Characterization of Enzymatic Antioxidants in the Lichen Ramalina lacera and Their Response to Rehydration

Lichens are slow-growing associations of fungi and green algae or cyanobacteria. This symbiotic association forms a common thallus that does not possess roots or a waxy cuticle and depends mainly on atmospheric input of mineral nutrients. The lifestyle of most lichens is composed of alternating periods of desiccation with low metabolic activity and hydration that induces increase in their metabolism. We have previously shown that rehydration of the naturally desiccated lichen Ramalina lacera resulted in a rapid increase in photosynthesis and was accompanied by a burst of intracellular production of reactive oxygen species and nitric oxide, as well as a transient decrease in water-soluble antioxidant capacity. We report here on enzymatic antioxidants of R. lacera and their response to rehydration. Native gel electrophoresis of crude extracts of R. lacera stained for superoxide dismutase (SOD) activity revealed four Fe-SOD and four Mn-SOD electromorphs that are synthesized by the alga, a Cu/Zn-SOD and a Mn-SOD that are the product of the fungus, and two catalases synthesized one by the fungus and the other by the algae. In addition, we detected glutathione reductase and glucose-6-phosphate dehydrogenase activities in crude extracts of R. lacera. Rehydration of the thalli resulted in a decrease in SOD activity of all forms, and a transient decrease in total catalase activity, as well as a decrease in the antioxidant auxiliary enzymes glutathione reductase and glucose-6-phosphate dehydrogenase.     

<i>In vitro</i> evaluation of antifungal activity of Agave (<i>Agave scabra</i>, Salm Dyck) extracts against post-harvest mushrooms

The agricultural sector, and particularly the horticultural production, has a singular importance in agriculture, considering that it ranks second on agricultural products, nationally and worldwide. Fungal diseases are one of the major causes of vegetable loss during storage, reducing their nutritional value, quality and sale price. Vegetables are usually exposed to diverse treatments with chemical products before storage; as a result, fungal populations develop an increased resistance over time becoming more difficult to control. Because of this, research efforts toward finding more suitable chemicals to control fungal diseases are needed. Natural extracts may be an alternative solve this problem. In the present investigation the fungicidal activity of aqueous and ethanol extracts of Agave scabra was evaluated on the growth of Botrytis cinerea, Mucor sp., Aspergillus niger, Fusarium sp. and Penicillium sp., whose strains were isolated from potato and tomato. To assess their effects, the agar-dilution and agar-well techniques were performed. The ethanol extract was more effective against Botrytis cinerea and Mucor sp. when the agar-well method was used. However, when using the agar-dilution method the ethanol extract of Agave scabra inhibited the growth of Botrytis cinerea, Mucor sp. and Penicillium sp.     

Endogenous ligands for sigma opioid receptors in the brain ("sigmaphin"): evidence from binding assays

Two endogenous ligands which interact preferentially with the sigma opioid receptors were identified from the guinea-pig brain extract in a Sephadex G-50 fractionation. These two ligands inhibited more potently the binding of [3H]SKF-10047 to sigma opioid receptors than [3H]naloxone to mu opioid receptors, [3H]ethylketocyclazocine to kappa opioid receptors and [3H]DADLE to delta opioid receptors. In the phencyclidine receptor assay, these two ligands were almost inactive. Incubation of these ligands with trypsin destroyed at least 50% of the activities in the sigma opioid receptor assay. Both ligands inhibited the sigma binding in a dose-dependent manner. The inhibition could be eliminated when the two ligands were removed from incubation media by extensive washings. It is therefore concluded that sigma opioid receptors are not phencyclidine receptors and that endogenous ligands for sigma opioid receptors may exist in the brain.     

Clonal analysis of mouse development reveals a polyclonal origin for yolk sac blood islands

Direct clonal analysis of tissue and organ maturation in vivo is a critical step in the interpretation of in vitro cell precursor-progeny relationships. We have developed a method to analyze clonal progenitor contributions in vivo using ES cells stably expressing separate fluorescent proteins and placed into normal blastocysts to form tetrachimeras. Here we applied this method to the analysis of embryonic yolk sac blood islands. In most vertebrates, yolk sac blood islands are the initial sites of appearance of hematopoietic and endothelial cells. It has been proposed that these lineages arise from a common clonal progenitor, the hemangioblast, but this hypothesis has not been tested directly in physiological development in vivo. Our analysis shows that each island has contributions from multiple progenitors. Moreover, contribution by individual hemangioblast progenitors to both endothelial and hematopoietic lineages within an island, if it happens at all, is an infrequent event.     

HIV Prevalence and Risks Associated with HIV Infection among Transgender Individuals in Cambodia

