The nature of inhibition of 3-hydroxy-3-methylglutaryl CoA reductase by garlic-derived diallyl disulfide

A concentration dependent inhibition of 3-hydroxy-3-methylglutaryl CoA (HMG CoA) reductase was found on preincubation of microsomal preparations with diallyl disulfide, a component of garlic oil. This inhibited state was only partially reversed even with high concentrations of DTT. Glutathione, a naturally occurring reducing thiol agent, was ineffective. The substrate, HMG CoA, but not NADPH, was able to give partial protection for the DTT-dependent, but not glutathione-dependent activity. The garlic-derived diallyl disulfide is the most effective among the sulfides tested for inhibition of HMG CoA reductase. Formation of protein internal disulfides, inaccessible for reduction by thiol agents, but not of protein dimer, is likely to be the cause of this inactivation.

Influence of Vitamin E on the Utilization of Carotene from Oils

IN a previous communication1, it was indicated that the variations in growth response of vitamin A-deficient rats to carotene dissolved in different oils may be due to the difference in vitamin E contents of the oils. The effect of equalizing the level of tocopherol in the supplements has since been studied and the results found to support the explanation.

In vitro protection of umbilical cord blood–derived primitive hematopoietic stem progenitor cell pool by mannose-specific lectins via antioxidant mechanisms

BACKGROUND: Earlier we reported that an oral administration of two mannose-specific dietary lectins, banana lectin (BL) and garlic lectin (GL), led to an enhancement of hematopoietic stem and progenitor cell (HSPC) pool in mice. STUDY DESIGN AND METHODS: Cord blood–derived CD34+ HSPCs were incubated with BL, GL, Dolichos lectin (DL), or artocarpin lectin (AL) for various time periods in a serum- and growth factor–free medium and were subjected to various functional assays. Reactive oxygen species (ROS) levels were detected by using DCHFDA method. Cell fractionation was carried out using lectin-coupled paramagnetic beads. RESULTS: CD34+ cells incubated with the lectins for 10 days gave rise to a significantly higher number of colonies compared to the controls, indicating that all four lectins possessed the capacity to protect HSPCs in vitro. Comparative analyses showed that the protective ability of BL and GL was better than AL and DL and, therefore, further experiments were carried out with them. The output of long-term culture-initiating cell (LTC-IC) and extended LTC-IC assays indicated that both BL and GL protected primitive stem cells up to 30 days. The cells incubated with BL or GL showed a substantial reduction in the ROS levels, indicating that these lectins protect the HSPCs via antioxidant mechanisms. The mononuclear cell fraction isolated by lectin-coupled beads got enriched for primitive HSPCs, as reflected in the output of phenotypic and functional assays. CONCLUSION: The data show that both BL and GL protect the primitive HSPCs in vitro and may also serve as cost-effective HSPC enrichment tools.

In vitro protection of umbilical cord blood-derived primitive hematopoietic stem progenitor cell pool by mannose-specific lectins via antioxidant mechanisms.

BACKGROUND: Earlier we reported that an oral administration of two mannose-specific dietary lectins, banana lectin (BL) and garlic lectin (GL), led to an enhancement of hematopoietic stem and progenitor cell (HSPC) pool in mice. STUDY DESIGN AND METHODS: Cord blood derived CD34+ HSPCs were incubated with BL, GL, Dolichos lectin (DL), or artocarpin lectin (AL) for various time periods in a serum- and growth factor free medium and were subjected to various functional assays. Reactive oxygen species (ROS) levels were detected by using DCHFDA method. Cell fractionation was carried out using lectin-coupled paramagnetic beads. RESULTS: CD34+ cells incubated with the lectins for 10 days gave rise to a significantly higher number of colonies compared to the controls, indicating that all four lectins possessed the capacity to protect HSPCs in vitro. Comparative analyses showed that the protective ability of BL and GL was better than AL and DL and, therefore, further experiments were carried out with them. The output of long-term culture-initiating cell (LTC-IC) and extended LTC-IC assays indicated that both BL and GL protected primitive stem cells up to 30 days. The cells incubated with BL or GL showed a substantial reduction in the ROS levels, indicating that these lectins protect the HSPCs via antioxidant mechanisms. The mononuclear cell fraction isolated by lectin-coupled beads got enriched for primitive HSPCs, as reflected in the output of phenotypic and functional assays.CONCLUSION: The data show that both BL and GL protect the primitive HSPCs in vitro and may also serve as cost-effective HSPC enrichment tools.

