Analysis of pipecolic acid in biological fluids using capillary gas chromatography with electron-capture detection and [H11]pipecolic acid as internal standard

A sensitive and accurate stable isotope dilution assay was developed for the measurement of pipecolic acid in body fluids using capillary gas chromatography with electron-capture detection. The method utilizes [²H₁₁]pipecolic acid as the internal standard. Sample preparation consisted of derivatization in aqueous solution (pH 11.5) of the amine moiety with methyl chloroformate to the N-methylcarbamate, followed by acidic ethyl acetate extraction at pH ≤ 2 and further derivatization of the carboxyl moiety with pentafluorobenzyl bromide, the excess of which was removed by solid-phase extraction. Control values have been determined in the plasma of at-term infants, age > 1 week (n = 21, mean = 1.36 μM, range = 0.47–3.27 μM). The utility of the method was demonstrated by quantitating pipecolic acid in biological fluids derived from patients with peroxisomal disorders. The method was validated against an established electron-capture negative ion mass fragmentographic technique.     

Interaction of small G proteins with photoexcited rhodopsin

Bovine rod outer segment (ROS) membranes contain in addition to the heterotrimeric G protein transducin, several small GTP-binding proteins (23-27 kDa). Furthermore, these membranes contain two substrate proteins (about 22 and 24 kDa) for botulinum C3 ADP-ribosyltransferase known to ADP-ribosylate small G proteins in any mammalian cell type studied so far. Most interestingly, [32P]ADP-ribosylation of ROS membrane small G proteins by C3 is regulated by light and guanine nucleotides in a manner similar to pertussis toxin-catalyzed [32P]ADP-ribosylation of the alpha-subunit of transducin. These findings suggest that not only the heterotrimeric G protein transducin but also the C3 substrate small G proteins present in ROS membranes interact with photoexcited rhodopsin and thus contribute to its signalling action.     

Neurosecretory cells in the circadian-clock system of the scorpion,Androctonus australis

Summary The somata of the efferent neurosecretory fibers that control the circadian sensitivity rhythm in the median eyes of the scorpion, Androctonus australis, were detected in the brain by retrograde labeling with Lucifer Yellow CH. A total of 20–40 neurons are arranged in two groups displaying a bilaterally symmetrical, marginal position near the circumesophageal connectives. Half the cells in each group send fibers into the ipsilateral optic nerve; the fibers from the other half enter the contralateral optic nerve.     

Human glioma tumour-associated antigens

Conclusion From this brief review it appears that at least three categories of human glioma-associated antigens may exist. The first seems to be restricted and common to gliomas. The second is shared between gliomas, normal adult brain, and fetal brain. The third is present on cells from adult and fetal tissue and on cells from tumours derived from the neural crest. The expression of glioma-associated antigens is highly variable from one tumour, or tumour cell line, to another, and reflects the phenotypic heterogeneity of the glioma group. Moreover, this heterogeneity has been found in different clones of individual glioma cell lines [1]. The fact that gliomas share some antigens with normal brain is of critical importance for immunodiagnosis or immunotherapy. It is evident that active immunotherapy for gliomas should be performed with cultured cells and not with tumour extracts, because such extracts may contain MBP.The exact nature of the various glioma-associated antigens remains to be clearly defined, however. They may belong to a group of surface glycoproteins such as those described by Lloyd et al. [24] for melanoma or more recently by Lubitz et al. [25] for glial cells.     

The overlooked role of a biotin precursor for marine bacteria - desthiobiotin as an escape route for biotin auxotrophy

