parallel afferent and efferent pathways to achieve cardiovascular homeostasis. It is surprising that so little appears in the literature about the integration of these neural reflexes in cardiovascular function. Thus, our purpose was to review the interplay between peripheral neural reflex mechanisms of arterial blood pressure and blood volume regulation in physiological and pathophysiological states. Special emphasis is placed on the experimental model of arterial hypertension induced by N-nitro-L-arginine methyl ester (L-NAME) in which the interplay of these three reflexes is demonstrable.

Towards Emotional Interaction: Using Movies to Automatically Learn Users' Emotional States

The HCI community is actively seeking novel methodologies to gain insight into the user’s experience during interaction with both the application and the content. We propose an emotional recognition engine capable of automatically recognizing a set of human emotional states using psychophysiological measures of the autonomous nervous system, including galvanic skin response, respiration, and heart rate. A novel pattern recognition system, based on discriminant analysis and support vector machine classifiers is trained using movies’ scenes selected to induce emotions ranging from the positive to the negative valence dimension, including happiness, anger, disgust, sadness, and fear. In this paper we introduce an emotion recognition system and evaluate its accuracy by presenting the results of an experiment conducted with three physiologic sensors.

A supervised versus home-based program effects on liver transplanted familial amyloidotic polyneuropathy patients: walking, fatigue and quality of life

Purpose: To evaluate the effects of a six months exercise training program on walking capacity, fatigue and health related quality of life (HRQL). Relevance: Familial amyloidotic polyneuropathy disease (FAP) is an autossomic neurodegenerative disease, related with systemic deposition of amyloidal fibre mainly on peripheral nervous system and mainly produced in the liver. FAP often results in severe functional limitations. Liver transplantation is used as the only therapy so far, that stop the progression of some aspects of this disease. Transplantation requires aggressive medication which impairs muscle metabolism and associated to surgery process and previous possible functional impairments, could lead to serious deconditioning. Reports of fatigue are common feature in transplanted patients. The effect of supervised or home-based exercise training programs in FAP patients after a liver transplant (FAPTX) is currently unknown.

Comparison of body fat content and distribution of familial amyloidotic polyneuropathy patients versus healthy subjects

The deposition of amyloid fibers at the peripheral nervous system can induce motor neuropathy in Familial Amiloidotic Polyneuropethy (FAP) patients. This produces progressive reductions in functional capacity. The only treatment for FAP is a liver transplant, followed by aggressive medication that can affect patients' metabolism. To our knowledge, there are no data on body fat distribution or comparison between healthy and FAP subjects, which may be important for clinical assessment and management of this disease. PURPOSE: To analyze body fat content and distribution between FAP patients and healthy subjects. METHODS: Body fat content and distribution were measured through Double Energy X-ray Densitometry (DXA) in two groups. Group 1 consisted of 43 Familial Amyloidotic Polyneuropathy patients (19 males, 32 + 8 Yrs, and 24 females, 37 + 5 yrs), who had liver transplant less than 2 months before. Group 2 consisted of 18 healthy subjects of similar age (8 males, 36 + 7 yrs, and 10 females, 39 + 5 yrs). RESULTS: Healthy subjects showed higher values than FAP patients for: BMI (24,2+2,3kg/m2 vs 22,3+3,8 kg/m2 respectively, p<0,05), % trunk BF (26,21+8,34kg vs 20,78+9,05kg respectively, p<0,05), % visceral BF (24,43+7,97% vs 19,21+9,30% respectively, p<0,05), % abdominal BF (26,63+8,51% vs 20,63+10,35% respectively, p<0,05) abdominal subcutaneous BF (0,533+0,421kg vs 0,353+0,257kg respectively, p=0,05), abdominal BF/BF ratio (0,09+0,02 vs 0,08+0,02 respectively, p<0,05) and abdominal BF/trunk BF ratio (0,19+0,03 vs 0,17+0,03 respectively, p<0,05). CONCLUSIONS: These results showed that FAP patients soon after liver transplantation exhibited a healthier body fat profile compared to controls. However, fat content and distribution varied widely in FAP subjects, suggesting an individualized approach for assessment and intervention rather than general guidelines. Future research is needed to investigate the long term consequences on body fat following liver transplant in this population.

Comparison of body fat content and distribution of familial amyloidotic polyneuropathy patients versus healthy subjects

The deposition of amyloid fibers at the peripheral nervous system can induce motor neuropathy in Familial Amiloidotic Polyneuropethy (FAP) patients. This produces progressive reductions in functional capacity. The only treatment for FAP is a liver transplant, followed by aggressive medication that can affect patients' metabolism. To our knowledge, there are no data on body fat distribution or comparison between healthy and FAP subjects, which may be important for clinical assessment and management of this disease.

Strength, muscle quality and functional capacity in liver transplanted familial amyloidotic polineuropathy patients

Liver transplantation is the unique treatment for several end-stage diseases. Familial Amiloidotic Polineuropathy (FAP) is a neurodegenerative disease related with systemic deposition of amyloidal fiber mainly on peripheral nervous system, clinically translated by an autonomous sensitive-motor neuropathy with severe functional limitations in some cases. The unique treatment for FAP disease is a liver transplant with a very aggressive medication to muscle metabolism and force production. To our knowledge there are no quantitative characterizations of body composition, strength or functional capacity in this population.

Peak force in familial amyloidotic polyneuropathy

Familial Amyloidotic Polyneuropathy FAP)- A neurodegenerative disease related with systemic deposition of amyloid fibers mainly at the level of the peripheral nervous system. Clinically, the disease is characterized by an autonomous sensitive-motor neuropathy, beginning nearly always in foot, and subsequently involving the hands. Purpose: Compare the levels of hand grip strength (peak force) in FAP patients with (FAPT) or without (FAPNT) liver transplant and in a healthy group (HG).

A supervised versus home-based program effects on liver transplanted familial amyloidotic polyneuropathy patients: walking, fatigue and quality of life

Familial amyloidotic polyneuropathy is a systemic deposition of amyloidal fibre mainly on peripheral nervous system (but also in other systems like heart, gastrointestinal tract, kidneys, etc) and mainly produced in the liver. Purpose of this study: to evaluate the effects of a six months exercise training program(supervised or home-based) on walking capacity, fatigue and health related quality of life (HRQL) on Familial Amyloidotic Polyneuropathy patients submitted to a liver transplant.