On the involvement of intramolecular protein disulfide in the irreversible inactivation of 3-hydroxy-3-methylglutaryl-CoA reductase by diallyl disulfide

Treatment with diallyl disulfide, a constituent of garlic oil, irreversibly inactivated microsomal and a soluble 50 kDa form of HMG-CoA reductase. No radioactivity was found to be protein-bound on treating the soluble enzyme with [35S]diallyl disulfide, indicating the absence of the mixed disulfide of the type allyl-S-S-protein. SDS-PAGE and Western blot analyses of the diallyl-disulfide-treated protein showed no traces of the dimer of the type protein-S-S-protein, but clearly indicated BME-reversible increased mobility, as expected of an intramolecular protein disulfide. The sulfhydryl groups, as measured by alkylation with iodo[2-14C]acetic acid, were found to decrease in the diallyl-disulfide-treated enzyme protein. Tryptic peptide analysis also gave support for the possible presence of disulfide-containing peptides in such a protein. It appears that diallyl disulfide inactivated HMG-CoA reductase by forming an internal protein disulfide that became inaccessible for reduction by DTT, and thereby retaining the inactive state of the enzyme.

Photon-photon total cross-section

We study the total inelastic gamma gamma cross-section and discuss predictions from different models, with a special attention to their dependence on the input parameters. In particular we examine the results from a simple extension of the Regge Pomeron exchange model and those from the eikonalized mini-jet model. We then compare both of them with recent LEP data.

Ethnobotany of the Monpa ethnic group at Arunachal Pradesh, India

Background: The present paper documents the uses of plants in traditional herbal medicine for human and veterinary ailments, and those used for dietary supplements, religious purpose, local beverage, and plants used to poison fish and wild animals. Traditional botanical medicine is the primary mode of healthcare for most of the rural population in Arunachal Pradesh. Materials and methods: Field research was conducted between April 2006 and March 2009 with randomly selected 124 key informants using semi-structured questionnaire. The data obtained was analyzed through informant consensus factor (F(IC)) to determine the homogeneity of informant's knowledge on medicinal plants. Results: We documented 50 plants species belonging to 29 families used for treating 22 human and 4 veterinary ailments. Of the medicinal plants reported, the most common growth form was herbs (40%) followed by shrubs, trees, and climbers. Leaves were most frequently used plant parts. The consensus analysis revealed that the dermatological ailments have the highest F(IC) (0.56) and the gastro-intestinal diseases have F(IC) (0.43). F(IC) values indicated that there was high agreement in the use of plants in dermatological and gastro-intestinal ailments category among the users. Gymnocladus assamicus is a critically rare and endangered species used as disinfectant for cleaning wounds and parasites like leeches and lice on livestocks. Two plant species (Illicium griffithii and Rubia cordifolia) are commonly used for traditional dyeing of clothes and food items. Some of the edible plants recorded in this study were known for their treatment against high blood pressure (Clerodendron colebrookianum), diabetes mellitus (Momordica charantia), and intestinal parasitic worms like round and tape worms (Lindera neesiana, Solanum etiopicum, and Solanum indicum). The Monpas of Arunachal Pradesh have traditionally been using Daphne papyracea for preparing hand-made paper for painting and writing religious scripts in Buddhist monasteries. Three plant species (Derris scandens, Aesculus assamica, and Polygonum hydropiper) were frequently used to poison fish during the month of June-July every year and the underground tuber of Aconitum ferrox is widely used in arrow poisoning to kill ferocious animals like bear, wild pigs, gaur and deer. The most frequently cited plant species; Buddleja asiatica and Hedyotis scandens were used as common growth supplements during the preparation of fermentation starter cultures. Conclusion: The traditional pharmacopoeia of the Monpa ethnic group incorporates a myriad of diverse botanical flora. Traditional knowledge of the remedies is passed down through oral traditions without any written document. This traditional knowledge is however, currently threatened mainly due to acculturation and deforestation due to continuing traditional shifting cultivation. This study reveals that the rural populations in Arunachal Pradesh have a rich knowledge of forest-based natural resources and consumption of wild edible plants is still an integral part of their socio-cultural life. Findings of this documentation study can be used as an ethnopharmacological basis for selecting plants for future phytochemical and pharmaceutical studies.

