A new naturally acetylated triterpene saponin from Nigella sativa

A new glycosylated triterpene 1 was identified as 3-O-[β-d-xylopyranosyl-(1→3)-α-l-rhamnopyranosyl-(1→4)-β-d-glucopyranosyl]-11-methoxy-16-hydroxy-17-acetoxy hederagenin from an ethanolic extract of seeds of Nigella sativa Linn. Identification of the naturally acetylated saponin was based on chemical and spectroscopic analyses including FABMS, ¹H, ¹³C, and 2D NMR and DEPT. The saponin was a penta hydroxy pentacyclic triterpene, in which one hydroxyl group was acetylated and other one was methylated naturally.     

The evolutionary origin of a second radiation of annual Castilleja (Orobanchaceae) species in South America: The role of long distance dispersal and allopolyploidy

Considerable attention has been directed toward understanding the wide gaps in range that are common among many groups of closely related organisms. By placing their biology and geography in a phylogenetic context, we may gain a broader knowledge of the series of historical events that have led to present species distributions. In addition to the North American annuals, a second radiation of annual Castilleja species is in Andean Peru and central Chile. Phylogenetic analyses of chloroplast and nuclear DNA regions revealed a complex history for the origin and diversification of annual Castilleja species in South America. In addition to at least three independent long-distance dispersal events from North America, allopolyploidy has played a significant role in this disjunct radiation. Only C. attenuata occurs in both California and South America, and these results support its recent arrival to central Chile. Two Peruvian species are inferred to be allopolyploids; hybridization between annual lineages derived from independent long-distance dispersal events from North America gave rise to C. profunda, and hybridization between South American annual and perennial species gave rise to C. cerroana. The relative importance these events are discussed with reference to the observed morphological, ecological, and distributional patterns.     

Associations between aortic calcification and components of body composition in elderly men

Objective: To investigate associations among body composition, cardiovascular risk factors, and atherosclerosis in middle‐aged and elderly men for the identification of potential pathogenic links. Research Methods and Procedures: The study included 168 white men 44 to 86 years old. Severity of aortic calcification (AC) was graded on lateral radiographs, and body fat and lean mass were measured by DXA. Information on demographic and lifestyle characteristics also was gathered. Results: A strong and independent inverse association was found between AC and peripheral lean mass (PLM), even after adjusting for age and BMI (p < 0.05). Independently of the influence of PLM, AC was directly correlated with truncal fat mass (p < 0.05). Furthermore, AC was inversely associated with tertiles of the free androgen index (p < 0.05). In a multiple regression model, age and serum cholesterol (p < 0.01) contributed directly, and truncal fat mass tended also to contribute directly (p = 0.09), whereas PLM contributed borderline inversely (p = 0.06) to the variation of AC (R = 0.635, p < 0.0001). Discussion: Severity of AC is strongly dependent on age and further modulated by an array of traditional cardiovascular risk factors. Sarcopenia and truncal fat mass are reciprocal correlates of atherosclerosis of borderline statistical significance in multivariate models. To clarify whether sarcopenia is an atherogenic risk factor or rather a parallel consequence of low‐grade inflammation also promoting atherogenic trends, further longitudinal studies in larger sample sizes of men and women are needed.     

Purification and characterization of rat pancreatic fatty acid ethyl ester synthase and its structural and functional relationship to pancreatic cholesterol esterase

