Attosecond real-time observation of electron tunnelling in atoms.

Atoms exposed to intense light lose one or more electrons and become ions. In strong fields, the process is predicted to occur via tunnelling through the binding potential that is suppressed by the light field near the peaks of its oscillations. Here we report the real-time observation of this most elementary step in strong-field interactions: light-induced electron tunnelling. The process is found to deplete atomic bound states in sharp steps lasting several hundred attoseconds. This suggests a new technique, attosecond tunnelling, for probing short-lived, transient states of atoms or molecules with high temporal resolution. The utility of attosecond tunnelling is demonstrated by capturing multi-electron excitation (shake-up) and relaxation (cascaded Auger decay) processes with subfemtosecond resolution.

Dual modes of aging in Mediterranean fruit fly females.

The life history of medflies is characterized by two physiological modes with different demographic schedules of fertility and survival: a waiting mode in which both mortality and reproduction are low and a reproductive mode in which mortality is very low at the onset of egg laying but accelerates as eggs are laid. Medflies stay in waiting mode when they are fed only sugar. When fed protein, a scarce resource in the wild, medflies switch to reproductive mode. Medflies that switch from waiting to reproductive mode survive longer than medflies kept in either mode exclusively. An understanding of the physiological shift that occurs between the waiting and reproductive modes may yield information about the fundamental processes that determine longevity.

Observation of a train of attosecond pulses from high harmonic generation.

In principle, the temporal beating of superposed high harmonics obtained by focusing a femtosecond laser pulse in a gas jet can produce a train of very short intensity spikes, depending on the relative phases of the harmonics. We present a method to measure such phases through two-photon, two-color photoionization. We found that the harmonics are locked in phase and form a train of 250-attosecond pulses in the time domain. Harmonic generation may be a promising source for attosecond time-resolved measurements.

Indoleamine 2,3-dioxygenase and foxp3 expression in skin rejection of human hand allografts.

Human hand transplantation is complicated by skin rejection. To better define the characteristics of infiltrating cells, biopsies from human hand transplants have been investigated for expression of Foxp3 and indoleamine 2,3-dioxygenase (IDO), a key regulatory enzyme to induce T-lymphocyte unresponsiveness. A total of 104 skin biopsies taken from three bilateral hand transplant recipients over 6 years posttransplant were assessed by hematoxylin-eosin histology (graded 1-4b) and immunohistochemistry for IDO and Foxp3 according to a three-grade classification and correlated with the grade of rejection as well as time after transplantation. Overall, rejection ranged between grades 0 and 4a with an average score of 0.94. IDO was expressed in the endothelium independent of rejection. Upon rejection, IDO staining within the cellular infiltrate was significantly increased. Foxp3 in regulatory T cells was mainly found in samples undergoing severe rejection. Expression of IDO and Foxp3 compared well to each other, although the overall expression of Foxp3 was lower when compared to IDO. An increased expression of IDO as well as Foxp3 during rejection late after transplantation was observed. Characteristics of the cellular infiltrate indicate tolerogenic properties of a proportion of the cells and therefore a tendency toward self-limitation of the alloimmune response during skin rejection after hand transplantation.

Unifying Amplitude and Phase Analysis: A Compositional Data Approach to Functional Multivariate Mixed-Effects Modeling of Mandarin Chinese.

Mandarin Chinese is characterized by being a tonal language; the pitch (or F0) of its utterances carries considerable linguistic information. However, speech samples from different individuals are subject to changes in amplitude and phase, which must be accounted for in any analysis that attempts to provide a linguistically meaningful description of the language. A joint model for amplitude, phase, and duration is presented, which combines elements from functional data analysis, compositional data analysis, and linear mixed effects models. By decomposing functions via a functional principal component analysis, and connecting registration functions to compositional data analysis, a joint multivariate mixed effect model can be formulated, which gives insights into the relationship between the different modes of variation as well as their dependence on linguistic and nonlinguistic covariates. The model is applied to the COSPRO-1 dataset, a comprehensive database of spoken Taiwanese Mandarin, containing approximately 50,000 phonetically diverse sample F0 contours (syllables), and reveals that phonetic information is jointly carried by both amplitude and phase variation. Supplementary materials for this article are available online.

Immunocytochemical localization of alkane-inducible cytochrome P-450 and its NADPH-dependent reductase in the yeast Candida maltosa.

