Ozone effects on trees, where uptake and detoxification meet

Ozone is the most important air pollutant and its concentration in ambient air is still rising. Ozone concentrations measured at reference height (50 m is EMEP ozone modelling height), do not reflect the real concentration at the top of the vegetative canopy and do not provide sufficient information about the ozone fluxentering the leaves. Modelling stomatal conductance is leading to estimations of cumulative ozone uptake and enables much better to evaluate the impact of ozone on trees. The negative impact of ozone exposure has a measurable effect on physiological processes such as stomatal conductance, photosynthesis and respiration. Disturbance of the basic physiological processes is leading to growth and wood production losses. There have been several attempts to establish critical levels (CL) for ozone effects on forest trees. Average concentrations and cumulative exposure indices are satisfactory to some extent, but do not fully describe the potential impact of ozone exposure. Much more promising is an evaluation based on the effective ozone flux, which is a function of the absorbed ozone flux and the defensive response. Ozone uptake takes place primarily through the stomata and reactions of ozone with hydrocarbons released by the plant cells and transformations of dissolved ozone in the apoplastic fluid create many reactive oxygen species of which free radicals are able to initiate membrane lipid peroxidation and destruction of cell membranes. The defence of a plant against absorbed ozone starts in the apoplastic fluid. Ascorbate is believed to be a very important radical scavenger avoiding detrimental effects of reactive oxygen species to the membranes. Other important antioxidants are phenolics. The defensive response can be linked to the abundance of ascorbate or the ability of the plants to regenerate (reduce) ascorbate from monodehydroascorbate and dehydroascorbate. The reduction of dehydroascorbate takes place in the symplast where ascorbate can be transported back through the plasma membrane into the apoplast. Ozone exposure also causes oxidative stress of the plant cell interior by the formation of reactive oxygen species. Plants can cope with those toxic substances in the symplast by using antioxidants such as ascorbate, -tocopherol, glutathione and carotenoids and enzymes such as superoxide dismutases, catalases and several peroxidases. The complexity of the apoplastic and symplastic antioxidative capacity with different turnover rates and transport of antioxidants makes it difficult to determine the total antioxidative power

Biotin- or digoxigenin-conjugated nucleotides bind to matrix vesicles in atherosclerotic plaques.

The present study analyzes the staining pattern of DNA in situ end-labeling techniques of human and rabbit atherosclerotic plaques. Both the terminal deoxynucleotidyl transferase end-labeling and the in situ nick translation technique detected, besides apoptotic nuclei, numerous round vesicles with diameters from 0.5 to 5 microns within the atherosclerotic plaques. These vesicles did not contain DNA but contained calcium. A pretreatment with EDTA or citric acid abolished the labeling of the vesicles but did not influence the detection of apoptotic nuclei. Ultrastructurally, the vesicles were of variable diameter and density, and their aspect was compatible with matrix vesicles, which are well known in epiphyses during bone formation. The larger vesicles contained cell organelles, and the small vesicles were very dense. X-ray microanalysis demonstrated high calcium and phosphorus levels within the most dense vesicles. Different stages of the process were present in the plaques. In this way we could demonstrate that cytoplasmic fragmentation of smooth muscle cells and subsequent formation of matrix vesicles are a frequent finding in atherosclerotic plaques. The association of apoptotic cell death and formation of matrix vesicles could be an interesting pathway in explaining calcification of atherosclerotic plaques. Both the terminal deoxynucleotidyl transferase end-labeling and the in situ nick translation technique detected simultaneously apoptotic nuclei and matrix vesicles if calcium is not removed from the sections

Laminin binding and internalization by human and murine mammary gland cell lines in vitro.

We have studied the binding and internalization of Engelbreth-Holm-Swarm mouse sarcoma laminin labeled with colloidal gold (LN-G40) by human and murine mammary gland cell lines. Interactions between the LN-G40 probe and the cells spread on a glass coverslip were monitored with video-enhanced contrast microscopy (Nanovid). Transmission electron microscopy allowed the quantitation of the LN-G40 probe at various cellular locations. During the first 15 min, a homogeneous binding of LN-G40 probe to the cell surface was observed with all cell lines. This binding did not occur with gold particles that were not conjugated to laminin. Then, the LN-G40 probe began to cluster on the cell surface and was, during the following 20 h, internalized by pits that were not coated. In the cells, the LN-G40 probe sometimes showed saltatory movements along linear tracks. The LN-G40 probe was intracellularly found in vesicles, multivesicular bodies, cisternal structures, and lysosomes, suggesting the degradation of the internalized laminin. However, not all cell surface-bound LN-G40 probe was internalized after 20 h. Differences between the cell lines were quantitative, but no clear correlation could be made between migration of cells on laminin and internalization of laminin