Fibroblast growth factor receptor expression in outer hair cells of rat cochlea

FIBROBLAST growth factors (FGFs) are critical for normal development of the organ of Corti, and may also protect hair cells from ototoxic damage. Four different fibroblast growth factors are known, three of which have different splice variants in the extracellular immunoglobin-like (Ig) III FGF-binding domain, giving different patterns of sensitivity to the different FGFs. Analysis of a cDNA library of rat outer hair cells by the polymerase chain reaction, using isoform specific primers, showed expression only of FGF receptor 3, splice variant IIIc. This allows us to predict the pattern of sensitivity to applied FGFs, may be useful in targeting outer hair cells selectively during an FGF-based strategy for cochlear therapy. (C) 1998 Lippincott Williams & Wilkins.

Role of the 6-20 disulfide bridge in the structure and activity of epidermal growth factor

Two synthetic analogues of murine epidermal. growth factor, [Abu6, 20] mEGF4-48 (where Abu denotes amino-butyric acid) and [G1, M3, K21, H40] mEGF1-48, have been investigated by NMR spectroscopy. [Abu6, 20] mEGF4-48 was designed to determine the contribution of the 6-20 disulfide bridge to the structure and function of mEGF The overall structure of this analogue was similar to that of native mEGF, indicating that the loss of the 6-20 disulfide bridge did not affect the global fold of the molecule. Significant structural differences were observed near the N-terminus, however, with the direction of the polypeptide chain between residues four and nine being altered such that these residues were now located on the opposite face of the main beta-sheet from their position in native mEGF Thermal denaturation experiments also showed that the structure of [Abu6, 20] mEGF4-48 was less stable than that of mEGF. Removal of this disulfide bridge resulted in a significant loss of both mitogenic activity in Balb/c 3T3 cells and receptor binding on A431 cells compared with native mEGF and mEGF4-48, implying that the structural changes in [Abu6, 20] mEGF4-48, although limited to the N-terminus, were sufficient to interfere with receptor binding. The loss of binding affinity probably arose mainly from steric interactions of the dislocated N-terminal region with part of the receptor binding surface of EGF [G1, M3, K21, H40] mEGF1-48 was also synthesized in order to compare the synthetic polypeptide with the corresponding product of recombinant expression. Its mitogenic activity in Balb/c 3T3 cells was similar to that of native mEGF and analysis of its H-1 chemical shifts suggested that its structure was also very similar to native.

Growth hormone binding protein correlates strongly with leptin and percentage body fat in GH-deficient adults, is increased by GH replacement but does not predict IGF-I response

GH-binding protein (GHBP) corresponds to the extracellular domain of the GH receptor (GHR) and has been shown to be closely related to body fat. This study aimed to examine the inter-relationship between GHBP, leptin and body fat, and to test the hypothesis that GHBP is modified by GH replacement in GH-deficient adults and predicts IGF-I response. Twenty adults, mean age 47 years (range 20-69) with proven GH deficiency were randomly allocated to either GH (up to 0.25 U/kg/week in daily doses) or placebo for 3 months before cross-over to the opposite treatment. Plasma GHBP and leptin were measured at baseline and 2, 4, 8 and 12 weeks after each treatment. Whole body composition was measured at baseline by dual-energy X-ray absorptiometry (DEXA). There was a strong correlation between baseline leptin and GHBP (r = 0.88, P < 0.0001) and between baseline GHBP and percentage body fat, (r = 0.83, P < 0.0001). Mean GHBP levels were higher on GH compared with placebo, 1.53 +/- 0.28 vs 1.41 +/- 0.25 nM, P = 0.049. There was no correlation between baseline IGF-I and GHBP (r = -0.049, P = 0.84), and GHBP did not predict IGF-I response to GH replacement. The close inter-relationship between GHBP, leptin and body fat suggests a possible role for GHBP in the regulation of body composition. GHBP is increased by GH replacement in GH-deficient adults, but does not predict biochemical response to GH replacement. (C) 1999 Churchill Livingstone.