Antibodies directed against cytochrome P-450Cm1 and the NADPH-cytochrome P-450 reductase were used to study the induction and intracellular localization of these components of the alkane monooxygenase system in the yeast Candida maltosa. Transition from glucose to n-hexadecane utilization resulted in an about 100-fold increase of the immunodetectable P-450 form whereas the reductase was only moderately induced by a factor of about 5. P-450 but not the reductase was further increased by oxygen limitation during cultivation on n-hexadecane. Using an immunogold technique on ultrathin cryosections, P-450 was found to be concentrated in the nuclear envelope during the early phase of the induction process. However, after maximal induction, the highest labeling was observed in membranes of the endoplasmic reticulum closely associated with the peroxisomes and the plasma membrane. Double-labeling experiments revealed that P-450 and its reductase were distributed in the same regions of the endoplasmic reticulum.

Comparison of two cytochromes P-450 from Candida maltosa: primary structures, substrate specificities and effects of their expression in Saccharomyces cerevisiae on the proliferation of the endoplasmic reticulum.

cDNAs were cloned, sequenced and expressed which encode two different cytochrome P-450 forms of the alkane-assimilating yeast Candida maltosa, designated as P-450Cm1 and P-450Cm2. The amino acid sequences deduced were about 55% identical. Expression in Saccharomyces cerevisiae resulted in the formation of intact microsomal P-450 systems catalyzing the hydroxylation of n-hexadecane and lauric acid with significantly different substrate preferences. A massive proliferation of the endoplasmic reticulum was observed in the S. cerevisiae cells which produced P-450. Depending on the P-450 form expressed, distinctly organized stacks of paired membranes appeared and occupied considerable areas of the cytoplasm. As shown by immunoelectron microscopy for P-450Cm1, the protein expressed was highly concentrated within these newly formed membrane structures.

Estimating the concentration of beta-carotene required for maximal protection of low-density lipoproteins in women.

The reportedly inconsistent antioxidant protective effect of beta-carotene on plasma LDL may depend on LDL's beta-carotene concentration. We measured carbonyl production by CuSO4-challenged LDL from nine healthy women living at the US Department of Agriculture-Western Human Nutrition Research Center and consuming a natural food diet that provided only 0.14 micromol beta-carotene/d for 120 d. During the first 60 d, four women received a placebo and the remaining five women received too small a supplement (0.93 micromol beta-carotene/d) to increase plasma or LDL beta-carotene; therefore, the data for all nine women during this time were pooled. From days 61 to 120, all subjects received the small supplement. From days 101 to 120 they all received an additional, larger, mixed carotenoid supplement (6.16 micromol beta-carotene/d). Plasma beta-carotene dropped from 0.76 +/- 0.21 micromol/L (x +/- SEM) on day 2 to 0.33 +/- 0.08 on day 60 (P = 0.035) and rose to 1.73 +/- 0.18 (P = 0.001) on day 120. LDL beta-carotene dropped from 1.67 +/- 0.53 micromol/g LDL protein on day 2 to 1.27 +/- 0.28 micromol/g LDL protein on day 60 (P = 0.650) and rose to 10.04 +/- 1.07 micromol/g LDL protein (P = 0.001) on day 120. Plasma lycopene dropped from 0.20 micromol/L on day 2 to 0.02 micromol/L on day 60 and did not increase by day 120. Carbonyl production rose from 24 +/- 6 micromol/g LDL protein on day 2 to 42 +/- 4 micromol/g LDL protein (P = 0.001) on day 60 and dropped to 6 +/- 1 micromol/g LDL protein (P = 0.001) on day 120. LDL seemed fully protected with 9.7 +/- 2.5 micromol beta-carotene/g LDL protein, or 2.3 +/- 1.8 micromol beta-carotene/L plasma.

Pretransplant systemic inflammation and acute rejection after renal transplantation.

BACKGROUND: There are presently no established pre-transplant tests that consistently identify patients who may be at increased risk for acute rejection episodes after renal transplantation. We studied whether pretransplant serum levels of C-reactive protein (CRP), a marker for the presence of systemic inflammation, would predict the occurrence of acute rejection episodes after renal transplantation. METHODS: Pretransplant serum was tested for CRP level in 97 consecutive renal transplant recipients. Time to acute rejection after transplantation was stratified by CRP level and compared using the Kaplan-Meier method. In addition, Cox regression multivariate analysis was performed to assess whether any pretransplant covariates could independently predict the subsequent occurrence of acute rejection episodes. RESULTS: Pretransplant mean CRP levels were higher in patients who subsequently had a rejection episode versus those who had no rejection (22.2+/-2.9 vs. 11.7+/-1.8 microg/ml, respectively, P=0.003). Patients less than the median CRP value had a significantly longer time to rejection compared to those with higher CRP levels (P=0.002). Similarly, patients within the lowest CRP quartile had longer times to rejection when compared with the highest quartile (P=0.006). Cox proportional hazards regression multivariate analysis identified CRP level as the only independent pretransplant risk factor for rejection identified (P=0.044). CONCLUSIONS: Pretransplant systemic inflammation as manifested by elevated serum CRP level independently predicts the risk of acute rejection after renal transplantation and may be useful in stratifying patients at the time of transplantation according to immunological risk. Thus, assessment of pretransplant systemic inflammatory status may be helpful in prospective individualization of immunosuppression therapy after renal transplantation.

