Moderate G6PD deficiency increases mutation rates in the brain of mice

Mice that harbored the x-ray-induced low efficiency allele of the major X-linked isozyme of glucose-6-phospate dehydrogenase (G6PD), Gpdx(a-m2Neu), and, in addition, harbored the transgenic shuttle vector for the determination of mutagenesis in vivo, pUR288, were employed to further our understanding of the interdependence of general metabolism, oxidative stress control, and somatic mutagenesis. The Gpdx(a-m2Neu) mutation conferred moderate G6PD deficiency in hemizygous males (Gpdx(a-m2Neu/y)) displaying residual enzyme activities of 27% in red blood cells and 13% in brain (compared to wild-type controls, Gpdx(a/y) males). In spite of this mild phenotype, the brains of G6PD-deficient males exhibited a significant distortion of redox control ( approximately 3-fold decrease in the ratio of reduced glutathione to oxidized glutathione), a considerable accumulation of promutagenic etheno DNA adducts ( approximately 13-fold increase in ethenodeoxyadenosine and approximately 5-fold increase in ethenodeoxycytidine), and a substantial elevation of somatic mutation rates ( approximately 3-fold increase in mutant frequencies in lacZ, the target and reporter gene of mutagenesis in the shuttle vector, pUR288). The mutation pattern in the brain was dominated by illegitimate genetic recombinations, a presumed hallmark of oxidative mutagenesis. These findings suggested a critical function for G6PD in limiting oxidative mutagenesis in the mouse brain.     

Kinetics and mechanism of long-chain fatty acid transport into phosphatidylcholine vesicles from various donor systems

The kinetics of long-chain fatty acid (FA) transfer from three different donor systems to unilamellar egg phosphatidylcholine (EPC) vesicles containing the pH-sensitive fluorophore pyranine in the vesicle cavity were determined. The transfer of long-chain FA from three FA donors, FA vesicles, unilamellar EPC vesicles containing FA, and bovine serum albumin-FA complexes to pyranine-containing EPC vesicles is a true first-order process, indicating that the FA transfer proceeds through the aqueous phase and not through collisional contacts between the donor and acceptor. A collisional mechanism would be at least bimolecular, giving rise to second-order kinetics. Evidence from stopped-flow fluorescence spectroscopy using the pyranine assay (as developed by Kamp, F., and Hamilton, J. A. (1992) Proc. Natl. Acad. Sci. U.S.A. 89, 11367-11370) shows that the transverse or flip-flop motion of long-chain FA (from 14 to 22 C atoms) is immeasurably fast in both small and large unilamellar EPC vesicles and characterized by half-times t(1/2) < 5 ms. The rate-limiting step of FA transfer from these different donor systems to pyranine-containing EPC vesicles is the dissociation or desorption of the FA molecule from the donor. The desorption of the FA molecule is chain-length-dependent, confirming published data (Zhang et al. (1996) Biochemistry 35, 16055-16060): the first-order rate constant k(1) decreases by a factor of about 10 with elongation of the FA chain by two CH(2) groups. Similar rates of desorption are observed for the transfer of oleic acid from the three donors to pyranine-containing EPC vesicles with rate constants k(1) ranging from 0.4 to 1.3 s(-1). We also show that osmotically stressed, pyranine-containing EPC vesicles can give rise to artifacts. In the presence of a chemical potential gradient across the lipid bilayer of these vesicles, fast kinetic processes are observed with stopped-flow fluorescence spectroscopy which are probably due to electrostatic and/or osmotic effects.ne     

Regulated phase transitions of bacterial chromatin: a non-enzymatic pathway for generic DNA protection

The enhanced stress resistance exhibited by starved bacteria represents a central facet of virulence, since nutrient depletion is regularly encountered by pathogens in their natural in vivo and ex vivo environments. Here we explore the notion that the regular stress responses, which are mediated by enzymatically catalyzed chemical transactions and promote endurance during the logarithmic growth phase, can no longer be effectively induced during starvation. We show that survival of bacteria in nutrient-depleted habitats is promoted by a novel strategy: finely tuned and fully reversible intracellular phase transitions. These non-enzymatic transactions, detected and studied in bacteria as well as in defined in vitro systems, result in DNA sequestration and generic protection within tightly packed and highly ordered assemblies. Since this physical mode of defense is uniquely independent of enzymatic activity or de novo protein synthesis, and consequently does not require energy consumption, it promotes virulence by enabling long-term bacterial endurance and enhancing antibiotic resistance in adverse habitats.     

Effect of continuous nutrient enrichment on microalgae colonizing hard substrates

In order to understand the effect of changing nutrient conditions on benthic microalgae on hard substrates, in-situ experiments with artificial substrates were conducted in Kiel Fjord, Western Baltic Sea. As an extension of previous investigations, we used artificial substrates without silicate and thus were able to supply nutrient media with different Si:N ratios to porous substrates, from where they trickled out continuously. The biofilm developing on these substrates showed a significant increase in biovolume due to N + P enrichment, while Si alone had only minor effects. The stoichiometric composition of the biomass indicated nitrogen limitation during most of the year. The C:N ratios were lowered by the N + P addition. The algae were dominated by diatoms in most cases, but rhodophytes and chlorophytes also became important. The nutrient treatment affected the taxonomic composition mostly at the species level. The significance of the results with regard to coastal eutrophication is discussed.     

Literature on archaeological remains of cultivated plants (1998/1999)

Publications on archaeological remains of cultivated plants have been collected, mainly from 1998, with some earlier ones, and some from 1999, A list is given of the finds according to taxon, country, site, and age     

Mechanisms of Hearing Loss after Blast Injury to the Ear

Given the frequent use of improvised explosive devices (IEDs) around the world, the study of traumatic blast injuries is of increasing interest. The ear is the most common organ affected by blast injury because it is the body’s most sensitive pressure transducer. We fabricated a blast chamber to re-create blast profiles similar to that of IEDs and used it to develop a reproducible mouse model to study blast-induced hearing loss. The tympanic membrane was perforated in all mice after blast exposure and found to heal spontaneously. Micro-computed tomography demonstrated no evidence for middle ear or otic capsule injuries; however, the healed tympanic membrane was thickened. Auditory brainstem response and distortion product otoacoustic emission threshold shifts were found to be correlated with blast intensity. As well, these threshold shifts were larger than those found in control mice that underwent surgical perforation of their tympanic membranes, indicating cochlear trauma. Histological studies one week and three months after the blast demonstrated no disruption or damage to the intra-cochlear membranes. However, there was loss of outer hair cells (OHCs) within the basal turn of the cochlea and decreased spiral ganglion neurons (SGNs) and afferent nerve synapses. Using our mouse model that recapitulates human IED exposure, our results identify that the mechanisms underlying blast-induced hearing loss does not include gross membranous rupture as is commonly believed. Instead, there is both OHC and SGN loss that produce auditory dysfunction.