Inhibition of the Ser-Thr phosphatases PP1 and PP2A by naturally occurring toxins

The okadaic acid class of naturally occurring toxins is a structurally diverse group of molecules that inhibit the protein phosphatases PP1 and PP2A. Studies providing information about the mode of binding between the toxins and the phosphatases contribute to an overall understanding of the signal transduction pathways in which the phosphatases are involved.     

Inhibition of murine hepatic cytochrome P450 activities by natural and synthetic phenolic compounds

1. The effect of the phenolic compounds protocatechuic acid, chlorogenic acid, tannic acid, gallates and silybin on ethoxyresorufin O-dealkylase (CYP1A1), methoxyresorufin O-dealkylase (CYP1A2) and pentoxy-O-dealkylase (CYP2B) was examined in mouse liver microsomes from induced animals. 2. All compounds tested could inhibit cytochrome P450-mediated enzyme activities, but to different extents. Tannic acid was the most potent inhibitor, especially toward EROD activity with an IC50=2.6 microM. Synthetic dodecyl gallate was also relatively selective toward this enzyme activity with an IC50=120 microM. 3. Protocatechuic acid, chlorogenic and silybin were more selective towards PROD and MROD activities. Their relative inhibitory potency for PROD activity was as follows: chlorogenic acid > protocatechuic acid > silybin > dodecyl gallate > propyl gallate. Protocatechuic acid was a more effective inhibitor of MROD activity than chlorogenic acid, and propyl gallate more effective than dodecyl gallate. Thus, no clear structure-activity or selectivity relationship was observed. 4. Analysis of the kinetics of inhibition revealed that the inhibition in most cases was non-competitive in nature.     

Transformational and altered signal transduction by a naturally occurring mutant EGF receptor

An amino-truncated variant form of the epidermal growth factor receptor (EGFRvIII) has been identified in human brain, breast, lung and ovarian tumors. We have found that overexpression of this mutant EGF receptor in NIH3T3 cells results in transformation as a result of the activation of the receptor kinase via ligand-independent dimerization. Transformation was correlated with tyrosine phosphorylation of only a subset of the proteins observed in cells overexpressing the normal EGF receptor. This suggested that further studies on cells expressing the EGFRvIII might provide insights into the pathways most relevant to transformation. In clones expressing high levels of mutant EGF receptor, the levels of both Grb2 and SHC were decreased. Despite this decrease, much of the endogenous Grb2 immunoprecipitated with EGFRvIII. Interestingly, no increase in ras-GTP loading was found in clones expressing the EGFRvIII and MAP kinase assays indicated only a small increase in activity. These results indicate that high-level expression of the EGFRvIII induces down-regulation of the ras-MAP kinase pathway and that other components involved in EGF receptor signal transduction may play a greater role in neoplastic transformation by the EGFRvIII.     

Two-dimensional protein crystallization via metal-ion coordination by naturally occurring surface histidines

A powerful and potentially general approach to the targeting and crystallization of proteins on lipid interfaces through coordination of surface histidine residues to lipid-chelated divalent metal ions is presented. This approach, which should be applicable to the crystallization of a wide range of naturally occurring or engineered proteins, is illustrated here by the crystallization of streptavidin on a monolayer of an iminodiacetate-Cu(II) lipid spread at the air-water interface. This method allows control of the protein orientation at interfaces, which is significant for the facile production of highly ordered protein arrays and for electron density mapping in structural analysis of two-dimensional crystals. Binding of native streptavidin to the iminodiacetate-Cu lipids occurs via His-87, located on the protein surface near the biotin binding pocket. The two-dimensional streptavidin crystals show a previously undescribed microscopic shape that differs from that of crystals formed beneath biotinylated lipids.     

Inhibition of human leukocyte elastase by chemically and naturally oversulfated galactosaminoglycans

Several samples of oversulfated chondroitin and dermatan were obtained by chemical sulfation and by SAX-HPLC enrichment. The starting products and oversulfated products were tested as potential inhibitors of human leukocyte elastase, an enzyme hypothesized to be involved in the etiology of diseases such as emphysema, atherosclerosis, and rheumatoid arthritis. Chemical oversulfation (SO3H/COOH 1.6-3.2), preferentially occurring at C-6 of galactosamine residues, was found generally to increase the inhibitory power on elastase. Chemically oversulfated galactosaminoglycans thus have potential as therapeutic agents, considering that they produce non-significant effects on the hemocoagulative system. Two naturally oversulfated dermatans sulfate (SO3H/COOH ca. 1.2), mainly oversulfated at C-2 of iduronic acid residues, showed comparatively higher anticoagulant activity (in the HC-II mediated thrombin inhibition test).     

