Adenosine-mediated inhibition of casein production by mouse mammary glands in culture

The present study was carried out to examine whether activation of adenosine receptors by adenosine analogues will affect casein production by mouse mammary epithelial cells. The morphogenesis and functions of epithelial tissue in the mammary gland are influenced by their surrounding adipocytes. Adipocytes are known to release adenosine into the extracellular fluid which can modulate cyclic-AMP levels in surrounding cells through binding to their adenosine receptors. To examine a possible paracrine effect of adenosine, the modulation of casein production in mammary explant culture and mammary epithelial cell (MEC) culture by adenosine receptor agonists has been investigated. We have observed that activation of the A₁-adenosine receptor subtype in mammary tissue by an adenosine analogue (—)N⁶-(R-phenyl-isopropyl)-adenosine (PIA) raised cAMP levels. PIA and another adenosine receptor agonist, isobutylmethylxanthine (IBMX), inhibited casein accumulation both in explants and in MEC cultures in the presence of lactogenic hormones, which suggests that PIA or adenosine can act directly on the epithelial cells. This inhibition does not appear to be caused by elevation of cAMP levels or phosphodiesterase activity. The inhibition of intracellular casein accumulation by PIA and IBMX in explant cultures can be reversed via treatment of pertussis toxin which is known to ADP-ribosylate GTP-binding Gαi-proteins, indicating that a Gi-protein-dependent pathway may be involved in this inhibition. The results also suggest that local accumulation of adenosine in the extracellular fluids of mammary glands is likely to inhibit the lactogenic response of mammary epithelial cells. © 1996 Wiley-Liss, Inc.     

Activation of PKC modulates blood-brain barrier endothelial cell permeability changes induced by hypoxia and posthypoxic reoxygenation

The blood-brain barrier (BBB) is a metabolic and physiological barrier important for maintaining brain homeostasis. The aim of this study was to determine the role of PKC activation in BBB paracellular permeability changes induced by hypoxia and posthypoxic reoxygenation using in vitro and in vivo BBB models. In rat brain microvessel endothelial cells (RMECs) exposed to hypoxia (1% O2-99% N2; 24 h), a significant increase in total PKC activity was observed, and this was reduced by posthypoxic reoxygenation (95% room air-5% CO2) for 2 h. The expression of PKC-betaII, PKC-gamma, PKC-eta, PKC-mu, and PKC-lambda also increased following hypoxia (1% O2-99% N2; 24 h), and these protein levels remained elevated following posthypoxic reoxygenation (95% room air-5% CO2; 2 h). Increases in the expression of PKC-epsilon and PKC-zeta were also observed following posthypoxic reoxygenation (95% room air-5% CO2; 2 h). Moreover, inhibition of PKC with chelerythrine chloride (10 microM) attenuated the hypoxia-induced increases in [14C]sucrose permeability. Similar to what was observed in RMECs, total PKC activity was also stimulated in cerebral microvessels isolated from rats exposed to hypoxia (6% O2-94% N2; 1 h) and posthypoxic reoxygenation (room air; 10 min). In contrast, hypoxia (6% O2-94% N2; 1 h) and posthypoxic reoxygenation (room air; 10 min) significantly increased the expression levels of only PKC-gamma and PKC-theta in the in vivo hypoxia model. These data demonstrate that hypoxia-induced BBB paracellular permeability changes occur via a PKC-dependent mechanism, possibly by differentially regulating the protein expression of the 11 PKC isozymes.     

Exposure of juvenile chinook and chum salmon to chemical contaminants in the Hylebos Waterway of Commencement Bay, Tacoma, Washington

The Hylebos Waterway is an industrialized waterway ofCommencement Bay, Tacoma, Washington, that is severelycontaminated with aromatic and chlorinatedhydrocarbons in the sediment. Juvenile chinook (Oncorhynchus keta) and chum salmon (O.tshawytscha) inhabit this waterway for a few days orweeks during their outmigration from freshwaterstreams to saltwater. The purpose of thisinvestigation was to determine to what degree juvenilechum and chinook salmon captured from the HylebosWaterway might bioaccumulate organic contaminants. These levels of exposure will be compared to previousstudies where such exposures have been linked tobiological dysfunction in juvenile salmon. Theresults showed that juvenile chum and chinook salmonfrom the Hylebos Waterway take up a wide range ofchemical contaminants, compared to fish fromhatcheries or reference estuaries. These contaminantsinclude high and low molecular weight polycyclicaromatic hydrocarbons (PAHs), polychlorinatedbiphenyls (PCBs, including the toxic congeners 105 and118), hexachlorobutadiene (HCBD), hexachlorobenzene(HCB), DDTs, heptachlor, and several pesticides. Immunohistochemical examination of the gill and gut injuvenile chum salmon from the Hylebos Waterway showedthe induction of the P450 metabolizing enzyme. Moreover, concentrations of contaminants in juvenilechinook and chum salmon from the Hylebos Waterway arecomparable to levels previously shown to be associatedwith biological injury in juvenile chinook salmon,such as impaired growth, suppression of immunefunction as demonstrated by reduced B cell function,and increased mortality following pathogen exposure.     