Growth and lactic acid production in batch culture of Lactobacillus rhamnosus in a defined medium

To facilitate metabolic analysis, batch fermentations of Lactobacillus rhamnosus were carried out in a new defined medium. Biomass at 10.5 g/l and lactic acid at 67 g/l with a Y-P/S of 0.84 were achieved. The maximum specific growth rate and the average productivity were 0.49/h and 2.48 g/l.h, respectively. These are comparable to those of this organism and related organisms in complex media. Preliminary amino acid studies were also conducted, highlighting the importance of serine, asparagine, glutamine and cysteine. Kinetic analysis revealed that lactic acid production was predominantly growth-associated with growth associated and non-growth associated lactic acid constants of 0.389 mol/g-cell and 0.0025 mol/g-cell.h, respectively. Finally a kinetic model has been included to describe the fermentation of L. rhamnosus.

Distribution of interleukin-2,-4,-10, tumour necrosis factor-alpha and transforming growth factor-beta mRNAs in oral lichen planus

In the present study. MRNA for the cytokines interleukin-2 (IL-2), IL-4, IL-10 tumour necrosis factor-alpha (TNF-alpha) and transforming growth factor beta-1 (TGF-beta-1) were investigated in oral lichen planus (OLP) lesions using in situ hybridization with S-35-labelled oligonucleotide probes on frozen tissue sections. In addition, the expression of interferon-gamma (IFN-gamma), IL-10 and IL-4 mRNAs was analysed in cultured lesional T lymphocytes from oral lichen planus by polymerase chain reaction. Cells expressing mRNA for IL-2, IL-4, IL-10, TNF-alpha and TGF-beta(1) were found in all the biopsies studied. Approximately 1-2% of the total number of infiltrating cells in the lesions were positive for each of the different cytokine mRNAs. Most biopsies contained basement membrane-oriented, mRNA-positive cells. In the cultured T-cell lines, message for IFN-gamma was detected in all the patients, IL-10 in all but one, and IL-4 in just one of the seven patients investigated. The results suggest that mRNA for both pro- and anti-inflammatory cytokines, i.e., mixed T-helper 1 (T(H)1) and T(H)2 cytokine profiles, are generated simultaneously by a limited number of cells in chronic lesions of OLP. (C) 1999 Elsevier Science Ltd. All rights reserved.

Errors in lamina growth of primary olfactory axons in the rat and mouse olfactory bulb

In the adult olfactory nerve pathway of rodents, each primary olfactory axon forms a terminal arbor in a single glomerulus in the olfactory bulb. During development, axons are believed to project directly to and terminate precisely within a glomerulus without any exuberant growth or mistargeting. To gain insight into mechanisms underlying this process, the trajectories of primary olfactory axons during glomerular formation were studied in the neonatal period. Histochemical staining of mouse olfactory bulb sections with the lectin Dolichos biflorus-agglutinin revealed that many olfactory axons overshoot the glomerular layer and course into the deeper laminae of the bulb in the early postnatal period. Single primary olfactory axons were anterogradely labelled either with the lipophilic carbocyanine dye, 1,1'-dioctodecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (DiI), or with horseradish peroxidase (HRP) by localized microinjections into the nerve fiber layer of the rat olfactory bulb. Five distinct trajectories of primary olfactory axons were observed in DLI-labelled preparations at postnatal day 1.5 (P1.5). Axons either coursed directly to and terminated specifically within a glomerulus, branched before terminating in a glomerulus, bypassed glomeruli and entered the underlying external plexiform layer, passed through the glomerular layer with side branches into glomeruli, or branched into more than one glomerulus. HRP-labelled axon arbors from eight postnatal ages were reconstructed by camera lucida and were used to determine arbor length, arbor area, and arbor branch number. Whereas primary olfactory axons display errors in laminar targeting in the mammalian olfactory bulb, axon arbors typically achieve their adult morphology without exuberant growth. Many olfactory axons appear not to recognize appropriate cues to terminate within the glomerular layer during the early postnatal period. However, primary olfactory axons exhibit precise targeting in the glomerular layer after P5.5, indicating temporal differences in either the presence of guidance cues or the ability of axons to respond to these cues. (C) 1999 Wiley-Liss, Inc.

