Characterization of human alpha fetoprotein charge microheterogeneity during fetal development.

Aliquotes of human amniotic fluid (AF), fetal serum (FS), and cord blood (CB) were obtained as by-products of routine clinical diagnostic procedures at term or in the second trimester of pregnancy. When samples of CB were applied to a pH 5.5-4 chromatofocusing gradient, three isoforms of AFP could be resolved; a pl 4.57 form (isoform IA, 52% AFP), a pl 4.27 form (isoform IB, 43% AFP), and one species that was bound to the column but could be eluted with 1.0 M NaCl (isoform II, pl less than 4.00, 5% AFP). Term AF displayed a profile similar to that observed in term CB. When samples of 15-20-week gestation AF were chromatofocused, the immunoreactive AFP recovered was distributed between isoform IA and IB (60%) and isoform II (40%). FS and AF obtained from same pregnancy (23-26 weeks) displayed an identical chromatofocusing profile. Aliquotes of AF subjected to conA revealed 83% reactive variants compared with greater than 95% reactive variants for CB. FS displayed a conA profile identical to CB. When individual CB charge isoforms were isolated and subjected to conA analysis, greater than 97% of the AFP bound to conA. In contrast, when AFP isoform IA and IB were isolated from midgestation AF, approximately 22% of the AFP did not bind to the lectin while 100% of isolated AFP isoform II eluted as the reactive variant. These data suggest that human AFP exists as at least three charge and two lectin variants and that the charge profile may change during fetal development.     

Growth kinetics of Lactobacillus acidophilus under ohmic heating

Lactobacillus acidophilus OSU133 was inoculated into MRS broth in a fermenter vessel and incubated at 30, 35, or 45 degrees C with agitation. Incubation temperatures were attained by conventional or ohmic heating. An electrical current at low (15 V) or high (40 V) voltage was used to heat the culture directly during fermentations under ohmic heating. The growth parameters (lag period, minimum generation time, and maximum growth) and changes in pH were determined during fermentation. Metabolic activities (consumption of glucose and production of lactic acid and bacteriocin) were determined during fermentation at 35 degrees C under both heating methods. Lag period for L. acidophilus was affected appreciably by the method of heating, but the magnitude of these changes depended on the fermentation temperature. When fermentation was done at 30 degrees C, lag period decreased by 94% under low-voltage ohmic, compared with conventional, heating methods. Ohmic heating did not change the generation time significantly and caused slight, but significant (p < 0.01) decrease in maximum growth. Therefore, the electric current enhances the early stages, but it inhibits the late stages of growth. Ohmic, compared with conventional, heating resulted in higher final pH and lower bacteriocin activity in the fermented medium. However, ohmic heating at 35 degrees C had minimal effect on glucose utilization and lactic acid production by L. acidophilus. Results show that measurement of the electric current when ohmic heating is done at a constant voltage may be used in monitoring such fermentations. In conclusion, ohmic heating is potentially useful in certain applications related to fermented foods. (c) 1996 John Wiley & Sons, Inc.     

Dendrochronological analysis of the canopy history of two Ohio old-growth forests

This study examined the temporal patterns of establishment, suppression, and release of major tree species in two old-growth Ohio forest remnants as a means to determine the past disturbance history of these forests. Increment cores were taken from a total of 154 trees from two well-drained, upland plots and two poorly-drained, bottomland plots in each of the two forested areas. Acer saccharum and Fagus grandifolia exhibited multiple episodes of suppression and release prior to becoming canopy trees, and could tolerate suppressions as long as 84 years. In contrast, Quercus macrocarpa, Q. muehlenbergii, Prunus serotina, and Acer saccharinum rerely exhibited any tolerance to suppression and appeared to have entered the canopy after single disturbances had opened large areas of canopy. There was clear synchrony in the temporal pattern of establishment and final release from suppression among trees from bottomland plots scattered throughout the stands, indicating that relatively large disturbances were important in these poorly-drained areas. In contrast, there was little synchrony among trees from well-drained upland plots, except in a single instance where selective cutting of Quercus trees opened the canopy. Thus, the canopy of upland site was likely subjected only to small disturbances resulting from the death of one or a few trees. At the whole of forest level, there was evidence of episodic recruitment of canopy trees in both forests. Establishment of Fraxinus spp. and Quercus spp. were particularly episodic, and few Fraxinus or Quercus trees alive today established during the last century. These data suggest that large disturbances have affected canopy dynamics of both upland and bottomland areas prior to 1900 and in bottomland forests through this century. In contrast, disturbances in upland areas during this century have been restricted to small, treefall-generated canopy gaps.     

