Validation of circulating histone detection by mass spectrometry for early diagnosis, prognosis, and management of critically ill septic patients.

Background: As leading contributors to worldwide morbidity and mortality, sepsis and septic shock are considered a major global health concern. Proactive biomarker identification in patients with sepsis suspicion at any time remains a daunting challenge for hospitals. Despite great progress in the understanding of clinical and molecular aspects of sepsis, its definition, diagnosis, and treatment remain challenging, highlighting a need for new biomarkers with potential to improve critically ill patient management.

Circulation of the plasma membrane in Dictyostelium.

We have developed a fluorimetric assay with the use of the dye FM1-43 to determine the rate at which Dictyostelium amoebae endocytose their surface membrane. Our results show that they do so about once each 4-10 min. A clathrin null mutant takes its surface up only approximately 30% more slowly, showing that this membrane uptake cannot be caused by clathrin-coated vesicles.

Membrane traffic during cell locomotion.

The key role played by a polarised endocytic cycle in extending the front of a eukaryotic cell is now becoming established. Highlights that have occurred during the past year include the visualisation of vesicles fusing with the advancing edge of Physarum, directly leading to extensions of the cell; furthermore, the fusion of vesicles at the leading edge in plant pollen tubes appears to be controlled by a small GTPase, Rop 1, which is a plant homologue of the mammalian Rac. In animal cells, Rac appears to help determine where exocytosis occurs on the surface of a polarised cell.

EGF induces recycling membrane to form ruffles.

KB cells are know to respond to epidermal growth factor (EGF) by producing prodigious ruffles in the plasma membrane within minutes. The signal transduction pathway underlying this effect in fibroblasts is mediated through Rac, a member of the Ras-like family of GTPases. As ruffles are rich in components of the cytoskeleton--particularly in actin and ezrin--it has been suggested that ruffles arise when activated Rac modulates the actin cytoskeleton to push out a membrane protrusion.

Dictyostelium myosin II null mutant can still cap Con A receptors.

Cross-linked antigens on the surface of a motile cell cap at the trailing end of the cell. In Dictyostelium discoideum, myosin II null mutants have previously been reported to be unable to cap Con A receptors, although they are able to locomote. This finding implicated myosin II as an essential component of the capping mechanism, although not of the machinery for locomotion.

NDP kinase can modulate contraction of Dictyostelium cytoskeletons.

Extraction of Dictyostelium amoebae with Triton X-100 produces robust cytoskeletons composed mainly of actin and myosin II. These cytoskeletons rapidly contract when mixed with Mg-ATP in simple buffers. The Triton-soluble fraction was found to contain a GTP-dependent activity that prevented contraction by Mg-ATP.

Arachidonic acid activation of a new family of K+ channels in cultured rat neuronal cells.

1. The presence and properties of K+ channels activated by arachidonic acid were studied in neuronal cells cultured from the mesencephalic and hypothalamic areas of rat brain. 2.

Isolation of myosin from Dictyostelium cytoskeletons.

Dictyostelium has several isoforms of "unconventional" single-headed myosins that do not assemble into filaments (myosin I). In contrast, there is only one form of conventional myosin (myosin II) that self-assembles into bipolar thick filaments, and molecular genetic studies have shown that this myosin is an essential motor protein for several cell functions, including cytokinesis. Myosin II is phosphorylated on both the heavy chain and light chain, and these modifications regulate assembly and ATPase activity.

Contraction of reconstituted Dictyostelium cytoskeletons: an apparent role for higher order associations among myosin filaments.

A large number of cellular functions require assembly of actin and myosin and coordinated interactions between the resulting filaments. To better understand the structure and function of one such contractile assembly, we have begun fractionation and reconstitution studies of Dictyostelium cytoskeletons. Isolated cytoskeletons rapidly contracted when mixed with Mg-ATP, and myosin II was essential for this since myosin-depleted (stripped) cytoskeletons failed to contract.