Properties of microtubule sliding disintegration in isolated tetrahymena cilia

Properties of the sliding disintegration response of demembranated tetrahymena cilia have been studied by measuring the spectrophotomeric response or turbidity of cilia suspensions at a wavelength of 350 nm relative to changes in the dynein substrate (MgATP(2-)) concentration. The maximum decrease in turbidity occurs in 20 muM ATP, and 90 percent of the decrease occurs in approximately 5.9 s. At lower ATP concentrations (1-20 muM), both the velocity and magnitude of the turbidity decreases are proportional to ATP concentration. The velocity data for 20 muM ATP permit construction of a reaction velocity curve suggesting that changes in turbidity are directly proportional to the extent and velocity of disintegration. At ATP concentrations more than 20 muM (50muM to 5mM), both velocity and magnitude of the turbidimetric response are reduced by approximately 50 percent. This apparent inhibition results in a biphasic response curve that may be related to activation of residual shear resistance or regulatory components at the higher ATP concentrations. The inhibitory effects of elevated ATP can be eliminated by mild trypsin proteolysis, whereupon the reaction goes to completion at any ATP concentration. The turbidimetric responses of the axoneme-substrate suspensions are consistent with the extent and type of axoneme disintegration revealed by electron microscope examination of the various suspensions, suggesting that the turbidimetric assay may prove to be a reliable means for assessing the state of axoneme integrity.     

Verification of biomechanical methods employed in a comprehensive study of mild traumatic brain injury and the effectiveness of American football helmets

Concussion, or mild traumatic brain injury, occurs in many activities, mostly as a result of the head being accelerated. A comprehensive study has been conducted to understand better the mechanics of the impacts associated with concussion in American football. This study involves a sequence of techniques to analyse and reconstruct many different head impact scenarios. It is important to understand the validity and accuracy of these techniques in order to be able to use the results of the study to improve helmets and helmet standards. Two major categories of potential errors have been investigated. The first category concerns error sources specific to the use of crash test dummy instrumentation (accelerometers) and associated data processing techniques. These are relied upon to establish both linear and angular head acceleration responses. The second category concerns the use of broadcast video data and crash test dummy head-neck-torso systems. These are used to replicate the complex head impact scenarios of whole body collisions that occur on the football field between two living human beings. All acceleration measurement and processing techniques were based on well-established practices and standards. These proved to be reliable and reproducible. Potential errors in the linear accelerations due to electrical or mechanical noise did not exceed 2% for the three different noise sources investigated. Potential errors in the angular accelerations due to noise could be as high as 6.7%, due to error accumulation of multiple linear acceleration measurements. The potential error in the relative impact velocity between colliding heads could be as high as 11%, and was found to be the largest error source in the sequence of techniques to reconstruct the game impacts. Full-scale experiments with complete crash test dummies in staged head impacts showed maximum errors of 17% for resultant linear accelerations and 25% for resultant angular accelerations.     

Protocol for Birmingham Atrial Fibrillation Treatment of the Aged study (BAFTA): a randomised controlled trial of warfarin versus aspirin for stroke prevention in the management of atrial fibrillation in an elderly primary care population [ISRCTN89345269]

Background

Statins effectively lower blood cholesterol and the risk of cardiovascular death. Immunomodulatory actions, independent of their lipid-lowering effect, have also been ascribed to these compounds. Since macrophages participate in several vascular pathologies, we examined the effect of statin treatment on the survival and differentiation of primary human monocytes.

Methods

Peripheral blood mononuclear cells (PBMCs) from healthy individuals were cultured in the presence or absence of mevastatin. Apoptosis was monitored by annexin V / PI staining and flow cytometry. In parallel experiments, cultures were stimulated with LPS in the presence or absence of mevastatin and the release of IL-1β and IL-1Ra was measured by ELISA.

Results

Among PBMCs, mevastatin-treated monocytes were particularly susceptible to apoptosis, which occurred at doses >1 microM and was already maximal at 5 microM. However, even at the highest mevastatin dose used (10 microM), apoptosis occurred only after 24 h of culture, possibly reflecting a requirement for cell commitment to differentiation. After 72 h of treatment the vast majority (>50%) of monocytes were undergoing apoptosis. Stimulation with LPS revealed that mevastatin-treated monocytes retained the high IL-1β output characteristic of undifferentiated cells; conversely, IL-1Ra release was inhibited. Concurrent treatment with mevalonolactone prevented the induction of apoptosis and suppressed both IL-1β and IL-1Ra release in response to LPS, suggesting a rate-limiting role for HMG-CoA reductase in monocyte differentiation.

Conclusions

Our findings indicate that statins arrest the functional differentiation of monocytes into macrophages and steer these cells into apoptosis, suggesting a novel mechanism for the vasculoprotective properties of HMG-CoA reductase inhibitors.     

Pulmonary granulocyte influx and impaired alveolar macrophage adenylyl cyclase responsiveness in developing respiratory distress

Alveolar macrophages have recently been postulated as being involved in the aetiology of adult respiratory distress syndrome (ARDS). To evaluate their role, basal cyclic AMP levels and responsiveness of adenylyl cyclase alveolar macrophages were determined at four intermediate stages of developing respiratory distress in piglets using a protocol with repeated lung lavage. Examination of alveolar cells recovered from the subsequent lavages reveals an influx of granulocytes (neutrophils and eosinophils) within 1 h of two intensive lung lavages. During the developing respiratory distress the basal cyclic AMPlevel of alveolar macrophages increases and adenylyl cyclase responsiveness to prostaglandin E(2) (PGE(2)) and isoprelanaline diminishes. The previously observed impairment of macrophage activity can then be explained at a subcellular level.     

SKPDB: a structural database of shikimate pathway enzymes

Background  

The functional and structural characterisation of enzymes that belong to microbial metabolic pathways is very important for structure-based drug design. The main interest in studying shikimate pathway enzymes involves the fact that they are essential for bacteria but do not occur in humans, making them selective targets for design of drugs that do not directly impact humans.