Cyclic phosphinamides and phosphonamides, novel series of potent matrix metalloproteinase inhibitors with antitumour activity

The design, synthesis, and structure–activity relationship (SAR) of a series of novel nonpeptidic cyclic phosphon- and phosphinamide-based hydroxamic acids as inhibitors of matrix metalloproteinases MMP-1, MMP-3, and MMP-9 are presented. Based on modelling studies and X-ray analysis, a model of the binding mode of these novel compounds in the MMP active site was obtained. This model provided a rational explanation for the observed SAR data, which included a systematic study of different S1′ directed substituents, zinc-complexing groups, chirality, and variation of the cyclic phosphon- and phosphinamide rings. The in vivo effect of four compounds in a human fibrosarcoma mouse model (HT1080) was evaluated and compared to that of a reference compound, Prinomastat. Inhibition of tumour growth was observed for all four compounds.     

Damaging effect of the fish pathogen <Emphasis Type="Italic">Aeromonas salmonicida</Emphasis> ssp. <Emphasis Type="Italic">salmonicida</Emphasis> on intestinal enterocytes of Atlantic salmon (<Emphasis Type="Italic">Salmo salar</Emphasis> L.)

In fish, bacterial pathogens can enter the host by one or more of three different routes: (a) skin, (b) gills and (c) gastrointestinal tract. Bacteria can cross the gastrointestinal lining in three different ways. In undamaged tissue, bacteria can translocate by transcellular or paracellular routes. Alternatively, bacteria can damage the intestinal lining with extracellular enzymes or toxins before entering. Using an in vitro (Ussing chamber) model, this paper describes intestinal cell damage in Atlantic salmon (Salmo salar L.) caused by the fish pathogen Aeromonas salmonicida ssp. salmonicida, the causative agent of furunculosis. The in vitro method clearly demonstrated substantial detachment of enterocytes from anterior region of the intestine (foregut) upon exposure to the pathogen. In the hindgut (posterior part of the intestine), little detachment was observed but cellular damage involved microvilli, desmosomes and tight junctions. Based on these findings, we suggest that A. salmonicida may obtain entry to the fish by seriously damaging the intestinal lining. Translocation of bacteria through the foregut (rather than the hindgut) is a more likely infection route for A. salmonicida infections in Atlantic salmon.Financial support from the Commission of the European Communities, quality of Life and Management of Living Resources programme, project Q5RT-2000-31656 Gastrointestinal Functions and Food Intake Regulation in Salmonids: Impact of Dietary vegetable Lipids (GUTINTEGRITY) and from Magnus Bergvalls Stiftelse for KS, is acknowledged.This work does not represent the opinion of the European Community, which is thus not responsible for any use of the data presented.     

A method to monitor replication fork progression in mammalian cells: nucleotide excision repair enhances and homologous recombination delays elongation along damaged DNA

The capacity to rescue stalled replication forks (RFs) is important for the maintenance of cell viability and genome integrity. Here, we have developed a novel method for monitoring RF progression and the influence of DNA lesions on this process. The method is based on the principle that each RF is expected to be associated with a pair of single-stranded ends, which can be analyzed by employing strand separation in alkali. This method was applied to examine the rate of RF progression in Chinese hamster cell lines deficient in ERCC1, which is involved in nucleotide excision repair (NER), or in XRCC3, which participates in homologous recombination repair, following irradiation with ultraviolet (UV) light or exposure to benzo(a)pyrene-7,8-diol-9,10-epoxide (BPDE). The endpoints observed were cell survival, NER activity, formation of double-strand breaks and the rate of RF progression. Subsequently, we attempted to explain our observation that cells deficient in XRCC3 (irs1SF) exhibit enhanced sensitivity to UV radiation and BPDE. irs1SF cells demonstrated a capacity for NER that was comparable with wild-type AA8 cells, but the rate of RF progression was even higher than that for the wild-type AA8 cells. As expected, cells deficient in ERCC1 (UV4) showed no NER activity and were hypersensitive to both UV radiation and BPDE. The observation that cells deficient in NER displayed a pronounced delay in RF progression indicates that NER plays an important role in maintaining fork progression along damaged DNA. The elevated rate of RF progression in XRCC3-deficient cells indicates that this protein is involved in a time-consuming process which resolves stalled RFs.     