Desmosomal-mitochondrial complexes in human nonpigmented ciliary and retinal pigment epithelia.

In the normal human nonpigmented ciliary epithelium structural associations between mitochondria and desmosomes are described electron-microscopically: either a single desmosome or a linear array of several desmosomes joined by filamentous bundles is associated with one or two mitochondria (19.6 +/- 5.4%; n = 29). The frequency of occurrence of these complexes was studied in five different regions of the ciliary body; analysis of covariance revealed a significantly increased number of associations in the ciliary processes. Further, the age dependence of their occurrence was examined in 29 different age classes (15 to 86 years); correlation analysis revealed no correlation between age and number of associations. Similar complexes occur, in addition, in the retinal pigment epithelium, but have not been observed in ciliary pigmented, lens, iris and corneal epithelia. Desmosomal-mitochondrial complexes are considered to be a characteristic feature of basic physiological significance of certain epithelia only. The cytochemical demonstration of calcium in the associated mitochondria provides support for the hypothesis that the mitochondria may serve as buffers for intracellular calcium by controlling the local calcium concentration, thus increasing the stability and functional integrity of desmosomal junctions in secretory or actively transporting epithelia with high endogenous calcium levels.

Reproductive potential predicts longevity of female Mediterranean fruitflies.

Reproduction exacts a price in terms of decreased survival. Our analysis of the interplay between age patterns of fecundity and mortality for individual female medflies (Ceratitis capitata) revealed that individual mortality is associated with the time-dynamics of the egg-laying trajectory. In a sample of 531 medflies, we found that each individual has a characteristic rate of decline in egg laying with age. This defines an individual's rate of reproductive exhaustion. This rate was shown to predict subsequent mortality The larger the remaining reproductive potential, the lower the subsequent mortality An increased mortality risk was seen in flies for which egg production declined rapidly early on, irrespective of the level of egg production. Thus, reproductive potential and lifetime are coupled in such a way that those flies which are able to profit most from an extended life span in terms of increased egg output are indeed likely to live longer.

Mortality oscillations induced by periodic starvation alter sex-mortality differentials in Mediterranean fruit flies.

Sex-specific mortality rates of medflies were monitored in cages containing individuals of both sexes and with food (either sugar-only or full diet) removed every 2nd, 3rd, or 4th day (plus ad libitum controls). The general finding is that periodic starvation led to marked oscillations in raw mortality rates. The specific findings are as follows: (i) female medflies live longer than male medflies when they are subjected to periodic starvation; (ii) male medflies maintained on a full diet experience a catastrophic increase in mortality (40%) on the first day food is removed. This mortality surge was not observed for females on either diet or for males maintained on a sugar-only diet; (ii) life expectancy is inversely related to the amplitude of mortality oscillations caused by food deprivation; and (iv) the large perturbations in mortality at younger ages caused by periodic starvation has little effect on the amplitude of mortality at older ages. In general, our data shed new light on the complexity of the mortality response of medflies to both the type and availability of food and thus provide a complimentary perspective to findings from dietary restriction studies on both vertebrate and invertebrate systems.

Relationship of age patterns of fecundity to mortality, longevity, and lifetime reproduction in a large cohort of Mediterranean fruit fly females.

Daily reproduction was monitored throughout the lives of 1000 individual female Mediterranean fruit flies (Ceratitis capitata). Inasmuch as the average female medfly lived 35.6 days and laid 740 eggs in her lifetime, the overall data set consisted of information on around 740,000 eggs distributed over 35,600 fly-days. Results described include the frequency distributions of eggs/day at young (< or =30 days), middle (31-60 days), and older (61-90 days) ages, the relationship between individual life span and lifetime reproduction, the distribution of deaths for non-egg layers versus egg layers, and density plots for daily egg production relative to both lifetime reproduction and life span. One of the more surprising results was the lack of correlation of the total number of eggs laid by females at younger ages (all ages under 30) and subsequent reproduction and life span. Technical and conceptual implications for analyses of reproductive data on other species and groups are briefly discussed.

