Endogenous phosphorylation of soluble enzymes in human red cells. Cyclic 3',5'-AMP-dependent phosphorylation of phosphofructokinase without detectable regulatory effect

ATP-depleted human red cells have been incubated in a glucose-containing medium with [32P]orthophosphate in the presence and in the absence of cyclic 3',5'-AMP and dibutyril cyclic 3',5'-AMP. Spectrin, pyruvate kinase, phosphofructokinase, glucose-6-phosphate dehydrogenase and hemoglobin A1 have been purified and analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Protein-bound radioactivity has been measured from the sodium dodecyl sulfate polyacrylamide gels and the trichloroacetic acid-precipitated proteins. In the cytosol, the most intense phosphorylation was found for pyruvate kinase whose, in the presence of cyclic AMP, specific radioactivity was comparable to that of the membrane protein and spectrin. In the absence of cyclic nucleotides it was five times less phosphorylated. Phosphofructokinase was only phosphorylated when the red cells were incubated with cyclic nucleotides; the extent of phosphorylation was four times less than for pyruvate kinase. Hemoglobin, glucose-6-phosphate dehydrogenase and a contaminant protein copurified with phosphofructokinase were not phosphorylated: the 'background' of the radioactivity found for these proteins was 100 times less than for pyruvate kinase and spectrin, and 20 times less than for phosphofructokinase (+cyclic AMP).     

Comparative Cost-Effectiveness of Drugs in Early versus Late Stages of Cancer; Review of the Literature and a Case Study in Breast Cancer

Background

Many oncological drugs that are being used in the adjuvant setting were first submitted for reimbursement in the metastatic stage, with differences in incremental cost-effectiveness ratios (ICERs) in both settings having potential implications for reimbursement and pricing. The aim of this study is to identify a possible trend in the cost-effectiveness for the early/adjuvant and late/metastatic stages of oncological drugs through review and case study.

Methods

We reviewed pairs of cost-effectiveness analyses of the same oncological drug in different stages for Scotland and the Netherlands. The case study in this report was directed at trastuzumab in the Dutch situation. Using a simplified Markov model, the cost-effectiveness in early and late stage of breast cancer was calculated and compared to the findings from the review.

Results

Comparable studies were found for cetuximab, bortezomib and bosutinib. Treatments in the late stage were found to be more expensive per QALY by a factor ranging from 1.5 to 12. The case study provided a similar result; late stage treatment was more expensive by a factor 10. Using, for example, a threshold of €80,000/QALY, the early stage of cetuximab, bosutinib and trastuzumab are deemed cost-effective, while their compared late stage is lifted over the threshold and potentially considered not cost-effective.

Conclusion

ICERs of oncological drugs used in different stages are more unfavourable in the late stage than in the early stage. Applying a reasonable threshold may result in early stage treatment being deemed cost-effective while late stage potentially not. Authorities should be aware of this when assessing oncological drugs and interpreting the corresponding ICERs, in the situation where oncological drugs are generally most submitted for reimbursement in the late stage initially.     

Fear and stress reactions in two species of duck and their hybrid

Susceptibility to fear and stress has often been studied in birds using selection experiments. Crosses between different species of the same genus have also been used to study the type of transmission observed for "fixed action patterns." In this experiment we used a cross between two species (male muscovy and female pekin ducks) to study the genetic transmission of various characteristics of fear reactions and adrenal activity. Males of the three genetic types (two parental species and the hybrid) were submitted to tonic immobility and fear of man tests. Blood was collected for corticosterone assay before and after physiological stress or injection of 5 microg/kg of immediate synacthen (ACTH agonist). The muscovy duck showed lower levels of fear reactions than the pekin duck in the majority of behavioral tests (six of eight). Corticosterone levels were also nearly always the lowest in muscovy ducks (five of six). Generally speaking, the muscovy duck appeared to be less fearful and less susceptible to stress than the pekin duck. The mule duck was similar to one parent, midway between the two, or showed heterosis for fear of man at 10 weeks of age (more fearful than the two parents). Corticosterone levels were midway between the two parents in the mule duck with only one exception, i.e., where the pekin duck was dominant. The existence of heterosis for fear is surprising, as this character is classically considered to be submitted to stabilizing selection. The transmission of corticosterone levels and other fear characteristics is, however, compatible with stabilizing selection. The results are discussed in terms of adaptability.     

