Susceptibility to invasive bacterial infections in children with sickle cell disease.

Individuals with sickle cell disease (SCD) demonstrate an increased susceptibility to invasive bacterial infections (IBI). The most common organisms causing IBI are Streptococcus pneumoniae, nontyphi Salmonella species and Haemophilus influenzae type b (Hib). IBI are the most common causes of death in children below 5 years of age with SCD. Increased susceptibility to IBI is because of several factors including dysfunctional antibody production and opsonophagocytosis as well as defective splenic clearance. Early diagnosis of Hib and pneumococcal infections combined with antibiotic prophylaxis and immunization programs, could lead to significant improvements in mortality, especially in Africa.

A comparison of the associations between risk factors and cardiovascular disease in Asia and Australasia.

BACKGROUND: Cardiovascular disease is already the leading cause of death in many Asian populations. Relationships between vascular risk factors and cardiovascular disease may differ in Asian and western populations. Previously, a lack of prospective data has prevented the reliable quantification of such differences, which, if they were shown to exist, would suggest that novel cardiovascular prevention and treatment strategies are required for Asia. DESIGN: An individual participant data meta-analysis of 32 studies from the Asia-Pacific region involving 331 100 subjects (75% from Asia; 25% from the predominantly Caucasian populations of Australia and New Zealand). METHODS: Outcomes were death from coronary heart disease, ischaemic and haemorrhagic stroke. Hazard ratios were estimated from Cox models for systolic blood pressure (SBP), total cholesterol, triglycerides, body mass index, diabetes and current cigarette smoking, stratified by study and sex and adjusted for age, the other risk factors and regression dilution. RESULTS: After an average period of follow-up of 4 years there were 2082 deaths from coronary heart disease, 600 from haemorrhagic stroke and 420 from ischaemic stroke. The direction and strength of the associations between risk factors and cardiovascular outcomes were similar in the two regions, although in two cases there were significant differences. Triglycerides were more strongly associated with coronary heart disease in Australia and New Zealand (P = 0.03), whereas SBP showed a stronger relationship with haemorrhagic stroke in Asia (P = 0.04). CONCLUSIONS: Classical vascular risk factors act similarly in Asian and Caucasian populations; prevention and treatment strategies should thus be similar. Blood pressure reduction should be particularly effective in Asia.

Analysis of the immunophenotypes of de novo acute lymphoblastic leukaemia (ALL) in the Sultanate of Oman.

Acute lymphoblastic leukaemia (ALL) immmunophenotypes were analysed by flow cytometry in 65 Omani patients (46 children, 19 adults). Common ("CALLA-positive") ALL was the most frequently encountered (70%) B-cell lineage immunophenotype. Among T-cell lineage ALL patients, mature T-cell ALL was the least frequent (7%). Expression of certain surface markers including CD20 and CD6 appears to have an effect on some clinical and haematological features but FAB morphology was not useful in predicting cell lineage immunophenotypes. Other interesting findings, currently of uncertain significance, include the sizeable proportions of pre-B-ALL (group V) and cortical thymocytic ALL (stage III). Results of this study should help strengthen the emerging leukaemia database of the recently established Oman National Cancer Register and thereby contribute to a successful global attack against the haematological malignancies.

Past, present and future experiments on muscle.

Since the basic outline of the sliding filament mechanism became apparent some 45 years ago, the principal challenge, an experimental one, has been to produce definitive evidence about the detailed molecular mechanisms by which myosin cross-bridges produce force and movement in a muscle. More recently, similar questions could be posed about other molecular motors, in non-muscle cells. This problem proved unexpectedly difficult to solve, in part because of the technical difficulty of obtaining the structural and mechanical information required about rapid events within macromolecules, especially in a working system, and this triggered many remarkable technical developments. There is now very strong evidence for a large change in shape of the myosin heads during ATP hydrolysis, consistent with a lever-arm mechanism. Whether this does indeed provide the driving force for contraction and movement--and, if so, exactly how--and whether some other processes could also play a significant role, is discussed in the light of the experimental and theoretical findings presented at this meeting, and other recent and long-term evidence.

Development of Synchrotron Radiation as a High-Intensity Source for X-ray Diffraction.

Interest in the molecular mechanism of muscle contraction led to the search for an intense source of X-rays of 1-2 A wavelength so as to be able to examine the rich X-ray diffraction patterns given by muscles during contraction. This led to the first X-ray diffraction experiments using synchrotron radiation, carried out by Holmes, Rosenbaum and Witz at DESY, Hamburg, in September 1970. In the following years, the EMBL Outstation, to utilize synchrotron radiation for biological structure determination, was established at DESY and preliminary experiments on muscle were also carried out at NINA (Daresbury). The development of time-resolved techniques for muscle diffraction was first started in the MRC Molecular Biology Laboratory in Cambridge, using rotating-anode X-ray tubes, and was then greatly extended at the EMBL Outstation, Hamburg, using the storage ring DORIS. This was a very successful venture, and helped to drive the whole technology development and to interest other potential users in the technique.