X-ray computed tomography to quantify tree rooting spatial distributions

Poor root development due to constraining soil conditions could be an important factor influencing health of urban trees. Therefore, there is a need for efficient techniques to analyze the spatial distribution of tree roots. An analytical procedure for describing tree rooting patterns from X-ray computed tomography (CT) data is described and illustrated. Large irregularly shaped specimens of undisturbed sandy soil were sampled from Various positions around the base of trees using field impregnation with epoxy resin, to stabilize the cohesionless soil. Cores approximately 200 mm in diameter by 500 mm in height were extracted from these specimens. These large core samples were scanned with a medical X-ray CT device, and contiguous images of soil slices (2 mm thick) were thus produced. X-ray CT images are regarded as regularly-spaced sections through the soil although they are not actual 2D sections but matrices of voxels similar to 0.5 mm x 0.5 mm x 2 mm. The images were used to generate the equivalent of horizontal root contact maps from which three-dimensional objects, assumed to be roots, were reconstructed. The resulting connected objects were used to derive indices of the spatial organization of roots, namely: root length distribution, root length density, root growth angle distribution, root spatial distribution, and branching intensity. The successive steps of the method, from sampling to generation of indices of tree root organization, are illustrated through a case study examining rooting patterns of valuable urban trees. (C) 1999 Elsevier Science B.V. All rights reserved.

Plant latex and first-instar monarch larval growth and survival on three North American milkweed species

First-instar larvae of the monarch butterfly, Danaus plexippus, a milkweed specialist, generally grew faster and survived better on leaves when latex flow was reduced by partial severance of the leaf petiole. The outcome depended on milkweed species and was related to the amount of latex produced. The outcome also may be related to the amount of cardenolide produced by the plants as a potential chemical defense against herbivory. Growth was more rapid, but survival was similar on partially severed compared with intact leaves of the high-latex/low-cardenolide milkweed, Asclepias syriaca, whereas both growth and survival were unaffected on the low-latex/low-cardenolide milkweed A. incarnata. On the low-latex/low-cardenolide milkweed A. tuberosa, both growth and survival of larvae were only marginally affected. These results contrast sharply to previous results with the milkweed, A. humistrata, in Florida, which has both high latex and high cardenolide. Larval growth and survival on A. humistrata were both increased by partially severing leaf petioles. Larval growth rates among all four milkweed species on leaves with partially severed petioles were identical, suggesting that latex and possibly the included cardenolides are important in first-instar monarch larval growth, development, and survivorship.

Administration of growth hormone or IGF-I to pregnant rats on a reduced diet throughout pregnancy does not prevent fetal intrauterine growth retardation and elevated blood pressure in adult offspring

Increasing evidence from human epidemiological studies suggests that poor growth before birth is associated with postnatal growth retardation and the development of cardiovascular disease in adulthood. We have shown previously that nutritional deprivation in the pregnant rat leads to intrauterine growth retardation (IUGR), postnatal growth failure, changes in the endocrine parameters of the somatotrophic axis, and to increased blood pressure in later life. In the present study, we investigated whether administration of insulin-like growth factor-I (IGF-I) or bovine growth hormone (GH) during pregnancy could prevent IUGR and/or alter long-term outcome. Dams h-om day 1 of pregnancy throughout gestation received a diet of nd libitum available food or a restricted dietary intake of 30% of ad libitum fed dams. From day 10 of gestation, dams were treated for 10 days with three times daily subcutaneous injections of saline (100 mu l), IGF-I (2 mu g/g body weight) or GH (2 mu g/g body weight). Maternal weight gain was significantly increased (P

The last glacial cycle from the humid tropics of northeastern Australia: comparison of a terrestrial and a marine record

A detailed pollen record from the Ocean Drilling Program Site 820 core, located on the upper part of the continental slope off the coast of northeast Queensland, was constructed to compare with the existing pollen record from Lynch's Crater on the adjacent Atherton Tableland and allow the production of a regional picture of vegetation and environmental change through the last glacial cycle. Some broad similarities in patterns of vegetation change are revealed, despite the differences between sites and their pollen catchments, which can be related largely to global climate and sea-level changes. The original estimated time scale of the Lynch's Crater record is largely confirmed from comparison with the more thoroughly dated ODP record. Conversely, the Lynch's Crater pollen record has assisted in dating problematic parts of the ODP record. In contrast to Lynch's Crater, which reveals a sharp and sustained reduction in drier araucarian forest around 38,000 yrs BP, considered to have been the result of burning by Aboriginal people, the ODP record indicates, most likely, a stepwise reduction, dating from 140,000 yrs BP or beyond. The earliest reduction shows lack of a clear connection between Araucaria decline and increased burning and suggests that people may not have been involved at this stage. However, a further decline in araucarian forest, possibly around 45,000 yrs BP, which has a more substantial environmental impact and is not related to a time of major climate change, is likely, at least partially, the result of human burning. The suggestion, from the ODP core oxygen isotope record, of a regional sea-surface temperature increase of around 4 degrees C between about 400,000 and 250,000 yrs BP, may have had some influence on the overall decline in Araucaria and its replacement by sclerophyll vegetation. (C) 2000 Elsevier Science B.V. All rights reserved.