Antimycin A treatment decreases respiratory internal rotenone-insensitive NADH oxidation capacity in potato leaves

Background  

The plant respiratory chain contains several energy-dissipating enzymes, these being type II NAD(P)H dehydrogenases and the alternative oxidase, not present in mammals. The physiological functions of type II NAD(P)H dehydrogenases are largely unclear and little is known about their responses to stress. In this investigation, potato plants (Solanum tuberosum L., cv. Desiree) were sprayed with antimycin A, an inhibitor of the cytochrome pathway. Enzyme capacities of NAD(P)H dehydrogenases (EC 1.6.5.3) and the alternative oxidase were then analysed in isolated leaf mitochondria.     

Local Adaptation and Genetics of Acid-Stress Tolerance in the Moor Frog, Rana arvalis

As potential to adapt to environmental stress can be essential for population persistence, knowledge on the genetic architecture of local adaptation is important for conservation genetics. We investigated the relative importance of additive genetic, dominance and maternal effects contributions to acid stress tolerance in two moor frog (Rana arvalis) populations originating from low and neutral pH habitats. Experiments with crosses obtained from artificial matings revealed that embryos from the acid origin population were more tolerant to low pH than embryos from the neutral origin population in embryonic survival rates, but not in terms of developmental stability, developmental and growth rates. Strong maternal effect and small additive genetic contributions to variation were detected in all traits in both populations. In general, dominance contributions to variance in different traits were of similar magnitude to the additive genetic effects, but dominance effects outweighed the additive genetic and maternal effects contributions to early growth in both populations. Furthermore, the expression of additive genetic variance was independent of pH treatment, suggesting little additive genetic variation in acid stress tolerance. The results suggest that although local genetic adaptation to acid stress has taken place, the current variation in acid stress tolerance in acidified populations may owe largely to non-genetic effects. However, low but significant heritabilities (h ² 0.07–0.22) in all traits – including viability itself – under a wide range of pH conditions suggests that environmental stress created by low pH is unlikely to lower moor frog populations' ability to respond to selection in the traits studied. Nevertheless, acid conditions could lower populations' ability to respond to selection in the long run through reduction in effective population size.     

Acidic proteome of growing and resting Lactococcus lactis metabolizing maltose

The acidic proteome of Lactococcus lactis grown anaerobically was compared for three different growth conditions: cells growing on maltose, resting cells metabolizing maltose, and cells growing on glucose. In maltose metabolizing cells several proteins were up-regulated compared with glucose metabolizing cells, however only some of the up-regulated proteins had apparent relation to maltose metabolism. Cells growing on maltose produced formate, acetate and ethanol in addition to lactate, whereas resting cells metabolizing maltose and cells growing on glucose produced only lactate. Increased levels of alcohol-acetaldehyde dehydrogenase (ADH) and phosphate acetyltransferase (PTA) in maltose-growing cells compared with glucose-growing cells coincided with formation of mixed acids in maltose-growing cells. The resting cells did not grow due to lack of an amino acid source and fermented maltose with lactate as the sole product, although ADH and PTA were present at high levels. The maltose consumption rate was approximately three times lower in resting cells than in exponentially growing cells. However, the enzyme levels in resting and growing cells metabolizing maltose were similar, which indicates that the difference in product formation in this case is due to regulation at the enzyme level. The levels of 30S ribosomal proteins S1 and S2 increased with increasing growth rate for resting cells metabolizing maltose, maltose-growing cells and glucose-growing cells. A modified form of HPr was synthesized under amino acid starvation. This is suggested to be due to alanine misincorporation for valine, which L. lactis is auxotrophic for. L. lactis conserves the protein profile to a high extent, even after prolonged amino acid starvation, so that the protein expression profile of the bacterium remains almost invariant.     