Female sensitivity to diet and irradiation treatments underlies sex-mortality differentials in the Mediterranean fruit fly.

Large-scale experiments on medflies that were subjected to sterilizing doses of ionizing radiation (plus intact controls) and maintained on either sugar-only or full, protein-enriched diets revealed that, whereas the mortality trajectories of both intact and irradiated male cohorts maintained on both diets are similar, the mortality patterns of females are highly variable. Mean mortality rates at 35 days in male cohorts ranged from 0.2 to 0.3 but in female cohorts ranged from 0.09 to 0.35, depending on treatment. The study reports three main influences: (a) qualitative differences exist in the sex-mortality response of medflies subjected to dietary manipulations and irradiation; (b) the female mortality response is linked to increased vulnerability due to the nutritional demands of reproduction; and (c) female sensitivity to environmental changes underlies the dynamics of the sex-mortality differential.

Evaluation of prognostic factors in cerebral arteriovenous malformations.

In a retrospective study of 48 patients who underwent elective surgery for cerebral arteriovenous malformations, a statistical analysis of demographic, clinical, and neuroradiological data was undertaken in order to discover the best predictors of operative morbidity. In addition, the predictive value of different clinical grading systems as applied to this series was compared. All patients had a computed tomographic scan and a positive angiogram before surgery. Complete resection was proven angiographically. The univariate Mann-Whitney-Wilcoxon rank sum test, the Fisher exact test, Spearman's rank correlation coefficient analysis, and multivariate logistic regression were used as statistical methods. Duration of surgery, the development, of either new deficits or an increase in the preoperative neurological signs immediately after surgery, and rehabilitation (as measured by the Karnofsky index) were taken as target variables for the difficulty of operation and for postoperative morbidity, respectively. The largest diameter of the nidus of the arteriovenous malformation, eloquence of the adjacent brain, and deep venous drainage showed the most consistent correlation with these target variables. Intracerebral hematoma and other single factors, such as the age of the patient or localization of the arteriovenous malformation did not affect the outcome. The clinical grading scale of Spetzler and Martin provided better prediction of surgical risks than other proposed systems.

Designing refoldable model molecules.

We report a numerical study of the design of lattice heteropolymers that can refold when the properties of only a few monomers are changed. If we assume that the effect of an external agent on a heteropolymer is to alter the interactions between its constituent monomers, our simulations provide a description of a simple allosteric transition. We characterize the free energy surfaces of the initial and the modified chain molecule. We find that there is a region of conformation space where molecules can be made to refold with minimal free energy cost. This region is accessible by thermal fluctuations. The efficiency of a motor based on such an allosteric transition would be enhanced by "borrowing" heat from the environment in the initial stages of the refolding, and "paying back" later. In fact, the power cycle of many real molecular motors does involve such a borrowing activation step.

Statistical tools for the analysis of nutrition effects on the survival of cohorts.

We discuss various methods which can be employed for the comparative analysis of samples of response curves. In the application discussed here, these curves are hazard functions, each generated by the survival data obtained for a cohort of experimental subjects which are fed a specific diet. It is demonstrated how comparisons of the effects of different diets on survival can be carried out by employing statistical techniques for inference on samples of curves. The methods are illustrated with data on the survival of large cohorts of male and female Mediterranean fruit flies under full diet and under protein deprivation. These statistical methods allow one to investigate differences between the samples of hazard functions generated by the four groups defined by combinations of sex and diet.

Statistical models for quantitative bioassay.

We discuss various statistical approaches useful in the analysis of nutritional dose-response data with a continuous response. The emphasis is on the multivariate case with several predictors. The methods which will be discussed can be classified into parametric models, including change-point models, and nonparametric models, which rely on smoothing methods such as weighted local linear fitting. The methods will be illustrated with the analysis of data generated from a folate depletion-repletion bioassay experiment conducted on rats, where the measured growth rate of the rate is the response variable. We also discuss the biological conclusions that can be drawn from applying various statistical methods to this data set.

Time-resolved holography with photoelectrons.

Ionization is the dominant response of atoms and molecules to intense laser fields and is at the basis of several important techniques, such as the generation of attosecond pulses that allow the measurement of electron motion in real time. We present experiments in which metastable xenon atoms were ionized with intense 7-micrometer laser pulses from a free-electron laser. Holographic structures were observed that record underlying electron dynamics on a sublaser-cycle time scale, enabling photoelectron spectroscopy with a time resolution of almost two orders of magnitude higher than the duration of the ionizing pulse.