Chemical synthesis, characterization and activity of RK-1, a novel alpha-defensin-related peptide

The 32-residue peptide, RK-1, a novel kidney-derived three disulfide-bonded member of the antimicrobial alpha-defensin family, was synthesized by the continuous now Fmoc-solid phase method. The crude, cleaved and S-reduced Linear peptide was both efficiently folded and oxidized in an acidic solution of aqueous dimethyl sulfoxide. Following purification of the resulting product, it was shown by a variety of analytical techniques, including matrix assisted laser desorption time of flight mass spectrometry, to possess a very high degree of purity. The disulfide bond pairing of the synthetic peptide was determined by H-1-NMR spectroscopy and confirmed to be a Cys(1)-Cys(6), Cys(2)-Cys(4), Cys(3)-Cys(5) arrangement similar to other mammalian alpha-defensin peptides. The synthetic RK-1 was also shown to inhibit the growth of Escherichia coli type strain NCTC 10418, Copyright (C) 2000 European Peptide Society and John Wiley & Sons, Ltd.

The salicylate-derived mycobactin siderophores of Mycobacterium tuberculosis are essential for growth in macrophages

Mycobacterium tuberculosis is an important pathogen of mammals that relies on 2-hydroxyphenyloxazoline-containing siderophore molecules called mycobactins for the acquisition of iron in the restrictive environment of the mammalian macrophage, These compounds have been proposed to be biosynthesized through the action of a cluster of genes that include both nonribosomal peptide synthase and polyketide synthase components. One of these genes encodes a protein, MbtB, that putatively couples activated salicylic acid with serine or threonine and then cyclizes this precursor to the phenyloxazoline ring system. We have used gene replacement through homologous recombination to delete the mbtB gene and replace this with a hygromycin-resistance cassette in the virulent strain of M. tuberculosis H37Rv, The resulting mutant is restricted for growth in iron-limited media but grows normally in iron-replete media. Analysis of siderophore production by this organism revealed that the biosynthesis of all salicylate-derived siderophores was interrupted. The mutant was found to be impaired for growth in macrophage-like THP-1 cells, suggesting that siderophore production is required for virulence of M. tuberculosis, These results provide conclusive evidence linking this genetic locus to siderophore production.

The influence of supra-optimal root-zone temperatures on growth and stomatal conductance in Capsicum annuum L.

Pepper (Capsicum annuum L.) plants were grown aeroponically in a Singapore greenhouse under natural diurnally fluctuating ambient shoot temperatures, but at two different root-zone temperatures (RZTs): a constant 20 +/- 2 degrees C RZT and a diurnally fluctuating ambient (A) (25-40 degrees C) RZT, Plants grown at 20-RZT had more leaves, greater leaf area and dry weight than A-RZT plants. Reciprocal transfer experiments were conducted between RZTs to investigate the effect on plant growth, stomatal conductance (g(s)) and water relations. Transfer of plants from A-RZT to 20-RZT increased plant dry weight, leaf area, number of leaves, shoot water potential (Psi(shoot)), and g(s); while transfer of plants from 20-RZT to A-RZT decreased these parameters. Root hydraulic conductivity was measured in the latter transfer and decreased by 80% after 23 d at A-RZT. Transfer of plants from 20-RZT to A-RZT had no effect on xylem ABA concentration or xylem nitrate concentration, but reduced xylem sap pH by 0.2 units. At both RZTs, g(s) measured in the youngest fully expanded leaves increased with plant development. In plants with the same number of leaves, A-RZT plants had a higher g(s) than 20-RZT plants, but only under high atmospheric vapour pressure deficit. The roles of chemical signals and hydraulic factors in controlling g(s) of aeroponically grown Capsicum plants at different RZTs are discussed.