The determination of stem cell fate by 3D scaffold structures through the control of cell shape

Stem cell response to a library of scaffolds with varied 3D structures was investigated. Microarray screening revealed that each type of scaffold structure induced a unique gene expression signature in primary human bone marrow stromal cells (hBMSCs). Hierarchical cluster analysis showed that treatments sorted by scaffold structure and not by polymer chemistry suggesting that scaffold structure was more influential than scaffold composition. Further, the effects of scaffold structure on hBMSC function were mediated by cell shape. Of all the scaffolds tested, only scaffolds with a nanofibrous morphology were able to drive the hBMSCs down an osteogenic lineage in the absence of osteogenic supplements. Nanofiber scaffolds forced the hBMSCs to assume an elongated, highly branched morphology. This same morphology was seen in osteogenic controls where hBMSCs were cultured on flat polymer films in the presence of osteogenic supplements (OS). In contrast, hBMSCs cultured on flat polymer films in the absence of OS assumed a more rounded and less-branched morphology. These results indicate that cells are more sensitive to scaffold structure than previously appreciated and suggest that scaffold efficacy can be optimized by tailoring the scaffold structure to force cells into morphologies that direct them to differentiate down the desired lineage. Published by Elsevier Ltd.

Crystal structure of a beta-prism II lectin from Remusatia vivipara

The crystal structure of a beta-prism II (BP2) fold lectin from Remusatia vivipara, a plant of traditional medicinal value, has been determined at a resolution of 2.4 A. This lectin (RVL, Remusatia vivipara lectin) is a dimer with each protomer having two distinct BP2 domains without a linker between them. It belongs to the ``monocot mannose-binding'' lectin family, which consists of proteins of high sequence and structural similarity. Though the overall tertiary structure is similar to that of lectins from snowdrop bulbs and garlic, crucial differences in the mannose-binding regions and oligomerization were observed. Unlike most of the other structurally known proteins in this family, only one of the three carbohydrate recognition sites (CRSs) per BP2 domain is found to be conserved. RVL does not recognize simple mannose moieties. RVL binds to only N-linked complex glycans like those present on the gp120 envelope glycoprotein of HIV and mannosylated blood proteins like fetuin, but not to simple mannose moieties. The molecular basis for these features and their possible functional implications to understand the different levels of carbohydrate affinities in this structural family have been investigated through structure analysis, modeling and binding studies. Apart from being the first structure of a lectin to be reported from the Araceae/Arum family, this protein also displays a novel mode of oligomerization among BP2 lectins.

A touch of history and a peep into the future of the lipid-quinone known as coenzyme Q and ubiquinone

Coenzyme Q (ubiquinone), a fully substituted benzoquinone with polyprenyl side chain, participates in many cellular redox activities. Paradoxically it was discovered only in 1957, albeit being ubiquitous. It required a person, F. L. Crane, a place, Enzyme Institute, Madison, USA, and a time when D. E. Green was directing vigorous research on mitochondria. Located at the transition of 2-electron flavoproteins and 1-electron cytochrome carriers, it facilitates electron transfer through the elegant Q-cycle in mitochondria to reduce O-2 to H2O, and to H2O2, now a significant signal-transducing agent, as a minor activity in shunt pathway (animals) and alternative oxidase (plants). The ability to form Q-radical by losing an electron and a proton was ingeniously used by Mitchell to explain the formation of the proton gradient, considered the core of energy transduction, and also in acidification in vacuoles. Known to be a mobile membrane constituent (microsomes, plasma membrane and Golgi apparatus), allowing it to reach multiple sites, coenzyme Q is expected to have other activities. Coenzyme Q protects circulating lipoproteins being a better lipid antioxidant than even vitamin E. Binding to proteins such as QPS, QPN, QPC and uncoupling protein in mitochondria, QA and QB in the reaction centre in R. sphaeroides, and disulfide bond-forming protein in E. coli (possibly also in Golgi), coenzyme Q acquires selective functions. A characteristic of orally dosed coenzyme Q is its exclusive absorption into the liver, but not the other tissues. This enrichment of Q is accompanied by significant decrease of blood pressure and of serum cholesterol. Inhibition of formation of mevalonate, the common precursor in the branched isoprene pathway, by the minor product, coenzyme Q, decreases the major product, cholesterol. Relaxation of contracted arterial smooth muscle by a side-chain truncated product of coenzyme Q explains its effect of decreasing blood pressure. Extensive clinical studies carried out on oral supplements of coenzyine Q, initially by K. Folkers and Y. Yamamura and followed many others, revealed a large number of beneficial effects, significantly in cardiovascular diseases. Such a variety of effects by this lipid quinone cannot depend on redox activity alone. The fat-soluble vitamins (A, D, E and K) that bear structural relationship with coenzyme Q are known to be active in their polar forms. A vignette of modified forms of coenzyme Q taking active role in its multiple effects is emerging.

