A reappraisal of the central effects of botulinum neurotoxin type A: by what mechanism?

Botulinum neurotoxin A (BoNT/A) is a metalloprotease that enters peripheral motor nerve terminals and blocks the release of acetylcholine via the specific cleavage of the synaptosomal-associated protein of 25-kDa. Localized injections of BoNT/A are widely employed in clinical neurology to treat several human diseases characterized by muscle hyperactivity. It is generally assumed that the effects of BoNT/A remain localized to the injection site. However, several neurophysiological studies have provided evidence for central effects of BoNT/A, raising the issue of how these actions arise. Here we review these data and discuss the possibility that retrograde axonal transport of catalytically active BoNT/A may explain at least some of its effects at the level of central circuits.     

On the mechanism of midtrimester abortions induced by the prostaglandin “impact”

Using Csapo's technique a single dose of 24.3±1.1mg PG F_2α had been delivered intraamniotically to 20 sedated 15.9±0.6 weeks pregnant patients, to provoke a “PG Impact” (PGI), a consequent progesterone (P) withdrawal and a conversion of the pharmacologically refractory normal pregnant uterus into a reactive organ. The side effects were occasional and acceptable and no further PG F_2α treatment was needed except in 4 cases (5–10mg). Only after the Oxytocin Test showed that the uterus is becoming reactive was 50mU/min oxytocin infused i.v., to facilitate the evolution of IUP to 93±3mm Hg and thus promote clinical progress. All the 20 patients aborted both the fetus and the placenta in 16.5±2.1 hours, but 8 women retained small placental residues to be removed by curettage. The Csapo Score was high, 92±2.As early as 3 hours after PGI, the plasma P levels already decreased significantly. They continued to decline throughout the IAT and reach a 72% withdrawal when the fetus was aborted. Fifteen patients, whose P-withdrawal was rapid aborted before the mean IAT, while those 5 women whose P-withdrawal was slow aborted after this time. Thus, the rate of P-withdrawal was directly, while parity and gestational age indirectly related to the IAT. Studies are in progress to elucidate further the abortifacient action of PG F_2α and through this knowledge promote predictable therapy.     

Processive proteolysis by γ-secretase and the mechanism of Alzheimer's disease

Abstract γ-Secretase is a membrane-embedded protease complex with presenilin as the catalytic component. Cleavage within the transmembrane domain of the amyloid β-protein precursor (APP) by γ-secretase produces the C-terminus of the amyloid β-peptide (Aβ), a proteolytic product prone to aggregation and strongly linked to Alzheimer's disease (AD). Presenilin mutations are associated with early-onset AD, but their pathogenic mechanisms are unclear. One hypothesis is that these mutations cause AD through a toxic gain of function, changing γ-secretase activity to increase the proportion of 42-residue Aβ over the more soluble 40-residue form. A competing hypothesis is that the mutations cause AD through a loss of function, by reducing γ-secretase activity. However, γ-secretase apparently has two types of activities, an endoproteolytic function that first cuts APP to generate a 48/49-residue form of Aβ, and a carboxypeptidase activity that processively trims these longer Aβ intermediates approximately every three residues to form shorter, secreted forms. Recent studies suggest a resolution of the gain-of-function vs. loss-of-function debate: presenilin mutations may increase the proportion of longer, more aggregation-prone Aβ by specifically decreasing the trimming activity of γ-secretase. That is, the reduction of this particular proteolytic function of presenilin, not its endoproteolytic activity, may lead to the neurotoxic gain of function.     

Is the fracto‐mechanoluminescence of ZnS:Mn phosphor dominated by charged dislocation mechanism or piezoelectrification mechanism?

Mathematical approaches made for both the charged dislocation model and piezoelectrically induced electron bombardment model of fracto‐mechanoluminescence (FML), the luminescence induced by fracture of solids, in ZnS:Mn phosphor indicate that the piezoelectrically induced electron bombardment model provides a dominating process for the FML of ZnS phosphors. The concentration of 3000 ppm Mn²⁺ is optimal for ML intensity of ZnS:Mn phosphor. The decay time of ML gives the relaxation time of the piston used to deform the sample and the time t_m of maximum of ML is controlled by both the relaxation time of the piston and decay time of charges on the newly created surfaces of crystals. As the product of the velocity of dislocations and pinning time of dislocations gives the mean free path of a moving dislocation. Both factors play an important role in the ML excitation of impurity doped II–VI semiconductors. The linear increase of total ML intensity I_T with the impact velocity indicates that the damage increases linearly with impact velocity of the load. Thus, the ML measurement can be used remotely to monitor the real‐time damage in the structures, and therefore, the ML of ZnS:Mn phosphor has also the potential for a structural health monitoring system. Copyright © 2015 John Wiley & Sons, Ltd.     