Binding of naturally occurring anti-DNA antibodies to estradiol

Binding of naturally occurring human anti-DNA autoantibodies to beta-estradiol and native DNA has been investigated. Sera from seven SLE patients were tested for their reactivity with beta-estradiol and native calf thymus DNA. Solid Phase enzyme immunoassay was performed using both colorigenic and fluorogenic substrates. The results show enhanced binding of human anti-DNA autoantibodies to beta-estradiol as compared to native DNA. This was further substantiated by increased binding of affinity purified SLE IgG to beta-estradiol. These results suggest a role for female sex hormone in the etiopathogenesis of SLE and are significant vis-a-vis female predominance of SLE.     

Inhibitory effects of naturally occurring coumarins on the metabolic activation of benzo[a]pyrene and 7,12-dimethylbenz[a]anthracene in cultured mouse keratinocytes

Several naturally occurring coumarins to which humans are routinely exposed have been previously found to be potent inhibitors and inactivators of cytochrome P450 (P450) 1A1-mediated monooxygenase in both murine hepatic microsomes and in a reconstituted system using purified human P450 1A1 [Cai et al. (1993) Chem. Res. Toxicol., 6, 872-879 and Cai et al. (1996) Chem. Res. Toxicol., 9, 729-736]. In the present study, several of these coumarins were investigated for their inhibitory effects on the metabolism and metabolic activation of benzo[a]pyrene (B[a]P) and 7,12-dimethylbenz[a]anthracene (DMBA) in cultured mouse keratinocytes. Initial analysis of B[a]P metabolism in cultured keratinocytes showed that imperatorin, isoimperatorin, coriandrin, and bergamottin, at concentrations of 2 nM equal with B[a]P, reduced the formation of water-soluble metabolites of B[a]P by 33% to 57%. Bergamottin and coriandrin were the most potent inhibitors of the compounds examined. HPLC analysis of organic solvent-soluble metabolites of B[a]P indicated that all the coumarins tested significantly reduced the formation of individual B[a]P metabolites (including phenols, diols and tetraols). However, the greatest effect was on the formation of B[a]P tetraols. Additional experiments determined the ability of selected coumarins to block covalent binding of B[a]P and DMBA to DNA in keratinocytes. Bergamottin preferentially inhibited the binding of B[a]P to DNA by 56%, while coriandrin preferentially inhibited the binding of DMBA to DNA by 48%. Notably, analysis of individual DNA adducts formed from B[a]P and DMBA indicated that both bergamottin and coriandrin specifically inhibited the formation of anti diol-epoxide DNA adducts derived from both hydrocarbons. The preferential inhibitory effect of bergamottin and coriandrin on the formation of anti diol-epoxide adducts derived from DMBA was further confirmed by separation of anti- and syn-diol-epoxide-DNA adducts using immobilized boronate chromatography. The current study demonstrates that certain naturally occurring coumarins inhibited metabolic activation of B[a]P and DMBA in cultured mouse keratinocytes and specifically inhibited the formation of DNA adducts derived from the anti diol-epoxide diastereomers from either hydrocarbon. The current data also suggest that certain naturally occurring coumarins may possess anticarcinogenic activity toward polycyclic aromatic hydrocarbons.     

Inhibition of murine neutrophil serine proteinases by human and murine secretory leukocyte protease inhibitor

Human secretory leukocyte protease inhibitor (SLPI) is a predominant physiologic inhibitor of elastase and cathepsin G, proinflammatory serine proteases released by activated neutrophils. In order to fully evaluate the potential pharmacologic efficacy of human SLPI in animal models of inflammation, it is critical to know the potency of the inhibitor for corresponding proteases from the species of interest. In this report, we compare the inhibitory activity of human and murine SLPI against elastase and cathepsin G from both species. Human and murine neutrophil elastase and cathepsin G display comparable Km values for their specific peptide substrates. Murine SLPI inhibits murine neutrophil elastase and cathepsin G with Ki values of 5 and 0.12 nM, respectively, while human SLPI inhibits the both murine serine proteases with Ki's of 0.02 nM. In contrast, murine SLPI inhibits human neutrophil elastase and cathepsin G with Ki values of 1.4 and 90 nM, respectively, while human SLPI inhibits the proteases with Ki's of 0.3 and 10 nM, respectively. These results demonstrate species-specific variations in the protease inhibitory activities of SLPI. Such variations should be considered in the evaluation of the activity of human SLPI in murine pharmacologic models.     