Biotin (vitamin B₇) is involved in a wide range of essential biochemical reactions and a crucial micronutrient that is vital for many pro- and eukaryotic organisms. The few biotin measurements in the world’s oceans show that availability is subject to strong fluctuations. Numerous marine microorganisms exhibit biotin auxotrophy and therefore rely on supply by other organisms. Desthiobiotin is the primary precursor of biotin and has recently been detected at concentrations similar to biotin in seawater. The last enzymatic reaction in the biotin biosynthetic pathway converts desthiobiotin to biotin via the biotin synthase (BioB). The role of desthiobiotin as a precursor of biotin synthesis in microbial systems, however, is largely unknown. Here we demonstrate experimentally that bacteria can overcome biotin auxotrophy if they retain the bioB gene and desthiobiotin is available. A genomic search of 1068 bacteria predicts that the biotin biosynthetic potential varies greatly among different phylogenetic groups and that 20% encode solely bioB and thus can potentially overcome biotin auxotrophy. Many Actino- and Alphaproteobacteria cannot synthesize biotin de novo, but some possess solely bioB, whereas the vast majority of Gammaproteobacteria and Flavobacteriia exhibit the last four crucial biotin synthesis genes. We detected high intra- and extracellular concentrations of the precursor relative to biotin in the prototrophic bacterium, Vibrio campbellii, with extracellular desthiobiotin reaching up to 1.09 ± 0.15*10⁶ molecules per cell during exponential growth. Our results provide evidence for the ecological role of desthiobiotin as an escape route to overcome biotin auxotrophy for bacteria in the ocean and presumably in other ecosystems.Subject terms: Water microbiology, Ecosystem ecology, Marine microbiology     

Receptor-mediated ADP-ribosylation of a phospholipase C-stimulating G protein

In membranes of myeloid differentiated HL 60 cells, the chemotactic peptide FMLP stimulates phospholipase C via a pertussis toxin-sensitive G protein. FMLP markedly stimulates the cholera toxin-dependent ADP-ribosylation of a 40 kDa protein in these membranes. This effect of FMLP is inhibited by GTP and GTP[S], and is almost completely abolished in membranes of pertussis toxin-pretreated HL 60 cells. Treatment of HL 60 membranes with cholera toxin and NAD markedly inhibits FMLP-stimulated high affinity GTPase. These results suggest that a 40 kDa G protein sensitive to both pertussis and cholera toxin functionally interacts with the formyl peptide receptor of HL 60 cells and, thus, very likely is the G protein that stimulates phospholipase C in this system.     

ASIC1a Deficient Mice Show Unaltered Neurodegeneration in the Subacute MPTP Model of Parkinson Disease

Inflammation contributes to the death of dopaminergic neurons in Parkinson disease and can be accompanied by acidification of extracellular pH, which may activate acid-sensing ion channels (ASIC). Accordingly, amiloride, a non-selective inhibitor of ASIC, was protective in an acute 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) mouse model of Parkinson disease. To complement these findings we determined MPTP toxicity in mice deficient for ASIC1a, the most common ASIC isoform in neurons. MPTP was applied i.p. in doses of 30 mg per kg on five consecutive days. We determined the number of dopaminergic neurons in the substantia nigra, assayed by stereological counting 14 days after the last MPTP injection, the number of Nissl positive neurons in the substantia nigra, and the concentration of catecholamines in the striatum. There was no difference between ASIC1a-deficient mice and wildtype controls. We are therefore not able to confirm that ASIC1a are involved in MPTP toxicity. The difference might relate to the subacute MPTP model we used, which more closely resembles the pathogenesis of Parkinson disease, or to further targets of amiloride.     

High water users can be drought tolerant: using physiological traits for green roof plant selection

Background and aims

Green roofs are often installed to reduce urban stormwater runoff. To optimally achieve this, green roof plants need to use water when available, but reduce transpiration when limited to ensure survival. Succulent species commonly planted on green roofs do not achieve this. Water availability on green roofs is analogous to natural shallow-soil habitats including rock outcrops. We aimed to determine whether granite outcrop species could improve green roof performance by evaluating water use strategies under contrasting water availability.

Methods

Physiological and morphological responses of 12 granite outcrop species with different life-forms (monocots, herbs and shrubs) and a common green roof succulent were compared in well watered (WW) and water deficit (WD) treatments.

Key results

Granite outcrop species showed a variety of water-use strategies. Unlike the green roof succulent all of the granite outcrop species showed plasticity in water use. Monocot and herb species showed high water use under WW but also high water status under WD. This was achieved by large reductions in transpiration under WD. Maintenance of water status was also related to high root mass fraction.

Conclusions

By developing a conceptual model using physiological traits we were able to select species suitable for green roofs. The ideal species for green roofs were high water users which were also drought tolerant.