Inactivation of the Porphyromonas gingivalis fimA gene blocks periodontal damage in gnotobiotic rats.

Fimbrial production by Porphyromonas gingivalis was inactivated by insertion-duplication mutagenesis, using the cloned gene for the P. gingivalis major fimbrial subunit protein, fimA. by several criteria, this insertion mutation rendered P. gingivalis unable to produce fimbrilin or an intact fimbrial structure. A nonfimbriated mutant, DPG3, hemagglutinated sheep erythrocytes normally and was unimpaired in the ability to coaggregate with Streptococcus gordonii G9B. The cell surface hydrophobicity of DPG3 was also unaffected by the loss of fimbriae. However, DPG3 was significantly less able to bind to saliva-coated hydroxyapatite than wild-type P. gingivalis 381. This suggested that P. gingivalis fimbriae are important for adherence of the organism to saliva-coated oral surfaces. Further, DPG3 was significantly less able to cause periodontal bone loss in a gnotobiotic rat model of periodontal disease. These observations are consistent with other data suggesting that P. gingivalis fimbriae play an important role in the pathogenesis of human periodontal disease.     

Changes in phosphatidylinositol metabolism in response to hyperosmotic stress in Daucus carota L. cells grown in suspension culture.

Carrot (Daucus carota L.) cells plasmolyzed within 30 s after adding sorbitol to increase the osmotic strength of the medium from 0.2 to 0.4 or 0.6 osmolal. However, there was no significant change in the polyphosphorylated inositol phospholipids or inositol phosphates or in inositol phospholipid metabolism within 30 s of imposing the hyperosmotic stress. Maximum changes in phosphatidylinositol 4-monophosphate (PIP) metabolism were detected at 5 min, at which time the cells appeared to adjust to the change in osmoticum. There was a 30% decrease in [3H]inositol-labeled PIP. The specific activity of enzymes involved in the metabolism of the inositol phospholipids also changed. The plasma membrane phosphatidylinositol (PI) kinase decreased 50% and PIP-phospholipase C (PIP-PLC) increased 60% compared with the control values after 5 min of hyperosmotic stress. The PIP-PLC activity recovered to control levels by 10 min; however, the PI kinase activity remained below the control value, suggesting that the cells had reached a new steady state with regard to PIP biosynthesis. If cells were pretreated with okadaic acid, the protein phosphatase 1 and 2A inhibitor, the differences in enzyme activity resulting from the hyperosmotic stress were no longer evident, suggesting that an okadaic acid-sensitive phosphatase was activated in response to hyperosmotic stress. Our work suggests that, in this system, PIP is not involved in the initial response to hyperosmotic stress but may be involved in the recovery phase.     

Localization of the Functional p34cdc2 Homolog of Maize in Root Tip and Stomatal Complex Cells: Association with Predicted Division Sites