Structural insight into the mechanism of activation of the Toll receptor by the dimeric ligand Spätzle

The Drosophila Toll receptor, which functions in both embryonic patterning and innate immunity to fungi and Gram-positive bacteria, is activated by a dimeric cytokine ligand, Sp?tzle (Spz). Previous studies have suggested that one Spz cross-links two Toll receptor molecules to form an activated complex. Here we report electron microscopy structures of the Toll ectodomain in the absence and presence of Spz. Contrary to expectations, Spz does not directly cross-link two Toll ectodomains. Instead, Spz binding at the N-terminal end of Toll predominantly induces the formation of a 2:2 complex, with two sites of interaction between the ectodomain chains, one located near to the N terminus of the solenoid and the other between the C-terminal juxtamembrane sequences. Moreover, Toll undergoes a ligand-induced conformational change, becoming more tightly curved than in the apo form. The unexpected 2:2 complex was confirmed by mass spectrometry under native conditions. These results suggest that activation of Toll is an allosteric mechanism induced by an end-on binding mode of its ligand Spz.     

Resistance to the third-generation cephalosporin ceftazidime by a deacylation-deficient mutant of the TEM β-lactamase by the uncommon covalent-trapping mechanism

The Glu166Arg/Met182Thr mutant of Escherichia coli TEM(pTZ19-3) β-lactamase produces a 128-fold increase in the level of resistance to the antibiotic ceftazidime in comparison to that of the parental wild-type enzyme. The single Glu166Arg mutation resulted in a dramatic decrease in both the level of enzyme expression in bacteria and the resistance to penicillins, with a concomitant 4-fold increase in the resistance to ceftazidime, a third-generation cephalosporin. Introduction of the second amino acid substitution, Met182Thr, restored enzyme expression to a level comparable to that of the wild-type enzyme and resulted in an additional 32-fold increase in the minimal inhibitory concentration of ceftazidime to 64 μg/mL. The double mutant formed a stable covalent complex with ceftazidime that remained intact for the entire duration of the monitoring, which exceeded a time period of 40 bacterial generations. Compared to those of the wild-type enzyme, the affinity of the TEM(pTZ19-3) Glu166Arg/Met182Thr mutant for ceftazidime increased by at least 110-fold and the acylation rate constant was augmented by at least 16-fold. The collective experimental data and computer modeling indicate that the deacylation-deficient Glu166Arg/Met182Thr mutant of TEM(pTZ19-3) produces resistance to the third-generation cephalosporin ceftazidime by an uncommon covalent-trapping mechanism. This is the first documentation of such a mechanism by a class A β-lactamase in a manifestation of resistance.     

On the mechanism of direct conversion of cellulose to ethanol by Fusarium oxysporum: effect of cellulase and β-glucosidase

Summary The effects of the three main enzymes involved in cellulose saccharification, namely cellobiohydrolase, carboxymethylcellulase and -glucosidase, on the direct conversion of cellulose to ethanol by Fusarium oxysporum F3 were investigated. Ethanol production was not affected when the activity of the former two enzymes was varied within a wide range. By contrast, -glucosidase markedly affected ethanol production showing an optimum level of 0.7–0.8 unit/ml growth medium. A significant decrease of cellulose bioconversion time to ethanol was obtained when -glucosidase activity was adjusted to this optimal level at the beginning of the fermentation process. Offprint requests to: B. J. Macris     

Focusing by shape change in the lens of the eye: a commentary on Young (1801) ‘On the mechanism of the eye’

In his Bakerian Lecture paper of 1801, Thomas Young provided the best account up to that time of the eye''s optical system, including refraction by the cornea and the surfaces of the lens. He built a device, an optometer, for determining the eye''s state of focus, making it possible to prescribe appropriate correction lenses. His main contribution, however, was to show that accommodation, the eye''s focusing mechanism, was not the result of changes to the curvature of the cornea, nor to the length of the eye, but was due entirely to changes in the shape of the lens, which he described with impressive accuracy. He was wrong, however, in believing that the reason the lens bulges when focusing on near objects was because it behaved as a contracting muscle. Half a century later, Helmholtz showed that the lens bulges not by its own contraction, but when it is relaxed as a result of contraction of newly discovered circular muscles in the ciliary body. This commentary was written to celebrate the 350th anniversary of the journal Philosophical Transactions of the Royal Society.     

