Purification and characterization of a growth inhibitory hepatic peptide. A preliminary note

A pentapeptide isolated from normal mouse liver seems to inhibit DNA synthesis (3H-thymidine incorporation into liver DNA and labeling indices) and the mitotic rate (G2-M cell flux) in regenerating mouse liver. The inhibitor is somewhat similar to the growth inhibitory pentapeptides previously reported for granulocytes and epidermis. It is active at very low dose levels, showing a bell-shaped dose-response curve.     

Characterization of a biologically active arabinogalactan from the leaves of Plantago major L.

PMIa is a Type II arabinogalactan with anti-complementary activity isolated from the leaves of Plantago major L. It has a molecular weight of 77000–80000 Da and consists of arabinose (38%), galactose (49%), rhamnose (6%), galacturonic acid (7%) and 1.5% protein with hydroxyproline, alanine and serine as the main amino acids. Characterization of PMIa by methylation and GC-MS, methanolysis and GC, Smith degradation, weak acid hydrolysis, ¹³C-NMR, ¹H-NMR, two-dimensional heteronuclear NMR and DEPT show that it consists of 1,3-linked galactan chains with 1,6-linked galactan side chains attached to position 6. The side chains are further branched in position 3 with 1,3-linked galactose residues which have 1,6-linked galactose attached to position 6; these 1,3- and 1,6-linked galactose chains altogether probably form a network. Terminal and 1,5-linked arabinose in furanose form are attached to the galactan mainly through position 3 of the 1,6-linked galactose side chains.     

Residual morphogenetic competence in the proximal core region of stage 25 chick limbs: Zwilling's hypothesis revisited

Morphogenetic competence (MC) exists in embryonic limb tissue once thought to have lost this property as a consequence of cytodifferentiation. By stage 25 of chick embryonic development, cells in the proximal core of the limb have committed to the cartilage phenotype and are producing their characteristic extracellular matrix. Recombinant limb-bud grafts constructed using isolated fragments of this tissue produce outgrowths with a limb-like skeletal pattern. Inclusion of proximal peripheral tissue in the grafts (with or without the polarizing tissue) inhibits outgrowth and skeletal morphogenesis, explaining the failure of earlier studies to reveal the MC of the proximal core (chondrogenic) cells. Since definitive chondroblasts express MC in more permissive surroundings, it appears that Zwilling's assertion, that the onset of cytodifferentiation causes the loss of MC, is an oversimplification and that complex tissue interactions are probably involved.     

Molecular Modeling of Cytochrome P450 1A1: Enzyme-Substrate Interactions and Substrate Binding Affinities

Abstract

Human cytochrome P450 1A1, which is present in lungs, plays an important role in the metabolic activation of chemical carcinogens, and in particular, is thought to be linked to lung cancer. The mechanism of carcinogenesis is related to the enzyme's ability to oxidize highly toxic compounds, such as polycyclic aromatic hydrocarbons (PAHs), to their carcinogenic derivatives.

In order to better understand P450 1A1 function, a homology model of this enzyme has been constructed. The model has been based on the structure of P450 2C5, the first mammalian P450 to be crystallized. The coordinates of the model have been calculated using a consensus strategy, and the resulting structure has been evaluated with the ProStat and Profiles-3D programs. P450 1A1 substrates, such as benzo[a]pyrene, ethoxyresorufin and methoxyre- sorufin, were then docked into the active site of the model, and key amino acid residues able to interact with the substrate, have been identified. The analysis of enzyme-substrate interactions indicated that hydrophobic interactions are mainly responsible for binding of these substrates in the active site. Moreover, the non-bond enzyme-substrate interaction energy for ethoxyresorufin was lower than that for methoxyresorufin, which is consistent with higher activity of 1A1 towards the former substrate. Key residue Val-382 may play an important role in these interactions. Additionally, we performed binding free energy calculations for the three substrates. The obtained values were similar to those observed experimentally, which suggests that this approach might be useful for prediction of binding constants.     

Phenotypic Profiling of Scedosporium aurantiacum,an Opportunistic Pathogen Colonizing Human Lungs

Genotyping studies of Australian Scedosporium isolates have revealed the strong prevalence of a recently described species: Scedosporium aurantiacum. In addition to occurring in the environment, this fungus is also known to colonise the respiratory tracts of cystic fibrosis (CF) patients. A high throughput Phenotype Microarray (PM) analysis using 94 assorted substrates (sugars, amino acids, hexose-acids and carboxylic acids) was carried out for four isolates exhibiting different levels of virulence, determined using a Galleria mellonella infection model. A significant difference was observed in the substrate utilisation patterns of strains displaying differential virulence. For example, certain sugars such as sucrose (saccharose) were utilised only by low virulence strains whereas some sugar derivatives such as D-turanose promoted respiration only in the more virulent strains. Strains with a higher level of virulence also displayed flexibility and metabolic adaptability at two different temperature conditions tested (28 and 37°C). Phenotype microarray data were integrated with the whole-genome sequence data of S. aurantiacum to reconstruct a pathway map for the metabolism of selected substrates to further elucidate differences between the strains.     

