Free and polymerized tubulin in cultured bone cells and Chinese hamster ovary cells: the influence of cold and hormones

A low pH method of liposome-membrane fusion (Schneider et al., 1980, Proc. Natl. Acad. Sci. U. S. A. 77:442) was used to enrich the mitochondrial inner membrane lipid bilayer 30-700% with exogenous phospholipid and cholesterol. By varying the phospholipid-to- cholesterol ratio of the liposomes it was possible to incorporate specific amounts of cholesterol (up to 44 mol %) into the inner membrane bilayer in a controlled fashion. The membrane surface area increased proportionally to the increase in total membrane bilayer lipid. Inner membrane enriched with phospholipid only, or with phospholipid plus cholesterol up to 20 mol %, showed randomly distributed intramembrane particles (integral proteins) in the membrane plane, and the average distance between intramembrane particles increased proportionally to the amount of newly incorporated lipid. Membranes containing between 20 and 27 mol % cholesterol exhibited small clusters of intramembrane particles while cholesterol contents above 27 mol % resulted in larger aggregations of intramembrane particles. In phospholipid-enriched membranes with randomly dispersed intramembrane particles, electron transfer activities from NADH- and succinate-dehydrogenase to cytochrome c decreased proportionally to the increase in distance between the particles. In contrast, these electron- transfer activities increased with decreasing distances between intramembrane particles brought about by cholesterol incorporation. These results indicate that (a) catalytically interacting redox components in the mitochondrial inner membrane such as the dehydrogenase complexes, ubiquinone, and heme proteins are independent, laterally diffusible components; (b) the average distance between these redox components is effected by the available surface area of the membrane lipid bilayer; and (c) the distance over which redox components diffuse before collision and electron transfer mediates the rate of such transfer.     

Burn Injury Reduces Neutrophil Directional Migration Speed in Microfluidic Devices

Thermal injury triggers a fulminant inflammatory cascade that heralds shock, end-organ failure, and ultimately sepsis and death. Emerging evidence points to a critical role for the innate immune system, and several studies had documented concurrent impairment in neutrophil chemotaxis with these post-burn inflammatory changes. While a few studies suggest that a link between neutrophil motility and patient mortality might exist, so far, cumbersome assays have prohibited exploration of the prognostic and diagnostic significance of chemotaxis after burn injury. To address this need, we developed a microfluidic device that is simple to operate and allows for precise and robust measurements of chemotaxis speed and persistence characteristics at single-cell resolution. Using this assay, we established a reference set of migration speed values for neutrophils from healthy subjects. Comparisons with samples from burn patients revealed impaired directional migration speed starting as early as 24 hours after burn injury, reaching a minimum at 72–120 hours, correlated to the size of the burn injury and potentially serving as an early indicator for concurrent infections. Further characterization of neutrophil chemotaxis using this new assay may have important diagnostic implications not only for burn patients but also for patients afflicted by other diseases that compromise neutrophil functions.     

Age-related variability in occlusal wear planes

Investigation of two populations of 136 individuals shows several patterns of occlusal wear plane change which are positively correlated with age. For individuals up to the age of 18, there is a characteristic pattern in which the occlusal wear planes of the mandibular teeth are lingually sloped and those of the maxillary dentition buccally sloped, with the exception of the maxillary premolars, which are also lingually sloped. The long axes of the mandibular teeth give them a lingual orientation relative to the maxillary teeth, and the long axes of the maxillary molars, by contrast, are buccally oriented. In the 18-30 age range for all sexes, the mandibular M1 becomes buccally sloped on its occlusal surface while the occlusal wear plane on the maxillary M1 becomes lingually sloped. Later age changes indicate a trend for the mandibular premolars to become buccally sloped, while the wear planes of the maxillary premolars remain lingually sloped. There is a corresponding tendency for the maxillary and mandibular second molars to undergo changes in the initial orientation of the occlusal wear planes.     

The role of lysyl oxidase and collagen crosslinking during sea urchin development

Lysyl oxidase, the only enzyme involved in collagen crosslinking, is shown to be present in embryos of the sea urchin Strongylocentrotus purpuratus. The enzyme specific activity increases over six-fold during development, showing the greatest rise during gastrulation and prism larva formation. The enzyme is inhibited by the specific inhibitor, beta-aminoproprionitrile (BAPN). Continuous BAPN treatment of S. purpuratus and Lytechinus pictus embryos from late cleavage stages onward increases the amount of noncrosslinked collagen present in prism larvae. When BAPN is added at the 128- or 256-cell stage it causes developmental arrest at the mesenchyme blastula stage. Embryos can be maintained in the arrested state for at least 96 h and will resume normal development and morphogenesis following BAPN removal. If BAPN is added after the mesenchyme blastula stage, it has little adverse effect on development; consequently nonspecific toxic effects of the drug are unlikely. The results suggest that lysyl oxidase and collagen crosslinking play a vital role in primary mesenchyme migration, gastrulation, and morphogenesis during sea urchin development and indicate that BAPN may be very useful in studying the extracellular matrix-cell interactions at the cellular and molecular level.