IntroductionRecognizing transgender individuals have a high risk of HIV acquisition, and to inform policies and programming, we conducted an HIV prevalence and risk behaviors survey among transgender individuals in Cambodia.MethodsCross-sectional survey using a respondent driven sampling method with self-administered audio-computer assisted interviews. HIV testing was performed prior to the questionnaire with results available immediately after. Eligible participants were ≥18 years, identified as male at birth and self-identified/expressed as a different gender, and reported having sex with at least one male partner in past year. From six major urban centers of Cambodia, 891 transgender individuals were recruited.ResultsThe majority of the 891 participants self-identified as third gender or female (94.5%), were young (median age 23, IQR [20–27]), had secondary education or higher (80.5%), not married (89.7%), and employed (90.2%). The majority had first sex before 18 years (66.8%), with a male (79.9%), 37.9% having been paid or paying for this first sex. The rate of HIV positivity among participants was found to be 4.15%. Consistent condom use with male and female partners was low with all partner types, but particularly low with male partners when paying for sex (20.3%). The majority of participants reported having experienced discrimination in their lifetime (54.8%) and 30.3% had been assaulted. Multivariate analysis revealed that older age (adjusted OR = 14.73 [4.20, 51.67] for age 35–44 and adjusted OR = 7.63 [2.55, 22.81] for age 30–34), only having a primary school education or no schooling at all (adjusted OR = 2.62 [1.18, 5.80], being a resident of Siem Reap (adjusted OR = 7.44 [2.37,23.29], receiving payment at first sex (adjusted OR = 2.26 [1.00, 5.11], having sex during/after using drugs (adjusted OR = 2.90 [1.09,7.73]), inconsistent condom use during last anal sex (adjusted OR = 3.84 [1.58, 9.33]), and reporting low self-esteem (adjusted OR = 3.25 [1.35,7.85]) were independently associated with HIV infection.ConclusionsThis study confirms transgender individuals as one of the highest-risk groups for HIV infection in Cambodia. It suggests the need for programmatic strategies that mitigate identified associated risks and facilitate access to HIV care for this population.     

Formation of leukotrienes and hydroxy acids by human neutrophils and platelets exposed to monosodium urate

Monosodium urate (MSU) crystals stimulate the production of arachidonic acid metabolites by human neutrophils and platelets. Neutrophils exposed to MSU generated leukotriene B₄(LTB₄). 6- -LTB₄, 12- -6- -LTB₄, and 5S, 12S DHETE from endogenous sources of arachidonate. In addition to these metabolites both monohyroxyeicosatetraenoic acids (i.e., 5-HETE) and w-oxidation products (i.e., 20-COOH LTB₄) were formed by neutrophils exposed to MSU. Addition of exogenous arachidonic acid led to increased formation of each of these metabolites. When neutrophils were treated with colchicine (10 uM), LTB₄ but 5-HETE formation was impaired. (1-¹⁴C) Arachidonate-labeled platelets exposed to MSU released (1-¹⁴C)-arachidonate. (¹⁴C)-12 HETE, (¹⁴C)-HHT and (¹⁴C)-thromboxane B₂. Results indicate that MSU stimulates arachidonic acid metabolism in both human neutrophils and platelets. Moreover, they suggest not only that metabolites of arachidonate may be considered as possible candidates for mediators of inflammation in crystal-associated diseases, but that colchicine blocks the formation of LTB₄.     

Identification of cis sequences controlling efficient position-independent tissue-specific expression of human major histocompatibility complex class I genes in transgenic mice.

We previously reported that genomic major histocompatibility complex class I human leukocyte antigen (HLA)-B7 gene constructs with as little as 0.66 kb of 5'- and 2.0 kb of 3'-flanking DNA were expressed efficiently and appropriately in transgenic mice. To identify and characterize the relevant cis-acting regulatory elements in more detail, we have generated and analyzed a series of transgenic mice carrying native HLA-B7 genes with further 5' truncations or intronic deletions and hybrid constructs linking the 5'-flanking region of B7 to a reporter gene. We were unable to detect a specific requirement for sequence information within introns 2 to 7 for either appropriate constitutive or inducible class I expression in adult animals. The results revealed the presence of cis-acting regulatory sequences between -0.075 kb and -0.66 kb involved in driving efficient copy number-dependent constitutive and gamma interferon-enhanced tissue-specific expression. The region from -0.11 to -0.66 kb is also sufficient to prevent integration site-specific "position effects," because in its absence HLA-B7 expression is frequently detected at significant levels at inappropriate sites. Conserved sequence elements homologous to the H-2 class I regulatory element, or enhancer A, and the interferon response sequence are located between about -151 and -228 bp of the B7 gene. Our results also indicate the existence of sequences downstream of -0.11 kb which can influence the pattern of tissue-specific expression of the HLA-B7 gene and the ability of this gene to respond to gamma interferon.     

Intensive nitrogen loss over the Omani Shelf due to anammox coupled with dissimilatory nitrite reduction to ammonium

A combination of stable isotopes (¹⁵N) and molecular ecological approaches was used to investigate the vertical distribution and mechanisms of biological N₂ production along a transect from the Omani coast to the central–northeastern (NE) Arabian Sea. The Arabian Sea harbors the thickest oxygen minimum zone (OMZ) in the world''s oceans, and is considered to be a major site of oceanic nitrogen (N) loss. Short (<48 h) anoxic incubations with ¹⁵N-labeled substrates and functional gene expression analyses showed that the anammox process was highly active, whereas denitrification was hardly detectable in the OMZ over the Omani shelf at least at the time of our sampling. Anammox was coupled with dissimilatory nitrite reduction to ammonium (DNRA), resulting in the production of double-¹⁵N-labeled N₂ from ¹⁵NO₂⁻, a signal often taken as the lone evidence for denitrification in the past. Although the central–NE Arabian Sea has conventionally been regarded as the primary N-loss region, low potential N-loss rates at sporadic depths were detected at best. N-loss activities in this region likely experience high spatiotemporal variabilities as linked to the availability of organic matter. Our finding of greater N-loss associated with the more productive Omani upwelling region is consistent with results from other major OMZs. The close reliance of anammox on DNRA also highlights the need to take into account the effects of coupling N-transformations on oceanic N-loss and subsequent N-balance estimates.