Mitochondrial mechanism of cardiomyocyte apoptosis induced by stress

Stress induced the serious disorder of cardiac function and cardiovascular diseases. Apoptosis is the cellular basis in stress induced cardiac injury. In our previous study we found that many stressors resulted in mitochondrial damage. It is certain that mitochondria is important mediator in triggering apoptotic cell death, but the mechanism, by which the stress induced mitochondrial injury leads to cardiomyocyte apoptosis, remains unclear. We designed the present study to investigate the changes of the mitochondria in cardiomyocytes undergoing stress and its role in inducing apoptosis. Here we reported that stress changed the membrane fluidity of mitochondria and induced the lipid peroxidation of mitochondrial membrane in     

Inhibition of neutrophil apoptosis by acrolein: a mechanism of tobacco-related lung disease?

Cigarette smoking is known to contribute to inflammatory diseases of the respiratory tract by promoting recruitment of inflammatory-immune cells such as neutrophils and perhaps by altering neutrophil functional properties. We investigated whether acrolein, a toxic unsaturated aldehyde found in cigarette smoke, could directly affect neutrophil function. Exposure of freshly isolated human neutrophils to acrolein markedly inhibited spontaneous neutrophil apoptosis as indicated by loss of membrane asymmetry and DNA fragmentation and induced increased neutrophil production of the chemokine interleukin-8 (IL-8). Acrolein (1--50 microM) was found to induce marked activation of extracellular signal-regulated kinase (ERK) and p38 mitogen-activated protein kinases (MAPKs), and inhibition of p38 MAPK activation by SB-203580 prevented acrolein-induced IL-8 release. However, inhibition of either ERK or p38 MAPK did not affect acrolein-dependent inhibition of apoptosis. Acrolein exposure prevented the activation of caspase-3, a crucial step in the execution of neutrophil apoptosis, presumably by direct inhibition of the enzyme. Our results indicate that acrolein may contribute to smoke-induced inflammatory processes in the lung by increasing neutrophil recruitment and reducing neutrophil clearance by apoptosis.     

Exploring the mechanism of β-amyloid toxicity attenuation by multivalent sialic acid polymers through the use of mathematical models

β-Amyloid peptide (Aβ), the primary protein component in senile plaques associated with Alzheimer's disease (AD), has been implicated in neurotoxicity associated with AD. Previous studies have shown that the Aβ-neuronal membrane interaction plays a role in the mechanism of Aβ toxicity. More specifically, it is thought that Aβ interacts with ganglioside rich and sialic acid rich regions of cell surfaces. In light of such evidence, we have used a number of different sialic acid compounds of different valency or number of sialic acid moieties per molecule to attenuate Aβ toxicity in a cell culture model. In this work, we proposed various mathematical models of Aβ interaction with both the cell membrane and with the multivalent sialic acid compounds, designed to act as membrane mimics. These models allow us to explore the mechanism of action of this class of sialic acid membrane mimics in attenuating the toxicity of Aβ. The mathematical models, when compared with experimental data, facilitate the discrimination between different modes of action of these materials. Understanding the mechanism of action of Aβ toxicity inhibitors should provide insight into the design of the next generation of molecules that could be used to prevent Aβ toxicity associated with AD.     

The mechanism of inactivation of S-adenosylhomocysteinase by 2′-deoxyadenosine

S-Adenosylhomocysteinase (SAHase) is irreversibly inactivated by 2′-deoxyadenosine (Hirshfield, M.S. (1979) J. Biol. Chem. $\text{254}$, 22–25). In the course of this inactivation, 2′dAd becomes tightly bound to the enzyme, i.e., cannot be removed by gel filtration or dialysis. Inactivation is accompanied by reduction of the enzyme bound NAD. When the inactivated enzyme is denatured, no 2′dAd is recovered. Adenine equivalent to about 80% of the bound 2′dAd is isolated. It is proposed that 2′-deoxyadenosine is first oxidized to 3′-keto-2-deoxyadenosine by enzyme bound NAD. The 3′keto group activates the hydrogen at C-2′ and facilitates elimination of adenine.     

Body temperature and sleep: Are they controlled by the same mechanism?