Searching for Optimal Sensory Signals: Iterative Stimulus Reconstruction in Closed-Loop Experiments

Shaped by evolutionary processes, sensory systems often represent behaviorally relevant stimuli with higher fidelity than other stimuli. The stimulus dependence of neural reliability could therefore provide an important clue in a search for relevant sensory signals. We explore this relation and introduce a novel iterative algorithm that allows one to find stimuli that are reliably represented by the sensory system under study. To assess the quality of a neural representation, we use stimulus reconstruction methods. The algorithm starts with the presentation of an initial stimulus (e.g. white noise). The evoked spike train is recorded and used to reconstruct the stimulus online. Within a closed-loop setup, this reconstruction is then played back to the sensory system. Iterating this procedure, the newly generated stimuli can be better and better reconstructed. We demonstrate the feasibility of this method by applying it to auditory receptor neurons in locusts. Our data show that the optimal stimuli often exhibit pronounced sub-threshold periods that are interrupted by short, yet intense pulses. Similar results are obtained for simple model neurons and suggest that these stimuli are encoded with high reliability by a large class of neurons.     

Rapid fluctuations in atrial fibrillatory electrophysiology detected during controlled respiration

Heart rate during sinus rhythm is modulated through the autonomic nervous system, which generates short-term oscillations. The high-frequency components in these oscillations are associated with respiration, causing sinus arrhythmia, mediated by the parasympathetic nervous system. In this study, we evaluated whether slow, controlled respiration causes cyclic fluctuations in the frequency of the fibrillating atria. Eight patients (four women; median age 63 yr, range 53-68 yr) with chronic atrial fibrillation (AF) and third-degree atrioventricular block treated by permanent pacemaker were studied. ECG was recorded during baseline rest, during 0.125-Hz frequency controlled respiration, and finally during controlled respiration after full vagal blockade. We calculated fibrillatory frequency using frequency analysis of the fibrillatory ECG for overlapping 2.5-s segments; spectral analysis of the resulting frequency trend was performed to determine the spectrum of variations of fibrillatory frequency. Normalized spectral power at respiration frequency increased significantly during controlled respiration from 1.4 (0.76-2.0) (median and range) at baseline to 2.7 (1.2-5.8) (P = 0.01). After vagal blockade, the power at respiration frequency decreased to 1.2 (0.23-2.8) (P = 0.01). Controlled respiration causes cyclic fluctuations in the AF frequency in patients with long-duration AF. This phenomenon seems to be related to parasympathetic modulations of the AF refractory period.     

A metamemory perspective on odor naming and identification

A metacognitive perspective is utilized to elucidate why it is so difficult to name common odors and what characterizes the subjective knowledge people have about their actual odor knowledge. Odor-naming failures are often accompanied by strong feelings of knowing (FOK) or feelings of imminent retrieval of what it is that smells. The paper's two experiments investigate FOK judgements and tip of the tongue (TOT) experiences for odor and person names. The data indicate that our inability to correctly name odors are typically not due to the often proposed uniquely poor association between odors and their proper names, but rather due to failures to identify the odors, that is, failures to know 'what it is'. It was also found that (i) TOT experiences are very unusual for odor names and more so than for person names; (ii) FOK judgements about odor names are significantly less predictive of later retrieval than equivalent judgements about names of persons; (iii) FOK judgements were highly correlated with the familiarity of the cue (odor or picture of famous person), rendering some support for the idea that FOK judgements are based on the perceived familiarity of the cue triggering the FOK; and (iv) the idea that FOK judgements are based on the amount of available information about the sought-for memory (accessibility theory) was also supported.