An investigation of naturally occurring episodes of self-controlled behaviors.

The naturally occurring (i.e., self-initiated) episode of self-controlled behavior is a common experience that has received little attention from researchers. Through a structured interview procedure, a demographic questionnaire, and a paper-and-pencil test (Rotter's Internal-External Locus of Control Scale), the present study determined those factors and methods that are crucial to successful self-management. 48 male and 48 female college students were interviewed about their attempts to self-control overeating, smoking, studying, or dating; 12 "successful" and 12 "unsuccessful" Ss were interviewed in each problem area. Results indicate the following: (a) Successful self-controllers reported using more techniques for longer periods of time. (b) The use of self-reinforcement procedures was an important discriminant of successful self-management. (c) The methods used by successful self-controllers seemed to vary according to the problem with which they were dealing. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Comparative effects of various naturally occurring cannabinoids on food, sucrose and water consumption by rats

The effects of intraperitoneally injected detla9-tetrahydrocannabinol (THC), cannabinol (CBN) and cannabidiol (CBD) were compared to d-amphetamine sulfate (d-AMP) on food and water consumption and intake of two different concentrations of sucrose solutions. Three groups of rats were given the following dietary regimens within a 6-hr feed period day: 1 - water and dry food; 2 - water, dry food and five percent sucrose solution; 3 - water, dry food and 20% sucrose solution. Food and water consumption were dramatically reduced by each test drug at feeding periods immediately following and in some instances up to 4 days after dosing in all 3 groups. However, sucrose consumption was much less affected by each cannabinoid, inidcating a preference for sweet calories, whereas d-AMP had an equal anorexic action on both food and sucrose consumption. These data suggest for the first time in rats that a preference for sweet calories occurs during an overall anorexic effect of THC, CBN and CBD.     

Behavioral response to naturally occurring stress in cyclothymia and dysthymia.

Outlines an approach to epidemiologic-type life-stress studies that involves the intensive study of smaller groups of well-defined disordered individuals across the course of the response. On the basis of their previous research, the authors suggest that cyclothymia may be characterized by poor regulation of biologic and behavioral variation and that this condition may be exaggerated by stress. To extend the evidence on poor regulation generated in laboratory studies of cortisol functioning, 31 cyclothyme patients and 24 normal controls (median age 19.3 yrs) were asked to provide daily ratings of moods and events over 28 consecutive days. Data show that Ss with cyclothymia showed prolonged duration of recovery of behavioral levels compared to controls. Findings support a notion of weak inhibitory modulation in certain CNS systems that control biologic and behavioral variables related to affective disorder. It is suggested that the prolonged duration of behavioral change in cyclothymia is unlikely to be the consequence of concurrent depression and that the nature of prolonged recovery to stress in cyclothymia reflects the operation of an endogenous regulatory process, as opposed to a cognitive appraisal bias. (33 ref) (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Induction of murine hepatic glutathione S-transferase by dietary dehydroepiandrosterone

The naturally occurring steroid dehydroepiandrosterone (DHEA), when administered as a supplement to the diet of mice and rats, produces alterations in the relative concentrations of specific liver proteins; among these, a protein of Mr approximately 28 K is markedly induced by DHEA action. In the present study we identified the murine hepatic approximately 28 kDa protein as glutathione S-transferase subtype GT-8.7. Glutathione S-transferases belong to a gene superfamily that encode closely related proteins which are induced in liver and other tissues by various chemicals, including carcinogens and chemoprotective agents such as dietary antioxidants. Based on the above finding, we evaluated glutathione S-transferase activity in cytosols and microsomes prepared from liver tissue of mice fed either a control diet or a DHEA-containing diet (0.45%, by weight). The specific activity of hepatic cytosolic glutathione S-transferase in mice treated with DHEA up to 7 days was either unchanged or slightly decreased when compared to controls; however, treatment for 14 days or longer resulted in significant increases in activity. The specific activity of microsomal glutathione S-transferase also was increased by long-term DHEA treatment; however, its activity was approximately one-tenth of that in corresponding cytosols.     

