Computed tomographic measurement of cortical bone geometry

In this study digital images of bone cross-sections obtained by computed tomography were analyzed with an automated outlining method. It was shown that unbiased cross-sectional geometric measurements of cortical bone could be obtained if the periosteal and endosteal surfaces were defined at separate thresholds. Use of different threshold levels for these two surfaces resulted in errors of 2.6% for periosteal diameters, 7.4% for endosteal diameters and 7.3% for cortical area. If incorrect thresholds were used, cortical thickness measurements can have errors as high as 30%. In addition, simulated variation in medullary fat content did not affect measurement of medullary dimensions.     

Inhibition of cholesterol esterases by Δ1-tetrahydrocannabinol

Δ¹-Tetrahydrocannabinol was found to inhibit the action of esterases derived from rat adrenal and luteinized ovary on exogenous cholesteryl palmitate. The drug was effective at a dose of 3.2μM causing greater than 30% inhibition; at 16μM almost complete inhibition occured. These findings are similar to those we have recently reported with mouse Leydig cells (1) showing that this is an effect common to steroidogenic tissues and raising the possibility that a variety of endocrine effects of this drug may be due to direct action on these tissues.     

Apoplastic extracts from a transgenic wheat line exhibiting lesion-mimic phenotype have multiple pathogenesis-related proteins that are antifungal

A transgenic wheat line constitutively expressing genes encoding a class IV acidic chitinase and an acidic beta-1,3-glucanase, showed significant delay in spread of Fusarium head blight (scab) disease under greenhouse conditions. In an earlier work, we observed a lesion-mimic phenotype in this transgenic line when homozygous for transgene loci. Apoplastic fluid (AF) extracted from the lesion-mimic plants had pathogenesis-related (PR) proteins belonging to families of beta-1,3-glucanases, chitinases, and thaumatin-like proteins (TLPs). AF had growth inhibitory activity against certain fungal pathogens, including Fusarium graminearum and Gaeumannomyces graminis var. tritici. Through a two-step ion-exchange chromatography protocol, we recovered many PR proteins and a few uncharacterized proteins. Three individual protein bands corresponding to a TLP (molecular mass, 16 kDa) and two beta-1,3-glucanases (molecular mass, 32 kDa each) were purified and identified by tandem mass spectrometry. We measured the in vitro antifungal activity of the three purified enzymes and a barley class II chitinase (purified earlier in our laboratory) in microtiter plate assays with macroconidia or conidiophores of F. graminearum and Pyrenophora tritici-repentis. Mixtures of proteins revealed synergistic or additive inhibitory activity against F. graminearum and P. tritici-repentis hyphae. The concentrations of PR proteins at which these effects were observed are likely to be those reached in AF of cells exhibiting a hypersensitive response. Our results suggest that apoplastic PR proteins are antifungal and their antimicrobial potency is dependent on concentrations and combinations that are effectively reached in plants following microbial attack.     

Metabolomics reveals novel pathways and differential mechanistic and elicitor-specific responses in phenylpropanoid and isoflavonoid biosynthesis in Medicago truncatula cell cultures

High-performance liquid chromatography coupled to ultraviolet photodiode array detection and ion-trap mass spectrometry was used to analyze the intra- and extracellular secondary product metabolome of Medicago truncatula cell suspension cultures responding to yeast elicitor (YE) or methyl jasmonate (MeJA). Data analysis revealed three phases of intracellular response to YE: a transient response in mainly (iso)flavonoid metabolites such as formononetin and biochanin-A that peaked at 12 to 18 h following elicitation and then declined; a sustained response through 48 h for compounds such as medicarpin and daidzin; and a lesser delayed and protracted response starting at 24 h postelicitation, e.g. genistein diglucoside. In contrast, most compounds excreted to the culture medium reached maximum levels at 6 to 12 h postelicitation and returned to basal levels by 24 h. The response to MeJA differed significantly from that to YE. Although both resulted in accumulation of the phytoalexin medicarpin, coordinated increases in isoflavonoid precursors were only observed for YE and not MeJA-treated cells. However, MeJA treatment resulted in a correlated decline in isoflavone glucosides, and did not induce the secretion of metabolites into the culture medium. Three novel methylated isoflavones, 7-hydroxy-6,4'-dimethoxyisoflavone (afrormosin), 6-hydroxy-7,4'-dimethoxyisoflavone (alfalone), and 5,7-dihydroxy-4',6-dimethoxy isoflavone (irisolidone), were induced by YE, and labeling studies indicated that the first two were derived from formononetin. Our results highlight the metabolic flexibility within the isoflavonoid pathway, suggest new pathways for complex isoflavonoid metabolism, and indicate differential mechanisms for medicarpin biosynthesis depending on the nature of elicitation.