Functional Analysis of the Genes Encoding Diaminopropionate Ammonia Lyase in Escherichia coli and Salmonella enterica Serovar Typhimurium

Diaminopropionate ammonia lyase (DAPAL) is a pyridoxal-5'phosphate (PLP)-dependent enzyme that catalyzes the conversion of diaminopropionate (DAP) to pyruvate and ammonia and plays an important role in cell metabolism. We have investigated the role of the ygeX gene of Escherichia coli K-12 and its ortholog, STM1002, in Salmonella enterica serovar Typhimurium LT2, presumed to encode DAPAL, in the growth kinetics of the bacteria. While Salmonella Typhimurium LT2 could grow on DL-DAP as a sole carbon source, the wild-type E. coli K-12 strain exhibited only marginal growth on DL-DAP, suggesting that DAPAL is functional in S. Typhimurium. The expression of ygeX in E. coli was low as detected by reverse transcriptase PCR (RT-PCR), consistent with the poor growth of E. coli on DL-DAP. Strains of S. Typhimurium and E. coli with STM1002 and ygeX, respectively, deleted showed loss of growth on DL-DAP, confirming that STM1002 (ygeX) is the locus encoding DAPAL. Interestingly, the presence of DL-DAP caused a growth inhibition of the wild-type E. coli strain as well as the knockout strains of S. Typhimurium and E. coli in minimal glucose/glycerol medium. Inhibition by DL-DAP was rescued by transforming the strains with plasmids containing the STM1002 (ygeX) gene encoding DAPAL or supplementing the medium with Casamino Acids. Growth restoration studies using media lacking specific amino acid supplements suggested that growth inhibition by DL-DAP in the absence of DAPAL is associated with auxotrophy related to the inhibition of the enzymes involved in the biosynthetic pathways of pyruvate and aspartate and the amino acids derived from them.

Ontology analysis of global gene expression differences of human bone marrow stromal cells cultured on 3D scaffolds or 2D films

Differences in gene expression of human bone marrow stromal cells (hBMSCs) during culture in three-dimensional (3D) nanofiber scaffolds or on two-dimensional (2D) films were investigated via pathway analysis of microarray mRNA expression profiles. Previous work has shown that hBMSC culture in nanofiber scaffolds can induce osteogenic differentiation in the absence of osteogenic supplements (OS). Analysis using ontology databases revealed that nanofibers and OS regulated similar pathways and that both were enriched for TGF-beta and cell-adhesion/ECM-receptor pathways. The most notable difference between the two was that nanofibers had stronger enrichment for cell-adhesion/ECM-receptor pathways. Comparison of nanofibers scaffolds with flat films yielded stronger differences in gene expression than comparison of nanofibers made from different polymers, suggesting that substrate structure had stronger effects on cell function than substrate polymer composition. These results demonstrate that physical (nanofibers) and biochemical (OS) signals regulate similar ontological pathways, suggesting that these cues use similar molecular mechanisms to control hBMSC differentiation. Published by Elsevier Ltd.

One-Carbon Metabolic Pathway Rewiring in Escherichia coli Reveals an Evolutionary Advantage of 10-Formyltetrahydrofolate Synthetase (Fhs) in Survival under Hypoxia

In cells, N-10-formyltetrahydrofolate (N-10-fTHF) is required for formylation of eubacterial/organellar initiator tRNA and purine nucleotide biosynthesis. Biosynthesis of N-10-fTHF is catalyzed by 5,10-methylene-tetrahydrofolate dehydrogenase/cyclohydrolase (FolD) and/or 10-formyltetrahydrofolate synthetase (Fhs). All eubacteria possess FolD, but some possess both FolD and Fhs. However, the reasons for possessing Fhs in addition to FolD have remained unclear. We used Escherichia coli, which naturally lacks fhs, as our model. We show that in E. coli, the essential function of folD could be replaced by Clostridium perfringens fhs when it was provided on a medium-copy-number plasmid or integrated as a single-copy gene in the chromosome. The fhs-supported folD deletion (Delta folD) strains grow well in a complex medium. However, these strains require purines and glycine as supplements for growth in M9 minimal medium. The in vivo levels of N-10-fTHF in the Delta folD strain (supported by plasmid-borne fhs) were limiting despite the high capacity of the available Fhs to synthesize N-10-fTHF in vitro. Auxotrophy for purines could be alleviated by supplementing formate to the medium, and that for glycine was alleviated by engineering THF import into the cells. The Delta folD strain (harboring fhs on the chromosome) showed a high NADP(+)-to-NADPH ratio and hypersensitivity to trimethoprim. The presence of fhs in E. coli was disadvantageous for its aerobic growth. However, under hypoxia, E. coli strains harboring fhs outcompeted those lacking it. The computational analysis revealed a predominant natural occurrence of fhs in anaerobic and facultative anaerobic bacteria.