Simultaneous changes in sleep and body temperature, produced either by lesion or by stimulation of the medial preoptic area (mPOA), have given reasons to suggest that thermoregulation and sleep regulation are controlled by the same set of neurons. The reasons for simultaneous changes in these parameters are discussed in the present paper with a view to explaining the relationship between thermoregulation and sleep regulation. Changes in body temperature and sleep on destruction of the preoptic area (POA) neurons and the sequence of these changes, suggest a separate control mechanism in the mPOA for regulation of sleep and body temperature. Evidence is put forward in the present paper to show that the mPOA is not involved in the downregulation or upregulation of changes in body temperature with alteration in the vigilance state. On the other hand, circadian modulation of body temperature is possibly involved in altering sleep propensity. A clear indication regarding separate control of sleep and body temperature came from the studies in which noradrenergic agents were applied into the mPOA of animals with and without lesion of the noradrenergic fibers projecting to the mPOA. Experiments in which sleep was analyzed after experimental manipulations of ambient temperature and body temperature, including peripheral, core and brain temperature, are presented here to show a close relationship between thermoregulation and sleep regulation. Various theories regarding the regulation of body temperature during slow wave sleep and rapid eye movement sleep are also discussed. The functional integrity of the mPOA may be essential not only for the regulation of body temperature and sleep-wakefulness but even for the homeostatic regulation of energy balance of the body in response to alterations in environmental temperature and sleep-wakefulness.

     

Cockle emergence at the sediment surface: 'favourization' mechanism by digenean parasites?

The aim of the present work was to assess the effect of digenean trematodes on indirect mortality of the cockle Cerastoderma edule, an infaunal bivalve. The tested hypothesis was that parasites altered the burrowing capacity of cockles and thus exposed them at the sediment surface, where they are more vulnerable to predators. If the predator is the final host, this mechanism, which drives the cockle out of the sediment, is considered as a 'favourization'. Cockle populations from 2 stations in Arcachon Bay (France)-Banc d'Arguin (oceanic situation) and La Canelette (lagoonal situation)--were sampled for 1 yr. At La Canelette, monitoring every 2 d showed that 50% of adult cockles regularly migrated to the sediment surface at a rate of 5 cockles m(-2) yr(-1) and disappeared in a few days. In the laboratory, 67% of these 'surface cockles' did not burrow again, suggesting that they would die in the field. Moreover, mortality measured after 7 d in the laboratory was 2 to 5 times higher than mortality of 'buried cockles', at both stations and particularly during summer. Species richness and abundance of digeneans from both stations were compared in 'buried' and 'surface' individuals to determine whether parasites played a role in cockle migration and mortality. Ten and 9 digeneans were found at Banc d'Arguin and La Canelette, respectively, with Himasthla quissetensis and Labratrema minimus being the most prevalent and abundant species at both stations. The abundance of H. quissetensis was slightly higher in surface cockles at Banc d'Arguin, but the difference fluctuated with station and cockle age (or size). L. minimus prevalence was only higher in surface cockles at La Canelette. In the latter station, we estimated that L. minimus and H. quissetensis were responsible for the emergence of 9 and 2%, respectively, of the buried cockles. Although this favourization mechanism may induce some mortality in cockles, it does not alone explain the magnitude of the observed mortalities (41 and 57% at La Canelette and Banc d'Arguin, respectively). A correspondence analysis did not show the presence of a particular parasite community in buried or surface cockles, which could explain these high surface cockle mortalities in association with the 2 dominant digeneans.     

Some aspects of mechanism of inhibition of cholesterol absorption by β-sitosterol

(1) Mixed bile salt micelle solubilized either cholesterol or β-sitosterol to a comparable extent. When added simultaneously, β-sitosterol restricted the micellar solubility of cholesterol. (2) β-Sitosterol also reduced the cholesterol content in the aqueous (micellar) phase of the intestinal contents of rats, the extent of reduction being comparable with that observed in vitro. The intestinal uptake of cholesterol in vivo was equivalent to the micellar incorporation of cholesterol both in vitro and in vivo. (3) β-Sitosterol had no inhibitory effect on cholesterol absorption from the micellar solution in jejunal loops in situ, whereas the rate of β-sitosterol uptake was only about one-fifth that of cholesterol. (4) The intestinal uptake of β-sitosterol intubated into the stomach of rats was about one-fifth that of cholesterol. The intestinal brush-border membrane discriminated these sterols. These results suggest that the restriction of the micellar solubility of cholesterol, rather than the inhibition of uptake from brush-border membrane, is the major determinant for the interference of β-sitosterol with cholesterol absorption.