Formation of fatty acid ethyl esters (FAEEs, catalyzed by FAEE synthase) has been implicated in the pathogenesis of chronic pancreatitis. In previous studies, we demonstrated that FAEE synthase, purified from rat liver microsomes, is identical to rat liver carboxylesterase (pI 6.1), and structurally and functionally different than that from pancreas. In this study, we purified and characterized rat pancreatic microsomal FAEE synthase, and determined its relationship with rat pancreatic cholesterol esterase (ChE). Since most of the serine esterases express p-nitrophenyl acetate (PNPA)-hydrolyzing activity as well as synthetic activity to form fatty acid esters or amides with a wide spectrum of alcohols and amines, respectively, we used PNPA-hydrolyzing activity to monitor the purification of FAEE synthase during various chromatographic purification steps. Synthesizing activity towards FAEEs, fatty acid methyl esters, and fatty acid anilides was measured only in the pooled fractions. At each step of purification (ammonium sulfate saturation, Q Sepharose XL, and heparin-agarose column chromatographies, and high performance liquid chromatography (anion exchange and gel filtration)) synthetic as well as hydrolytic activities copurified. Using ethanol, methanol, or aniline as substrates, the ester or anilide synthesizing activity of the purified protein was found to be 8709, 13000, and 2201 nmol/h/mg protein, respectively. The purified protein displayed a single band with an estimated molecular mass of approximately 68 kD upon SDS-PAGE under reduced denaturing conditions, cross-reacted with antisera against rat pancreatic ChE and showed 100% N-terminal sequence homology of the first 15 amino acids to that of rat pancreatic ChE. These results suggest that the purified protein has broad substrate specificity towards the conjugation of endogenous long chain fatty acids with substrates having hydroxyl and amino groups and is identical to ChE.     

Factors affecting ammonium uptake in streams – an inter-biome perspective

1. The Lotic Intersite Nitrogen eXperiment (LINX) was a coordinated study of the relationships between North American biomes and factors governing ammonium uptake in streams. Our objective was to relate inter‐biome variability of ammonium uptake to physical, chemical and biological processes. 2. Data were collected from 11 streams ranging from arctic to tropical and from desert to rainforest. Measurements at each site included physical, hydraulic and chemical characteristics, biological parameters, whole‐stream metabolism and ammonium uptake. Ammonium uptake was measured by injection of ¹⁵N‐ammonium and downstream measurements of ¹⁵N‐ammonium concentration. 3. We found no general, statistically significant relationships that explained the variability in ammonium uptake among sites. However, this approach does not account for the multiple mechanisms of ammonium uptake in streams. When we estimated biological demand for inorganic nitrogen based on our measurements of in‐stream metabolism, we found good correspondence between calculated nitrogen demand and measured assimilative nitrogen uptake. 4. Nitrogen uptake varied little among sites, reflecting metabolic compensation in streams in a variety of distinctly different biomes (autotrophic production is high where allochthonous inputs are relatively low and vice versa). 5. Both autotrophic and heterotrophic metabolism require nitrogen and these biotic processes dominate inorganic nitrogen retention in streams. Factors that affect the relative balance of autotrophic and heterotrophic metabolism indirectly control inorganic nitrogen uptake.     

Serum placental protein 14: a novel marker of selective oestrogen receptor modulator action on the postmenopausal endometrium.

The primary objective of this study was to investigate whether changes in the serum level of an endometrial secretory protein, placental protein 14 (PP14), can reflect endometrial adverse events induced by selective oestrogen receptor modulators (SERMs). A randomized, double-blind, placebo-controlled trial was used. Participants were healthy postmenopausal women aged 45-65 years, who received either various doses of raloxifene (30, 60 or 150 mg day-1) or levormeloxifene (1.25, 5, 10 or 20 mg day-1) or placebo for 12 months. Serum PP14 and endometrial thickness (ET) were monitored by radio-immunoassay and transvaginal ultrasonography, respectively. In the levormeloxifene trial, endometrial status at 12 months was assessed by hysteroscopy. Raloxifene induced only slight increases in serum PP14 and ET. Levormeloxifene, however, induced marked increases in both study parameters at all the does tested. The 6 month changes in PP14 showed a positive correlation with both the 6 and 12 month changes in ET (P < 0.001). Marked stromal oedema, pseudocysticity with or without hypervascularity and endometrial proliferation were seen on hysteroscopy in those showing the largest increases in serum PP14. These results suggest that the PP14 assay used on a group basis may provide useful information on the endometrial effects of SERMs administered in a given dose range, and thereby could assist future clinical trials aiming to find the optimal dose range of new SERMs.     