Effects of age and naturally occurring experience on spatial visualization performance.

A questionnaire designed to assess experience with activities presumed to require spatial visualization abilities, and psychometric tests of these abilities, were administered to 383 adults ranging from 20 to 83 years of age. Although research participants varied considerably in the amount of self-reported experience, statistical control of experience resulted in relatively modest attenuations of the relations between age and spatial visualization performance. These findings seem inconsistent with a strong disuse interpretation of cognitive aging phenomena and suggest that at least some age-related differences in cognitive functioning are independent of the amount of experience with relevant activities. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Canine hepatozoonosis in Brazil: description of eight naturally occurring cases

Eight cases of canine hepatozoonosis were diagnosed at the Veterinary Hospital (Faculdade de Medicina Veterinária e Zootecnia, Universidade Estadual Paulista, Campus de Botucatu), between October 1993 and April 1994. Clinical signs included anorexia, pale mucous membranes, weight loss, pain, diarrhoea, vomit, gait abnormalities, fever, polyuria and polydipsia. Haematologic findings revealed anaemia in seven cases, leucocytosis with neutrophilia in three cases, lymphopenia in three cases and monocytosis in four cases. Serum biochemistries included alterations in many parameters. The micrometry of Hepatozoon canis gametocytes ranged from 6.8 x 4.0 microns to 7.5 x 4.5 microns. Parasitaemia ranged from less than 0.5% to 2%. In all the cases reported other concurrent diseases were present. Diagnosis of canine hepatozoonosis was made by identifying H. canis gametocytes within leucocytes in stained blood smears.     

A naturally occurring mouse model of X-linked congenital stationary night blindness

PURPOSE: To describe a naturally occurring X-linked recessive mutation, no b-wave (nob), that compromises visual transmission between photoreceptors and second-order neurons in mice. METHODS: Affected mice were identified by recording the light-evoked response of the retina, the electroretinogram (ERG). To evaluate visual transmission, cortical potentials were recorded with a scalp electrode. The inheritance pattern for nob was defined by breeding nob animals with normal mice. Retinal histologic analysis was performed by light microscopy. RESULTS: Although the photoreceptor-mediated ERG component (a-wave) was normal in nob mice, the major response component reflecting postreceptoral neuronal activity (b-wave) was missing. Visually-driven cortical activity was also abnormal in nob animals. At the light microscopic level, the nob retina appeared to have a normal cytoarchitecture. CONCLUSIONS: These findings suggest that the nob defect interferes with the transmission of visual information through the retina and that these mice are a useful model for the study of outer retinal synaptic function. In addition, this mutant mouse seems to provide an animal model for the complete form of congenital stationary night blindness, a human disorder in which patients have a profound loss of rod-mediated visual sensitivity.     

Antitumor, immunomodulatory and anti-HIV effect of mangiferin, a naturally occurring glucosylxanthone

We proposed previously a cyclic code made of 22 triplets, which we now call the AB code. It is made up of the following chain: AUGGUGCCAUUCAAGACUAUGA. The letters A, U, C, G represent the classical symbols of the (purine and pyrimidine) bases of the genetic code. This chain presents the following features: (1) when it is in cyclic form, it begins with the initiation codon AUG, ends with the termination codon UGA, and it can be read triplet after triplet by choosing 1 and only 1 representative of each synonymy class in the classical degenerate genetic code made of 64 triplets. The chain, therefore, possesses 1 and only 1 codon for each amino-acid; (2) except for the doublet CG, triplets of the chain begin with the 15 other possible doublets of bases (satisfying the "wobble" hypothesis presented by Crick); (3) it corresponds (except for 1 base) to the "loop" part of the CEnothera mitochondrial Gly-tRNA; (4) it can be modified, without loss of the properties (1) and (2), in such a way as to have 15 bases in common with the loop part of other mitochondrial tRNA's considered as primitive, like Ala-, Pro- and Arg-tRNA; (5) it contains the most frequent triplets, but not the most rare ones, appearing in the genome of numerous species; (6) it exhibits a coherent internal structure with respect to the molecular weight of its triplets. This structure, also found in the loop part of mitochondrial tRNA's, contains an excess of AU bases with respect to GC bases. This fact has no explanation in the classical probabilistic model of the tRNA's. Therefore, we propose the cyclic AB code as a primitive genetic structure with the essential coding properties of the present genetic code.