Efficiently Tracking Selection in a Multiparental Population: The Case of Earliness in Wheat

Multiparental populations are innovative tools for fine mapping large numbers of loci. Here we explored the application of a wheat Multiparent Advanced Generation Inter-Cross (MAGIC) population for QTL mapping. This population was created by 12 generations of free recombination among 60 founder lines, following modification of the mating system from strict selfing to strict outcrossing using the ms1b nuclear male sterility gene. Available parents and a subset of 380 SSD lines of the resulting MAGIC population were phenotyped for earliness and genotyped with the 9K i-Select SNP array and additional markers in candidate genes controlling heading date. We demonstrated that 12 generations of strict outcrossing rapidly and drastically reduced linkage disequilibrium to very low levels even at short map distances and also greatly reduced the population structure exhibited among the parents. We developed a Bayesian method, based on allelic frequency, to estimate the contribution of each parent in the evolved population. To detect loci under selection and estimate selective pressure, we also developed a new method comparing shifts in allelic frequency between the initial and the evolved populations due to both selection and genetic drift with expectations under drift only. This evolutionary approach allowed us to identify 26 genomic areas under selection. Using association tests between flowering time and polymorphisms, 6 of these genomic areas appeared to carry flowering time QTL, 1 of which corresponds to Ppd-D1, a major gene involved in the photoperiod sensitivity. Frequency shifts at 4 of 6 areas were consistent with earlier flowering of the evolved population relative to the initial population. The use of this new outcrossing wheat population, mixing numerous initial parental lines through multiple generations of panmixia, is discussed in terms of power to detect genes under selection and association mapping. Furthermore we provide new statistical methods for use in future analyses of multiparental populations.     

Boron and blue light reduce responsiveness of <Emphasis Type="Italic">Arabidopsis</Emphasis> hypocotyls to exogenous auxins

We reported earlier that boron stimulates hypocotyl growth in several Arabidopsis ecotypes but not in the boron-deficient mutant bor1-1. Others have shown that boron influences the metabolism and transport of the plant hormone auxin. We investigated how boron, in interaction with light, influences Arabidopsis hypocotyl growth responses to the exogenous auxins 1-NAA, 2,4-D and IAA. In either light condition, 1-NAA similarly inhibited hypocotyl growth in bor1-1 and the corresponding WT (Col-0), while in both genotypes, boron did not essentially affect the extent of the inhibition. Whatever the light conditions and in the absence of boron, 2,4-D inhibited hypocotyl elongation in WT, while in BL seedlings, high responsiveness to 2,4-D vanished when boron was added to the culture medium. Hypocotyl of bor1-1 seedlings in all boron concentrations tested and grown in the dark or RL responded to the auxin similar to WT plants. In BL, the mutant hypocotyls retained full sensitivity to 2,4-D at 0.1 mM H₃BO₃ but lost that sensitivity by 2 mM. In both genotypes tested, in the dark or RL, IAA inhibited hypocotyl growth. Conversely, IAA stimulated hypocotyl elongation in both genotypes developed in BL at 0.1 mM H₃BO₃. That stimulation disappeared when the boron supply increased to 2 mM. Our results suggest that specifically in BL, boron reduces hypocotyl responsiveness to auxins 2,4-D or IAA via the functional transporter BOR1. Our results lead to a discussion of how BL and BOR1 influence the mechanisms of auxin transport into and out of the cell.     

Playback Interactions with Great Crested Flycatchers,Myiarchus crinitus (Aves,Tyrannidae)

Interactive playback was used to test observational findings that different vocalizations uttered by singing great crested flycatchers, Myiarchus crinitus (Aves, Tyrannidae), each provide distinctive information about behavior. We present the results of these tests and interpret their significance in combination with the observations. The following predictions were confirmed: 1. Few weep or weeuh songs, and no thrr, were uttered by subjects that approached playback; 2. Churr, common during observations of more active and changing behavior, often predominated during subjects' initial approaches and searches for simulated intruders; 3. Churr was succeeded by wit, which had been found during confrontational behavior, and weihp, which came with movement near opponents or playback; and 4. Quick answers to vocalizations of mates, opponents, and playback were with rreet, also uttered during attack behavior. Functionally, birds uttering the unassertive vocalizations (weep, weeuh, and thrr) may have been taking only minimal initiative to interact, and simply advertising their presence and potential responsiveness. With increasing numbers of churr, subjects maintained social contact with mates or opponents or probed for responses from quieter birds. Wit and weihp may strongly provoke opponents to respond. Further escalation involved met, weed, and wi. In contrast, ch-ee was a defensive threat. This species sings with more vocalizations than do other tyrannids we have studied. Its vocalizations correlate with relatively fine distinctions among behavioral categories. Yet the vocalizations, like those of the other species, provide information about different extents of initiative that a singer will show in interacting. Such information could be fundamentally important in shaping and stabilizing social relationships, not just of tyrannids but also of many other kinds of animals that use singing to interact with one another while at a distance.     

Radiation induced peroxidation of polyunsaturated fatty acids: recent results on formation of hydroperoxides

Aqueous solutions of linoleic acid were irradiated in air with gamma-rays of 137Cs. High pressure liquid chromatography (HPLC) was been used to separate and measure the production of hydroperoxides. The results obtained after reverse phase chromatography, associated with a microperoxydase for hydroperoxide detection, indicate the presence of two different hydroperoxides. One type of hydroperoxide was the major product obtained when the initial linoleic concentrations were below the critical micellar concentration (2 mM), and the second type was produced when the concentrations were above 2 mM. A further separation carried out on the second hydroperoxide by direct phase HPLC showed that it contains three compounds, mainly HPODE 9 and 13.