Diversity and ecology of algae from the Nahal Qishon river,northern Israel

In 59 samples of periphyton and phytoplankton collected in 2002 - 2003 from the Nahal Qishon (Qishon River), northern Israel, we found 178 species from seven divisions of algae and cyanoprocaryotes. Diatoms, clorophytes, and cyanoprocaryotes prevail. Nitzschia and Navicula (Bacillariophyta) are the most abundant. Most of the species are cosmopolitan or widespread, except Lagynion janei (Chrysophyta), which is endemic for the Mediterranean Realm. About 17% of species (26) are new for Israel and five of them represent the first recorded genera: Crinalium endophyticum Crow, Actinocyclus normanii (Gregory) Hustedt, Rhizoclonium hieroglyphicum (Agardh) Kütz (Chlorophyta), Lagynion janei Bourelly, and Stylococcus aureus Chodat. Most of them come from a rare riverine assemblage with red alga Audouinella pygmea, as well as from the estuarine assemblage. Alkaliphiles predominate among the indicators of acidity, with few acidophiles confined to the communities under the impact of industrial wastes. Among the indicators of salinity, most numerous are the oligohalobien-indifferents and species adapted to a moderate salinity level. The relative species richness of ecological groups and the indices of saprobity are correlated with changes in conductivity, pH, and N-nitrate concentration. Indicators of organic pollution fall in the range of betameso- to alfamesosaprobic self-purification grades. Our studies show ecological significance of the Nahal Qishon as a model for a strongly disturbed aquatic ecosystem in the coastal zone of eastern Mediterranean.     

Comparsion of staining characteristics of Toto bodies

Toto bodies are eosinophilic structures that resemble the cells of the superficial cell layer of the oral epithelium. Toto bodies commonly are associated with inflammatory gingival and other mucosal lesions including pyogenic granuloma, irritational fibroma, epulis fissuratum, peripheral giant cell granuloma and inflammatory hyperplastic gingivitis. We evaluated staining characteristics of Toto bodies to establish their origin and to identify their significance in lesions. We investigated pyogenic granuloma, fibroma and leukoplakia with epithelium that exhibited Toto bodies after hematoxylin and eosin (staining. Sections were stained with Alcian blue, periodic acid-Schiff and Ayoub-Shklar stains to evaluate staining intensity and distribution. More Toto bodies were found in pyogenic granuloma than in fibroma and leukoplakia. PAS and Alcian blue staining exhibited mild intensity and did not establish the origin of Toto bodies. High staining intensity and diffuse distribution of stain was observed using Ayoub-Shklar staining, which indicated that Toto bodies originate from keratin.     

Design and synthesis of highly selective,orally active Polo-like kinase-2 (Plk-2) inhibitors

Polo-like kinase-2 (Plk-2) is a potential therapeutic target for Parkinson’s disease and this Letter describes the SAR of a series of dihydropteridinone based Plk-2 inhibitors. By optimizing both the N-8 substituent and the biaryl region of the inhibitors we obtained single digit nanomolar compounds such as 37 with excellent selectivity for Plk-2 over Plk-1. When dosed orally in rats, compound 37 demonstrated a 41–45% reduction of pS129-α-synuclein levels in the cerebral cortex.     

Design and synthesis of thiophene dihydroisoquinolines as novel BACE1 inhibitors

Utilizing a structure based design approach, combined with extensive medicinal chemistry execution, highly selective, potent and novel BACE1 inhibitor 8 (BACE1 Alpha assay IC₅₀ = 8 nM) was made from a weak μM potency hit in an extremely efficient way. The detailed SAR and general design approaches will be discussed.     

Full-sequence computational design and solution structure of a thermostable protein variant

Computational protein design procedures were applied to the redesign of the entire sequence of a 51 amino acid residue protein, Drosophila melanogaster engrailed homeodomain. Various sequence optimization algorithms were compared and two resulting designed sequences were experimentally evaluated. The two sequences differ by 11 mutations and share 22% and 24% sequence identity with the wild-type protein. Both computationally designed proteins were considerably more stable than the naturally occurring protein, with midpoints of thermal denaturation greater than 99 degrees C. The solution structure was determined for one of the two sequences using multidimensional heteronuclear NMR spectroscopy, and the structure was found to closely match the original design template scaffold.