Epitope Mapping of Form-Specific and Nonspecific Antibodies to Acetylcholinesterase

We have mapped the epitopes to which two monoclonal antibodies against acetylcholinesterase (AChE) from Torpedo californica are directed. One antibody, 2C9, has equivalent affinity for both the 5.6S (amphiphilic) and 11S (hydrophilic) enzyme forms; the other, 4E7, recognizes only the amphiphilic form and has been shown previously to require an N-linked oligosaccharide residue on the protein. Isolation of cyanogen bromide peptides from the amphiphilic form and assay by a competition ELISA for 2C9 and by a direct binding ELISA for 4E7 identified the same peptide, residues 44–82, as containing epitopes against both antibodies. The epitope for 4E7 includes the oligosaccharide conjugated to Asp⁵⁹, an N-linked glycosylation site not present in mouse AChE. A 20-amino-acid synthetic peptide, RFRRPEPKKPWSQVWNASTY, representing residues 44–63, was synthesized and found to inhibit completely 2C9 binding to 5.6S enzyme at molar concentrations comparable to those of the cyanogen bromide peptide. It was unreactive with 4E7. Fractionation of the synthetic peptide further localized the 2C9 epitope. Peptides RFRRPEPKKPW and KPWSGVWNASTY both reacted but less so than the entire synthetic peptide at equivalent molar concentrations, whereas the peptide RPEPKKPWSGVWNASTY was as effective as the larger synthetic peptide. The crystal structure of AChE shows the peptide to be on the surface of the molecule as part of a convex hairpin loop starting before the first α-helix.     

The effect of healing time on the proximodistal organization of double-half forelimb regenerates in the axolotl,Ambystoma mexicanum

The effect of healing on the proximodistal organization of regenerates from double-half forelimbs was studied. Double-anterior and double-posterior upper forelimbs were prepared surgically and amputated at 5, 10, 15, 20, 30, and 60 days after grafting. All experimental groups regenerated hypomorphic skeletal patterns. Double-half forelimbs amputated at Days 5 and 10 regenerated more distally complete skeletal patterns than did limbs amputated at Days 30 and 60. The mean numbers of skeletal elements regenerated were seen to decrease as a function of time after grafting, with the maximal suppression of skeletal patterns observed to occur when limbs were amputated 30 days following grafting. There was no appreciable difference between limbs amputated at Days 30 and 60. These results suggest that healing time has a profound effect on the proximodistal organization of limbs regenerated from double-half forelimb stumps.     

Morphogenetic interactions occurring between blastemas and stumps after exchanging blastemas between normal and double-half forelimbs in the axolotl, Ambystoma mexicanum.

Regeneration blastemas were exchanged between surgically constructed forelimbs comprised of symmetrical tissues (double-anterior and double-posterior) and normal, unoperated forelimbs. Normal blastemas grafted at the stage of medium bud (MB) onto double-half forelimb stumps regenerated normal skeletal patterns in nearly all cases. Double-half blastemas transplanted at the stage of MB onto normal forelimb stumps did not regenerate complete limb patterns. These results indicate that a double-half blastema cannot be “rescued” by transplantation to a normal stump and that a double-half limb stump does not interfere with the ability of a normal blastema to distally transform. The regeneration blastema possesses sufficient positional information at the stage of MB to permit it to develop autonomously. Supernumerary forelimbs resulted from several types of graft-stump combinations. The location and handedness of these supernumerary limbs are predicted by the rules of a recently presented model for pattern regulation in epimorphic fields [French, V., Bryant, P. J., and Bryant, S. V. (1976). Science193, 969–981].     

Persistent neural activity: prevalence and mechanisms

Persistent neural activity refers to a sustained change in action potential discharge that long outlasts a stimulus. It is found in a diverse set of brain regions and organisms and several in vitro systems, suggesting that it can be considered a universal form of circuit dynamics that can be used as a mechanism for short-term storage and accumulation of sensory or motor information. Both single cell and network mechanisms are likely to co-operate in generating persistent activity in many brain areas.