Osteopontin Is Induced by TGF-β2 and Regulates Metabolic Cell Activity in Cultured Human Optic Nerve Head Astrocytes

The aqueous humor (AH) component transforming growth factor (TGF)-β2 is strongly correlated to primary open-angle glaucoma (POAG), and was shown to up-regulate glaucoma-associated extracellular matrix (ECM) components, members of the ECM degradation system and heat shock proteins (HSP) in primary ocular cells. Here we present osteopontin (OPN) as a new TGF-β2 responsive factor in cultured human optic nerve head (ONH) astrocytes. Activation was initially demonstrated by Oligo GEArray microarray and confirmed by semiquantitative (sq) RT-PCR, realtime RT-PCR and western blot. Expressions of most prevalent OPN receptors CD44 and integrin receptor subunits αV, α4, α 5, α6, α9, β1, β3 and β5 by ONH astrocytes were shown by sqRT-PCR and immunofluorescence labeling. TGF-β2 treatment did not affect their expression levels. OPN did not regulate gene expression of described TGF-β2 targets shown by sqRT-PCR. In MTS-assays, OPN had a time- and dose-dependent stimulating effect on the metabolic activity of ONH astrocytes, whereas TGF-β2 significantly reduced metabolism. OPN signaling via CD44 mediated a repressive outcome on metabolic activity, whereas signaling via integrin receptors resulted in a pro-metabolic effect. In summary, our findings characterize OPN as a TGF-β2 responsive factor that is not involved in TGF-β2 mediated ECM and HSP modulation, but affects the metabolic activity of astrocytes. A potential involvement in a protective response to TGF-β2 triggered damage is indicated, but requires further investigation.     

On the chemical stability of beta-hydroxyphenylalkylhydroxylamines and its relevance to the metabolism of amphetamines and ephedrines.

β, N-bis (hydroxy) phenylalkylamines rapid oxidative decomposition to benzaldehyde and an oxime in the presence of small quantities of Cu(II). The reaction occurs in aqueous solution at pH 7.4 so that products of metabolic transformation reactions involving the oxidation of such hydroxylamines would be expected to decompose in this way. N-Oxidation of norephedrine would result in benzaldehyde by this mechanism, and since benzaldehyde is a precursor to benzoic acid, it is proposed that the N-hydroxylation pathway of arylalkylamine metabolism instead of carbon oxidation could lead to benzoic acid. This acid is a major metabolite of compounds such as amphetamine and ephedrine in some species.     

Capitalizing on multi-element interactions through bal-anced nutrition——A pathway to improve nitrogen use efficiency in China,India and North America

A viable option for increasing nitrogen (N) use efficiency and mitigation of negative impacts of N on the environment is to capitalize on multi-element interactions through implementation of nutrient management programs that provide balanced nutrition. Numerous studies have demonstrated the immediate efficacy of this approach in the developing regions like China and India as well as developed countries in North America. Based on 241 site-years of experiments in these countries, the first-year N recovery efficiency (RE) for the conventional or check treatments averaged 21% while the balanced treatments averaged 54% RE, for an average increase of 33% in RE due to balanced nutrition. Effective policies to promote adoption are most likely those that enable site-specific approaches to nutrient management decisions rather than sweeping, nation-wide incentives supporting one nutrient over another. Local farmers, advisers and officials need to be empowered with tools and information to help them define necessary changes in practices to create more balanced nutrient management.     

Adenomatous polyposis coli influences micronuclei induction by PhIP and acrylamide in mouse erythrocytes

Micronucleus (MN) induction in erythrocytes of multiple intestinal neoplasia (Min) mice with heterozygous Apc mutation was measured after s.c. injections of acrylamide, glycidamide, 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) and colchicine, and compared with wild-type (wt) mice. Since Apc influences microtubule dynamics, we wanted to test whether Min-mice were more sensitive to the production of MN than wild-type mice. We also examined the effect of pre-treatment with cytosine β-D-arabinofuranoside (Ara C) and hydroxyurea, which inhibit ligation of DNA strand breaks in the repair of DNA adducts. All compounds induced a significant increase in MN in both strains of mice with the following potencies: acrylamide < glycidamide < PhIP. No difference in the induction of MN was seen between Min-mice and wt-mice exposed to acrylamide, glycidamide or colchicine without pre-treatment. However, in Min-mice, PhIP treatment induced much less MN than in wt-mice, with about four- and six-fold increase in MN in Min-mice and wt-mice, respectively. A reduced ability to repair PhIP adducts may be the reason for the lower induction of MN in Min-mice. Treatment with Ara C and hydroxyurea, to increase sensitivity, gave more than a four-fold increase in MN, but strongly reduced proliferation. Pre-treatment with Ara C and hydroxyurea made the Min-mice slightly more sensitive to MN induction by glycidamide compared to wt-mice. We conclude that Min-mice are less sensitive than wt-mice to MN induction by PhIP that forms bulky DNA adducts, while Min-mice and wt-mice are equally sensitive to MN induction by acrylamide and glycidamide that form DNA base adducts.