A personal view of muscle and motility mechanisms.

This is a personal account of some of the successive steps in our understanding of the structural mechanism of muscle contraction during the last 45 years. It describes how I, as an ex-physicist, came to be studying muscle by X-ray diffraction in 1949; how the concepts of the double array of actin and myosin filaments and, later, the overlapping filament model and the sliding filament mechanism were developed; and how further electron microscope findings of the structural polarity of muscle filaments led to the suggestion that analogous structures and mechanisms might be involved in cellular motility. The article describes briefly how synchrotron radiation has made it possible to obtain detailed structural information about contracting muscle with millisecond time resolution and discusses some of the recent major advances in the field and the prospects of reaching a full understanding of the contraction mechanism.

The working stroke of myosin crossbridges.

Recent x-ray diffraction measurements of the axial periodicities of the actin and myosin filaments in contracting muscles show that they are stretched by small but significant amounts by the developed tension, so that at least one half, possibly more, of the observed compliance of a sarcomere must reside in the filaments themselves. This implies that the movement steps of a crossbridge deduced from quick-release experiments may be shorter than some previous estimates, necessitating at least two steps to account for current in vitro single-molecule measurements. Intensity measurements of the wider angle x-ray reflections also show some unexpected features.

X-ray diffraction measurements of the extensibility of actin and myosin filaments in contracting muscle.

We have used a small angle scattering system assembled on the high flux multipole wiggler beam line at CHESS (Cornell) to make very accurate spacing measurements of certain meridional and layer-line reflections from contracting muscles. During isometric contraction, the actin 27.3 A reflection increases in spacing from its resting value by approximately 0.3%, and other actin reflections, including the 59 and 51 A off-meridional reflections, show corresponding changes in spacing. When tension is augmented or diminished by applying moderate speed length changes to a contracting muscle, changes in spacing in the range of 0.19-0.24% (when scaled to full isometric tension) can be seen. The larger difference between the resting and isometric spacings suggests either nonlinearity at low tension levels or the presence of a component related to activation itself. Myosin filaments also show similar increases in axial period during slow stretch, in addition to the well known larger change associated with activation. An actin spacing change of 0.25-0.3% can also be measured during a 2 ms time frame immediately after a quick release, showing that the elastic behavior is rapid. These observations of filament extensions totaling 2-3 nm per half-sarcomere may necessitate some significant revision of the interpretation of a number of mechanical experiments in muscle, in which it has usually been assumed that virtually all of the elasticity resides in the cross-bridges.

Ultrastructure of skeletal muscle fibers studied by a plunge quick freezing method: myofilament lengths.

We have set up a system to rapidly freeze muscle fibers during contraction to investigate by electron microscopy the ultrastructure of active muscles. Glycerinated fiber bundles of rabbit psoas muscles were frozen in conditions of rigor, relaxation, isometric contraction, and active shortening. Freezing was carried out by plunging the bundles into liquid ethane. The frozen bundles were then freeze-substituted, plastic-embedded, and sectioned for electron microscopic observation. X-ray diffraction patterns of the embedded bundles and optical diffraction patterns of the micrographs resemble the x-ray diffraction patterns of unfixed muscles, showing the ability of the method to preserve the muscle ultrastructure. In the optical diffraction patterns layer lines up to 1/5.9 nm-1 were observed. Using this method we have investigated the myofilament lengths and concluded that there are no major changes in length in either the actin or the myosin filaments under any of the conditions explored.

A stopped-flow/rapid-freezing machine with millisecond time resolution to prepare intermediates in biochemical reactions for electron microscopy.

We have developed an instrument capable of freezing transient intermediates in rapid biochemical reactions for subsequent freeze-fracturing, replication, and viewing by transmission electron microscopy. The machine combines a rapid mixing unit similar to one widely used in chemical kinetics (Johnson, 1986) with a propane jet freezing unit previously used to prepare static samples for freeze-fracturing (Gilkey and Staehelin, 1986). The key element in the system is a unique thin-walled flow cell of copper that allows for injection and aging of the sample, followed by rapid freezing. During freeze-fracturing, a tangential cut is made along the wall of the flow cell to expose the sample for etching and replication. The dead time required for mixing and injection of the reactants into the flow cell is less than 5 ms. Electronic controls allow one to specify, on a millisecond time scale, any time above 5 ms between initiation of the reaction and quenching by rapid freezing.