We have used an antibody against the functional homolog of the cdc2 kinase of maize to localize the p34cdc2 protein within dividing cells of the root apex and the stomatal complex of leaf epidermis. The microtubule cytoskeletal structure of plant cells was visualized concomitantly with a monoclonal antibody specific for [alpha]-tubulin. We found that the cdc2 protein is localized mainly to the nucleus in plant cells at interphase and early prophase. This finding contrasts markedly with the predominantly cytoplasmic staining obtained using antibody to the PSTAIRE motif, which is common to cdc2 and numerous cdc2-like proteins. In a subpopulation of root cells at early prophase, the p34cdc2 protein is also distributed in a band bisecting the nucleus. Double labeling with the maize p34cdc2Zm antibody and tubulin antibody revealed that this band colocalizes with the preprophase band (PPB) of microtubules, which predicts the future division site. Root cells in which microtubules had been disrupted with oryzalin did not contain this band of p34cdc2 protein, suggesting that formation of the microtubule PPB is necessary for localization of the p34cdc2 kinase to the plane of the PPB. The p34cdc2 protein is also localized to the nucleus and PPB in cells that give rise to the stomatal complex, including those cells preparing for the highly asymmetrical divisions that produce subsidiary cells. Association of the p34cdc2 protein with the PPB suggests that the cdc2 kinase has a role in establishing the division site of plant cells and, therefore, a role in plant morphogenesis.     

Fine-scale mapping of physicochemical and microbial landscapes of the coral skeleton

The coral skeleton harbours a diverse community of bacteria and microeukaryotes exposed to light, O₂ and pH gradients, but how such physicochemical gradients affect the coral skeleton microbiome remains unclear. In this study, we employed chemical imaging of O₂ and pH, hyperspectral reflectance imaging and spatially resolved taxonomic and inferred functional microbiome characterization to explore links between the skeleton microenvironment and microbiome in the reef-building corals Porites lutea and Paragoniastrea benhami. The physicochemical environment was more stable in the deep skeleton, and the diversity and evenness of the bacterial community increased with skeletal depth, suggesting that the microbiome was stratified along the physicochemical gradients. The bulk of the coral skeleton was in a low O₂ habitat, whereas pH varied from pH 6–9 with depth. Physicochemical gradients of O₂ and pH of the coral skeleton explained the β-diversity of the bacterial communities, and skeletal layers that showed O₂ peaks had a higher relative abundance of endolithic algae, reflecting a link between the abiotic environment and the microbiome composition. Our study links the physicochemical, microbial and functional landscapes of the coral skeleton and provides new insights into the involvement of skeletal microbes in the coral holobiont metabolism.     

Lisofylline prevents leak, but not neutrophil accumulation, in lungs of rats given IL-1 intratracheally

Hybertson, Brooks M., Stuart L. Bursten, Jonathan A. Leff,Young M. Lee, Eric K. Jepson, Chris R. Dewitt, John Zagorski, Hyun G. Cho, and John E. Repine. Lisofylline prevents leak, but not neutrophil accumulation, in lungs of rats given IL-1intratracheally. J. Appl. Physiol.82(1): 226-232, 1997.Interleukin-1 (IL-1) is increased in lunglavages from patients with the acute respiratory distress syndrome, andadministering IL-1 intratracheally causes neutrophil accumulation and aneutrophil-dependent oxidative leak in lungs of rats. In the presentstudy, we found that rats pretreated intraperitoneally with lisofylline[(R)-1-(5-hydroxyhexyl)-3,7-dimethylxanthine (LSF)], an inhibitor of lysophosphatidic acid acyl transferase, which reduces the production of unsaturated phosphatidic acid species,did not develop the lung leak or the related ultrastructural abnormalities that occur after intratracheal administration of IL-1.However, rats pretreated with LSF and then given IL-1 intratracheally did develop the same elevations of lung lavage cytokine-induced neutrophil chemoattractant (CINC) levels and the same increased numbersof lung lavage neutrophils as rats given IL-1 intratracheally. Lungs ofrats given IL-1 intratracheally also had increased unsaturated phosphatidic acid and free acyl (linoleate, linolenate) concentrations compared with untreated rats, and these lipid responses were prevented by pretreatment with LSF. Our results reveal that LSF decreases lungleak and lung lipid alterations without decreasing neutrophil accumulation or lung lavage CINC increases in rats given IL-1 intratracheally.